Journal of Ornamental Plants - Webs3)/J...132 Journal of Ornamental Plants, Volume 5, Number 3: 131-138, September, 2015 INTRODUCTION One of the most important environmental factors

Screening of Eleven Festuca arundinacea Native Populations for NaCl Tolerance in Order toUse in Green Space................................................................................................................131-138Azadeh Mousavi Bazaz, Ali Tehranifar, Mohammad Kafi, Ali Gazanchian and Mahmood Shoor

The Effect of Magnetic Water and Irrigation Intervals on the Amount of the Nutrient Elementsin Soil and Aerial Parts of Periwinkle (Catharanthus roseus L.).........................................139-149Davood Hashemabadi, Fatemeh Zaredost and Maryam Jadid Solimandarabi

Turf Quality Characteristics of Crested Wheatgrass (Agropyron cristatum (L.) Gaertner.)Specimens Native to Iran......................................................................................................151-157Hassan Bayat, Hossein Nemati, Ali Tehranifar and Ali Gazanchian

Indirect Shoot Regeneration in Anthurium andreanum ‘Clisto’ from Leaf Explant.......159-166Maryam Ajdarbin , Mohsen Kafi, Masoud Mirmasoumi and Pejman Azadi

Exogenous Putrescine Delays Senescence of Lisianthus Cut Flowers..............................167-174 Davoud Ataii, Roohangiz Naderi and Azizollah Khandan-Mirkohi

Spatial Distribution Pattern of Tetranychus urticae Koch (Acari: Tetranychidae) on DifferentRosa Cultivars in Greenhouse Tehran..................................................................................175-182Fatemeh Bidarnamani, Elham Sanatgar and Mehdi Shabanipoor

Pathogenicity of Alternaria Species Isolated from Chamaecyparis lawsonia In Vitro.......183-188Mohammad Reza Safari Motlagh, Fatemeh Ramezani Rad and Shahram Sedaghathoor

Effect of Gibberellic Acid Pulsing and Sucrose Continuous Treatment on Some QualitativeCharacteristics of Cut Rose Flower cv. Velvet......................................................................189-195Zahra Ahmadi and Rahim Naghshiband Hassani

Volume 5, Number 3September 2015

Journal of Ornamental Plants

Scientific-Research


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Screening of Eleven Festuca arundinacea Native Populations for NaCl Tolerance in Order to Use inGreen Space.................................................................................................................................131-138

The Effect of Magnetic Water and Irrigation Intervals on the Amount of the Nutrient Elements in Soiland Aerial Parts of Periwinkle (Catharanthus roseus L.)..........................................................139-149

Turf Quality Characteristics of Crested Wheatgrass (Agropyron cristatum (L.) Gaertner.) SpecimensNative to Iran..............................................................................................................................151-157

Indirect Shoot Regeneration in Anthurium andreanum ‘Clisto’ from Leaf Explant..................159-166

Exogenous Putrescine Delays Senescence of Lisianthus Cut Flowers........................................167-174

Spatial Distribution Pattern of Tetranychus urticae Koch (Acari: Tetranychidae) on Different RosaCultivars in Greenhouse Tehran.................................................................................................175-182

Pathogenicity of Alternaria Species Isolated from Chamaecyparis lawsonia In Vitro...............183-188

Effect of Gibberellic Acid Pulsing and Sucrose Continuous Treatment on Some Qualitative Characteristicsof Cut Rose Flower cv. Velvet....................................................................................................189-195

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Journal of Ornamental Plants, Volume 5, Number 3: 131-138, September, 2015 131

Screening of Eleven Festuca arundinaceaNative Populationsfor NaCl Tolerance in Order to Use in Green Space

Keywords: Growth Parameter, Native Population, Salt tolerance, Tall Fescue.

Azadeh Mousavi Bazaz 1*, Ali Tehranifar 1, Mohammad Kafi 2, Ali Gazanchian 3 and Mahmood Shoor 11 Department of Horticultural Science, Faculty of Agriculture, Ferdowsi University of Mashhad, I.R. Iran2 Agronomy Department, Faculty of Agriculture, Ferdowsi University of Mashhad, I.R. Iran3 Resources Research Center of Khorassan. I.R. Iran

*Corresponding author,s email: [email protected]

Abstract

In the turfgrass industry, the need for salinity tolerant turfgrasses isincreasing because of the increased use of saline and non-potable water.Greenhouse container experiments were conducted to determine the relativesalinity tolerance and growth responses of eleven native populations of tallfescue (Festuca arundinacea Schrub) (TF), including: Semirom, Mashhad,Sanandaj, Yasuj, Yazd Abad, Daran, Kamyaran, Gandoman, Borujen, NasirAbad and Alborz to 8 weeks of salinity stress. Also, commercial TF was usedas control. Four salinity levels of irrigation water (0, 45, 90, 135 mM NaCl)were applied to turfgrasses. Results showed shoot and root dry weight, totalleaf area, leaf length, leaf width and leaf firing percentage was significantlyaffected by salinity for all turfgrasses. The lowest leaf firing percentage at 90and 135 mM, was related to Sanandaj population and commercial TF, andthe highest leaf firing percentage was related to Alborz at 45 mM andGandoman population at 90 and 135 mM. Shoot dry weight, total leaf areaand leaf length of Sanandaj population was less affected by salinity comparedto other populations. Based on data on growth parameters, the salinitytolerance ranking of selected populations was: Sanandaj >Daran >Yasuj>Kamyaran >Nasir Abad >Semirom >Mashhad >Alborz >Yazd Abad >Borujen>Gandoman . These results showed the potential and competetive role of TFnative populations compared to commercial TF.

132 Journal of Ornamental Plants, Volume 5, Number 3: 131-138, September, 2015

INTRODUCTIONOne of the most important environmental factors limiting plant growth is salinity (Bayat et

al., 2013). Worldwide, more than one-third of irrigated land is salinized (Diedhiou et al., 2009;Chen et al., 2009). On the other hand, rapidly expanding population growth is occurring in manyarid regions, where soil and water salinity are problems and there are increased demands on limitedfresh water resources (Marcum, 2006 ).

The United Nations predicts that 2.7 billion people will suffer severe water deficiency by2025 if consumption continues at current rates (Montaigne, 2002). In the turfgrass industry, theincreased use of saline and non-potable water, the development of turfgrass landscapes in arid andseashore regions where saline soil is common, and the use of salt for deicing roadways, the needfor salinity tolerant turfgrasses is so important. (Chen et al., 2009). The major compound con-tributing salinity in soils is sodium chloride (NaCl), and salt-tolerant turfgrasses are required tocope this problem (Uddin et al., 2012). Under soil salinity, high concentration of Na+ competeswith the uptake of other nutrients, especially K+ as a necessary element (Sharbatkhari et al., 2013).The earliest plant response of salt stress is a reduction in leaf surface expansion, followed by ces-sation of expansion as the stress intensifies (Wu et al., 2006). However, little studies have beenreported regarding leaf growth and shoot density. Under salinity stress, shoot biomass is often re-duced. As salinity stress level increases further, root growth is also decreased (Adavi et al., 2006;Alshammary et al., 2004; Marcum and Murdoch, 1994; Marcum, 1999). Other detrimental effectsof salinity on turfgrass growth include: osmotic stress, nutritional disturbances, and ion toxicity.Salt tolerant plants have the ability to decrease these detrimental effects by producing a series ofanatomical, physiological, and morphological adaptations such as an extensive root system andsalt secreting glands on the leaf surface (Alshammary et al., 2004).

Differences in salt tolerance between turfgrasses have been demonstrated in many studies(Horst and Taylor, 1983; Marcum and Murdoch, 1994; Qian et al., 2001; Alshammary et al., 2004;Chen et al., 2009; Uddin et al., 2012). Much research has been conducted on the physiological re-sponses and growth to salinity stress in warm season turfgrasses (Adavi et al., 2006; Alshammaryet al., 2004; Marcum and Murdoch, 1994; Marcum, 1999; Chen et al., 2009; Uddin et al., 2012).But there are some reports on responses cool season turfgrasses to salinity (Alshammary et al.,2004; Simkunas et al., 2007; Dianati Tilaki et al., 2010).

Tall fescue (Festuca arundinacea Schrub.) is an important perennial cool-season grass intemperate regions and it is widely used for both forage and turf purposes (Zhao et al., 2007; Wuet al., 2006; Alshammary et al., 2004). It has a high degree of self in compatibility, which makesconventional breeding quite difficult (Barnes, 1990), including breeding for salt tolerance (Zhaoet al., 2007). On the other hand, tall fescue native populations are reach sources of variation in-cluding salt tolerance, and there is no study about evaluation of salinity on tall fescue native pop-ulations in Iran. The major objective of this study was to determine the relative salt tolerance andgrowth response of native populations of tall fescue to salinity in sand culture system.

MATERIALS AND METHODSIn this experiment seeds of some native populations of tall fescue (TF) (F. arundinacea

Schreb) including: Semirom, Mashhad, Sanandaj, Sanajan, Yasuj, Yazd Abad, Daran, Kamyaran,Gandoman, Borujen, Nasir Abad, Alborz and commercial TF (C. TF) seeds were used. Seeds weregerminated on pots filled with a mix of 1/3 soils: 1/3sand: 1/3 humus (v/v/v). After germination,ten seedling of TF were transplanted into plastic pots (20 cm diameter) in a greenhouse at FerdowsiUniversity of Mashhad and grown for 8 weeks with non-saline irrigation water. Pure sand wasused as the growing media and Hoagland solution was used as nutrient. Grasses were clippedthroughout the experiment to 5 cm.

Irrigation waters of different salinities were prepared by the addition of NaCl to tap water.Saline waters of 45, 90, and 135 mM along with tap water as the control treatment were applied

133Journal of Ornamental Plants, Volume 5, Number 3: 131-138, September, 2015

to TF. To avoid salinity shock, salinity levels were gradually increased by daily increments of 22.5mM. The amount of water applied including 30% excess water as leaching requirement (400 ml).The irrigation waters were applied on every other day basis for a period of 8 weeks.

The experimental treatments were set up by following a factorial experiment based on com-pletely randomized design and each treatment had three replications. During the salinity treatmentperiod, data were collected on leaf firing (in three stage: after 30, 45 days and at the end of salinitystress), shoot (green leaves) and root dry weight, leaf length, leaf width and total leaf area. Leaffiring was measured as the total percentage of chlorotic leaf area in plants at first and second stage,and percentage of chlorotic leaf dry weight in third stage of measuring. At the end of the experi-ment, shoots and roots were harvested separately. Both shoots and roots were washed with deion-ized water, and dried at 70 ◦C for 48 h to determine shoot and root dry weight. Leaf width wasmeasured by calipers. Also, leaf area was measured by area measurement Delta-T device LTD.For relative water content (RWC) excised leaves from each pot (0.2 g) were measured for freshweight, and leaf samples were rehydrated in a water-filled petri dish for 4 h at room temperature.Turgor weight was measured by allowing full rehydration, removing all water from leaf surface,and weighted. Leaf dry weights were recorded after oven drying for one week at 60 ◦C. The leafrelative water content was determined using the following formula (Whetherly, 1950): RWC =(Fresh weight − Dry weight) / (Fully turgid weight − Dry weight) × 100. The experimental datawere analyzed using JMP 8 software. Treatment means were separated by Fisher’s protected LSD.

RESULTSLeaf firing percentage

Interaction of salinity and population had a significant effect on leaf firing. Leaf firing per-centage for all populations increased as salinity increased. After 30 days (Fig. 1. A) of salinitytreatment, reaching ~ 25% for Nasir Abad population, 30% for commercial TF, ~ 30% for Kam-yaran, Yasuj, Semirom and Sanadej population and ~ 35% for remaining population at 45 mM.Gandoman population had greatest leaf firing percentage between all turfgrasses at 90 and 135mM. Among all of populations, Sanandaj population had lowest leaf firing at 90 and 135 mM, re-spectively. Results showed after 45 days use of NaCl, at 45 mM, commercial TF and Sanadaj pop-ulation, at 90 mM, Sanadaj population and at 135 mM, Daran population were least affected bysalinity (Fig. 1. B). On the other hand Alborz population, Alborz and Borujen populations, andBorujen and Gandoman populations were most affected at 45, 90 and 135 mM, respectively. Atthe end of the study at 45 mM, Alborz, at 90 mM, Gandoman population and at 135 mM, Gando-man population had the highest leaf firing percentage (Table 1). Also, the lowest leaf firing per-centage was related to Sanandaj and commercial TF at 90 and 135 mM, and there were nosignificant difference between them (Table 1).

Fig. 1. Leaf firing percentage of Festuca arundinacea populations after 30 (A) and 45 (B) days salinitystress (1. Yazd Abad, 2. Kamyaran, 3. Yasuj, 4. Semirom, 5. Gandoman, 6. Borujen, 7. Mashhad, 8.

Daran, 9. Nasir Abad, 10. Sanandaj, 11. Alborz TF native populations, and 12. Commercial TF). Verticalbars indicating standard error.

Journal of Ornamental Plants, Volume 5, Number 3: 131-138, September, 2015134

Shoot (green leaf) dry weightInteraction effect of salinity and population was significant on shoot dry weight. Shoot dry

weights of turfgrass populations decreased as the level of salinity increased (Table 2). Commercial TFand Sanandaj population were less affected compared to other turfgrasses, and at 45 mM there were30% and 37% decrease, respectively in shoot dry weight compared to control treatment. At 90 mMNaCl, decrease in shoot dry weight reached 63% for commercial TF and 65% for Sanadaj population.Also, at 135 mM NaCl, decrease in shoot dry weight reached 73% for commercial TF and 84% forSanadaj population. Among native populations, after Sanadaj population, Daran population had greatestshoot dry weight at different salinity levels compared to control. As it was shown in Table 2, Gandomanpopulation had lowest shoot dry weight among all TF populations at all different salinity levels.

Root dry weightAs salinity increased from control to 135 mM, root mass decreased (Table 2). At 45 mM

and 90 mM salinity levels, Sanadaj population produced much higher root than other populations.At 45 mM salinity level (Table 2), decrease in root mass reached to 51% and 42% for commercial

TF populations Salinity levels ( mM )

0 45 90 135

Yazd AbadKamyaranYasujSemiromGandomanBorujenMashhadDaranNasir AbadSanandajAlborzC. TF

13 .0 no9.1 o13.0 no5.9 o12.1 no8.7 o11.1 no2.6 o7.2 o4.2 o11.1 no8.2 o

62.4 gh66.3 gh54.0 hij43.1 jk85.1 cde69.5 fg41.6 kl37.8 kl65.5 gh37.5 kl86.1 cde22.0 mn

88.5 a-e88.7 a-e81.0 ef80.1 ef99.7 a88.2 a-e83.1 de58.1 gh94.9 a-d44.5 ijk94.9 a-d37.9 kl

95.6 abc86.8 cde84.0 cde89.9 a-e100 a99.2 ab89.2 a-e69.7 fg87.1 b-e61.8 gh93.6 a-d58.7 hi

The data with the same letter are not significantly different at P<1%.

Table 1. Leaf firing percentage of Festuca arundinacea populations at the end of salinity stress.

Treatments

Salinity levels (mM)

Shoot dry weight (g/pot) Root dry weight (g/pot)

0 45 90 135 0 45 90 135


5.89 fgh6.18 efg6.59 de6.42 ef5.71 gh6.37 ef7.85 c7.10 d6.37 ef8.62 b9.04 ab9.32 a

2.13 lm2.15 lm2.32 kl2.73 jk0.58 p-t1.36 no2.71 jk3.30 i1.40 no5.41 h0.91 op6.51 e

0.31 q-u0.27 q-u0.75 pqr0.67 p-s0.01u0.36 q-u0.80 pq1.67 mn0.20 stu2.98 ij0.29 q-u3.41 i

0.10 tu0.23 r-u0.20 stu0.16 stu0.01 u0.02 u0.12 tu0.68 p-s0.28 q-u1.36 no0.36 q-u2.46 jkl

3.5 ef2.9 hi3.4 efg3.2 fgh4.2 bc3.7 de2.9 hi3.0 ghi4.0 cd4.2 bc4.5 ab4.8 a

1.3 lmn0.97 l-p1.3 lm0.97 l-p1.2 l-o1.7 k0.95 m-p1.3 lmn1.3 l lmn2.0 j1.1 l-o2.9 i

0.69 tuv0.45 uv1.2 l-o0.83 pq0.83 pq0.72 tuv1.2 l-p0.67 tuv0.82 qr1.9 k1.2 l-o2.3 j

0.76 s-v0.58 tuv0.71 tuv0.75 s-v0.80 r-u0.69 tuv0.45 uv0.40 v0.90 nop0.80 r-u0.67 tuv0.85 op


Table 2. Shoot and root dry weight of Festuca arundinacea populations after salinity stress.


TF and Sanadaj population, respectively compared to 0 mM salinity. Also, at 90 mM salinity, de-crease in root mass reached to 53% and 56% for commercial TF and Sanadaj population, respec-tively. At 135 mM, the lowest decrease in root mass belonged to Semirom population and reachedto 77%, and highest decrease for this factor belonged to Semirom, Kamyaran and Daran popula-tions, at 45, 90 and 135, respectively (Table 2).

Leaf areaInteraction effect of salinity and population was significant on leaf area. Leaf area of turf-

grass populations decreased as the level of salinity increased (Table 3), and was significantly de-creased in all populations compared to control treatment (Table 3). Among native populations, indifferent salinity levels, the greatest leaf area was belonged to Sanandaj population. On the otherhand, there were no significant difference between Sanandaj population with commercial TF inall levels of salinity. Also, Gandoman population was more affected by NaCl treatment, and de-crease in leaf area reached 95%, 99%, and 100% at 45, 90, and 135 mM, respectively (Table 3).

Leaf relative water content (RWC)Relative water content (RWC) of all turfgrass populations was significantly influenced by salinity

(Table 3). As salinity increased, RWC decreased. (Table 3). Relative water content significantly decreasedat 90 mM salinity level, except for commercial TF,Sanandaj, Daran, Yasuj and Yazd Abad populationscompared to 45 mM salinity level. At 135 mM salinitylevel, commercial TF, Sanandaj and Kamyaran popu-lations had higher RWC compared to other populations(Table 3).

Leaf widthInteraction effect of salinity and population

was not significant for leaf width. Leaf width of allturfgrass populations was significantly influencedby salinity. Among all populations, the greatest andlowest leaf width was related to Semirom and Gan-doman populations, respectively (Fig. 2).

TF populations Salinity levels (mM)

Leaf relative water content Leaf area

0 45 90 135 0 45 90 135


92 ab94 a93 a93 a89 a-e92 ab92 ab91 abc92 ab93 a93 a90 a-d

82 c-h87 a-f83 b-h77 g-j85 a-g81 e-j80 e-j85 a-g81 d-i88 a-e82 c-i83 b-h

78 f-j83 b-h80 e-j58 lm48 n42 n76 h-k81 e-j63 l88 a-e75 h-k83 b-h

67 kl81 e-j72 jk46 n51 mn44 n75 h-k78 f-j44 n86 a-f73 ijk88 a-e

724 fg1254 ab929 de1004 cd792 ef977 cd1027 cd1205 ab1129 bc1289 a1267 ab1218 ab

161 i-p202 i-l317 hi167 i-o37 m-q116 j-q262 ij441 h117 j-q629 g47 l-q620 g

19 opq50 l-q89 k-q27 n-q0.6 q19 opq24 n-q104 k-q14 opq187 ijk17 opq231 ijk

3 pq15 opq7 pq4 pq0.1 q1 q6 pq42 m-q14 opq122 j-q21 n-q180 i-n


Table 3. Leaf relative water content and leaf area of festuca arundinacea populations at the end of salinity stress.

Fig. 2. Leaf width of Festuca arundinacea popula-tions after 30 day salinity stress (1. Yazd abad, 2.Kamyaran, 3. Yasuj, 4. Semirom, 5. Gandoman, 6.Borujen, 7. Mashhad, 8. Daran, 9. Nasir abad, 10.Sanandaj, 11. Alborz TF native populations, and12. Commercial TF). (The data with the same let-

ter are not significantly different at P<1%).


Leaf lengthInteraction of salinity and population had no significant effect on leaf length. At first stage

of measuring (after 15 day salinity stress) (Fig. 3. A), turfgrasses that had the greatest leaf lengthincluded: Sanandaj and Daran populations, whereas Semirom population had the lowest leaf lenght(Fig. 3. A). All turfgrasses had lower leaf lenght at stage 2 of measuring (after 45 day salinitystress) compared to stage 1 (Fig. 3. B). Gandoman population had the lowest leaf length, whereasturfgrasses that had the greatest leaf lenght included: Sanandaj and Daran populations (Fig. 3.B).

DISCUSSIONGrowth parameters have been reported to be excellent criteria to determine salinity tolerance

among turfgrasses (Alshammary et al., 2004). Based on data on growth parameters (shoot and rootmass, leaf firing, leaf width and length and leaf area) the salinity tolerance ranking of selectedgrasses was: Commercial tall fescue >Sanandaj >Daran >Yasuj >Kamyaran >Nasir Abad >Semi-rom >Mashhad >Alborz >Yazd Abad >Borujen >Gandoman TF native populations.

Our results show that salinity declined root and shoot mass of turfgrasses. Alshammary etal. (2004) reported that yield production of turfgrasses was reduced significantly when the salinitylevel increased from 4 to 12 dS m−1. Dean et al. (1996) found that turf quality and shoot growth ofturfgrasses were reduced as the salinity level increased from 1.1 to 6 dS m−1. Exposure of plantroots to salinity altered the shape, dimensions and volume of cortical and epidermal cells and thuslimit water uptake (Alshammary et al., 2004). Plant biomass production depends on the presenceof carbon products through photosynthesis, and salinity stress can reduced photosynthetic capacityof many plant species, which is associated with stomatal closure, increased mesophyll resistancefor CO2 diffusion, reduced efficiency of Rubisco for carbon fixation, damage to photosyntheticsystems by excessive energy, and structural disorganization (Lee et al., 2004). In addition to re-duced photosynthesis, the negative effect of salinity on plant growth is attributed to the decreasedosmotic potential of the growing medium, specific nutrient ion deficiency, and ion toxicity (Luoet al., 2005).

Leaves are the main organs of plant photosynthesis, and keeping leaf area stable undersalinity stress can reduce the effect on photosynthesis (Chen et al., 2009). In this experiment, com-mercial TF, Sanandaj and Daran TF populations, maintained the higher shoot biomass among 12turfgrasses at high salinity levels, Sanandaj and Daran populations were also the grasses thatshowed the greater quantity in leaf length and leaf area, but all of them decreased in leaf lengthand leaf area compared to control. The leaves of some glycophyte, such as Sorghum bicolor (L.),Zea mays L., Triticum aestivum L., and four turfgrasses (Zoysia matrella L., Zoysia japonicaSteud., Cynodon dactylon L., and Paspalum vaginatum Sw.) often become short in length, narrowin width, light in weight, and reduced in number per shoot under salinity stress (Hu and Schmid-halter, 2007; Neves-Piestun and Bernstein, 2005; Chen et al., 2009).These studies suggest thatsalinity modified growth through its effects on the leaf expansion. Indeed, several authors have

Fig. 3. Leaf length of Festuca arundinacea populations after 15 (A) and 45 (B) dayssalinity stress (1. Yazd abad, 2. Kamyaran, 3. Yasuj, 4. Semirom, 5. Gandoman, 6. Borujen, 7. Mashhad,8. Daran, 9. Nasir abad, 10. Sanandaj, 11. Alborz TF native populations, and 12. Commercial TF). The

data with the same letter are not significantly different at P<1%).


reported the strong implication of leaf expansion in glycophyte as well as in halophyte responseto salinity (Tarchoune et al., 2010). Salt-induced growth reduction might be related to salt osmoticeffects, which affect expansion and cell turgor. Differently, it has reported that salinity affectedinitiation of new leaves without reducing cell expansion (Ben Amor et al., 2005). In this experi-ment, result showed over time leaf firing was increased. It may be explained with the fact that atfirst salinity affects growth with a short-term effect, including osmotic effects and, second long-term effects, including excessive salt uptake, which caused ionic stress, take part afterwards(Munns, 2002). High sodium levels disturb potassium (K+) nutrition and when accumulated incytoplasm it inhibits many enzymes, and such secondary stresses as oxidative stress linked to theproduction of toxic reactive oxygen intermediates (Iqbal et al., 2006). This leads to leaf firingwhich follows a reduction in the available photosynthetic area necessary to maintain growth. (Tar-choune et al., 2010).

CONCLUSIONIn conclusion, according to salinity problem and importance of tall fescue in the world, it

is necessary to find salinity tolerance grasses in this species. Based on the present results, it is con-cluded that commercial tall fescue, Sanandaj and Daran tall fescue populations can be used undersaline condition. These results showed the importance and competitive role of TF native popula-tions compared to commercial seeds. In addition to detailed field experiments are needed to confirmthis conclusion.

ACKNOWLEDGMENTSWe would like to thank Ferdowsi University of Mashhad for the financial support of this project.

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139Journal of Ornamental Plants, Volume 5, Number 3: 139-149, September, 2015

The Effect of Magnetic Water and Irrigation Intervals onthe Amount of the Nutrient Elements in Soil and AerialParts of Periwinkle (Catharanthus roseus L.)

Keywords: Irrigation intervals, Magnetic field, Ornamental plant, Water quality.

Davood Hashemabadi1*, Fatemeh Zaredost 2 and Maryam Jadid Solimandarabi 21 Department of Horticulture, Rasht Branch, Islamic Azad University, Rasht, Iran2 Yanug Researchers and Elite Club, Rasht Branch, Islamic Azad University, Rasht, Iran


Abstract

The periwinkle with the scientific name of Catharanthus roseus isone of the most important ornamental plants of the Apocynaceae family. Inorder to evaluate the effect of different waters on the amount of the nutrientelements in soil and aerial parts of Catharanthus roseus, a factorial experimentbased on completely randomized design was conducted in 3 replications.Experimental treatments were including: type of water (magnetized tapwater, tap water, magnetized well water and well water) and irrigationintervals (2, 4, 6 and 8 days). In this study, the attributes such as display life, plant height, leaf number, the amount of nitrogen, phosphorus and potassiumof the soil and the plant were evaluated. According to results, the maximumdisplay life (42.23 days), plant height (21.71 cm) and leaf number (165.88)were related to the treatment of irrigation with the magnetized tap waterwith2 days interval. The maximum amount of nitrogen of the plant wasrelated to the treatment of irrigation with the magnetized well water +2 daysinterval (0.32 mgl-1). The treatments of irrigation with the non- magnetizedtap and well water with 8 days interval had the maximum amount of nitrogenof the soil. The maximum amount of potassium of the plant with 56.21 mgl-1was obtained in the treatment of irrigation with the magnetized well waterwith 2 days interval. The maximum amount of phosphorus of the plant with39.5 mgl-1and then 38.8 mgl-1were related to the treatments of irrigationwith the magnetized well water + 2 days interval and irrigation with the mag-netized tap water+ 2 days interval.


INTRODUCTIONCatharanthus roseus is a perennial or annual ornamental plant which normally is cultivated

in gardens as an flowering plant. Moreover, this plant is one of the most important pharmaceuticalplants of the Apocynaceae family which is containing more than 400 types of terpenoid indole al-kaloids such as vinblastine and vincristine (Aslam et al., 2010; Faheem et al., 2011; Kalidass etal., 2010; Loyola-Vargas et al., 2007).

An important point in the production and cultivation of ornamental plants is increasingflowering by using non-chemical methods. Nowadays, researches of agricultural sciences are goingtowards impact of non-chemical factors like the ionizing, laser and ultraviolet rays, electric ormagnetic field on the yeild of different plants (Faqenabi et al., 2009; Feizi and Rezvani Moghad-dam, 2011).

One of the harmless technologies which is considered in recent years by the researchers ofthe agricultural science to increase plant yeild and also increase the water productivity through amagnetic field before irrigation (Maheshwari and Grewal, 2009; Lin and Yotvat, 1990; Xiao-Fengand Bo, 2008; Panda et al., 2004).

The magnetic water is a type of water that passes through a constant magnetic field. Irriga-tion water filtration by magnetic field causes positive changes in physical and chemical propertiesof the water such as pH, electrical conductivity, interfacial tension, solubility of salts and minerals,wetting properties and so causes increasing of the water quality (Xiao-Feng and Bo, 2008; Samad-yar et al., 2014). Researchers believe that by passing the water through a magnetic field, the com-plex structure of the water is converted to a simple structure. The force of interfacial tension ofthe water is reduced and freedom of action, fluidity and wetting properties of the water moleculesare increased. Thus, the magnetic water is absorbed by the plant more easily compared with non-magnetic water and causes increasing of the growth and performance of the plan by increasingand improving nutrient absorption and soluble minerals in the soil and the water (Xiao-Feng andBo, 2008; Ran et al., 2009).

Ran et al. (2009) reported that passing the water through a magnetic field increases thenumber of water molecules in the volume unit and increases the ability of water molecules toabsorb nutrients. These researchers believe that irrigation with the magnetic water increases theabsorption of minerals and nutrients by the plants and as a result increases the growth and yeild.

In addition to the water quality, the quantity of water also affects the quantitative andqualitative properties of plants. Tuzel et al. (2001) investigated the effect of irrigation periodsof 1, 2 and 4 times a day on the bag culture of tomato. The obtained results of these researchersshowed that by increasing the irrigation period from 1 to 4 times per day, yeild, number andaverage weight of the fruits were increased. Fakhraei Lahiji et al. (2011) reported that irriga-tion period of 10 days provides required water of the plant better than 15 days interval and asa result, reduces the soil evaporation and by supplying the required water of the plant, im-proves the vegetative and reproductive properties of the Gladiolus' Rose Supreme'. Saliha(2005) investigated the effect of magnetic water on the physical and chemical properties ofthe soil and reported the positive effect of magnetic water on the solubility and leaching ofthe soil's minerals. He said that using the magnetic water is suitable for improving the qualityof irrigation water and soil properties for agricultural purposes. In a research, Nashir (2008)evaluated that using the magnetic water has a positive effect on culturing pea and stated thatit is because of increasing of the solving power of the magnetic water and absorbing more nu-trients from the soil.

Due to the economic importance of ornamental – pharmaceutical plants of Catharanthusroseus (Fig. 1a) and also the necessity of using non-chemical methods in the production of the dif-ferent plants, the purpose of this study is investigating the effect of magnetic water on the growingproperties and nutrients elements of the soil and aerial parts of Catharanthus roseus.


MATERIALS AND METHODSIn order to investigate the effect of magnetic water treatment and irrigation interval on peri-

winkle, the factorial experiment was carried out in a randomized complete design with two factorsincluding the type of water (magnetized tap water, tap water, magnetized well water and wellwater) and irrigation period (2, 4, 6 and 8 days) with three replicates and 16 treatments.

Seeds of periwinkle was purchased from Farid Institute of Tehran. Two seeds were plantedin transplant pots. The bed used in current study is a mixture of garden soil + leaf composts + sand(1: 1: 1) that its physical and chemical properties has been given in Table 1. 45 days after plantingthe seeds, seedlings with 4 to 6 leaf were transferred to larger pots.

Irrigating plants was performed from seed planting to a week after transplanting the seedlingper day with water corresponding to each treatment. Water magnetized device of AQUA made inGermany was used in order to prepare magnetic (Fig. 1). For this purpose, used water was passedthrough the machine before the irrigation and was used immediately. Characteristics of used waterin has been given in Table 2.

One week after transferring seedling, the effect of irrigating period was applied. Irrigationperiod was determined using a digital tensiometer. Plant nutrition was performed with completefertilizer 20-20-20 once every two weeks. In this study, features such as display life, plant height,leaf number, nitrogen, phosphorus and potassium of plant and soil were analyzed. Display lifewas obtained by counting days from the appearance of the first bud until wilting 50% of flowersof the plant. The number of leaves was measured by counting leaves and plant height in cm at theend of display life. To measure nutrients, the soil was sampled by DTPA and plant was sampledwith a mixture of acids (100 ml of sulfuric acid + 6 g of salicylic acid + 18 ml of hydrogen perox-

Fig. 1. a: periwinkle; b: water magnetized device of AQUA

Available K(mg/kg)

Available P(mg/kg)

Total N(%)

EC(dS m-1) pH

Mixture of garden soil + leafcomposts + sand (1: 1: 1)

45.52 24 1.8 0.018 6.69

Table 1. Physicochemical properties of the media used for experimental.

Na+(ppm)

Mg2+(ppm)

Ca2+(ppm)

HCO3-(ppm)

Cl-meq/lit

EC(dS m-1)

pH

Magnetized Tap Water (W1)Tap Water (W2)Magnetized Well Water (W3)Well Water (W4)

55.7164.2853.5762.14

48602040

220240220230

109.8125.543.291.5

42.649.778.195.4

0.5840.6190.6470.656

77.256.97

7

Table 2. Physicochemical properties of the water used for experimental.

a b


ide) and digestion was performed by heating. Then, nitrogen content of the soil and the plant wasmeasured by Kjeldahl method and titration with sulfuric acid and finally, nitrogen content of thesoil and the plant was calculated using the following formula in percent and it was reported:

a-b)×V/W×100/(D.M))×t× 0.56=N%t: The concentration of used acid for titration in mole per liter; a: The volume of consumed

acid for sample in milliliter; b: The volume of acid consumed for control the amount in milliliter;V: The volume of extract resulted from digestion in milliliter; W: The weight of the plant to digestin gram; D. M.: The percentage of dry matter.

The amount of phosphorus of plant and soil samples was measured by spectrophotometrymethod at a wavelength of 470 nm. Finally, the amount of phosphorus was calculated by using stan-dard curve and was reported in mgl-1. Potassium were measured by flame photometery method. Inthis way the number of potassium of prepared sample was read by flame photometer and then theamount of potassium was calculated using the standard curve in mgl-1 and was reported. Data analy-sis was performed using MSTATC software and comparison of data was performed using LSD test.

RESULTSDisplay life

The interaction effect of different levels of water type and irrigation intervals on displaylife of periwinkle was significant at 5% level (Table 3). The mean comparison showed that mag-netic water had a greater impact on increasing display life compared with the non-magnetic water,as the highest display life with 42.23 days was observed in treatment of irrigation with magnetictap water 2days interval and then was 38.73 days in treatment of irrigation with magnetic wellwater for 2 days interval. Minimum display life with7.96 days was related with treatment of irri-gation with tap water for 8 days interval that had no significant difference with the treatment of ir-rigation with tap water for 8 days interval(9.06 days) (Fig.2).

Source of variance

df Plantpottasium

Soilpottasium

Plantphosphorus

Soilphosphorus

Plant nitrogen

Soilnitrogen

Plantheight

Numberof leaf

Displaylife

Water type(W)Irrigation intervals(T)W*TErrorCV (%)

339

326.05

11.82*

111.7**

64.61**4.004.13

316**

693**

6.74*

3.4176.05

270**

629**

133**

4.0010.27

123.25**

1647**

39.82**

0.9642.90

0.1155*0.124*0.312*0.01048.83

0.859**

6.88**

0.338**

0.0023.63

5.83**

13.84**

1.416*

0.4833.76

711*

3456**

1297**

164.8310.36

187**

1453**

8.47*

3.0987.90

Table 3. Analysis of variance (ANOVA) of the effect of different treatments on traits.

**: Significant at α= 1%, *: Significant at α= 5%, ns= Not significant

Fig.2. Effect of different treatments on display life of periwinkle.

Irrigation intervalsIrrigation interval 2 days (T1)Irrigation interval4 days (T2)Irrigation interval 6 days (T3)Irrigation interval8 days (T4)

Water typeMagnetized tap water (W1)

Tap water (W2)Magnetized well water (W3)Magnetized tap water (W4)


The number of leavesThe interaction effect of different levels of water type and irrigation intervals on the number

of leaves was significant at level of 1% (Table 3). According to the results of comparing the meanof data, the number of leaves was decreased with increasing interval of irrigation period. It shouldbe noted that the applied magnetic treatments on water increased the number of leaves comparedwith non-magnetic water. As irrigation treatment of magnetic tap water two days interval with 165.9leaves had the highest number of leaves between the treatments. Irrigation with magnetized wellwater + two days interval with 154.8leaves had the second place among the top treatments (Fig.3).

Plant heightA significant difference was observed between various levels of interaction effect of water

type and irrigation intervals for plant height at level of 5% (Table 3). The results of the mean com-parison of data show that plant height of plants irrigated with magnetized water is higher than ofthose irrigated with normal water, so that irrigation of magnetized tap water 2 days interval with21.71cm increases height compared with not irrigation of magnetized well water +2 days intervalwith 19.93 cm and magnetized tap water (19.83cm) increases height compared with irrigation ofnot magnetized well water2 days interval (18.16cm). The minimum height of plant was observedfor the irrigation of well water + 8 days interval (16.57cm) (Fig.4).

Fig.3. Effect of different treatments on the number of leaves of periwinkle.




Fig.4. Effect of different treatments on plant height of periwinkle.





Soil nitrogenThe ANOVA showed that the effect of different treatments on the amount of soil nitrogen

is statistically significance at 1% level (Table 3).The maximum amount of soil nitrogen is obtainedin irrigation of well water 8 days interval (2.3%) and the irrigation of tap water 8 days interval(2%).The minimum amount of soil nitrogen is related with the irrigation treatment of magnetized wellwater two days interval by 0.44 percent (Fig.5).

Shoot nitrogenThe effect of different treatments on nitrogen percentage of shoot is significant at 5% level

(Table 3). The results of the mean comparison showed that in all treatments as irrigation intervalsincreased from 2 days to8 days, the amount of nitrogen is gradually decreased. Magnetized wellswater 2 days interval with 0.32 % had the maximum percentage of nitrogen and magnetized tapwater 2 days interval with 0.29 %had the maximum percentage of shoot nitrogen. The minimumpercentage of nitrogen of plant was observed for well water 8 days interval (1.0%) and tap water8 and 6 days intervals with 0.10 % and 0.11 %, respectively (Fig.6).

Fig.5. Effect of different treatments on soil nitrogen of periwinkle.




Fig.6. Effect of different treatments on shoot nitrogen of periwinkle.





Soil phosphorusThe effect of different treatments on soil phosphorus was significant at 1 % level (Table 3).

The results of mean comparison show that the minimum amount of soil phosphorus (17.50 mgl-1)and then (18.95 mgl-1) belong to irrigation treatments of magnetized well water and magnetizedtap water 2 days interval, respectively. Irrigation with well water (49.66 mgl-1), tap water (47.33mgl-1), and magnetic well water (47.23 mgl-1) 8 days interval had the maximum amount of soilphosphorus (Fig.7).

Shoot phosphorus The effect of different treatments on shoot phosphorus is significant at1% level (Table 3).

As seen in Fig. 8,in all treatments as irrigation interval is increased, the amount of phosphorus ofshoot is decreased. Irrigation treatments with magnetic well water and magnetic tap water 2 daysinterval with 39.5and 38.8 mg l-1, respectively had the maximum amount of phosphorus of plant.Irrigation with magnetic well water 8 days interval with 9.8 shoot phosphorus and next, irrigationwith tap water and magnetic tap water 8 days interval had the minimum amount of phosphorousof plant that were not significantly different (Fig. 8).

Fig.7. Effect of different treatments on soil phosphorus of periwinkle.




Fig. 8. Effect of different treatments on shoot phosphorus of periwinkle.





Soil potassiumThe interaction between type of water and irrigation intervals on soil potassium was sig-

nificant at 1% level (Table 3). Based on the results of mean comparison obtained from effect ofdifferent treatments on soil potassium, using magnetic water for irrigation, especially in lesser ir-rigation intervals, causes more depletion of the potassium from the soil and the amount of potas-sium in these treatments is lower than irrigation treatments with large interval periods and normalwater. So that the minimum amount of potassium in soil is related with irrigation treatments ofmagnetized tap water 4 and 2 days intervals with 18.68and 19.21mgl-1, respectively. The maximumamount of potassium in soil is related with irrigation treatments of well water and tap water 8 daysinterval with 43.94and 42.36 mgl-1, respectively (Fig.9).

Shoot potassiumThe interaction effect between type of water and irrigation interval on soil potassium of plant was

significant at 1% level (Table 3). Mean comparison of shoot potassium shows that unlike potassium ofsoil, potassium content of plants is decreased with decreasing irrigation intervals, so that the minimumamount of potassium of plant is related with irrigation treatment of every 4 types of water 8 days interval

Fig.9. Effect of different treatments on soil potassium of periwinkle.




Fig.10. Effect of different treatments on shoot potassium of periwinkle.





and tap water 8 days interval with 40.70 mgl-1.Potassium uptake by plants treated with magnetic waterwith smaller intervals was more so that among the all treatments, irrigation treatment of magnetized wellwater 2 days interval with 56.21 mg l-1 had more potassium compared with other treatments (Fig.10).

DISCUSSIONShortage and poor quality of water is the most important limiting factor for plant growth.

Most of the nutrients in the soil are not absorbed by plants because by irrigation with tap wateronly a small amount of nutrients is dissolved in water which is absorbable for plants. However,several reports have shown that water magnetic treatment increases the solubility of water.

In fact, with the induction of electric charge on the water, water ions with opposite chargerepel each other and are absorbed by magnetic field ions with opposite charge, so the circles ofwater molecules are increased, thereby its solubility is increased, more nutrients are dissolved inthis kind of water and are available for plant, and increases plant growth and yield (Bogatin, 1999;Saliha, 2005; Zangane Youse fabadi et al., 2012). As mentioned in the statement of results, mag-netic water increases the solubility of soil nitrogen or in other word it increases the discharge ofnitrogen from the soil, resulting in its accumulation in the plant. In this respect, the same resultsare reported by Lin and Yotvat (1990), Maheshwari and Grewal (2009) and Nashir (2008) thatshows the use of magnetic water causes the discharge of nutrients (nitrogen, phosphorus, and cal-cium) from soil and more absorption by the plant, and increased growth and yeild of the plant andthese results are in accordance with the results of this study.

Researchers studied the effects of magnetic water on minerals in the soil and concludedthat the concentration of nitrogen, potassium, phosphorus and magnesium + calcium in the soil ir-rigated with magnetic water is different from that in the soil irrigated with conventional water.They stated that magnetic water by accelerating the processes of crystallization and sedimentationof nutrient dissolved in the soil, reduces the movement of the mineral toward down and as a resultgreater amount of these elements are adsorbed by plants (Noran et al., 1996). Maheshwari andGrewal (2009) reported that magnetic water with organic compounds by affecting on organic com-pounds causes more solubility and accessible of nutrients for plants and thus improves growth andyeild. Ahmadi (2010) believes that by passing water through a magnetic field, absorption of min-erals, useful salts and elements in soil and water is increased because of more solubility and free-dom of water molecules. Several researchers believe that the accumulation of potassium in theplant by preventing the destruction of cells against active oxygen species, increasing activity ofantioxidant enzymes and increasing water use efficiency, causes the maintenance of the cell turgorand improves plant morphological and physiological features (Nandwal et al., 1998; Zheng et al.,2008; Hu and Schmidhalter, 2005). Grewal and Maheshwari (2011) reported the increase of potas-sium in pea plants treated with magnetic water.

The use of magnetic water causes the discharge of phosphorus from the soil and more ab-sorption of it by the periwinkle plant. Durate Diaz et al. (1997) reported the increase of nutrientsin tomatoes by treating the plants with a magnetic field. Lin and Yotvat (1990) stated that magneticwater increases the absorption of phosphorus and calcium of the soil that by plant and increasesgrowth and yield of the plant. The use of magnetic water for irrigation of celery and pea increasesthe concentration of calcium and phosphorus in the shoot (Maheshwari and Grewal, 2009). Nashir(2008) reports the increase of the solubility of elements in soil and their absorption by the peaplant that is in accordance with our results.

In current study, irrigation with magnetic water compared with non-magnetic water causesto improve the growing traits that this can be attributed to more accessible nutrients for bushes ofperiwinkle plants irrigated with magnetic water. In addition, irrigation periods had a significantimpact on increasing the display life and mentioned traits, so that as the interval of irrigation isincreased, the display life and the number of leaves and plant height is decreased that this trend


was less in magnetic water compared with non-magnetic water.Researchers believe that reducing the electrical conductivity of the magnetic field leads to

breaking of water structure, and by reducing the surface tension of water causes more freedomand mobility of water molecules, and the solubility of available nutrients for plants is increased.Then, it increases the photosynthesis and food production ability of the plant by increasing the up-take of water and nutrients by the roots of the plant, and these factors increase the vegetative andreproductive growth and yield of the plants (Nashir, 2008; Ran et al., 2009; Hozayn and AbdulQados, 2010; Nikbakht et al., 2013). Nashir (2008) found that the magnetic water increases theheight of a pea for 2.67 cm compared to the control. He stated that the reason of this fact is the in-crease of the solving power of magnetic water and provide more water for the plant. Similar resultsfor the lentil are reported by Abdul Qados and Hozayn (2010) that are corresponded with the resultsof this study. Finally, it can be said that the magnetic water with suitable interval for irrigation, bymore and better providing nutrients, provides better conditions for the plant compared with non-magnetic water and causes to maintain and increase the quality and quantity of the periwinkleplant. Therefore, the use of this water for irrigation of periwinkle is recommended.

ACKNOWLEDGMENTThe authors greatly appreciate from the Vice Chancellery of Research of Islamic Azad Uni-

versity, Rasht Branch for their financial support of this study.

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Keywords: Cluster analysis, Correlation, Low-maintenance, Mowing quality, Native grass.

Hassan Bayat 1, Hossein Nemati 1*, Ali Tehranifar 1 and Ali Gazanchian 2

1 Department of Horticultural Science, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad,Iran2 Agricultural and Natural Resources Research Center of Khorassan Razavi, Mashhad, Iran


Abstract

Crested wheatgrass (Agropyron cristatum (L.) Gaertner.) is a potentialsource of low-maintenance turf species for use in areas with less water. Themain goal of this study was to evaluate turf quality characteristics of 24crested wheatgrass specimens collected from different locations of Iran aslow-maintenance turfgrass. The experimental design was a randomizedcomplete block design with 3 replications. The results showed significantvariation among specimens for turf quality and turf quality components andthere were 15 specimens with an average turf quality rating of 6.0 or higher,indicating adequate quality performance. Mowing quality were higher than 6for all specimens and ranging from 6.30 (210 M) to 7.28 (4049). Habitats ofsamples varied in performance for all traits and different geographical regionsmay offer germplasm with high turf quality performance. There were significantpositive correlation between turf quality and mowing quality (r = 71**) andalso turf quality and tillering (r = 57**). Based on the results of clusteranalysis, the specimens were assigned to 3 clusters and the single specimenfrom cluster 2 had the highest ratings for turf quality (7.95), mowing quality(7.28), color (7.27) and leaf texture (7.10). These results indicated that crestedwheatgrass specimens native to Iran could be an important resource for use aslow-maintenance turfgrass.


INTRODUCTIONWater is becoming a more and rarer resource in arid and semi-arid regions in Iran which is

characterized by little rainfall, high solar radiation and high temperatures in summer (Madani,2014). In recent years turf grass has been extensively used in urban landscape and water demandfor the irrigation of turfgrass will also increase substantially. One strategy to mitigate irrigationdemands for turfgrass is to identify species and cultivars that maintain better qualities with lesswater for low-maintenance situations (Feldhake et al., 1983; Clark and Watkins, 2010).

Extensive landscape areas planted with high-maintenance turf could be replaced with low-maintenance turf species or cultivars (Wu and Harivandi, 1988). Low-maintenance turf is a relativeterm describing areas that receive reduced or no inputs of irrigation, fertilizer, herbicides, andmowing and can withstand weed invasion (Meyer, 1989; Dernoeden et al., 1994) and thus helpconserve natural resources and reduce pollutants. Moreover, maintenance costs of turfgrass inmany areas could be reduced by planting low-maintenance turfgrass that need limited irrigationand mowing (Feldhake et al., 1983; Clark and Watkins, 2010).

Crested wheatgrass (Agropyron cristatum) is a potential source of turf germplasm for aridand semi-arid regions. The genus is originated from central Asia, including parts of Iran, Turkey,Afghanistan, Russia, and China (Dewey and Asay, 1975) and includes 150 species, of which morethan 20 species are distributed in different areas in Iran (Mozaffarian, 1996). Crested wheatgrassis a persistent, long-lived perennial and its properties include excellent seeding and well droughtand cold resistance. It withstands weed competition, tolerates insect depredation, establishes easilyand is adapted to a wide variety of soils (Asay and Jensen, 1996; Asay et al., 1999). Althoughcrested wheatgrass is normally bunch-type growth habit. Specimens with varying degrees of rhi-zome development have also been acquired from Iran and Turkey (Hanks et al., 2005).

Several studies have investigated the potential (Robins et al., 2006; Bushman et al., 2007;Robins and Waldron, 2007; Watkins et al., 2011) and breeding (Asay et al., 1999; Hanks et al.,2005) of crested wheatgrass for turf use. Robins and Waldron (2007) reported that crested wheat-grass had a high genetic variation and heritability estimates for turf quality and revealed that crestedwheatgrass might have promise for low maintenance situations. Hanks et al. (2005) identified therapid spring growth and reduced summer turf quality as the most limiting characteristics of crestedwheatgrass for turfgrass.

Development of turf-type crested wheatgrass cultivars may be possible by selection for turfquality traits including color, leaf texture, density and uniformity, as well as, drought toleranceand persistence. Genetic differences among cultivars could be employed by turf breeders as selec-tion process in breeding programs. There was no information about the potential of native Iranianspecimens of crested wheatgrass for use as low-maintenance turf species. The objectives of thisstudy were to (1) evaluate the specimens for turf quality traits under low-maintenance conditionsand (2) identify key geographical locations for future germplasm collection efforts and breedingprograms.

MATERIALS AND METHODSPlant material and experimental design

Seeds of twenty four native Iranian specimens of crested wheatgrass (Agropyron cristatum)(Table 3) were collected from different geographical regions by the Rangelands and Forestry Re-search Institute and the experiment was conducted at research farm of Department of HorticulturalScience in Ferdowsi University Mashhad, Mashhad (36º16N, 59º38E; 989 m latitude), Iran during2013-2014. The meteorological data of the experimental site were shown in Table 1. The grassseeds were sowed directly on 7 October 2013. The experimental design was a randomized completeblock design with 3 replications. Individual plots consisted of 10 plants and spaced 50 cm betweenrows and 30 cm between plants within a row. The experiment soil was a loamy texture, pH= 7.1and 0.9% organic matter. Throughout the study, irrigation occurred weekly from May to September


at 60% evapotranspiration (ET0) replacement. Plots were weeded by hand or using hoes as neededthroughout growing season. Following establishment, plots were mowed twice weekly to 8.0 cm.

MeasurementsTurf characteristics including turfgrass quality, color, summer color, fall color and leaf tex-

ture were rated visually using a 1-9 scale (9 = best turf quality) following the guidelines providedby National Turfgrass Evaluation Program (NTEP) (Turgeon, 1996). A rating of 6.0 or higher in-dicates acceptable performance for traits. A similar scale was utilized to score mowing quality, i.e.the amount of leaf shredding (9 = no shredded leaf tips). Lateral spread (diameter [cm] at 4 cmabove the soil) was measured at the widest expanse of the crown in one direction. Numbers oftillers were also recorded.

Statistical analysisAnalysis of variance (ANOVA) for all the variables was carried out using the JMP8 software

and LSD test at 5% levels was used for mean comparison. Ward’s method based on Euclidean distancegrouped all 24 specimens using measured characters (Ward, 1963) made by the Minitab software.

RESULTS AND DISCUSSIONThe results of analysis of variance showed significant differences (P<0.01) for turfgrass

quality, color, summer and fall color, leaf texture, lateral spread and number of tillers within spec-imens (Table 2). Block was not significant for all of the traits except number of tillers (Table 2).

Mean separation analysis for measured traits was shown in Table 3. Specimens 4049 (7.90)and 210 M (4.67) had the highest and lowest turf quality, respectively. There were 15 specimensthat received an average turf quality rating 6.0 or higher, indicating adequate turf quality perform-ance. Mowing quality were higher than 6 for all specimens and ranging 6.30 (210 M) to 7.28(4049). The highest ratings for leaf color (7.28) and texture (7.10) were obtained from 4049. Thehighest ratings for summer (6.78) and fall (6.37) color were observed from 4049 M and 2854, re-spectively (Table 3).

These results are similar to Cook (2000), Robins et al. (2006), Bushman et al. (2007), Robinsand Waldron (2007) and Watkins et al. (2011) evaluations of turf-type crested wheatgrasses foroverall and seasonal patterns of turf quality and color. The range of variation among specimens in-dicates that there is potential for developing crested wheatgrass specimens for use as low-input turf.The number of individual specimens with adequate turf performance and other traits suggests thatthese species should be the target of germplasm improvement efforts for use as a low-input turf.

Year Month Min. averagetemperature (◦C)

Max. averagetemperature (◦C) Precipitation (mm)

2013

2014

OctoberNovemberDecemberJanuaryFebruary

MarchAprilMayJuneJuly

AugustSeptember

11.34.30.8

- 3.5- 4.09.05.9

14.917.520.919.516.0

27.116.211.87.99.4

15.018.228.932.636.435.631.5

26.43.27.25.02.93.0

35.122.44.30.00.00.0

Table 1. Monthly average temperatures and precipitation at the experimental station duringOctober 2013 to December 2014.


Specimens 619 (47.90 cm), 208 M (47.65 cm), 2854 (47.62 cm) and 4050 (47.10 cm) hadhigher lateral spread among specimens (Table 3). Lateral spread is a measurement of plant vigor andan indicator of the ability to adequately cover the soil surface as a uniform turf. This trait is desirablein turfgrass to fill in gaps of a turf canopy caused by damage or wear. The highest tiller number wasobserved in 3029 (Table 3). Plant density appeared to be influenced by environmental factors and bygenetic factors such growth habit (Clark and Watkins, 2010). The specimens differed substantiallyin tillering density that will be useful in selecting genotypes with the high density ratings.

Means separation analysis on origin basis showed that collection regions differed in per-formance for all traits (Table 4). The single accession from Hamedan (208 MH) performed thehighest turf quality (7.44), mowing quality (7.10) and color (7.19). All collection origins had ac-ceptable turf quality (higher than 6) except Lorestan (5.90), Hamand (4.67) and Golestan (5.47).The lowest turf quality rating was obtained from Hamand (4.67). The finest (6.62) and coarsest(5.58) leaf texture were observed in Kerman and Bojnord, respectively. The lateral spread andtillering were the highest for collections from Arak (47.62 cm) and Bojnord (81.49), respectively.The single accession from Arak (208 M) performed the highest ratings for summer (6.41) and fall

Precipitation(mm)

df Turfquality

Mowingquality

Leaftexture

Color Summercolor

Fallcolor

Lateralspread

Lateralspread

ReplicationSpecimensError

22346

0.05 ns†

2.43**0.14

0.04 ns

0.20*0.10

0.10 ns

0.54**0.08

0.22 ns

0.30**0.10

0.09 ns

0.61**0.04

0.04 ns

1.30**0.31

46.21 ns

135.13**16.78

438.05*565.22**104.91

Table 2. Analysis of variance for trial characteristics of crested wheatgrass specimens.

† *, ** and ns indicate significance at P < 0.05, P < 0.01 levels and non-significance, respectively.

Specimen OriginTurf

qualityMowingquality

Leaf texture

Color Summercolor

Fallcolor

Lateralspread

Numberof tiller

1-9 1-9 1-9 1-9 1-9 1-9 cm n

30291727 P101727 M208 M405628541727 P121550619 S208 P24049208 P13619 M4056 M210 M619619 P13208 MH1727 P11727209 M40504049 M1551

BojnordGolestanGolestanEsfahanChadeganArakGolestanBojnordEsfahanEsfahanKermanEsfahanEsfahanChadeganHamandEsfahanEsfahanHamedanGolestanGolestanEsfahanLorestanKermanBojnordLSD (0.05)

7.346.455.086.697.316.304.736.796.415.737.906.696.566.724.676.586.807.445.905.205.105.906.004.980.61

6.926.376.357.007.206.536.386.896.616.807.286.866.676.876.306.666.777.106.717.006.666.826.676.560.53

5.446.006.766.436.316.136.945.825.826.627.106.026.156.215.635.766.006.006.456.435.816.016.155.480.47

7.237.056.337.047.226.306.487.126.967.007.276.616.966.926.796.846.877.196.546.786.207.006.676.330.55

6.045.634.706.706.386.416.346.455.776.675.666.116.136.005.926.006.106.115.356.216.006.346.786.000.33

5.376.335.675.335.676.375.805.736.175.333.005.805.635.575.376.005.005.336.075.775.536.335.335.670.90

44.5343.5146.4747.6544.0647.6236.3136.7444.7335.8123.7142.2040.3144.6238.0347.9043.4346.3542.2421.6044.6747.1044.0040.676.99

97.1340.4941.5767.8375.7056.7832.7369.8474.5364.8053.8475.2658.3260.3938.3768.6155.2381.2736.0838.2957.3755.2259.6777.5018.36

Table 3. Origin and mean separation analysis of turf quality, mowing quality, leaf texture, color, summercolor, fall color, lateral spread and number of tiller for crested wheatgrass specimens native to Iran.


(6.33) color. Clark and Watkins (2010) revealed that significant differences among collection regionsof prairie junegrass specimens for most traits like turf quality, mowing quality and color that weresimilar to present results. The analysis of collection regions allowed for the identification of futuregermplasm collection sites and planning for future turf breeding programs. Collection of specimensfrom different geographical regions of Iran may offer germplasm with high turf quality performance.

Correlations based on the 24 specimen means were calculated among the turf quality traits(Table 5). As expected, there were generally positive correlations between overall turfgrass qualityand the turfgrass quality components. There were significant positive correlation between turfquality and mowing quality (r = 71**) and between turf quality and tillering (r = 57**). Positivecorrelation was occurred between mowing quality and tillering (r = 0.47*). There were a negativecorrelation between lateral spreading and leaf texture (r = -0.42*) and significant positive corre-lation between lateral spreading and fall color (r= 0.51**). Hanks et al. (2005) reported that sig-

Origin No. specimenTurf


Leaf texture

Color Summercolor

Fallcolor

Lateralspread

Numberof tiller

1-9 1-9 1-9 1-9 1-9 1-9 cm n

BojnordGolestanEsfahanChadeganArakKermanHamandHamedanLorestanLSD (0.05)

358212111

6.375.476.327.026.306.954.677.445.901.21

6.796.566.757.046.536.976.307.106.820.56

5.586.526.086.266.136.625.636.006.010.62

6.896.646.817.076.306.976.797.197.000.65

6.165.646.186.196.416.225.926.116.340.72

5.595.935.605.626.334.175.375.336.281.06

41.1337.7343.3444.3947.6233.8638.0346.3547.1011.61

81.4937.5664.6966.5256.7856.7538.3781.2755.2218.89

Table 4. Means separation analysis of turf quality, mowing quality, leaf texture, color, summer color, fallcolor, lateral spread and number of tiller by origin of crested wheatgrass specimens native to Iran.

Turf quality

Mowingquality

Color Leaftexture

Lateralspread

Summercolor

Numberof tiller

Fallcolor

Turf qualityMowing qualityColorLeaf textureLateral spreadSummer colorNumber of tillerFall color

0.71**†

0.73**

- 0.00 ns

0.10 ns

0.12 ns

0.57**

- 0.37 ns

0.63**

0.18 ns

- 0.27 ns

0.31 ns

0.47*

- 0.50*

0.00 ns

- 0.15 ns

0.22 ns

0.38 ns

- 0.34 ns

- 0.42*

- 0.11 ns

- 0.51*

- 0.38*

- 0.02 ns

0.34 ns

0.51*0.32 ns

0.03 ns - 0.06 ns

Table 5. Correlation coefficients among several turf quality characteristics of crested wheatgrassspecimens native to Iran.

† *, ** and ns indicate significance at P < 0.05, P < 0.01 levels and non-significance, respectively.

ClusterTurf


Leaf texture

Color Summercolor

Fallcolor

Lateralspread

Numberof tiller

1-9 1-9 1-9 1-9 1-9 1-9 cm n

123

6.647.905.38

6.847.286.54

6.057.106.18

6.977.276.53

6.265.665.84

5.613.005.84

43.5323.7140.04

67.8253.8444.81

Table 6. Means of traits used in the identification of three clusters formed from 24 crestedwheatgrass specimens native to Iran.


nificant positive correlation between turf quality and mowing quality in population of CWG-Rcrested wheatgrass in agreement with the present results.

On the basis of the results of cluster analysis, the specimens were assigned to 3 clusters(Fig. 1). Cluster 1, the largest group, included 14 specimens and had the highest lateral spread(43.53 cm), number of tillers (67.82) and summer color (6.26) (Table 6). In cluster 2, containedthe single accession and had the highest ratings for turf quality (7.95), mowing quality (7.28), color(7.27) and leaf texture (7.10). Cluster 3 included 9 specimens and fall color rating (5.84) was thehighest (Table 6). Cluster analysis has been used to group specimens in genetic diversity studiesand examine the relationships among specimens of diverse germplasm (Garcia et al., 1997;Bayuelo-Jimenez et al., 2002; Gazanchian et al., 2006; Oliveira et al., 2008). Cluster 2 had singleaccession (4049) collected from Kerman was superior for turf quality, mowing quality and coloramong all of specimens that could be used in future breeding programs. Genetic diversity of cluster1 could be used for selecting resistance specimens to summer dormancy and reduction of greenness.

CONCLUSIONIn the present study, several superior specimens had higher turf quality ratings than the

overall mean of the specimens, further indicating significant potential improvement from selection.The variability in turf quality traits observed in the specimens supports their use in a breeding pro-gram for the development of low-input turf. The specimens with high mowing quality performancewill be important in germplasm improvement of native plant material which does not tolerate mow-ing. The specimens collected from different geographical regions of Iran may be germplasm withhigh turf quality and mowing quality ratings. It will be necessary to examine the better performingspecimens for turf quality in seeded turf plots.

ACKNOWLEDGMENT This study was supported by the Ferdowsi University of Mashhad, Mashhad, Iran (No.

28735). The authors wish to acknowledge Gene Bank of the Rangelands and Forestry ResearchInstitute of Iran for providing seeds.

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grasses. Agron Monorg. 34. ASA, Madison, WI.

Fig. 1. Ward’s cluster analysis classifications of 24 crested wheatgrassspecimens native to Iran.


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Indirect Shoot Regeneration in Anthurium andreanum‘Clisto’ from Leaf Explant

Keywords: Anthurium, BA, Shoot multiplication, Tissue culture, 2,4-D.

Maryam Ajdarbin 1*, Mohsen Kafi 2, Masoud Mirmasoumi 3 and Pejman Azadi 41 Department of Horticultural Sciences, Karaj Branch, Islamic Azad University, Karaj, Iran2 Department of Horticultural Sciences, Faculty of Agricultural Sciences and Natural Resources, Universityof Tehran, Karaj, Iran3 Department of Plant Physiology, Faculty of Biology, University of Tehran, Tehran, Iran4 Agricultural Biotechnology Research Institute of Iran (ABRII), Mahdasht Road, Karaj, Iran


Abstract

Anthurium is commercially produced as cut flowers and potted plantsand known for its colorful spathes. It is traditionally propagated by stem cuttingand suckers, so micropropagation is an alternative production method. In thisexperiment we carried out shoot regeneration in Anthurium andreanum ‘Clisto’.The leaf explant was used and after sterilization, they cultured on two differentmedia (half-strength MS and Nitsch’s media) which supplemented with differentcompositions of 2,4-D (0, 0.25, 0.5 and 0.75 mg/L) and BA (0, 0.5, 1 and 1.5mg/L). After three months callus size and percentage of explant that producedcallus were measured and callus transferred to MS media with different concen-trations of BA (0, 0.25, 0.5 and 0.75 mg/L). After 6 months, number of shoot,shoot length and leaf number was recorded. The highest callus proliferation wasobtained in Nitsch’ media supplemented with 0.25 mg/L 2,4-D and 0.25 or 0.5mg/L BA. The callus was creamy color and compact. The highest shootregeneration was observed in media with 0.75 mg/L BA. We cloud achieved 31shoot per callus segment in the best treatment. Rooting was easily performed inpeat: perlite (1:2) pots and they well acclimated and transferred to greenhouseand 99% plants were survived.


INTRODUCTIONAnthuriums are known as an exotic ornamental crop. The commercial flower is a combi-

nation of a colorful modified leaf, termed spathe, subtending bisexual flowers carried in a spiralfashion on a spadix (Kuehnle et al., 2001). Anthurium species and hybrids belongs to Araceae fam-ily have an importance in monocotyledonous ornamental plants and they are commercially pro-duced as cut flowers and potted plants in tropical and subtropical countries and also in the globalmarket. Anthurium cultivars with valued flowers are the second beside the Orchids among tropicalcut flowers (Dufour and Guerin, 2008; Gantait and Sinniah, 2011).

Anthurium is conventionally propagated by seeds and, this method is associated with someproblems due to the inherent heterozygosity. Anthurium traditionally is propagated via stem cuttingand suckers and these are they are tedious and not practical when carried out on a large scale (Pu-chooa, 2005).

In recent years, propagation of a wide range of ornamental plants by tissue culture has be-come accepted commercial practice and the rapid strides were achieved in the field of microprop-agation of ornamental plants (Read and Preece, 2009). Ornamental industry has applied immenselyin vitro propagation approach for large-scale plant multiplication of elite superior varieties (Routet al., 2006). Tissue culture offers an alternative tool for rapid multiplication of selected clones ina short period (Bejoy et al., 2008).

The tissue culture of Anthurium was first reported by Pierik et al., (1974) and they achievedregeneration of Anthurium andreanum through adventitious shoots formation from callus.

Many researchers studied on optimization of Anthurium tissue culture. Micropropagationof Anthurium is performed via adventitious shoot regeneration from callus (Kuehnle and Sugii,1991; Beyramizade et al., 2008; Khorrami Raad et al., 2012) and direct shoot regeneration (Martinet al., 2003; Islam et al., 2010). The rapid and efficient tissue culture protocols are important formicropropagation of Anthurium as much as in other plants. The success of plant tissue culture de-pends on the composition of the medium used (Atak and Çelik, 2012). Kuehnle and Sugii (1991)explained callus proliferation in seven cultivars of Hawaiian anthuriums with leaf and petiole ex-plants. They concluded that callus proliferation was best with modified Pierik medium containing0.36 µM 2,4-D and 4.4 µM BA for leaf explant and Pierik modified Pierik, and Finnie and vanStaden media for petiole explant and also for adventitious plantlets, callus was subcultured on aKunisaki medium containing 2.2 or 22 µM BA. Beyramizade et al. (2008) investigated micro-propagation of Anthurium andreanum ‘Tera’. They achieved best result for callus induction onhalf strength MS medium containing 0.08 mg/L 2,4-D and 1 mg/L BA, whereas medium withoutphytohormones induced the highest number of shoots from the callus. Khorrami Raad et al., (2012)reported that callus production in Anthurium andreanum L. different cultivars was best in mediumcontaining 0.5 mg/L NAA + 3 mg/L BA in dark conditions. Then, the best proliferation of shootsper callus (22.83 shoots per cm2 of callus) was observed on medium supplemented with 0.01 mg/LNAA + 1 mg/L BA after 8 weeks in a 16/8 h light and dark cycle under a photoperiod of 50µmol/m2/s. Afterward, in root induction media, the largest number of root (11.50 roots per plantlets)was obtained on medium supplemented with 1 mg/L IBA + 0.2 mg/L KIN.

The aim of this study was to investigate the effects of culture media and different plantgrowth regulators (BA and 2,4-D) and their concentrations on callus induction, and adventitiousshoot formation of Anthurium, approaching an efficient Anthurium micropropagation system.

MATERIALS AND METHODSThe 3-4 year old Anthurium andreanum ‘Clisto’ was commercially selected from Ashianeh

Sabz greenhouse used as donor plant. The research was conducted in Plant Physiology Tissue Cul-ture Laboratory, College of Science, University of Tehran. Young and fresh leaves were taken andused as plant material. Leaf segments were pre-washed in 0.1% commercial dish washing and luke


warm water solution for 5 min and rinsed with running tap water then divided to smaller segmentand were surface sterilized with solution of 10% (v:v) sodium hypochlorite (NaOCl) with twodrops of Tween-20 for 20 min and finally rinsed 3 times in sterile distilled water. The aseptic ex-plant were cut into 1 cm × 1 cm segment and placed on Petri dishes containing callus inductionmedia. The media was half-strength Murashige and Skoog medium (Murashige and Skoog, 1962;MS) or Nitsch‘s medium (Nitsch and Nitsch, 1969) with 30 g/L sucrose and solidified with 7 g/Lagar (Agar-agar 1614, Merck, Darmstadt, Germany) and supplemented with different compositionof 6-Benzyladenine (BA) with concentrations 0, 0.5, 1 and 1.5 mg/L and 2,4-dichlorophenoxyaceticacid (2,4-D) with concentrations 0, 0.25, 0.5 and 0.75 mg/L. The pH of all medium was adjustedto 5.8 prior to autoclaving. The cultures were incubated at 25 ± 1ᵒC and darkness. They were sub-cultutred per 20 days. After 3 months, callus size (observational) and percentage of explant thatproduced callus were measured. Produced callus were divided to same size segment and transferredto shoot media regeneration. This media was MS that supplemented with BA (0, 0.25, 0.5 and 0.75mg/L) and 6 g/L agar. Six callus segments were plated in a 300 ml glass jar with 33 ml media andmaintained in growth room with 25 ± 2ᵒC temperature and 16/8 photoperiod with florescent lamp.After 6 months, number of shoot, shoot length and leaf number was recorded. Regenerated shootswere removed from calli and rinsed with sterilized water and planted in plastic pot filled withsterile peat: perlite (1:2) and covered with a transparent plastic container in order to moister preser-vation and maintained in growth room under 25 ± 1ᵒC temperature and photoperiod 16/8 for twomonths. Plantlets acclimated and finally transferred to greenhouse.

This study was conducted in a factorial experiment based on completely randomized design.Each experiment included 3 replications. Data were analyzed using the SAS software (version9.1). Percent data were arcsine transformed before performing ANOVA. The mean values werecompared using Duncan’s Multiple Range Tests (P<0.05).

RESULTS AND DISCUSSIONCallus induction was performed from cutedge of the explants which was most reactive

zones. The 25-30 days after culture of explant on media creamy color and compact callus was in-duced. Smilar to our results, Kuehnle and Sugii (1991) reported that Anthurium cultivars were pro-duced callus along cut edges of both leaf and petiole explants for all cultivars tested and it waspale yellow and firm. But they cited that callus produced after 2 to 3 months on a modified Pierikmedium. Also we observed that callus induction was more from cut edge and midrib. KhorramiRaad et al. (2012) explained that lamina explants exhibited more potential for callus formationwhen they contained midrib.

Plant growth regulators (PGRs) composition showed significant effect on callus size andpercentage of explant that produced callus (Table 1). Exogenous application of auxin and cytokinininduces callus in various plant species. Generally speaking, an intermediate ratio of auxin and cy-tokinin promotes callus induction, while a high ratio of auxin-to-cytokinin or cytokinin-to-auxin

ns, nonsignificant; **, significant at P<0.01.

Table 1. Analysis of variance for the effect of culture media and PGR concentrations on callusinduction trait in Anthurium andreanum ‘Clisto’.

S.O.V dfMean square

Callus size Callus percent

Media culture (M)PGR composition (P)M×PerrorCV (%)

1151564

4.55 **3.86 **0.76 ns

0.4929

14.89 ns

62.24 **5.59 ns

4.5528


induces root and shoot regeneration, respectively (Skoog and Miller, 1957). The highest percentageof explant that produced callus was obtained with 0.25 mg/L 2,4-D + 0.5 mg/L BA, however itwas not statistically different from 0.25 mg/L 2,4-D + 1 mg/L BA. The lowest percentage of explantthat produced callus was observed in media without BA (Fig. 1). These results show that BA hasan important role in callus induction of this plant.

The results for callus size determined same as callus production percent and the media cul-ture without BA showed lower callus size (Fig. 2). Our result was in agreement with Bejoy et al.(2008). They concluded that PGR balance in culture regulated explant response and they foundthat BAP along with 2,4-D were better in respect to rate of responsiveness and extent of callus de-velopment. They observed best dedifferentiation in 1 mg/L BAP and 0.5 mg/L 2,4-D in 6 week.Beyramizade et al. (2008) reported that best result for callus induction was obtained on half strengthMS medium containing 0.08 mg/L 2,4-D and 1 mg/L BA.

Culture media significantly influenced callus size (Table 1) and Nitsch’s medium producedhigher callus size in comparison to ½ MS (Fig. 3). In the study of Puchooa and Sookun (2003) tis-sue culture response of three variety of Anthurium andraeanum (Nitta, Osaki and Anouchka) ontwo different media (modified MS and modified Nitsch) for callus induction were evaluated. Nitschmedium with reduced ammonium nitrate concentration (200 mg/L), proved to be the best for callus

Fig. 1. Effect of PGR composition on percentage of explant thatproduced callus per treatment in callus induction stage of Anthuriumandreanum ‘Clisto’. Means with different letters were significantly different

at the 5% level as determined by Duncan’s multiple range test (DMRT).

Fig. 2. Effect of PGR composition on callus size per treatment incallus induction stage of Anthurium andreanum ‘Clisto’. Means withdifferent letters were significantly different at the 5% level as deter-

mined by Duncan’s multiple range test (DMRT).


induction. The media was supplemented with BA at 1 mg/L and 0.1 mg/L 2,4-D.Callus was transferred to shoot formation media and light condition changed to green color

and started to shoot formation and after six months data collected and shoots were removed andplanted in pots. Jahan et al. (2009) explained that developed calli by leaf and spadix explants whichmaintained in dark condition for three months, no multiple shoots were regenerated and so, inorder to induction of multiple shoots, callus was cultured on MS medium supplemented with dif-ferent concentrations of BAP and KIN by shifting them from dark to light condition.

In shoot regeneration, BA concentration treatment showed significant effect on number ofshoot, shoot length and number of leaf (Table 2). The tallest shoot length determined with mediumwithout BA and 0.25 mg/L BA and with 0.5 and 0.75 mg/L shoot length was shorter (Fig. 4).

Fig. 3. Effect of media culture type on callussize per treatment in callus induction stage ofAnthurium andreanum ‘Clisto’. Means with dif-ferent letters were significantly different at the5% level as determined by Duncan’s multiple

range test (DMRT).

ns, nonsignificant; **, significant at P<0.01.

Table 2. Analysis variance for effect of BA concentration on shoot regeneration trait inAnthurium andreanum ‘Clisto’.

S.O.V dfMean square

Shoot length Number of shoot Number of leaf

BAerrorCV (%)

320

8.93 *2.5125.3

11.36 *3.7027.3

4.88 *1.33

26.78

Fig. 4. Effect of BA concentration on shoot length inshoot regeneration stage of Anthurium andreanum

‘Clisto’. Means with different letters were significantlydifferent at the 5% level as determined by Duncan’s

multiple range test (DMRT).


The highest number of shoot was achieved with medium with 0.75 mg/L BA (31 shootper callus) and it was not significantly different from 0.25 and 0.5 mg/L BA. The lowestnumber of shoot was observed with medium without BA (Fig. 5). Budiarto (2008) reportedthat presence of BA affected on the number of newly developed shoots. They found that thehighest shoot formation was in Nitsch and Nitsch media supplemented with 2 and 3 mg/LBA in several potted Anthurium accessions. Viegas et al. (2007) observed shoot formationof Anthurium andraeanum ‘Flamingo’ during 70 day culture and it is increase positivelywhen MS media supplemented with 0.0, 0.5 and 1.0 mg/L BA. They concluded that a mediumcontaining this cytokinin resulted in constant regeneration and growth of shoot until 10thweek of culture.

Viegas et al. (2007) reported that each shoot produced 3 to 4 leaves throughout the 70 daysof observation and these leaves were morphologically normal, dark green and cordiform. It wassimilar to our results. The results presented that the highest leaf number observed with mediumwithout BA and it was statistically not different from 0.25 mg/L BA. The lowest number of leafwas determined with 0.5 and 0.75 mg/L BA (Fig. 6).

Whereas this plant easily is rooted, we directly transferred multiplied shoots to pots andafter acclimation period transferred them to greenhouse. We could obtained an effective acclimationand 99% plantlet rooted and survived. Directly rooted shoots in soil show higher survival rate inthe field than rooted under in vitro conditions. Therefore, there are several methods to high thesurvival rate of in vitro rooted shoots (Atak and Celik, 2012).

Fig. 5. Effect of BA concentration on number of shoot inshoot regeneration stage of Anthurium andreanum

‘Clisto’. Means with different letters were significantly dif-ferent at the 5% level as determined by Duncan’s multiple

range test (DMRT).

Fig. 6. Effect of BA concentration on number of leaf inshoot regeneration stage of Anthurium andreanum

‘Clisto’. Means with different letters were significantly dif-ferent at the 5% level as determined by Duncan’s multiple

range test (DMRT).


CONCLUSIONIn this experiment, we successfully regenerated shoot from leave explants of Anthurium

andreanum ‘Clisto’. Callus induction was the best in 0.25 mg/L 2,4-D + 0.5 mg/L BA and Nitsch’smedium. In shoot regeneration phase also results was excellent and 31 shoot per callus segmentwere obtained. The highest shoot multiplication was achieved in MS media supplemented with0.75 mg/L BA. Regenerated plantlet showed admirable adaptation and subsequent well growth.We could optimize shoot regeneration protocol for Anthurium andreanum ‘Clisto’.

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Fig. 7. A: Callus induction from cut edge of leaf explant. B:Callus proliferation with subculturing. C: Transfering callusto light condition for shoot regeneration. D: Shoot regener-

ation in different BA concentration 0, 0.25, 0.5 and 0.75mg/L from left to right. E: Shoot proliferation. F: Removingshoot from callus. G: Planting shoot in pot and rooting. H:

Plantlet acclimation in greenhouse.


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Exogenous Putrescine Delays Senescence of LisianthusCut Flowers

Keywords: Antioxidant enzyme, Lipoxygenase, Lisianthus, Putrescine, Vase life.

Davoud Ataii*, Roohangiz Naderi , Azizollah Khandan-MirkohiDepartment of Horticultural Sciences, Faculty of Agricultural Sciences and Natural Resources, Universityof Tehran, Karaj, Iran.


Abstract

The anti-senescence biogenic polyamines (PAs) putrescine (Put), sper-midine (Spd) and spermine (Spm) are involved in plant growth processes andresponses to stress. The physiological mechanism of exogenous Put to affectthe senescence of cut lisianthus flowers during vase life was investigated.Fresh cut lisianthus flowers were treated with distilled water (control), 0.5, 1and 2 mM Put and then held at 25 ◦C up to 12 days. Exogenous Put supply at2 mM extended vase life, which was associated with reduced electrolyteleakage and MDA content. Put treatment also reduced activity of lipoxygenase(LOX), is responsible for membrane lipid peroxidation. Put treatment also en-hanced activities of catalase (CAT) and ascorbate peroxidase (APX) anddecreased H2O2 accumulation during vase life. Thus, exogenous Put supplycould maintain membrane integrity by increasing antioxidant system activity,thereby retarding the senescence of cut lisianthus flower during vase life.


INTRODUCTIONDue to the ephemeral nature of the different tissues, the high respiratory activity and the

low carbohydrate content of reserves flowers are classified as highly perishable products (Nowakand Rudnicki., 1990). Cut Eustoma flowers have many cultivars with variations in flower colour,size and shape. There are several opened flowers and buds in Eustoma inflorescence and for theextending the vase life of the inflorescence the longevity of each flower and the rate of bud openingare important (Shimizu-Yumoto and Ichimura., 2005). Thus, for the extending the vase life of cutEustoma flowers preservatives containing sugars are effective (Cho et al., 2001). Increasing eth-ylene production during petal senescence indicating the Eustoma flower is ethylene-sensitive andethylene involvement in flower senescence (Ichimura et al., 1998). Some physiological and bio-chemical changes occurred during senescence such as loss of water from the senescing tissue, leak-age of ions generation of reactive oxygen species (ROS), increase in membrane fluidity andperoxidation, hydrolysis of proteins, nucleic acids, lipids and carbohydrates (Tripathi and Tuteja,2007). Polyamines (PAs) are ubiquitous and biogenic amines that have been implicated in cellularfunctions in living organisms. In plants they have been implicated in a wide range of biologicalprocesses including cell division, cell elongation, senescence, embryogenesis, root formation, floralinitiation and development, fruit development and ripening, pollen tube growth and plant responsesto biotic and abiotic stress (Hussain et al., 2011). In plant tissues, the main PAs are putrescine(Put), spermidine (Spd) and spermine (Spm). Exogenous application of putrescine improvementof physiological status of Helianthus annuus and stimulated catalase activity (Rubinowska andMichałek., 2009). Pre- and post-harvest putrescine applications extend the vase life of cut alstroe-meria flowers (Soleimany-Fard et al., 2014). Conditioning of cut stems of rose ‘Red Berlin’ in thesolution of putrescine at a concentration of 2 mmol resulted in the highest vase life (Rubinowskaet al., 2012). Exogenous spermidine delays senescence and extend vase life of cut Dianthuscaryophyllus flowers (Tassoni et al., 2006). In the present study, we investigated the effects ofpulse treatment with Put on the vase life of cut lisianthus flowers, as well as physiological andbiochemical changes during its petal senescence.

MATERIAL AND METHODS Flowers and treatment

Cut flowers of lisianthus (Eustoma grandiflorum) ‘Miarichi Grand White’ were obtainedfrom a commercial greenhouse and were re-cut under tap water to have uniform length of 30 cm.Flowers were then placed in a holding solution containing distilled water (control), 0.5, 1 and 2mM Put. All treatments were kept at 25 ± 1°C under a 16:8 h light/dark cycle and 60 ± 5% RH for24 hours. Subsequently, flowers were transferred to flasks containing only 200 mL distilled waterand were kept at 25 ± 1°C under a 16:8 h light/dark cycle and 60 ± 5% RH for 12 days. The endof vase life was evaluated as the time which 50% of the open flowers had wilted (Cho et al., 2001).

Membrane integrity evaluation Membrane permeability, expressed by relative electrolyte leakage rate, was measured by

the method of Jiang and Chen (1995). Thirty petal discs were immersed in 20 mL of 0.3 M mannitolsolution at 25 ◦C, followed by shaking for 30 min. Electrolyte leakage was determined with a con-ductivity meter. Total electrolyte leakage was determined after boiling the samples for 10 min andcooling to 25 ◦C. Relative electrolyte leakage rate was expressed as a percentage of total electrolyteleakage. MDA content was measured according to the method of Heath and Parker (1968). Frozenpetal tissues (1 g) from 10 flowers were ground finely in liquid nitrogen, then homogenized in 15mL of 10% trichloroacetic acid (TCA) and finally centrifuged at 5000 × g for 10 min. The super-natant phase was then collected. MDA content was determined by adding 5 mL of 0.5% thiobar-bituric acid (dissolved in 10% TCA) to 0.5 mL supernatant. The solution was heated at 95 ◦C for


20 min, quickly cooled, and centrifuged at 10,000 × g for 10 min to clarify precipitation. Ab-sorbance at 532 nm was measured and subtracted from the non-specific absorbance at 600 nm.The concentration of MDA was calculated with an extinction coefficient of 1.55 n mol L−1m−1.MDA content was expressed as n mol g−1 fresh weight (FW).

According to method of Doderer et al.,(1992), for analysis of LOX activity, frozen petaltissues (1 g) from 10 flowers were ground finely in liquid nitrogen and then homogenized in 15mL of 50 mM phosphate buffer (pH 7.0). After centrifugation at 10,000 × g and 4 ◦C for 20 min,the supernatant was collected and then used as the crude enzyme extract. LOX activity was assayedat 25 ◦C by monitoring the formation of conjugated dienes from linoleic acid at 234 nm accordingto the method of Axelrod et al., (1981). The reaction mixture (3 mL) contained 2.8 mL of 50 mMsodium phosphate buffer (pH 7.0), 0.1 mL of 10 mM sodium linoleic acid solution and 0.1 mL ofthe crude enzyme solution. One unit of LOX activity was defined as a change of 0.01 in absorbanceper minute at 25 ◦C. The specific LOX activity was expressed as U mg−1 protein.

Antioxidant system activity evaluation Frozen petal tissues (2 g) from 10 flowers were ground finely in liquid nitrogen and then

homogenized in 15 mL of 50 mM potassium phosphate buffer (pH 7.0) containing 1% (w/v) PVP.The homogenate was centrifuged at 10,000 × g for 15 min at 4 ◦C and then the supernatant wasused to determine activities of CAT and APX. CAT activity was assayed by measuring the disap-pearance of hydrogen peroxide (H2O2) according to the method of Oracz et al., (2009). The assaymixture (3 mL) contained 2.95 mL of 44.25 M H2O2 in 50 mM phosphate buffer (pH 7.0) and 0.05mL of enzyme extract. CAT activity was calculated by a decrease in absorbance at 240 nm for 3min at 25 ◦C. One unit of CAT activity was defined as the amount of the enzyme that caused achange of 0.001 in absorbance per minute and the specific activity was expressed as U mg−1 protein.APX activity was determined by the method of Nakano and Asada (1981). The reaction mixture(3 mL) consisted of 1.5 M ascorbic acid, 0.3 M EDTA and 0.3 M H2O2 solution in 50 mM phos-phate buffer (pH 7.0) and 0.1 mL of enzyme extract. Ascorbate concentration was followed by thedecrease in absorbance at 290 nm (extinction coefficient 2.8 mM−1 cm−1). One unit of APX activitywas defined as 1 M ascorbate oxidized per minute at 290 nm and the specific activity was expressedas U mg−1 protein. The protein concentration of petal extracts was estimated using the method ofBradford (1976) by BSA as standard. The H2O2 content measured according to Patterson et al., (1984).Frozen petal tissues (1 g) from 10 flowers were homogenized with 10 ml of acetone at 0 °C. After cen-trifugation for 15 min at 6000 × g at 4 °C, the supernatant was collected. The supernatant (1 ml) wasmixed with 0.1 ml of 5% titanium sulphate and 0.2 ml ammonia, and then centrifuged for 10 min at6000 × g and 4 °C. The pellets were dissolved in 3 ml of 10% (v/v) H2SO4 and centrifuged for 10min at 5000 × g. Absorbance of the supernatant phase was measured at 410 nm. H2O2 content wascalculated using H2O2 as a standard and then expressed as µmol g−1 fresh weight (FW).

For physiological parameters, results were expressed as mean ± SE from 3 replications.Statistical significance between mean values was assessed using one way analysis of variance withSAS (Version 9.1) statistical software. Means were compared using the LSD test.

RESULTS AND DISCUSSION Vase life

Putrescine treatment increased significantly the vase life cut lisianthus flowers, as comparedto control treatment during vase life. As shown in Fig. 1, treatment with postharvest Put at 2 mM re-sulted in a higher lisianthus cut flowers vase life (P<0.01). Based on these results, 2 mM Put forpostharvest treatment was chosen for further analyses. Exogenous putrescine treatment increase vaselife of cut alstroemeria and chrysanthemum flowers (Soleimany-Fard et al., 2014; Kandil et al., 2011).Effect of putrescine on the extending vase life of cut lisianthus flower is might be due to suppressedwater loss, inhibiting ethylene action and decrease in transpiration rate (Pandey et al., 2000).


Putrescine treatment and membrane integrity Electrolyte leakage of the lisianthus cut flowers increased during vase life (Table. 1). The

electrolyte leakage of lisianthus cut flowers treated with 2 mM Put at postharvest stage remainedlower than that in untreated control flowers (P<0.01; Table. 1). As well, during vase life, the MDAcontent in the lisianthus cut flowers increased (Table. 1). Compared to the controls, a lower contentof MDA was found in the lisianthus cut flowers treated with postharvest 2 mM Put (P<0.01; Table.1). There was a significant increase in the activity of LOX in lisianthus cut flowers during vaselife (Table. 1). The treatment with Put at 2 mM caused reduction in LOX activity in comparisonto the control for the whole 12 days of vase life (P<0.01; Table. 1).

Increased the electrolyte leakage is one of the first symptom of correlated with advancingsenescence (Shahri and Tahir., 2011). Exogenous application of putrescine decrease electrolyteleakage during leaf senescence of Helianthus annuus potted plants (Rubinowska and Michałek.,2009). Polyamines increase of plant’s cellular membranes resistance by inhibition of lipid perox-idation process (Roy et al., 2005). Application of polyamines decrease electrolyte leakage duringthe vase life of cut rose ‘Red Berlin’ (Rubinowska et al., 2012). Inhibited protease and RNA-seactivities by exogenous spermidine was reported, which probably supported the membrane stabilityand protected plant cell against injuries due to a stress (Kubiś., 2006). The increment in MDA hasbeen described as a biomarker of lipid peroxidation (Bailly et al., 1996) and thus decreased itslevel in lisianthus cut flowers treated with Put indicates reduced lipid peroxidation. Reduced lipid

Fig.1. Effects of Put treatment at 0.05, 1 and 2 mM on vaselife of lisianthus cut flowers.

Time (day) Treatment Membrane integrity

Put (mM) EL (%) MDA (n mol g−1 FW) LOX (U mg−1 protein)

3

6

9

12

SignificantTreatmentTimeT × T

0 2 0 2 0 2 0 2 df133

18.55 ± 1.231 c

19.63 ± 0.559 c

22.15 ± 0.342 b

19.41 ± 0.787 c

26.48 ± 0.472 ab

20.43 ± 1.481 c

37.43 ± 1.652 a

23.45 ± 0.772 b

*****

2.125 ± 0.751 d1.450 ± 0.164 e3.172 ± 0.282 c1.481 ± 0.425 de

5.353 ± 0.354 b2.628 ± 0.185 cd

7.785 ± 0.425 a5.225 ± 0.896 b

*****

0.908 ± 1.531 d0.701 ± 0.659 e1.876 ± 0.382 c0.671 ± 0.707 e2.650 ± 0.452 b1.275 ± 1.182 c3.589 ± 1.128 a1.459 ± 0.275 b

****

Table. 1. Effect of postharvest Put treatment at 2 mM on electrolyte leakage, MDA content and LOXenzyme activity of lisianthus cut flowers for 12 days.


peroxidation participates to decreased electrolyte leakage in response to Put treatment. Levels ofmalondialdehyde (MDA) in Lilium measured during senescence in inner and outer tepals, and cutflowers and flowers on the plant (Arrom and Munné-Bosch., 2010). Levels of MDA in outer tepalsof cut flowers fell throughout development and senescence. In contrast, levels of MDA in innertepals rose in both flowers still attached to the plant and in cut flowers during development.Ramezanian and Rahemi (2011) reported that the combination of calcium chloride and spermidinetreatment reduce the MDA content in pomegranate fruit. In a number of flowers including carnation(Lynch and Thompson., 1984), day lily (Panavas and Rubinstein., 1998) and rose (Fukuchi-Mizu-tani et al., 2000), lipid peroxidation and ion leakage precedes by increase in LOX activity. lipidperoxidation levels in day lily increased even before flower opening and were associated with anincrease in LOX (Panavas and Rubinstein., 1998). LOX activity and lipid peroxidation may beconsequences of senescence in some species (Rogers., 2012). Polyamines due to its antioxidantproperty mainly prevent lipid peroxidation, and thus protect the membrane lipid from being con-version in physical state (Mirdehghan et al., 2007). According to our results, Put might extendvase life through improving membrane permeability and decreasing of lipid peroxidation. Sincelipid peroxidation is mediated by ROS (Kellogg., 1975), therefore Put may either be directly scav-enging ROS and thus decreasing lipid peroxidation, or it may be modulating the activity of an-tioxidant enzymes. Senescing plant tissue also experiences an increase in LOX activity, which alsopromotes the process of membrane polyunsaturated fatty acid peroxidation (Lynch and Thompson.,1984). Similar to lipid peroxidation (MDA content), Put caused a decrease in LOX activity duringvase life (Table. 1). An increase in LOX activity has been correlated with an increase in cell mem-brane permeability and senescence in Daylily and Rose (Panavas and Rubinstein 1998; Fukuchi-Mizutani et al. 2000). ROS accumulation may cause oxidative damage to lipids, forming toxicproducts, such as MDA, a secondary end product of polyunsaturated fatty acid oxidation. Accu-mulation of MDA is often taken into consideration as an indicator of senescence, because degra-dation of polyunsaturated fatty acids produces peroxide ions and MDA production (MacRae andFerguson., 1985). Thus, MDA build-up is usually considered as an indicator of plant oxidativestress and also as the degree of damage to the structural integrity of cell membranes of plants underpostharvest senescence (Posmykt et al., 2005).

Putrescine treatment and antioxidant system activityPlants cells are usually protected against ROS by antioxidant system such as CAT and APX

enzymes activity which scavenging ROS produced during oxidative stress. As shown in Table. 2,

Time (day) Treatment Antioxidant system activity

Put (mM) CAT(U mg−1 protein)

APX(U mg−1 protein)

H2O2(µ mol g−1 FW)

3

6

9

12

SignificantTimeTreatmentT × T

02 02 02 02 df313

0.921 ± 0.151 a1.382 ± 0.330 a0.865 ± 0.381 b0.998 ± 0.391 c0.785 ± 0.175 c0.826 ± 0.123 cd

0.687 ± 0.156 d0.965 ± 0.524 c

*****

2.43 ± 0.234 a2.97 ± 0.688 a1.36 ± 0.564 b2.69 ± 0.342 a1.22 ± 0.457 c1.83 ± 0.218 b0.88 ± 0.456 d1.63 ± 0.178 b

******

59.44 ± 7.235 a51.38 ± 5.526 a65.46 ± 3.875 b54.82 ± 3.977 a70.63 ± 7.789 c62.50 ± 9.789 ab

92.66 ± 5.724 d84.68 ± 15.452 cd

****

Table. 2. Effect of postharvest Put treatment at 2 mM on antioxidant enzymes CAT and APX activityand H2O2 accumulation of lisianthus cut flowers for 12 days.


lisianthus cut flowers treated with Put at 2 mM showed higher activities of CAT and APX associ-ated with lower H2O2 accumulation during vase life (P<0.01; Table. 1). The activities of antioxidantenzymes are considered as a response against oxidative stress (Zhou et al., 2014). Put treatmentsenhanced the activities of antioxidant enzymes which scavenge the ROS, as indicated by the de-creased level of MDA (Table 1 and 2). Exogenous application of putrescine decrease catalase ac-tivity during leaf senescence of Helianthus annuus potted plants (Rubinowska and Michałek,2009). Pomegranate fruit treated with combination of calcium chloride and spermidine had sig-nificantly higher activities in CAT and SOD and a lower activity in peroxidase (POX) (Ramezanianand Rahemi., 2011). in senescent petals of several species such as day lily (Panavas and Rubin-stein., 1998), Iris (Bailly et al., 2001), Gladiolus (Yamane et al., 1999) and carnation (Zhang etal., 2007). CAT activity has also been measured., CAT activity increases during flower developmentup to flower opening in all these flowers and there is then a fall in CAT activity during senescence.However, in Iris and day lily, levels appear to rise even during senescence. APX activity was gen-erally high in young flowers and fell during floral senescence in Gladiolus (Hossain et al., 2006).As senescence progresses slightly earlier fall in the activity of APX compared to the other ROS-scavenging enzymes (Rogers., 2012). There is an increase in petal or tepal ROS levels duringsenescence. In Lilium both inner and outer tepals increase in H2O2 levels was much greater in de-tached flowers compared to flowers still attached to the plant (Arrom and Munné- Bosch., 2010).PAs have multifaceted nature, and working as an antioxidant, a free radical scavenger and a mem-brane stabilizer (Hussain H2O22011).

Cellular membranes are highly prone to ROS such as H2O2 attack, and it is reasonable topropose that progressive decline in membrane stability assayed by MDA content is probably theconsequence of enhanced ROS attack under decreasing antioxidant activity such as CAT and APXenzymes activity during vase life (Table. 2). In the present study, the decline in membrane integrityof lisianthus cut flowers was alleviated by treatment with Put, which was associated with an in-crease in CAT and APX activity in treated flower. Put has a role in the induction of antioxidant en-zymes and/or might also be acting as a scavenger of ROS (Magda et al., 2015) thus maintainingmembrane integrity for extended period.

CONCLUSION In conclusion, the study was an attempt to investigate the potential roles of Put in delaying

the senescence of cut lisianthus flowers. Put was able to prolong the vase life and delay flowersenescence by maintaining membrane integrity, which was result from decreasing LOX enzymeactivity as responsible for membrane lipid peroxidation and increasing the antioxidant enzymesCAT and APX activities, which was led to diminishing H2O2 accumulation. The effects of Puttreatment on retarding flower senescence was due to increased antioxidant enzyme activities andthus reduced lipid peroxidation and maintained membrane stability, assayed by electrolyte leakageand MDA content.

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Spatial Distribution Pattern of Tetranychus urticae Koch(Acari: Tetranychidae) on Different Rosa Cultivars inGreenhouse Tehran

Keywords: Density, Rosa, Spatial distribution, Tetranychus urticae.

Fatemeh Bidarnamani 1*, Elham Sanatgar 2 and Mehdi Shabanipoor 31 Institute of Agricultural Science, University of Zabol, Zabol, Iran 2 Department of Entomology, Faculty of Agriculture, Islamic Azad University, Arak .Iran.3 Ph.D. Student in Agricultural Entomology Arak, Arak Iran


Abstract

The two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetrany-chidae), is an economically important pest of ornamental plant in Iran. Thepopulation abundance and spatial distribution were studied on five Rosa cultivarsincluding Maroussia, Wendela, Elderado, Wenedetta, and Hot Lady during twogrowing seasons of 2011 and 2012 in greenhouse. The k parameter, index of dis-persion, Lloyd’s mean crowding, Morisita’s index and regression methods(Taylor and Iwao) were used to estimate the spatial distribution pattern ofmite. There was improved sampling program by k parameter and coefficientsof regression models. The result indicated that the highest population densityof T. urticae was early of July (2.47 per leaf) and early of September (4.11 perleaf). Also, a sequential sampling plan was developed using the fixed-precisionmethod of Green for estimating the density of the mite. Populations on differentRosa cultivars, especially Maroussia, were aggregated during most of thegrowing season and negative binomial models fit data sets better than thePoisson series. The mean number of two-spotted spider mite per plant had sig-nificantly difference between all plant strata. The fitness percentage of cultivarMaroussia Rosa revealed a slight tendency towards the Poisson distribution, sothat 58 and 14.1 percent of data sets from T. urticae on cultivar Wenedettapopulations fit the Poisson distribution. These results fully support findings ofdistribution indices, indicating an aggregated distribution. Determining samplingprogram and spatial distribution pattern of the pest can effectively help to designand perform of IPM.


INTRODUCTIONThe two-spotted spider mite, Tetranychus urticae, is one of the main pests of agricultural

crops due to its broad host range. This polyphagous species feeds on more than 1,100 plant species,from which about 150 are of great economic value. Thus, it represents a very important pest forfield and greenhouse crops, ornamentals, annual and perennial plants especially shrub Rosa, allover the world (Grbic et al., 2011).

Roses, Rosa spp. (Rosaceae) are an exceptionally valuable decorative element of greenareas in the cities and important ornamental shrubs. It is the most popular perennial flowering plantin almost all countries. Some species represent an important aromatic plant providing volatile oilswhich are used for the production of medicines, perfumes, cosmetics and other aromatherapy prod-ucts. It is cultivated as an agricultural crop in various countries of the northern hemisphere suchas Bulgaria, China, Egypt, France, India, Iran, Morocco and Turkey (Demirozer, 2012).

The methods for estimating population densities in arthropods constitute the cornerstoneof basic research in agricultural ecosystems and are the principal tool for implementation of pestmanagement programs (Kogan and Herzog, 1980). At this estimating plan a reliable sampling pro-gram along with suitable techniques should bead opted (Pedigo and Buntin, 1994; Southwood andHenderson, 2000). A reliable sampling program includes an identification of the appropriate sam-pling time, sampling unit, a determination of the spatial distribution of sampling units as well assample size (Pedigo and Buntin, 1994).

Spatial distribution is a behavioral response of the individuals of a species to habitat (Youngand Young, 1998; Southwood, 1995). The information of spatial distribution (i.e., regular, randomor aggregated) can determine what sampling program must be carried out, especially sequentialsampling (Feng et al., 1993).

In ecology, it is often difficult to study the movement of individual animal directly, becauseit is small in size and numerous in populations. Ecologists studied the spatial pattern of individualsof a particular species to infer the underlying behavior rules that govern their movement (Greig-Smith, 1952; Taylor, 1986). Spatial distribution pattern of a pest and its predator is essential infor-mation needed in order to analysis their relationships (Hassell and May, 1974).

There are various studies that described the spatial distribution and population density ofT. urticae. Aggregative spatial distribution of T. urticae was reported in different crop such as soy-bean (Sedaratian et al., 2008), bean (Ahmadi et al., 2005; Mehrkhou et al., 2008), strawberry(Greco et al., 1999), pear (Takahashi et al., 2001) and apple (Slone and Croft, 1998). Despite theimportance of two-spotted spider mite in ornamental greenhouse, adequate information on the spa-tial distribution and reliable sampling plans has not been developed for this two-spotted spidermite in Rosa greenhouse. The main objectives of this study were: (1) to determine the populationdensity and seasonal dynamics of T. urticae on Rosa in greenhouse tehran, (2) to describe thewithin-field distribution characteristics of the Rosa two-spotted spider mite population and the ef-fects of some cultivars of roses (Rosa spp.) (3) to develop fixed-precision sequential samplingplans (Green, 1970) for estimating T. urticae density on ornamental Rosa.

MATERIALS AND METHODSSampling program

Samples were taken twice weekly on Pakdasht in Tehran Province on 34 dates from 4 Julyto 29 September 2011 and 26 dates from 12 July to 20 September 2012 on five Rosa cultivars in-cluding Maroussia, Wendela, Elderado, Wenedetta, Hot Lady. The greenhouse were divided into10 blocks of the same size; then inside each block, plants were randomly selected along a diagonaltransect and three leaves from the upper, middle and bottom strata of the plants were taken ran-domly on side branches from each plant. The number of leaves (n) sampled on each sampling datewas calculated using equation 1: (Southwood, 1995)


(1)

Where N, t, s, d and m are sample size, t-student, standard the individuals of a species tohabitat (Southwood, 1995; Young and Young, 1998). The deviation, desired fixed proportion ofthe mean and the information of special distribution (i.e., regular, random or mean of primary data,respectively (Southwood, 1995).

Relative variation (RV) was used to compare to be carried out, especially sequential sam-pling (Feng et al., 1993). Efficiency of various sampling methods (Hillhouse and Pitre, 1974). TheRV was a successful management of two-spotted spider mite strongly calculated using equation 2:

(2)

Where SE and m are the standard error of the mean and the need to be considered (Pedigo,1994). For example, compared with mean of primary sampling data, respectively. Leaves weretransported to the laboratory and the number of two-spotted spider mite was counted separatelyunder the stereomicroscope.

Spatial distributionDistribution of population was useful for estimating two-spotted spider mite density on five

cultivars Rosa including Maroussia, Wendela, Elderado, Wenedetta, Hot Lady classified using cal-culation of the variance to mean, which in turn, would be valuable for (S2 / m). Departure from therandom distribution was then ecological and pest management studies. Tested by calculating theindex of distribution (Id), as The objectives of the present study were to determine using equation 3:

(3)Where n denotes the number of samples (Southwood and Hendrson, 2000). This index was

tested by Z values to determine using equation 4:

(4)

Morisita’s coefficient of distribution: The uneven distribution coefficient (Iϭ) was calculatedthrough the equation 5:

(5)

Where n, xi and N are the number of sample units, the i number of individuals in each sam-ple unit and total number of individuals in n samples, respectively. To determine whether the sam-pled population was significantly different from random distribution; the large sample test ofsignificance was applied using Z values as follows equation 6:

(6)

Where m and n are the mean population density per leaf in each sampling date and the num-ber of sample units, respectively (Pedigo and Buntin, 1994).

The spatial distribution af the T. urticae was evaluated by using the parameters of Taylor’spower law. This law describes the regression between logarithem of population variance and log-arithm of population mean according to the equation 7:

(7)


Where, S2 is the population variance, X ̅ is population mean, a is the Y-intercept and b isslope of regression line. This equation can transform as euation 3:

(8)Iwao’s patchiness regression models: Iwao’s patchiness regression method was applied

to quantify the relationship between mean crowding index (m*) and mean (m) using the follow-ing equation 9:

m*=α+ βm (9)Where α and β refer to the tendency to crowding/repulsion and the distribution of population

on space.

The value of F and P acquired from regression equation were used to test whether the Tay-lor’s (b) and Iwao’s (β) coefficients were significantly different from 0. In addition, to test for theirdifference from 1, the satistic t (as t=slope-1/ SEslope) was used. Here, slope and SEslope are Taylor’sor Iwao’s coefficient and their standard errors in regression equations, respectively. Since Taylor’sand Iwao’s coefficient were estimated by two-year, the difference between year’s distribution co-efficients, were tested by the statistic t equation 10 (Feng et al., 1993; Iwao and Kuno, 1968):

(10)

Here, b1(and SE1) and b2(and SE2) are the Taylor’s and Iwao’s coefficient(and its standarderror) for the first and the second year, respectively.

The data of two years were integrated and a total distribution coefficient was estimatedonly when the difference between coefficients of two years was not significant.

RESULTS AND DISCUSSIONData sets from counts of different cultivars Rosa including Maroussia, Wendela, Elderado,

Wenedetta, Hot Lady, of T. urticae fit the negative binomial better than the Poisson series (Table1). When T-statistics were used for spatial analysis, 62 percent of total population data sets fit thenegative binomial distribution. The percentage fit of the negative binomial was obviously reduced(28 percent) when the Chi-square test was used to test the fit between observed and expected fre-quencies. The fitness percentage of cultivar Maroussia Rosa revealed a slight tendency towardsthe Poisson distribution, so that 58 and 14.1 percent of data sets from T. urticae on cultivarWenedetta populations fit the Poisson distribution.

Cultivars

Goodness of fit test

Chi-square T-statistics

NBa Pb NBa Pb

MaroussiaWendelaElderadoWenedettaHot ladyTotal

625856284728

12.121.226.12.17.31.3

515448344531

10.812.118.36.28.29.5

Table 1. Percentage fit of T. urticae different cultivars Rosa to negativebinomial and Poisson frequency distributions, using two Chi-square and

T-statistics goodness of fit tests.

a Negative binomial, b Poisson


Dispersion indicesThe results of primary sampling showed that the reliable sample size with maximum vari-

ation of 35% was 38 and 36 for 2011-2012 growing seasons, respectively. The relative variation(RV) of the primary sampling was 9.1 and 8.6 for the previous growing seasons, respectively. TheseRVs were very appropriate for the sampling program (Table 1). The Taylor’s equations for thegrowing seasons were obtained as log S2 = 0.256 + 1.421 log m (F38 = 1204.7, P< 0.05; Table 3)and log S2 = 0.311 + 1.345 log m (F38 = 644.9, P< 0.05), both with a great degree of fit (> 0.90).In addition, the coefficient b was significantly greater than 1 (2011: t 41= 11.25, P< 0.05; 2012: t38 = 8.51, P< 0.05; Table 3), implying an aggregated distribution. The Iwao’s equation for thegrowing seasons were obtained as m* = 0.458 + 1.367m (F41 = 1448.4, P< 0.05; Table 4) and m*= 0.378 + 1.356m (F38 = 709. 4, P< 0.05), both with a great degree of fit (> 0.90). In addition, thecoefficient was significantly greater than 1 (2011: t41= 11.20, P< 0.05; 2012: t38= 10.85, P< 0.05;Table 4), implying an aggregated distribution.

Previous studies have been stated an aggregated form for the spatial distribution pattern ofT. turicae. Population on other crops such as cotton (Ahmadi et al., 2005; Mehrkhou et al., 2008)and strawberry (Greco et al., 1999). Here, the estimated Taylor index b was between 1.36 and 1.35.In other studies, the estimated values of this index has been ranged from 1.4 to 1.6, for example1.3 on apple (Slone and Croft, 1998), 1.4 on pear (Takahashi et al., 2001) 1.5 T. urticae on soybean(Sedaratian et al., 2008).

The interaction between plant strata and sampling dates significantly influenced the meannumber of two-spotted spider mite per strata Rosa plant (F = 12.43, df = 79, p = 0.002). The meannumber of two-spotted spider mite per plant between all plant strata was significantly different (F= 127.32, df = 79, p = 0.0006). The lower stratum had the highest mean number of two-spotted

Growing season n a Sbc Sd c RV d m e d f N g

20112012

4141

0.0410.035

0.180.11

9.18.6

0.310.23

0.350.35

3836

Table 2. Estimated parameters from primary sampling of T. urticae onshrub rosa during 2011-2012.

(a) Number of samples; (b) Standard error of the mean; (c) Standard deviation;(d) Relative variation; (e) Mean of primary data(f) Desired fixed proportion of the mean, (g) Sample

Growing season b +_SE Loga±SE R2 F T Df

20112012Oveall

1.321±0.0571.345±0.0361.421±0.047

0.345±0.0470.311±0.0780.451±0.023

0.9730.9680.962

1204.7**644.9**

231.451**

11.25*8.51*4.21*

413879

Table 3. Regression statistics of Taylor’s power law for of T. urticae populations onshrub Rosa during growing season 2011-2012.

*and** show significant difference at 0.05 level with 0 and 1, respectively.

Growing season b +_SE Loga±SE R2 F T Df

20112012Oveall

1.367±0.0431.356±0.0511.231±0.061

0.458±0.0490.378±0.0660.523±0.051

0.9300.9440.951

1448.324**709.413**961.330**

11.20*10.85*22.29*

413879

Table 4. Regression statistics of Iwao’s power law for of T. urticae populations onshrub Rosa during growing season 2011-2012.

*and** show significant difference at 0.05 level with 0 and 1, respectively.


spider mite per plant and was significantly different (p <0.05) than the middle and upper strata.The number of two-spotted spider mite per plant was also found to differ significantly (F = 84.39,df = 79, p = 0.0011) between the sampling dates.

To explain these differences, some researchers believe that the spatial distribution ofT. urticae on other crop aggregated than on other host plants. But considering the result of similarstudies in various parts of the world, It might be concluded that the differences are at least partlycaused by the different host plants, pest population density and environmental conditions such asweather and pesticide applications (Zenjun, 1997; Yan, 1998).

The density of per stratum per plant was significantly higher at the lower stratum (p <0.05)than the other two strata throughout the cropping period except at 79, where the number of two-spotted spider mite at the middle stratum was higher than the other two strata. This indicated thattwo-spotted spider mite generally prefer the lower leaves of plant than leaves of middle and upperstrata of the plant. Similar result was also reported by (Pakyari, 2012) who evaluated similar aspectson several bean varieties.

Spatial distribution modelsWhen the spatial distribution models were fitted on the population density of T. urticae,

negative binomial model showed the best fit (2012: χ2=21.08, P>0.95; 2013:χ2=14.51, P>0.95 ).These results fully support findings of distribution indices, indicating an aggregated distribution.Although distribution indices showed random distribution in some planting, total population dis-tribution during growing season matched the aggregated model. The aggregated distribution of apopulation is likely to be confirmed by aggregated indices, but its frequencies might not be corre-spondent to negative binomial distribution. Further studies are needed to evaluate T. urticae pop-ulation distribution fitting is by other aggregated distributions than negative binomial distribution.In this case, the recommendations about the kind of spatial distribution and proper test for theirfitting are more reliable. There are some studies that describe the spatial distribution of T. urticaeon rose and other crops, using dispersion indices that only show the aggregation or randomness ofbehavior of the two-spotted spider mite population. In this study, we examined in addition the fit-ness of the population data of the T. urticae to the negative binomial and poisson frequency distri-bution in order to obtain the mathematical distribution models.

Spatial distributions is one of the most important ecological characters of a population thatcan be used in extended sampling programs for pest managements (Kuno, 1991). In an extendedsampling which is a quick and precise method for estimating mean population or decision of controltime, spatial distribution data is crucial in determination of equations and necessary sample sizefor the decision (Young and Young, 1998).

CONCLUSIONOur study indicated that the spatial distribution of T. urticae in Rosa greenhouse was of ag-

gregated from and the fixed precision sampling scheme developed using Green´s method was ac-ceptable for estimating two-spotted spider mite densities in commercial Rosa greenhouses.Therefore, the sampling strategies provided here can be used to obtain a rapid estimate of two-spotted spider mite densities with minimal effort. In addition, the knowledge of density level ofT. urticae would provide the solid basis for optimal decision- making in IPM programs.

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Pathogenicity of Alternaria Species Isolated fromChamaecyparis lawsonia In Vitro

Keywords: Alternaria spp., Chamaecyparis lawsonia, Fungi, Pathogenicity.

Mohammad Reza Safari Motlagh 1*, Fatemeh Ramezani Rad 2 and Shahram Sedaghathoor 31 Department of Plant Pathology, Faculty of Agriculture, Rasht Branch, Islamic Azad University, Rasht,Guilan Province, PO box 41335-3516, Iran2 Master of Horticulture, Rasht Branch, Islamic Azad University, Rasht, Iran3 Department of Horticulture, Faculty of Agriculture, Rasht Branch, Islamic Azad University, Rasht, Iran


Abstract

Chamaecyparis lawsonia, which brings beauty in parks and green space,is attacked by a variety of pathogenic agents specially fungi. In this study, someChamaecyparis lawsonia available in parks that had disease symptoms weresampled. After sample collection, in order to isolate fungal pathogens, cuts ofthe infected plant tissues were placed on PDA (potato dextrose agar) and laterwere put on WA (water agar) for identifying. Then test of pathogenicity of theseisolates on Chamaecyparis lawsonia was done inside the box with plasticstopper. To do this, cuts of colonies of 4 day fungi obtained on PDA mediumwere put on leaves. The final evaluation was performed after 10 days. This ex-periment was performed in a completely randomized design with 3 replicationsand 7 treatments. The results showed that isolated fungi are in 2 groups asfollowing: Alternaria franseriae and Alternaria tenuissima. According to theresults, both species were pathogenic on host. Totally, A. franseriae showedmore disease severity compared to A. tenuissima on C. lawsonia.


INTRODUCTIONChamaecyparis lawsonia is one of important conifers in gardening, planting and growing

of ornamental plants and due to its branches form has many fans among designers and gardeners(Zare, 2002). Fungi are the most important factors that affect the quantity and quality conifers (Ja-farpour, 1994). Agents of disease of shed leaves of conifers (Needle cast) are species of Lopho-dermium, Scirriha, Hypodermella, Adelopus, Rhabdocline and other related genus that removedleaves from host plant and fall. At first, symptoms of this disease appear as bright green or yellowspots on needle leaves that finally turn to brown or red (Jafarpour, 1994).

Pine dieback disease (Macrophoma die-back) was collected and reported for the first timein 1970 from Noshahr from Pinus eldarica. The agent of this disease was Diplodia sp. (Ershad,1978). According to the research done, various pathogenic fungi have been reported on conifersaround the world. In 1985, in Florida, Pinus spp. were exposed many root infecting fungi (Barnardet al., 1985). One of fungi resident in root is Ophiostoma sp. (Barnard and Meeker, 1995).

Root rot of Annosum that is caused by Heterobasidium annosum is one the most importantand devastating diseases that affect the conifers of world in northern temperate regions. About 200forest species including several conifers are host of H. annosum; the most common genus areAbies, Juniperus, Larix and Pinus (Barnard, 1999). Among the research carried out in Iran, iden-tifying pathogenic soilborne fungi in hand planting forest of conifers in Fars province can be re-ferred (Zarghani et al., 2010). The field observations of different hand planting forests in Farsprovince's geographic area, number of fungi were identified, which only two isolates from Fusar-ium. Sambucinum, Rhizoctonia. solani and Pythium okanoganens were pathogenic. P. okanoganenshad relatively high pathogenicity, while other fungi had relatively mild pathogenicity (Zarghani etal., 2010). Also, the causes of death in conifer seedlings in nursery of Lakan was studied (Herfe-hdoost et al., 2009). Damping off is one of the common diseases of forest plant nurseries that im-poses a lot of damage to plant nurseries. In another study, infected seedlings were studied afterbeing collected and transported to the laboratory, and then pathogens obtained after doing patho-genicity tests were identified as Rhizoctonia solani, Pythium sp., Fusarium. oxysporum, F. semi-tectum, F. solani and Fusarium sp. (Herfehdoost et al., 2009).

Chamaecyparis lawsonia, are used a lot in greenery and landscape design of urban parksdue to its beauty value, variety of colors and their ever greenness in the family of conifers, (Zare,2002). Therefore, objectives of this study were to study of pathogenicity of Alternaria species iso-lated from C. lawsonia that damages it at different stages of its growth.

MATERIALS AND METHODSSample collection

Samples were taken from different infected parts of plants with diseases symptoms (SafariMotlagh, 2000). Infected samples were placed individually in plastic bags and transported to the lab-oratory immediately for isolation of pathogens and laboratory operations were performed on them.

Identification of fungiIdentification of fungi was done using morphological characteristics as shape of colony,

color of colony, mycelial growth mode, conidiophore’s being single or group, conidiophore’s sizeand color, conidia’s length and width, the number of conidia’s septa, and so on. For this purpose,keys of identifying fungi such as Simmons, 2007; Ellis, 1971; Leslie and Summerell, 2006; NagRaj, 1993 were used.

Pathogenicity testPathogenicity tests were performed in plastic box with stopper in dimensions of 5 × 15 ×

24 cm. For inoculating on leaves, a piece of dimensions 3 × 2 mm from 4-day cultured fungus onculture medium PDA, was put on leaves. Then, boxes were placed in incubator at 26°C for 10


days. Ten days after inoculation, leaf appearance symptoms were assessed (Kamran and Bani-Hashemi, 1995; Yousefi and Hagian Shahri, 2009).

The measurement was based on visual observations of symptoms. Description of symptomsand grading was done as follows: 1= the leaves were healthy and asymptomatic, 2= creation ofsmall and undeveloped spots on leaves, 3= creating medium and developed spots on leaves, 4=complete blight (Safari Motlagh, 2011). Finally, severity of disease in each treatment was calculatedbased on the number of spots on the leaves according to this formula:

RESULTS AND DISCUSSIONA total of 50 isolated sampled fungi were obtained and after morphological evaluation, two

fungal groups were identified as follows:

First group: Alternaria franseriae E.G. SimmonsColonies were gray to white velvet. Conidiophores

were simple, with a single conidiogenous cell and wereproliferate in curve mode that had a terminal helium in eachshort development. Conidia were oval and elongated ovalwith a round tip, in some cases without the tip, approxi-mately oval or elliptical in golden brown, smooth and indimensions 12-8 × 40-30 µm, and had 5-8 septa and some-times longitudinal (Fig. 1). Characteristics of this group ofisolates were consistent with Alternaria franseriae (Sim-mons, 2007).

Second group: Alternaria tenussima (Kunze)Wiltshire

Colonies were blackish brown with fast growth.Conidiophores were simple or branched individually or insimple or branched groups, straight or curved groups, al-most cylindrical, septate, light yellow or light brown,smooth, up to 115 µm in length and 4.6 µm in thickness(Fig. 2). Conidia were present individually or in shortchains, straight or curved, bent spindle or oval shape thatgradually narrows towards the tip (Fig. 2).Characteristicsof this group of isolates were consistent with Alternariatenussima (Ellis, 1971)

In pathogenicity test, by comparing the averages ofdisease rating caused by the fungi under study in Chamaecyparis lawsoniana, compared with con-trol, it can be concluded that both isolated fungi species were pathogenic (Figs. 3 and 4).

In a study, Phytophthora lateralis was isolated from head dried, died and infected parts ofChamaecyparis lawsoniana in eight forest areas, protected areas and tourist cottages in England,Scotland and Northern Ireland (Green et al., 2013). In this study, P. lateralis was isolated fromyoung seedlings of C. lawsoniana and Thuja occidentalis that have disease symptoms. This wasthe first report of presence of this fungus on this plant (Green et al., 2013). In 1949 in Russia wasreported that seedling damping off, wilting and dying of pine trees are the most important diseaseswhich caused pine seedlings to be damaged in 30 Russian nurseries (Ankudinov, 1950). After in-vestigations carried out on seedlings, the most important cause of pine seedlings death was reported

Fig. 1. Conidia and conidiophores ofAlternaria franseriae (×1200).

Fig. 2. Conidia and conidiophores ofAlternaria tenussima (×1200).


as Fusarium spp. and then some species of Alternaria were identified (Ankudinov, 1950). In astudy on seeds, seedlings and nursery pine bark of pine trees in south Georgia, Huang and Kuhlman(1990) identified 41 species of 23 fungi genus and 12 species of fungi identified from pineseedlings seed were selected to demonstrate the pathogenicity. Among them, Alternaria alternata,Fusarium moniliforme var. moniliforme, and Penicillum expansum caused seedling damping off.

According to the results obtained in this study it can be concluded that each year manyconifers in the parks and forests around the world are attacked by a variety of pathogenic fungicausing their destruction.

According to analysis of variance, there was a significant difference in the disease severityof tested fungi level at probability 5%; and was observed that severity of Alternaria franseriaedisease was more of Alternaria tenuissima.

In Australia, was reported leaf blight of grey mangrove tree caused by Alternaria alternata(Chandrashekar and Ball, 1980). In this study, severe blight caused by Alternaria alternata, affectedleaves, flowers and cuttings of grey mangrove trees in the southern coast of Australia.

Asdaghi et al. (2014) investigated diseased trees of Populus euphratica in Khuzestanprovince and leaves with symptoms were collected and transported to the laboratory. Results ofpathogenicity test was creating spots on health leaves similar to spots on the leaves of infectedtrees. By doing these experiments was diagnosed pathogen Alternaria alternata. This was the firstreport of A. alternata on Populus euphratica in Iran. Kamalakannan et al. (2008) investigated dis-ease of leave spot of Aloe vera in the state of Tamil Nadu, India with symptoms including ovalshaped circles with dark brown necrotic spots, mostly located at the tips of the leaves. Alternariaalternata was reported as pathogen. This was the first report of leaf spot disease caused by Al-ternaria alternata in Aloe vera in India. In another study, blight symptom was observed in Incar-villea emodi cultivated in India that was caused by Alternaria sp. Symptoms of this disease wereobserved on leaves, flowers and twigs that caused loss of flowers in cool months. This was thefirst report of Alternaria sp. from I. emodi around the world (Shanmugan, 2011).

Fig. 3. Diagram of the comparison of A. franseriaemean disease rating in treatment and control.

Fig. 4. Diagram of the comparison of A. tenuissimamean disease rating in treatment and control.

Fungi host Mean disease severing

Alternaria franseriaeAlternaria tenuissima

Chamaecyparis lawsonianaC. lawsoniana

3.761.23

Table 1. Mean disease severity of fungi tested on Chamaecyparis lawsoniana.

SOV df Squares Mean

Treatment ErrorC.V. (%)

614

6.63**0.0589.46

Table 2. Variance analysis of disease rating in pathogenicity test.


In the summers of 2007-2008 when the temperature was increased in an unusual way, Al-ternaria alternata caused decay of cluster of grapes in Slovakia (Kakalikova et al., 2009). Thiswas the first report of this disease in Slovakia. Soleimani and Esmailzadeh (2007) investigateddiseased leaves of apple trees in the northeastern Iran. Symptoms of this disease include black anddark brown spots on the leaves of the apple tree. Alternaria mali was isolated from diseased leaves.This is the first report of Alternaria mali which causes leaf spot disease on apple trees in Iran.

CONCLUSIONAccording to the results obtained in this research and previous studies, Alternaria spp. were

pathogenic on different plants and therefore, further studies in particular greenhouse studies needto be done.

ACKNOWLEDGMENTSThis experiment was supported by the Islamic Azad University, Rasht Branch, Iran.

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Effect of Gibberellic Acid Pulsing and Sucrose ContinuousTreatment on Some Qualitative Characteristics of CutRose Flower cv. Velvet

Keywords: Carbohydrate, Cut rose, Plant growth regulators, Vase life, Water uptake.

Zahra Ahmadi 1 and Rahim Naghshiband Hassani 2*

1 Graduated student of MSc. degree of Horticultural Sciences, Faculty of Agriculture, University of Tabriz. 2 Assistant Professor, Department of Horticulture Sciences, Faculty of Agriculture, University of Tabriz.


Abstract

The aim of this study was the better assessment of the relationship ofpulsing treatment of gibberellic acid as an anti-senescence agent and holdingin sucrose as a carbohydrate supply resource on improving some qualitativeand physiological attributes of cut rose flower cv. Velvet. Hence, anexperiment involving the pulsing treatment with gibberllic acid (GA3) at 0,20, 40 and 60 mg L-1, for 24 hours, and holding them in sucrose at 0, 2 and3% with 250 mg L-1 of 8- HQS as an antimicrobial agent for all holdingtreatments was conducted. The study was performed as a factorial experimentbased on a randomized completely design (RCD) with three replication foreach combination treatment. Applying GA3 pulse treatment alone at all con-centrations increased significantly vase life and its effect enhanced withsucrose holding at 2 and 3% compared with control. The effect of GA3

pulsing in increasing of stem relative fresh weight (RFW) and solutionuptake (SU) was hastened by sucrose holding treatment at 2 and 3%.Highest amount of flower opening and petal water content during vase lifeperiod was observed in 60 mg L-1 of GA3 pulsing and sucrose 3% holdingtreatment which had been longer flower diameters and greater petals.Gibberellic acid pulsing alone and along with sucrose holding treatment atall concentrations caused to prevention of leaf chlorophyll degradationcompared with control. In conclusion, GA3 pulsing at 40 mg L-1 along withsucrose 2% holding treatment had a significant effect on improving vase lifeand other qualitative attributes of cut rose flower cv. Velvet.


INTRODUCTIONSenescence of cut flowers is under hormonal control and related to the changes in the car-

bohydrate status of the petals (Halevy and Mayak, 1981). The GA3 is considered to be a senes-cence- delaying plant growth regulator (Arteca, 1996). Pulsing alstromeria flowers for 24 h witha 0.01 mmol GA3 solution increased longevity of cut flowers (Jordi et al., 1995). Sabehat and Zies-lin (1995) also noted that GA3 treatment increased the vase life of roses. Hunter et al. (2004) foundthat treatment with GA3 repressed accumulation of the seven senescence associated transcripts indaffodil. It has recently been shown that treatment of flower with cytokinins and gibberellins candelay the senescence of cut flowers. Results of Ganelevin and Zieslin (2002) showed that it is pos-sible that sepals are as a source of gibberellic acid during flower bud development. Removingsepals, reduces fresh and dry weight, with the buds and peduncle length. Gibberellic acid changescritical rate of ethylene that increase vase life (Saks and Staden, 1993). Singh et al. (2008) demon-strated that the vase solution treatment combinations of GA3 and benzyladenine with sucrose sig-nificantly increased the vase life of cut spikes of gladiolus as compared to the sucrose alonetreatment or the control. However, the results obtained have been variable. Boose and van Staden(1989) demonstrated that the efficiency of these compounds depends on the mode of applicationas well as the type and concentration of a cytokinin used. The aim of the present work was to de-termine the effect of benzyladenine and gibberellic acid at different concentrations, used in a pulsetreatment solution, on the longevity of rose ‘Red One’.

Cut flower longevity has been associated with the concentration of carbohydrates in the cutflowers (Halevy and Mayak, 1981). The final stages of flower development are characterized by a de-cline in the content of carbohydrates and dry weight of petals. The gradual decline during respirationin aging flowers may be caused by short supply of readily respirable substrates, mainly sugars. It wassuggested that the content of these substrates may indicate the potential life of the flower at a specifictemperature (Nichols, 1973). Translocation within the flower from the petals to the ovary was alsodemonstrated in senescing flowers (Nichols, 1976). Sucrose mostly is common as a pulsing treatmentfor cut flowers. By treatment of cut flowers with external carbohydrates, maintains dry matter contentand level respiration, in result increases longevity cut flowers (Kuiper et al., 1995). Also, sugars im-proves water balance in the plant and are effective in the regulation of stomata and through this reduceswater evaporation (Sarka, 2004). Different concentrations of sucrose had been investigated by Butt(2005) on two cultivars of Rosa hybrida and results showed that sucrose at 25 g L−1 extended the vaselife by 8.2 days in var. Whisk Mac and 7.5 days in var. Triska as compared to 5.3 days in control. Theaim of this study was the better assessment of the relationship of pulsing treatment of gibberellic acidas an anti-senescence agent and holding in sucrose as a carbohydrate supply resource on improvingsome qualitative and physiological attributes of cut rose flower cv. Velvet.

MATERIALS AND METHODSPlant material, experimental design and treatments

Cut rose flowers cv. Velvet (Rosa hybrida) was prepared from Mahan commercial green-house in Tabriz Iran. This experiment was performed in October 2013 in the postharvest biologyLaboratory of a Department of Horticultural Sciences of University of Tabriz based as a factorialexperiment based on randomized completely design (RCD) consisting of 12 treatments and 3 repli-cations with 4 cut stems in each treatment combination. As soon as the flowers arrived to the lab,their thorn and lower leaves of the cut stem was removed gently. Prior to treatment, rose stemswere trimmed to a length of 50 cm and then they pulse treated with gibberellic acid (GA3) solutionat four levels (0, 20, 40 and 60 mg/l) for 24 hours in glass containers and then held in vases includedsucrose solution at 3 levels (0, 2 and 3 %) with 250 mg/l of the 8-hydroxyquinoline sulfate (8-HQS). Cut flowers were kept at room temperature (23 ±2°C), relative humidity 60±5% and thelight intensity of 12 µmol/m-2.s-1of cool white fluorescent lamps with 12 hours of light until theend of vase life.


MeasurementsVase life

During the vase-life period, the visual quality of cut flowering stems was inspected daily.In our study, vase-life was defined as the period from the time of cutting to the time when 50% offloret petals wilted or abscised or floret necks bent as described by Liao et al. (2000).

Solution uptake, relative fresh weight and flower diameterThe cut flowers fresh weight and the solution uptake rate were measured daily. The weight

of vases with and without cut flowers was recorded daily. Mean daily solution uptake (g stem−1

day−1) was computed using the formula (St−1−St), where St is the weight of vase solution (g) att=day 1, 2, 3,…, and n. Relative fresh weight (RFW) of stems was computed using the formulaRFW (%)=(Wt/W0)×100, where, Wt is the weight of stem (g) at t=day 0, 1, 2…, and n, and W0is the weight of the same stem (g) at t=day 0 (He et al., 2006). Flower diameter was measured asan index for petals expanding rate. The outer diameter of opened flowers was measured by a VernierCaliper (mm).

Leaf chlorophyll index, dry weight and petal water contentChlorophyll Index was measured by a SPAD-502 (Minolta Co., Japan). All readings were

carried out between the tip and the base of fully expanded leaves in each sample. Fresh weight ofpetals was recorded and petal dry weight was recorded after drying at 105 °C for 48 h in an elec-trical oven until constant weight was obtained. Petal water content was determined as the percent-age of total petal weight [(FW – DW)/FW×100] by weighing samples of all petals from a singleflower.

Statistical analysisThe recorded data were subjected to analysis of variance (one-way ANOVA) using the gen-

eral linear model program of SPSS software (SPSS Ver. 16). Means were compared by the leastsignificant difference (LSD) test at the 0.05 probability level.

RESULTSVase life

Sucrose holding of cut stems alone had a significant effect (p<0.05) on increasing vase lifecompared with control. Gibberellic acid (GA3) pulsing at all concentrations had significantly(p<0.05) longer vase life than the control. Pulsing by gibberellic acid alone at all concentrationsgave a vase life improvement compared with control and sucrose alone treatments (2% and 3%).However, the combination treatments of gibberellic acid and sucrose holding showed that the con-centration 40 mg/l of gibberellic acid pulsing alone and its combination with sucrose (2% and 3%)had longer vase life values compared with other levels (Table 1).

Treatments SucroseVase life (days)

0% 2% 3%

ControlGA3 20 mg L-1

4060LSD 0.05= 1.6 (n= 3)

8.4 d13.66 b14.8 ab

14.5 ab

10.5 c14.83 ab

16.6 a15.8 a

12 c14 b

16.9 a15.7 a

Each value represents a mean of three replicates. Means followed by the same letterswere not significantly different at 5% level of significance.

Table 1. Effect of Gibberellic acid (GA3) pulsing and sucrose holding treatmenton vase life of cut rose cv. Velvet.


Solution uptake, relative fresh weight and flower diameterCut stems pulsed with GA3 at all concentrations (20, 40 and 60 mg L-1) and holding at 2

and 3 % of sucrose solution showed markedly relative fresh weight amounts compared with control.However holding them at sucrose solution (2 and 3%) alone had a significant effect in increasingRFW values in comparison to control (Table 2).

Results showed that the increasing effect of RFW values of cut stems dependent to the su-crose concentration of the vase solution alone. Solution uptake of cut stems, improved significantlyby gibberellic acid pulsing and sucrose alone treatments. The maximum amount of solution uptake(3.88 g stem-1 day-1) was observed at all concentrations of GA3 pulsing with sucrose 3% holdingtreatment (Table 2). Flower diameter in all treatments, significantly increased by GA3 pulsing andsucrose holding as its maximum flower bud opening (55 mm) was observed at GA3 60 mg L-1

pulsing with sucrose 3% holding treatment and its minimum rate was belong to control (38.2 mm)(Table 3).

Leaf chlorophyll index, flower petal dry weight and water contentOur results showed that leaf chlorophyll index values of cut rose cv. Velvet increased

significantly by GA3 pulsing alone with maximum value (62.3) in concentration of 60 mg L-

1. Sucrose holding treatment of 3% with GA3 pulsing treatment (40 and 60 mg L-1) significantlyhad higher levels of leaf chlorophyll index compared with 20 mg L-1 and control (Table 3).There was no significant difference among flower petal dry weight values for both treatments.Higher amounts of flower petal water content was observed in all concentrations of GA3 puls-ing with all concentrations of sucrose holding treatment compared with control. The maximumrate of flower petal content (83.3%) was observed in 60 mg L-1 with 3% of sucrose vase so-lution (Table 4).

Treatments SucroseRFW (%) SU (g stem-1 day-1)

0% 2% 3% 0% 2% 3%


4060LSD 0.05 (n= 3)

96.3 c96.9 c95.2 c95.1 c

105.85 ab

104.95 b

105.6 ab

107.7 a2.17

107.65 a105.7 ab

107.05 a109.33 a

1.25 c2.43 b2.54 b2.76 b

2.82 b3.05 a3.27 a3.55 a0.95

3.41 a3.87 a3.83 a3.88 a

Table 2. Effect of Gibberellic acid (GA3) pulsing and sucrose continuous treatment on relativefresh weight (RFW), solution uptake (SU) of cut rose cv. Velvet.

Each value represents a mean of three replicates. Means with the same letter were not significantlydifferent at 5% level of significance (P < 0.05).

Treatments SucroseLCI FD (mm)

0% 2% 3% 0% 2% 3%


4060LSD 0.05 (n= 3)

48.3 c53.2 b58.2 ab

62.3 a

50.8 c

54.95 b58.6 ab

62.7 a3.85

52. 5 c56.6 ab

59.8 a63.3 a

38.2 g40.2 f42.2 e45.5 d

40.3 f42.5 e48.4 c52.6 b1.78

43.6 ed

44.3 d

53.5 b55.3 a

Table 3. Effect of gibberellic acid (GA3) pulsing and sucrose holding on leaf chlorophyll index(LCI) and flower diameter (FD) of cut rose cv. Velvet.

Each value represents a mean of three replicates. Means with the same letter were not significantly dif-ferent at 5% level of significance (P<0.05).


DISCUSSIONOne of the main problems in the postharvest of cut rose flowers is stem collapse, known as

bent neck (Dole and Wilkins, 1999). Bent neck is caused by insufficient flower stem hardening,maturation of the stem tissue below the harvested flower or low levels of dry matters, water contentof cut flowers (Nowak and Rudnicki, 1990). It has been reported that GA3 delays wilting and senes-cence as associated proteolysis (Eason, 2002). In an experiment on 5 rose cultivars which was per-formed by Goszczynska et al. (1990), vase life of cv. Mercedes increased significantly as indetached petals by gibberellic acid treatment. Our results revealed that using GA3 pulsing at 40and 60 mg L-1 significantly increased vase life of cut stems than the control and delaying bent neckof cut rose cv. Velvet (Table 1), by improving flower petal water content and solution uptake (Tables2 and 4). Similar results have been reported in gerbera cut flowers with GA3 treatment (Emongor,2004; Danaee et al., 2011).

The GA3 along with sucrose has been suggested to induce water uptake in cut flowers ofgerbera (Emongor, 2004). In fact gibberellic acid caused negative osmotic potential cell and in-crease water uptake by hydrolysis of starch and sucrose (Goszczynska and et al., 1990). Singh andet al. (2008) reported that 50-500 mg/l gibberellic acid spray over cut roses increases water uptake.Water shortages caused when the amount of transpiration is more than water uptake (Nowak andRudnicki, 1990). Also gibberellins continue to modulate growth of flowers after harvest as evi-denced by increased fresh weight of cut roses upon application of GA3 (Sabehat and Zieslin, 1995).Our results showed that with increasing sucrose concentration the amount of relative fresh weightof cut stems and solution uptake enhanced (Table 2). In a study by Elgimabi and Ahmad (2009)observed that the 3% sucrose treatment had more longevity than sucrose 2% and 1%, which similarto our results which obtained in this study. Improvement of the postharvest life of flowers by sugarshas been demonstrated for many years and most pulsing and holding solutions applied to cut flow-ers include sucrose (Halevy and Mayak, 1981). It is widely thought that sugar treatment prolongsthe vase life by increasing the levels of respiratory substrate (van Doorn, 2001). Apparently, thissugar provides energy for fundamental cellular processes, such as maintenance of the structureand function of mitochondria and other organelles (Capdeville et al., 2003). Increased sucroseconcentration enhanced petal growth in detached flowers of Eustoma grandiflorum (Kawabata etal., 1995). It is known that sucrose improves water balance in cut flowers (Halevy and Mayak,1974). After the supplied sugar reached the flower head, an improvement in water balance wasobserved (Borochov et al., 1976).

Our results showed that with increasing concentration of gibberellic acid pulsing treatmentflower opening process is increased too as it caused to higher values of flower diameter than control(Table 3). Skutnik et al. (2001) showed that a pulse treatment with GA3 greatly improved thepostharvest performance of Zantedeschia aethiopica leaves and dramatically reduced the normalincrease in pH and conductivity of the cell sap. The role of GA in petal growth has been demon-

Treatments SucrosePDW (g) PWC (%)

0% 2% 3% 0% 2% 3%


4060LSD 0.05 (n= 3)

0.17 0.17 0.18 0.18

0.18 0.2

0.25 0.28 ns

0.200.270.270.28

65.2 e68.2 de

70.2 d71.5 cd

68.3 de

77.5 b75.4 c78.6 b2.48

73.6 c79.3 b78.5 b83.3 a

Table 4. Effect of gibberellic acid (GA3) pulsing and sucrose holding on flower petal dry weight(PDW) and flower petal water content (PWC) of cut rose cv. Velvet.

Each value represents a mean of three replicates. Means with the same letter were not significantlydifferent at 5% level of significance (P<0.05) , ns: LSD-test was not significant (P<0.05).


strated in many plants and seems to be a general phenomenon (Pharis and King, 1985). Gibberellintreatments increased the size of flowers. The possibility is discussed that GA, which is exogenouslysupplied, enhances flower dimensions and keep flower pigmentation by drawing photosynthatesto the flower as a consequence of intensification of the sink (Zieslin et al., 1974).

GA significantly protects the chlorophyll in plants and prevents leaf yellowing duringpostharvest period, in some plants such as lily, Alstroemeria which can be stored for a long timeand caused to increase the vase life (Lukaszewska, 1995). In Easter lily leaves, the senescence de-laying effect of GA3 was associated with depression of the respiration rate (Han, 1995). Saks andvan Staden (1993) reported that GA3 treatment reduced levels of ACC and ethylene production. Inour research higher values of leaf chlorophyll index was observed in GA3 pulsing treatment of 40and 60 mg L-1 at the end of vase life of cut rose cv. Velvet than control (Table 3). This hormoneprevents the degradation of chlorophyll in plants (Ichimura and Goto, 2000), this may be due todecrease in pH cell sap and to prevent of degradation chlorophyll that protects chlorophyll (Skutinket al., 2001). Gibberellic acid decrease chlorophyll degradation and loss during the senescenceprocess, because of its role strengthening in the membrane of chloroplasts.

CONCLUSIONIn conclusion, GA3 pulsing at 40 mg L-1 along with sucrose 2% holding treatment had a

significant effect on increasing vase life of cut rose cv. Velvet and improved their flower qualityby increasing solution uptake, fresh weight, flower diameter, and leaf chlorophyll index thereforeenhancing flower quality and delaying senescence.

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type, concentration and mode of application on flower longevity. Journal of Plant Physiology, 135:155-159.

Borochov, A., Tirosh, T. and Halevy, A.H. 1976. Abscisic acid content of senescing petals on cut rose flower as affected by sucrose and water stress. Plant Physiology, 58:175-178.

Butt, S.J .2005. Extending the vase life of roses (Rosa hybrida) with different preservatives. Journal of Agriculture Biology, 7: 91-99.

Capdeville, G.D., Maffia, L.A., Finger, F.L. and Batista, U.G. 2003. Gray mold severity and vase life of rose buds after pulsing with citric acid, salicylic acid, calcium sulfate, sucrose and silver thiosulfate. Fitopatologia Brasileria, 28:380-385.

Danaee, E., Mostofi, Y. and Moradi, P. 2011. Effect of GA3 and BA on postharvest quality and vase life of gerbera (Gerbera jamesonii. cv. Good Timing) cut flowers. Horticulture Environment and Biotechnology, 52:140-144.

Dole, J.M. and Wilkins, H.F. 1999. Floriculture: principles and species. Prentice-Hall, inc. New Jersey. 613P.

Eason, J.R. 2002. Sandersonia aurantiaca: An evaluation of postharvest pulsing solution to maximize cut flower quality. New Zealand Journal of Crop and Horticultural Sciences, 30:273-279.

Elgimabi, M.N. and Ahmed, O.K. 2009. Effects of bactericides and sucrose-pulsing on vase life of rose cut flowers (Rosa hybrida). Botany Research International, 2: 164-168.

Emongor, V.E. 2004. Effects of gibberellic acid on postharvest quality and vase life of gerbera cut flowers (Gerbera jamesonii). Journal of Agronomy, 3:191-195.

Goszczynska, D.M., Zieslin. N., Mor, Y. and Halevy, A.H. 1990. Improvement of postharvest keeping quality of ‘Mercedes’ roses by gibberellins. Plant Growth Regulation, 9: 293-303.

Halevy, A.H. and Mayak, S. 1974. Improvement of cut flower quality opening and longevity by pre-shipment treatments. Acta Horticulturae, 43:335-347.


Halevy, A.H. and Mayak, S. 1981. Senescence and postharvest physiology of cut flowers –part II. Horticultural Reviews, 3: 59–143.

Han, S.S. 1995. Growth regulators delay foliar chlorosis of Easter lily leaves. Journal of American Society for Horticultural Sciences, 120:254-258.

He, S., Joyce, D.C., Irving, D.E. and Faragher, J.D. 2006. Stem end blockage in cut Grevillea ‘Crimson Yul-lo’ inflorescences. Postharvest Biology and Technology, 41: 78–84.

Hunter, D.A., Yi, M., Xu, X. and Reid, M.S. 2004. Role of ethylene in perianth senescence of daffodil (Narcissus pseudonarcissus L. ‘Dutch Master’). Postharvest Biology and Technology, 32:269-280.

Ichimura, K. and Goto, R. 2000. Effects of gibberellin (GA3) on yellowing and vase life of Narcissus tazetta var. Chinensis flowers. Journal of Japanese Society for Horticultural Sciences, 69: 423-427.

Jordi, W., Stoopen, M., Kelepouris, K. and Van Der Krieken, M.W. 1995. Gibberellin induced delay of leaf senescence of Alstroemeria cut flowering stems is not caused by an increase in the endogenous cytokinin content. Journal of Plant Growth Regulation, 14:121-127.

Kawabata, S., Ohta, S.M., Kusuhara, Y. and Sakiyama, R. 1995. Influences of low light intensities on the pigmentation of Eustoma grandiflorum flowers. Acta Horticulturae , 405: 173–178.

Kuiper, D., Van Reenen, H.S. and Ribot, S.A. 1995. Effect of gibberellic acid on sugar transport in to petals of ‘Madelon’ rose flowers during bud opening. Acta Horticulturae, 298:93–95.

Liao, L.J., Lin Yu, H., Huang, K.L., Chen, W. S. and Cheng Yi, M. 2000. Postharvest life of cut rose flowers as affected by silver thiosulfate and sucrose. Botanical Bulletin of Academia Sinica, 41:299-303.

Lukaszewska, A.J. 1995. Distribution of sugars in tulip flower parts as affesctd by ethereal and GA3

in the holding solution. Acta Horticulture, 405: 45-50.Nichols, R. 1973. Senescence of cut carnation flower: Respiration and sugar status. Journal of Horticultural

Sciences, 48:111-121.Nichols, R. 1976. Cell enlargement and sugar accumulation in the gynoecium of the glasshouse

carnation (Dianthus caryophyllus L.) induced by ethylene. Planta 130:47-52.Nowak, J. and Rudnicki, R.M. 1990. Postharvest handling and storage of cut flowers, florist greens

and potted plants. Timber Press. Oregon, U.S.A. pp-25-26. Pharis, R.P. and King, R.W. 1985. Gibberellins and reproductive development in seed plants. Annual

Review of Plant Physiology, 36: 517–568.Sabehat, A. and Zieslin, N. 1995. Promotion of postharvest increase in weight of rose (Rosa hybrida)

petals by gibberellin. Journal of Plant Physiology, 145:296-298.Saks, Y. and van Staden, J. 1993. Evidence for the involvement of gibberellins in developmental

phenomena associated with carnation flower senescence. Plant Growth Regulation, 12:105-110.Sarka, L. 2004. Yield, quality and vase life of cut roses in year-round greenhouse production. University

of Helsinki, Department of Applied Biology, Publication no. 23.Singh, A., Kumar, J. and Kumar, P. 2008. Effects of plant growth regulators and sucrose on postharvest

physiology, membrane stability and vase life of cut spikes of gladiolus. Plant Growth Regulation, 55:221-229.

Skutnik, E., Lukaszzewska, A., Serek, M. and Rabiza, J. 2001. Effect of growth regulators on postharvest characteristics of Zantedeschia aethiopica. Postharvest Biology and Technology, 21:241-246.

Van Doorn, W.G. 2001. Role of soluble carbohydrates in flower senescence: A survey Horticulturae, 543:179-183.

Zieslin, N., Biran, I. and Halevy, A.H. 1974. The effect of growth regulators on the growth and pigmentation of ‘Baccara’ rose flowers. Plant and Cell Physiology, 15:341-349.

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Chamaecyparis بیماری زایی گونه های آلترناریای جدا سـازی شـده از In Vitro در شرایط lawsonia

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کلیــد واژگــان: Alternaria spp.، بیماری زایی، شبه سروالوسون، قارچ ها.

7 مجله گیاهان زینتی، سال پنجم، شماره 3، )1394(

دهــیـکــــی را در ســـطح پارک هـــا چ ـــرگ Chamaecyparis lawsonia کـــه زیبای ـــاه ســـوزنی ب گی

ــون ــامری زای گوناگـ ــل بیـ ــیله ی عوامـ ــی آورد بوسـ ــان مـ ــه ارمغـ ــبز بـ ــای سـ و فضـ

بویـــژه قارچ هـــا مـــورد حملـــه قـــرار می گیـــرد. در ایـــن تحقیـــق از تعـــدادی

ـــامری ـــم بی ـــه دارای عالئ Chamaecyparis lawsonia موجـــود در ســـطح پارک هـــا ک

بودنـــد منونه بـــرداری انجـــام شـــد. پـــس از جمـــع آوری منونه هـــا، بـــه منظـــور

ــی روی ــوده ی گیاهـ ــای آلـ ــی از بافت هـ ــی، قطعاتـ ــای قارچـ ــازی پاتوژن هـ جداسـ

ـــایی روی ـــه منظـــور شناس ـــدی ب ـــت و در مراحـــل بع ـــرار گرف ـــط کشـــتPDA ق محی

محیـــط کشـــت WA قـــرار گرفتنـــد. ســـپس آزمایـــش بیامری زایـــی ایـــن جدایه هـــا

روی Chamaecyparis lawsonia در داخـــل جعبه هـــای پالســـتیکی درپـــوش دار

انجـــام گرفـــت. بـــرای ایـــن کار قطعاتـــی از کلنی هـــای 4 روزه ی قارچ هـــای بدســـت

ــا قـــرار داده شـــدند. ارزیابـــی آمـــده روی محیـــط کشـــت PDA بـــر روی برگ هـ

ـــال ـــرح کام ـــک ط ـــب ی ـــش در قال ـــن آزمای ـــت. ای ـــام گرف ـــس از 10 روز انج ـــی پ نهای

تصادفـــی بـــا 3 تکـــرار و 7 تیـــامر انجـــام شـــد. نتایـــج نشـــان داد کـــه قارچ هـــای

Alternaria franseriae :جـــدا شـــده در 2 گـــروه قـــرار می گیرنـــد کـــه عبارتنـــد از

ــر روی ــا بـ ــه ی آلرتناریـ ــر دوگونـ ــج هـ ــق نتایـ ــر طبـ و Alternaria tenuissima. بـ

ـــه واریانـــس اختـــالف معنـــی داری ـــر اســـاس جـــدول تجزی ـــد. ب ـــان بیـــامری زا بودن میزب

در شـــدت بیـــامری قارچ هـــای مـــورد مطالعـــه در میزبـــان مشـــاهده گردیـــد.

ـــه ـــبت ب ـــرتی را نس ـــامری بیش ـــدت بی ـــه Alternaria franseriae ش ـــی ک ـــن معن بدی

Alternaria tenuissima بـــر روی شـــبه رسوالوســـون نشـــان داد.

کنه دو نقطه ای )Tetranychus urticae Koch )Acari: Tetranychidae یکی

توزیع و جمعیت فراوانی می باشد. ایران در زینتی گیاهان اقتصادی مهم آفات از

Hot Maroussia، Wendela، Elderado، Wenedetta و فضایی پنج رقم رز شامل

Lady در طول دو فصل رشد در سال های 1390 و 1391 در گلخانه رز پاکدشت تهران

و موریسیتا شاخص لیود، تراکم میانگین پراکندگی، شاخص ،k پارامرت شد. مطالعه

روش های رگرسیون )Taylor و Iwao( برای تخمین الگوی توزیع فضایی بکار برده شد.

k و رضایب مدل های رگرسیون وجود پارامرت برنامه های اصالحی منونه برداری توسط

تیر )2/47 در اوایل T. urticae تراکم جمعیتی نتایج نشان داد که بیشرتین داشت.

هر برگ( و اوایل شهریور )4/11 در هر برگ( بودند. هم چنین یک برنامه منونه برداری

استفاده کنه تراکم تخمین برای گرین مدل از استفاده با گسرتده بصورت دنباله ای

اکرث فصل رشد بصورت Maroussia، در به ویژه ارقام مختلف رز شد. جمعیت روی

تجمعی بود و مدل های دو جمله ای منفی مجموعه داده ها را بهرت از مدل پویسون

تطبیق دادند. میانگین تعداد کنه دو نقطه ای در هر گیاه بین قسمت های مختلف آن

اختالف معنی داری داشت. درصد تطبیق رقم رز Maroussia نشان داد گرایش کمی

به سمت مدل پویسون هم دارد. هم چنین 58 و 14/1 درصد از مجموعه داده های

این داشت. پویسون توزیع با بیشرتی تطبیق Wenedetta رقم روی بر T. urticae

توزیع نشان دهنده و می کند تأیید را پراکنش شاخص یافته های کامل بطور نتایج

تجمعی است. تعیین برنامه منونه برداری و الگوی توزیع فضایی آفت بر روی طراحی

و اجرای مدیریت تلفیقی آفت می تواند اثرگذار باشد.

دهــیـکـچ

Tetranychus urticae Koch الگـوی توزیع فضایی کنـه دو نقطـه ایAcari: Tetranychidae روی ارقـام مختلـف رز در گلخانه تهران

فاطمه بیدار نمانی 1*، الهام صنعتگر 2 و مهدی شعبانی پور 3موسسه علوم کشاورزی، دانشگاه زابل، زابل، ایران

گروه حشره شناسی دانشکده کشاورزی، دانشگاه آزاد اسالمی واحد اراک، اراک، ایراندانشجوی دکتری حشره شناسی، اراک، ایران

تاریخ تایید: 18 مرداد 1394 تاریخ دریافت: 28 فروردین 1394 [email protected] :ایمیل نویسنده مسئول *

.Tetranychus urticae ،کلیــد واژگــان: تراکم، توزیع فضایی، رز




پلی آمین هــای پوترســین، اســپرمیدین و اســپرمین مــواد ضــد پیــری هســتند

کــه در فرآیندهــای رشــد گیــاه و در پاســخ بــه تنش هــا نقــش دارنــد. مکانیســم

ــوس را در طــول ــده لیزیانت ــری گل شــاخه بری ــرد پوترســین پی ــه کارب فیزیولوژیکــی ک

عمــر گلجایــی تحــت تاثیــر قــرار می دهــد مــورد بررســی قــرار گرفتــه اســت. گل هــای

ــا آب مقطــر )شــاهد(، 0/5، 1 و 2 میلــی مــوالر پوترســین ــده لیزیانتــوس ب شــاخه بری

ــرد ــداری شــدند. کارب ــا 12 روز نگه ــای 25 دجــه ســانتی گراد ت ــامر شــدند و در دم تی

پوترســین بــا غلظــت 2 میلــی مــوالر عمــر گلجایــی را توســعه داد کــه بــا کاهــش نشــت

یونــی و محتــوای مالــون دی آلدئیــد )MDA( مرتبــط بــود. تیــامر پوترســین فعالیــت

آنزیــم لیپوکســی ژناز)LOX( را کــه مســئول پراکسیداســیون چربی هــای غشــاء اســت

را کاهــش داد. تیــامر پوترســین همچنیــن فعالیــت آنزیم هــای کاتــاالز )CAT( و

H( را 2O

2ــدروژن ) ــید هی ــع پراکس ــش و تجم ــیداز )APX( را افزای ــکوربات پراکس آس

در طــول عمــر گلجایــی کاهــش داد. بنابرایــن کاربــرد پوترســین می توانــد نفوذپذیــری

غشــاء را بــه وســیله افزایــش فعالیــت سیســتم آنتــی اکســیدانی حفــظ کنــد و در نتیجــه

پیــری گل شــاخه بریــده لیزیانتــوس را در طــول عمــر گلجایــی بــه تاخیــر بینــدازد.

دهــیـکـچ

ــده ــری گل بری ــن پی ــر انداخت ــه تاخی ــی در ب ــین خارج ــر پوترس تاثیــوس لیزیانت

داوود عطایی *، روح انگیز نادری و عزیزاهلل خندان میرکوهیگروه علوم باغبانی، دانشکده منابع طبیعی و علوم کشاورزی تهران، کرج، ایران

تاریخ تایید: 5 مرداد 1394 تاریخ دریافت: 25 اردیبهشت 1394 [email protected] :ایمیل نویسنده مسئول *

کلیــد واژگــان: آنزیم های آنتی اکسیدان، پوترسین، عمر گلجایی، لیپوکسی ژناز، لیزیانتوس.



آنتوریــوم بــه عنــوان یــک گل بریــده و گلدانــی پــرورش داده می شــود و بخاطر اســپات های

ــا قلمــه ســاقه و تقســیم پاجــوش ــاه بطــور ســنتی ب ــن گی رنگــی اش معــروف اســت. ای

تکثیــر می شــود و ریزازدیــادی می توانــد یــک روش جایگزیــن باشــد. در ایــن آزمایــش، مــا

باززایــی شــاخه در Anthurium andreanum رقــم ‘Clisto’ را مــورد بررســی قــرار دادیــم.

ریزمنونــه بــرگ مــورد اســتفاده قــرار گرفــت کــه بعــد از ضدعفونــی شــدن روی دو نــوع

محیــط کشــت مختلــف )MS 1/2 و محیــط کشــت Nitsch( کشــت شــدند ایــن محیــط

کشــت بــا ترکیب هــای مختلفــی از تنظیــم کننده هــای رشــد توفــوردی بــا غلظت هــای 0،

0/25، 0/5 و 0/75 میلــی گــرم بــر لیــرت و BA بــا غلظت هــای 0، 0/5، 1 و 1/5 میلــی گــرم

بــر لیــرت تکمیــل می شــدند. ســه مــاه بعــد انــدازه کالــوس و درصــد ریزمنونه هــای کالــوس

،0/25 ،0( BA بــا غلظت هــای مختلــف MS زا انــدازه گیــری شــدند و بــه محیــط کشــت

0/5 و 0/75 میلی گــرم بــر لیــرت( بــرای شــاخه زایی منتقــل شــدند. بعــد از 6 مــاه، تعــداد

شــاخه، طــول شــاخه و تعــداد بــرگ ایجــاد شــده ثبــت شــدند. بیشــرتین تولیــد کالــوس

ــا 0/5 ــر لیــرت توفــوردی و 0/25 ی ــا 0/25 میلی گــرم ب ــا محیــط کشــت Nitsch همــراه ب ب

میلی گــرم بــر لیــرت BA بــه دســت آمــد. باالتریــن باززایــی شــاخه نیــز در محیــط کشــت

بــا 0/75 میلی گــرم بــر لیــرت BA حاصــل شــد. مــا 31 شــاخه در هــر قطعــه کالــوس کشــت

شــده در بهرتیــن تیــامر مــورد آزمایــش بــه دســت آوردیــم. ریشــه زایی شــاخه های تولیــد

شــده بــه آســانی در گلدانــی بــا بســرتی از پیــت: پرلیــت )2:1( انجــام شــد و آن هــا ســازگار

شــده و بــه گلخانــه انتقــال داده شــدند و 99% گیاهــان انتقــال یافتــه زنــده ماندنــد.

دهــیـکـچ

باززایی غیرمستقیم شاخساره آنتوریوم رقم ‘Clisto’ از ریز نمونه برگمریم اجداربین1*، محسن کافی2، مسعود میرمعصومی3 و پژمان آزادی4

1 گروه علوم باغبانی، دانشگاه آزاد اسالمی، واحد کرج، کرج، ایران

2 گروه علوم باغبانی، دانشکده منابع طبیعی و علوم کشاورزی تهران، کرج، ایران

3 گروه فیزیولوژی گیاهی، دانشکده زیست شناسی، دانشگاه تهران، تهران، ایران

4 موسسه تحقیقات بیوتکنولوژی کشاورزی ایران، کرج، ایران

تاریخ تایید: 14 شهریور 1394 تاریخ دریافت: 4 مرداد 1394 [email protected] :ایمیل نویسنده مسئول *

کلیــد واژگــان: آنتوریوم، ازدیاد گیاه، کشت بافت، BA، توفوردی.




اگروپیـرون ).Agropyron cristatum )L.( Gaertner( گیاهـی اسـت کـه پتانسـیل

تبدیـل شـدن بـه چمـن کـم هزینه و اسـتفاده در مناطق کـم آب را دارد. هـدف اصلی این

تحقیـق بررسـی خصوصیـات کیفی چمنـی 24 منونه اگروپیرون، جمع آوری شـده از مناطق

مختلـف ایـران، بـرای اسـتفاده بـه عنـوان چمـن کـم هزینـه بـود. آزمایـش در قالـب طرح

کامـال تصادفـی بـا 3 تکـرار انجـام شـد.. نتایـج نشـان داد کـه تنوع معنـی داری بیـن منونه

هـا در صفـات کیفـی چمنـی و اجزای آن وجـود دارد و میانگین کیفیـت چمنی 15 منونه 6

یـا باالتـر بـود کـه بیانگـر عملکرد کیفـی چمنی مطلـوب اسـت. کیفیت رسزنی بـرای همه

منونـه هـا باالتـر از 6 بـود و بیـن M(210( 6/30( تـا 7/28 )4049( متغیـر بـود. عملکـرد

مناطـق جمـع آوری بـذور بـرای همه صفات مورد بررسـی متنوع و متفاوت بـود و مناطق

مختلـف جغرافیایـی ایـران پتانسـیل معرفـی ژنوتیـپ هـای بـا کیفیـت چمنـی مطلـوب را

)r =71**( داشـتند. همبسـتگی مثبـت معنـی داری بیـن کیفیـت چمنـی و کیفیت رسزنـی

و بیـن کیفیـت چمنـی و پنجـه زنـی )**r =57( وجـود داشـت. بـر اسـاس نتایـج حاصل از

تجزیـه خوشـه ای، منونـه هـا در 3 گروه دسـته بندی شـدند و تک منونه خوشـه 2 باالترین

کیفیـت چمنـی )7/95(، کیفیـت رسزنـی )7/28(، رنـگ )7/27( و بافـت بـرگ )7/10( را

داشـت. ایـن نتایـج نشـان مـی دهـد کـه منونـه هـای بومـی اگروپیـرون ایـران می تواننـد

ذخیـره مهمـی بـرای اسـتفاده به عنـوان چمن کـم هزینه باشـند.

دهــیـکـچ

Agropyron cristatum (L.)( ایـران بومـی گیـاه انحصـاری و کیفـی خصوصیـات چمـن عنـوان بـه ).Gaertner

حسن بیات 1، حسین نعمتی 1*، علی تهرانی فر 1 و علی گزانچیان21 گروه علوم باغبانی، دانشکده کشاورزی، دانشگاه فردوسی مشهد، مشهد، ایران

2 مرکز تحقیقات منابع طبیعی و کشاورزی خراسان رضوی، مشهد، ایران

تاریخ تایید: 24 شهریور 1394 تاریخ دریافت: 15 اردیبهشت 1394 [email protected] :ایمیل نویسنده مسئول *

کلیــد واژگــان: آنالیز خوشه ای، حداقل نگهداری، کیفیت چمن زنی، نازک برگ بومی، همبستگی.



پروانــش بــا نــام علمــی Catharanthus roseus یکــی از مهمرتیــن گیاهــان

ــف روی ــای مختل ــر آب ه ــی تاثی ــور بررس ــه منظ ــت. ب ــره اس ــواده خرزه ــی خان زینت

ــه ــر پای ــزان عنــارص غذایــی خــاک و انــدام هوایــی پروانــش، آزمایــش فاکتوریــل ب می

طــرح کامــال تصادفــی در 3 تکــرار اجــرا شــد. تیامرهــای آزمایشــی شــامل: نــوع آب )آب

شــهر مغناطیــس شــده، آب شــهر، آب چــاه مغناطیــس شــده و آب چــاه( و دورهــای

آبیــاری )2، 4 ،6 و 8 روز یکبــار( بودنــد. در ایــن مطالعــه صفاتــی از قبیــل عمــر

گلدانــی، ارتفــاع بوتــه، تعــداد بــرگ، مقــدار ازت، فســفر و پتاســیم خــاک و گیــاه مــورد

ــاع ــی)42/23 روز(، ارتف ــج بیشــرتین عمــر گلدان ــق نتای ــر طب ــت. ب ــرار گرف ــی ق ارزیاب

ــاری 2 روز ــامر آبی ــه تی ــوط ب ــرگ )165/88( مرب ــه )21/71 ســانتی مرت( و تعــداد ب بوت

یکبــار بــا آب شــهر مغناطیــس شــده بــود. بیشــرتین مقــدار ازت گیــاه نیــز بــه تیــامر

آبیــاری 2 روز یکبــار بــا آب چــاه مغناطیــس شــده )0/32 میلی گــرم در لیــرت( اختصــاص

داشــت. تیامرهــای آبیــاری 8 روز یکبــار بــا آب چــاه و شــهر مغناطیــس نشــده بیشــرتین

ــا 56/21 میلی گــرم در ــاه ب مقــدار ازت خــاک را داشــتند. بیشــرتین مقــدار پتاســیم گی

لیــرت در تیــامر آبیــاری 2 روز یکبــار بــا آب چــاه مغناطیــس شــده بدســت آمــد. بیشــرتین

مقــدار فســفر گیــاه بــا 39/5 و پــس از آن 38/8 میلی گــرم در لیــرت به ترتیــب مربــوط

ــاری دو روز ــس شــده و آبی ــا آب چــاه مغناطی ــار ب ــاری دو روز یکب ــه تیامرهــای آبی ب

یکبــار بــا آب شــهر مغناطیــس شــده بــود

دهــیـکـچ

تاثیـر آب مغناطیسـی و دورهـای آبیاری روی میـزان عناصر غذایـی خاک و )Catharanthus roseus( اندام هوایـی پروانـش

داود هاشم آبادی1*، فاطمه زارع دوست2 و مریم جدید سلیماندارابی2 1 استادیار گروه باغبانی، واحد رشت، دانشگاه آزاد اسالمی، رشت، ایران

2 عضو باشگاه پژوهشگران جوان و نخبگان، واحد رشت، دانشگاه آزاد اسالمی رشت، ایران

تاریخ تایید: 17 شهریور 1394 تاریخ دریافت: 20 تیر 1394 [email protected] :ایمیل نویسنده مسئول *

کلیــد واژگــان: دورهای آبیاری، میدان مغناطیسی، گیاه زینتی، کیفیت آب.




نیاز به تولید گراس های متحمل به دلیل استفاده فزاینده از آب شور و غیرقابل مرصف

در حال افزایش است. آزمایش های گلدانی به صورت گلخانه ای جهت تعیین تحمل نسبی و

)Festuca arundinacea Schrub( پاسخ های رشدی به شوری 11 توده بومی فستوکا بلند

شامل: سمیرم، مشهد، سنندج، یاسوج، یزد آباد، داران، کامیاران، گندمان، بروجن، نصیرآباد و

الربز به مدت 8 هفته مورد ارزیابی قرار گرفتند. همچنین فستوکا تجاری به عنوان شاهد مورد

استفاده قرار گرفت. چهار سطح آب شور )0، 45، 90 و 135 میلی موالر NaCl( در گراس های

کشت یافته در گلدان های پالستیکی پر شده از ماسه خالص به عنوان بسرت، مورد استفاده

قرار گرفت و محلول هوگلند جهت تغذیه به کار رفت. نتایج نشان داد که صفات بیوماس

قسمت هوایی، بیوماس ریشه، سطح برگ کل، طول برگ، عرض برگ و درصد خسارت برگ

به طور معنی داری با افزایش شوری، افزایش یافت. کمرتین درصد خسارت برگی در سطوح

90 و 135 میلی موالر مربوط به توده سنندج و فستوکا تجاری بود، همچنین بیشرتین درصد

خسارت برگی در سطوح 45، 90 و 135 میلی موالر به ترتیب مربوط به توده های الربز،

گندمان و گندمان بود. بیوماس قسمت هوایی، سطح برگ کل و طول برگ توده سنندج در

مقایسه با سایر توده های بومی، کمرت تحت تاثیر شوری قرار گرفت. بر اساس داده های حاصل

از پارامرتهای رشدی )بیوماس ریشه و قسمت هوایی، خسارت برگی، عرض برگ، طول برگ و

سطح برگ(، رتبه بندی توده های بومی برای تحمل به شوری، عبارتند از: سنندج، داران، یاسوج،

کامیاران، نصیرآباد، سمیرم، مشهد، الربز، یزدآباد، بروجن و گندمان. این نتایج بیانگر پتانسیل و

توان رقابتی توده های بومی با نوع تجاری آن می باشد.

دهــیـکـچ

غربالگـری توده هـای بومـی Festuca arundinacea در ایـران بـرای تحمـل بـه NaCl بـا هـدف کاربـرد در فضای سـبز

آزاده موسوی بزاز 1*، علی تهرانی فر 1، محمد کافی 2، علی گزانچیان 3 و محمود شور 11 گروه علوم باغبانی، دانشکده کشاورزی، دانشگاه فردوسی مشهد، مشهد، ایران

2 گروه زراعت، دانشکده کشاورزی، دانشگاه فردوسی مشهد، مشهد، ایران

3 مرکز تحقیقات منابع طبیعی و کشاورزی خراسان رضوی، مشهد، ایران

تاریخ تایید: 17 مرداد 1394 تاریخ دریافت: 15 خرداد 1394 [email protected] :ایمیل نویسنده مسئول *

کلیــد واژگــان: پارامتر رشد، تحمل شوری، توده بومی، فستوکا بلند.



www.jornamental.comThe Journal of Ornamental Plants, is an open access journal that provides rapid publication of manuscripts

on Ornamental plants, Floriculture and Landscape. Journal of Ornamental Plants is published in English,

as a printed journal and in electronic form.

All articles published in Journal of Ornamental Plants are peer-reviewed. All manuscripts should convey im-

portant results that have not been published, nor under consideration anywhere else. Journal of Orna-

mental Plants will be available online around the world free of charge at http://www.jornamental.com.

In addition, no page charge are required from the author(s). The Journal of Ornamental Plants is pub-

lished quarterly by Islamic Azad University, Rasht Branch, Rasht, Iran.

Manuscript Submission

Please read the “Instructions to Authors” before submitting your manuscript. Submit manuscripts as e-

mail attachment to Dr. Ali Mohammadi Torkashvand, Executive Director of Journal of Ornamental Plants,

at [email protected]. Electronic submission of manuscripts is strongly encouraged, provided that

the text, tables, and figures are included in a single Microsoft Word 2003 file. A manuscript acknowledg-

ment including manuscript number will be emailed to the corresponding author within 72 hours.

Please do not hesitate to contact meif you have any questions about the journal. We look forward to

your participation in the Journal of Ornamental Plants.

Address: Islamic azad University, Rasht Branch

Horticultural Department,

Agriculture Faculty,

Rasht,

Iran.

P.O.Box 41335-3516

Email: [email protected]

URL: http:// www.jornamental.com

Topics and Types of PaperJournal of Ornamental Plants is an international journal to the publication of original papers and reviews

in the Ornamental plants, Floriculture and Landscape fields. Articles in the journal deal with Ornamental

plants, Floriculture and Landscape. The scope of JOP includes all Ornamental plants, Floriculture and

Landscape. The journal is concerned with Ornamental plants, Floriculture and Landscape and covers

all aspects of physiology, molecular biology, biotechnology, protected cultivation, and environmental areas

of plants. The journal welcomes the submission of manuscripts that meet the general criteria of signif-

icance and scientific excellence, and will publish:

● Research articles

● Short Communications

● Review

Papers are welcome reporting studies in all aspects of Ornamental plants, Floriculture and Landscape

including:

Any Novel Approaches in Plant Science

Biotechnology

Environmental Stress Physiology

Genetices and Breeding

Photosynthesis, Sources-Sink Physiology

Postharvest Biology

Seed Physiology

Soil-Plant-Water Relationships

Modelling

Published by:Islamic Azad University, Rasht Branch, Iran


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Journal of Ornamental Plants - Webs3)/J...132 Journal of Ornamental Plants, Volume 5, Number 3: 131-138, September, 2015 INTRODUCTION One of the most important environmental factors