Genetic Resources and Crop Evolution 45: 917, 1998. 9c
1998 Kluwer Academic Publishers. Printed in the Netherlands.
Diversity for abiotic and biotic stress resistance in the wild annual Cicerspecies
K.B. Singh1, B. Ocampo1 & L.D. Robertson21Germplasm Program & 2Genetic Resources Unit, International Center for Agricultural Research in the DryAreas (ICARDA), P.O. Box 5466, Aleppo, Syria
Received 24 February 1997; accepted 10 June 1997
Key words: chickpea, Cicer arietinum, disease, genetic diversity, genetic resources, multiple resistance, nematode,pest, wild species
Data on 228 accessions of eight annual wild Cicer species and 20 cultivated chickpea check lines were evaluated fordiversity in response to six of the most serious biotic and abiotic stresses which reduce crop yield and productionstability of chickpea, i.e., ascochyta blight, fusarium wilt, leaf miner, bruchid, cyst nematode, and cold. Relativefrequencies of score reactions to the above six stresses were recorded from all the annual wild Cicer species andthe cultivated taxon. Patterns of distribution and amount of variation of the resistance reactions differed betweenstresses and species. C. bijugum, C. pinnatifidum and C. echinospermum showed accessions with at least one sourceof resistance (1 to 4 score reactions) to each stress. Overall, C. bijugum showed the highest frequencies of the highestcategories of resistance. Next in performance was C. pinnatifidum followed by C. judaicum, C. reticulatum and C.echinospermum. Furthermore, C. bijugum had the highest number of accessions with multiple resistance to the sixstresses; two accessions were resistant to five stresses and 16 to four. According to the Shannon-Weaver diversityindices (H0), five species showed discrete mean diversity indices which varied from 0.649 in C. pinnatifidum to0.526 in C. judaicum, whereas C. chorassanicum, C. cuneatum and C. yamashitae showed the lowest H0s, whichwere respectively 0.119, 0.174 and 0.216. Pair-wise correlations among the six biotic and abiotic stresses showedthe possibility of combining these resistances. Interestingly, multiple resistant accessions were predominantly ofTurkish origin.
Despite large breeding efforts to improve chickpea(Cicer arietinum L.), lack of stable production contin-ues to be a major concern. This condition is aggravatedby the continuous shift of the crop to marginal landswhere it faces a multitude of stresses. Improvementin the current crop status requires resistance to bioticand abiotic stresses which are the fundamental caus-es for unstable production. The cultivated taxon lacksadequate sources of resistance to these stresses. Thesuccess of the large-scale adoption of winter sowing inthe Mediterranean region, which has shown the poten-
Joint publication from ICARDA and ICRISAT (InternationalCrops Research Institute for the Semi-Arid Tropics), PatancheruP.O., A.P. 502324, India
tial to double chickpea yields (Hawtin & Singh, 1984),is crucially dependent on resistance to ascochyta blight(caused by Ascochyta rabiei [Pass.] Lab.). Worldwide,ascochyta blight and fusarium wilt (caused by Fusar-ium oxysporum Schlecht. emend. Synd. & Hans. f.sp.ciceri [Padwick] Synd. & Hans.) are the most seriousdiseases of chickpea. Leaf miner (Liriomyza cicerinaRond.) and cold are important stresses in the Mediter-ranean region. Wild relatives of chickpea have beenshown to be promising sources for genes for resistanceto the major biotic and abiotic stresses in chickpea(Singh et al., 1994). There are many examples of theexploitation of wild relatives for improvement of othercrops (Stalker, 1980; Prescott-Allen, 1988).
The genus Cicer comprises 43 species made upof 33 perennials, nine annuals including the culti-
Table 1. Country of origin of the annual wild Cicer species accessions and C. arietinumlines
Cicer Country of originAFGa ETH JOR LBN PAL SYR TUR Total
no. of accessionsarietinum 4 2 3 1 2 7 1 20bijugum 4 33 37chorassanicum 5 5cuneatum 4 4echinospermum 11 11judaicum 2 2 8 17 9 22 7 67pinnatifidum 1 1 5 8 34 50breticulatum 51 51yamashitae 3 3
Total 14 9 11 19 16 41 137 248
a AFG= Afghanistan, ETH= Ethiopia, JOR= Jordan, LBN= Lebanon, PAL= Pales-tine, SYR= Syria, TUR = Turkey.b Country of origin is unknown for one accession.
vated chickpea and one unspecified (van der Mae-sen, 1987). The International Center for AgriculturalResearch in the Dry Areas (ICARDA), Aleppo, Syr-ia, maintains the worlds largest collection of annualwild Cicer species (270 accessions), and of the Kabulitype chickpea (9870 accessions). Extensive collectionsof wild Cicer are also maintained at the InternationalCrops Research Institute for the Semi-Arid Tropics(ICRISAT), India and at the USDA Regional PlantIntroduction Station at Pullman, Washington, USA.
The ICARDA world collection of annual wild Cicerspecies has been evaluated for ascochyta blight, fusari-um wilt, leaf miner, bruchid (Callosobruchus chinensisL.), cyst nematode (Heterodera ciceri Vovlas, Grecoet Di Vito), and cold (Singh et al., 1994). The morpho-physiological features of the annual wild Cicer speciesaccessions of ICARDA have been catalogued (Robert-son et al., 1995).
To date, no studies have been conducted on thecombined variability and diversity of responses to biot-ic and abiotic stresses in the annual wild Cicer species.This paper examines the patterns of distribution and thevariation in response to the above mentioned adverseconditions in a collection of 228 accessions of 8 annualwild Cicer species maintained by ICARDA. The objec-tive of this work was to estimate their relative diversity[Shannon-Weaver information index (H0) based on fre-quency data] for response to six stresses. This proce-dure has been used earlier to measure genetic diversitybased on biotic and abiotic stress responses (Singh &
Jana, 1993) as well as on quantitative traits (Porceddu,1976; Holcomb et al., 1977; Peeters, 1988; Jana et al.,1990; Pecetti et al., 1992).
Material and methods
The materials of this study consisted of 228 accessionsof the eight annual wild Cicer species from the ICAR-DA genebank, C. bijugum K.H. Rech., C. chorassan-icum (Bunge) M. Popov, C. cuneatum A. Rich., C.echinospermum Davis, C. judaicum Boiss., C. pin-natifidum Jaub. & Spach, C. reticulatum Ladiz. andC. yamashitae Kitamura, and 20 cultivated chickpealines. The cultivated accessions were chosen in orderto cover the geographical distribution of the annualwild Cicer species. Table 1 shows the number of acces-sions per species used in this study and their countryof origin. The accessions were screened for resistanceto ascochyta blight (Ascochyta rabiei), fusarium wilt(Fusarium oxysporum f.sp. ciceri), leaf miner (Liri-omyza cicerina), bruchid (Callosobruchus chinensis),cyst nematode (Heterodera ciceri), and cold resistance.
All evaluations, except for fusarium wilt, were car-ried out at Tel Hadya (36 350E, 36 510N and 284 ma.s.l.), Syria, the main research station of ICARDA.This site has a semi-arid continental Mediterraneanclimate with cold and relatively wet winters, dry andhot summers, and an average of 16 seasons precipi-
tation of about 330 mm. Evaluation for fusarium wiltwas done at Monterotondo, Rome, Italy.
Screening started in the 1987/88 season. The fol-lowing procedures were used. Ascochyta blight resis-tance (AB); was scored on a scale of 1 to 9, where1 = plants free from any damage and 9 = all plantskilled, following the technique and the scale describedby Singh et al. (1981). Each accession was sown in asingle 2 m row plot with inter- and intra-row spacing of45 and 10 cm, respectively (20 plants). Fusarium wiltresistance (FW); was scored on a scale of 1 to 9, where1 = plants free from any damage and 9 = all plantskilled, following the technique described by Infantinoet al. (1996) and a scale developed by Singh (unpub-lished): 10 to 15 plants per accession were assayed.Leaf miner resistance (LM); was scored on a scale of1 to 9, where 1 = plants free from any damage and9 = leaves severely mined, many dried and droppedfrom the plant, following the technique and the scaledescribed by Singh & Weigand (1994). Each accessionwas sown in a single 2 m row plot with inter- and intra-row spacing of 45 and 10 cm, respectively (20 plants).Bruchid resistance (CC); was scored on a scale of 1 to9, where 1 = seeds free from any damage and 9 = allseeds infested by beetles, following the technique andthe scale described by Weigand & Tahhan (1990); 50seed were assayed per accession. Cyst nematode resis-tance (CN); was scored on a scale of 1 to 9, where 1=plants free from any damage and 9 = highly suscepti-ble, following the technique described by Di Vito et al.(1988) and a scale developed by Singh (unpublished).Two pots of five seed each were grown per accessionto provide two replications. Cold resistance (CO); wasscored on a scale of 1 to 9, where 1 = plants freefrom any damage and 9 = all plants killed, followingthe technique and the scale described by Singh et al.(1989). Each accession was sown in two replicates ina randomized complete block design with each plot asingle 2 m row using inter- and intra-row spacing of 45and 10 cm, respectively (20 plants per replicate).
The screening for ascochyta blight resistance wascarried out in an ascochyta blight nursery, wheredisease was created artificially through inoculationwith diseased plant debris and with spore-suspensionsprays. The screening for fusarium wilt resistance wascarried out in a greenhouse at the Istituto Sperimentaleper la Patologia Vegetale, Rome. The screening forleaf miner resistance was carried out by growing theaccessions during spring under natural insect infesta-tion. The screening for bruchid infestation was carriedout by artificially releasing the bruchid in the Ento-
mology Laboratory of ICARDA to seed samples. Cystnematode screening was conducted in the greenhouseby growing accessions in plastic pots containing cystnematode infested soils. The screening for cold toler-ance was conducted at the Tel Hadya farm by advanc-ing the sowing date to early October. Evaluations weredone in systematic screening nurseries with repeatedchecks, except for fusarium wilt. Frequency distribu-tion, mean, standard deviation, coefficient of variationand skewness were calculated for each stress for eachspecies, and for the overall annual wild Cicer speciescollection using SPSSr for WindowsTM, Release 6.0(SPSS, 1993).
The measurements of genetic diversity for biot-ic and abiotic stress reactions were assessed by theShannon-Weaver information index (H0) (Shannon &Weaver, 1949) given as:
H0 = s
where s is the number of reaction scores showed byaccessions of a species to a given stress and p
is theproportion of the total number of accessions with theith score reaction. To confine the information index inthe range 0 to 1, each value of H0 was normalized bydividing it by its maximum value, i.e., log s (Tolbertet al., 1979). The mean information index (H0) of aspecies was estimated as the arithmetic average of theH0s values over all stresses.
Simple correlation coefficients were computedfrom stress scores for all pairwise combinations ofstresses for the entire wild Cicer germplasm collec-tion using SPSSr for WindowsTM, Release 6.0 (SPSS,1993).
Distributions of relative frequencies (%) of reactionsof the annual Cicer species to the six stresses: ascochy-ta blight, fusarium wilt, leaf miner, bruchid, cystnematode and cold, are given in Table 2. Overall,the six stresses differed in their patterns of distribu-tion and amount of variation. For instance, distributionfor ascochyta blight and bruchid were skewed towardsresistance; that for fusarium wilt was skewed towardssusceptibility; whereas distributions for leaf miner andcold were nearly normal. Species-wise distribution ofscore reactions were similar for ascochyta blight, fusar-
Table 2. Distribution of relative frequencies (%) of reactions (19 scale) of annual wild Cicer species to six biotic andabiotic stresses
Cicer speciesScale aria ret ech jud pin bij cho yam cun
Ascochyta blight1 (free from damage) 2 (highly resistant) 9.1 3.5 8.9 5.4 3 (resistant) 2.2 13.5 4 (moderately resistant) 3.5 5.4 5 (tolerant) 15.8 9.1 8.8 26.7 18.9 6 (moderately susceptible) 5.3 4.1 27.3 10.5 20.0 35.1 7 (susceptible) 15.8 10.2 18.2 12.3 13.3 13.5 25.08 (highly susceptible) 15.8 30.6 9.1 8.8 15.6 2.7 9 (all plants killed) 47.4 55.1 27.3 52.6 13.3 5.4 100.0 100.0 75.0
Skewness 0.88 1.24 0.98 1.27 0.47 0.17 2.00No. of accessions 19 49 11 57 45 37 1 3 4
Fusarium wilt1 (free from damage) 10.0 23.4 44.4 43.9 18.8 81.0 2 (highly resistant) 12.8 11.1 5.3 6.3 6.1 66.73 (resistant) 10.0 10.6 1.8 3.0 33.34 (moderately resistant) 10.0 31.9 22.2 26.3 15.6 3.0 5 (tolerant) 6.4 11.1 12.3 6.3 6 (moderately susceptible) 10.0 12.8 11.1 8.8 9.4 6.1 7 (susceptible) 10.0 15.6 8 (highly susceptible) 20.0 2.1 12.2 9 (all plants killed) 30.0 1.8 15.6
Skewness 0.89 0.29 0.52 0.58 0.31 2.92 1.73No. of accessions 10 47 9 57 32 33 0 0 3
Leaf miner1 (free from damage) 2 (highly resistant) 9.1 29.0 8.5 2.8 100.0 100.03 (resistant) 8.3 27.3 35.5 8.5 5.6 4 (moderately resistant) 18.8 14.6 27.3 11.3 19.1 16.7 5 (tolerant) 6.3 35.4 17.7 38.3 33.3 33.3 6 (moderately susceptible) 6.3 20.8 27.3 3.2 17.0 5.6 7 (susceptible) 18.8 20.8 9.1 3.2 8.5 30.6 33.3 8 (highly susceptible) 6.3 9 (very highly susceptible) 43.8 5.6 33.3
Skewness 0.60 0.18 0.30 0.88 0.39 0.14 0.00 No. of accessions 16 48 11 62 47 36 1 3 4
ium wilt and leaf miner, and different for bruchid, cystnematode and cold.
C. bijugum, C. pinnatifidum and C. echinospermumshowed accessions with at least one source of resis-tance (1 to 4 score reactions) to each stress (Table 2).The highest levels of resistance within this collectionwere found for fusarium wilt, bruchid and cyst nema-tode. The least number of resistance sources werefound for ascochyta blight. The highest relative as well
as absolute number of accessions resistant to ascochy-ta blight, fusarium wilt, cyst nematode and cold (1to 2 score reactions) were found in C. bijugum. C.chorassanicum and C. cuneatum showed the highestpercentage (%) of accessions highly resistant to leafminer but they numbered merely one and four acces-sions respectively. On the other hand, 18 (29%) ofthe accessions of C. judaicum were highly resistant toleaf miner. Similarly, for bruchids, all accessions of
Table 2. Continued
Cicer speciesScale aria ret ech jud pin bij cho yam cun
Bruchid1 (free from damage) 5.0 100.0 12.8 42.9 2 (highly resistant) 5.0 8.5 6.9 23.8 33.33 (resistant) 5.0 14.3 4 (moderately resistant) 5.0 3.4 4.8 33.35 (tolerant) 5.0 2.1 6 (moderately susceptible) 15.0 4.3 14.3 7 (susceptible) 10.0 45.0 12.8 33.3 33.38 (highly susceptible) 30.0 15.0 51.1 72.4 66.6 100.0 9 (very highly susce...