Microbial production of plant hormones

Plant and Soil 133: 1-8, 1991. © 1991 Kluwer Academic Publishers. Printed in the Netherlands. PLSO BARC31

Microbial production of plant hormones*

M U H A M M A D ARSHAD and W.T. FRANKENBERGER Jr* Department of Soil and Environmental Sciences, University of California, Riverside, CA 92521, USA

Key words: Azotobacter, ectomycorrhizae, plant growth regulating substances, precursor-inoculum interactions

Abstract

Our laboratory has focused on microbial production of plant hormones and precursor-inoculum interactions. Indole-3-acetic acid was detected in soils incubated with L-tryptophan (L-TRP). Inocula- tion with the ectomycorrhizae, Pisolithus tinctorius significantly stimulated the growth of Douglas fir when supplied with low concentrations of L-TRP to soil. Among three Azotobacter spp. and two Pseudomonas spp., the most prolific producer of cytokinins was A. chroococcum and among the precursors tested, adenine (ADE) and isopentyl alcohol (IA) were the most effective. Corn rhizosphere was found to be quite rich with microflora capable of producing ethylene from L-methionine (L-MET). Amino acids, carbohydrates and organic acids typically found in root exudates, were stimulatory to ethylene biosynthesis in soil. Etiolated pea seedlings exhibited the classical 'triple' response when L-MET and Acremonium falciforme were applied in combination to sterile soil or when L-MET was added to nonsterile soil.

Introduction

To improve crop yields, inoculation with specific microbial preparations is a common practice in many regions of the world including the Soviet Union, New Zealand, Australia, Belgium, Netherlands, India and U.S.A. In 1895, for the first time, an inoculant 'nitragin' containing rhizobia was marketed as a commercial product. Now rhizobial inoculants are well established as an agricultural product in U.S.A., Europe, Au- stralia and India. In 1937, an Azotobacter prepa- ration, under the trade name 'Azotogen' and later renamed 'Azotobaktrin' was commercial- ized in the Soviet Union for soil and seed treatment. Increased yields were reported for veget- ables and cereals. Later, Soviet microbiologists introduced another product 'phosphobakterin' consisting of Bacillus megaterium var. phos-

* First published in The Rhizosphere and Plant Growth. Eds. D L Keister and P B Gregan. Kluwer Academic Publishers, Dordrecht.

phaticum which was thought to enhance phos- phorus availability. The application of Azospiril- lure brasilense in Brazil (D6bereiner et al., 1976) led to its use as an inoculum for cereals. Today, there are a number of cases where other microbial preparations commercially available under different trade names may benefit crop yield.

It has been assumed that inoculation with Rhizobium, Azotobacter and Azospirillum enhanced plant growth as a result of their ability to fix nitrogen. However, despite extensive re- search efforts, only rhizobia have been shown to increase yields from dinitrogen fixation. In many cases, nitrogen levels in plant tissues and soils do not increase upon inoculation with these bacteria. Growth promotion may be attributed to other mechanisms such as production of plant growth regulating substances, PGRS (e.g. plant hormones) in the rhizosphere. Plants may show a response to inoculation even in nitrogen-rich conditions. Thus, an inoculum should be screened for its ability to synthesize PGRS.

Up to now, an exceedingly large number of

2 Arshad and Frankenberger

studies have indicated the involvement of plant hormones produced by a microsymbiont in altering plant growth and development. Brea and Brown (1974) observed increased growth of several plants including Paspalurn notatum, without concomitant nitrogen fixation upon inoculation with cultures of Azotobacter paspali. Similarly, Hussain et al. (1987) reported a yield increase of 16% in maize due to inoculation with Azotobac- ter chroococcurn in field studies receiving heavy applications of nitrogen fertilizer. This yield increase was attributed to the bacterial production of plant hormones. Many workers have reported production of plant hormones by Azotobacter (Brea and Brown, 1974; Brown and Burlingham, 1968; Jagnow, 1987; Lee et al., 1970; Nieto and Frankenberger, 1989a), Azospirillum (Hubbell et al., 1979; Tien et al., 1979), and Rhizobium (Phillips and Torrey, 1970; Puppo and Rigaud, 1978) and have considered their physiological action in altering plant growth and development.

Although several studies have shown significant increases in crop yields resulting from the addition of microbial cultures to the rhizosphere, inconsistencies in plant growth are often noted and in most cases lacking in reproducibility. To eliminate this inconsistency and to enhance their reproducibility, precursor-inoculum interactions have proven successful. The production of a specific plant hormone via a microbial trans- formation of an available precursor can be con- trolled to which a plant may show a response. In our laboratory, we have successfully established this concept.

Auxins

Indole-3-acetic acid (IAA) is one of the most physiological active auxins. IAA is a common product of L-tryptophan (TRP) metabolism by soil fungi (Gruen, 1959) and bacteria including those which can stimulate plant growth (Lynch, 1985). Frankenberger and Brunner (1983) detected three separate indoles in soil by high performance liquid chromatography (HPLC)- mass spectrometry, including indole-3-acetamide (IAM), indole-3-pyruvic acid (IPyA) and IAA when incubated with L-TRP. Based upon these results, they suggested that IAA is synthesized by two distinct pathways: (i) TRP is converted to

IAA by deamination to IPyA followed by de- carboxylation to indole-3-acetaldehyde (IAAld) which is further oxidized to IAA and (ii) TRP is decarboxylated to IAM which is hydrolyzed to IAA (Fig. 1). Since IAAld is the immediate substrate of IAA (Libbert and Brunn, 1961), the precursor preceding IAAId could either be tryp- tamine (TAM) or IPyA. TAM has been reported in some microbial cultures (Perley and Stowe, 1966a,b). IPyA is the most probable intermediate arising from L-TRP by an a-ketoglutarate-dependent transaminase reaction (Frankenberger and Poth, 1988; Gamborg and Wetter, 1963; Kaper and Veldstra, 1958; Sembdner et al., 1980; Truelsen, 1972). Since this reaction is one of the first essential steps in the synthesis of auxins, any parameter which affects the transaminase rate would have an overall influence on the auxins released as sec- ondary metabolites. Frankenberger and Poth (1988) separated and purified a soluble a-ketoglutarate-dependent L-TRP transaminase enzyme from cell-free extracts of a rhizobacterial isolate associated with Festuca octoflora Walt. This enzyme catalyzed the conversion of L-TRP to IPyA. Using L-TRP as the substrate in the presence of a-ketoglutarate, the average K m was 1.22mM and the activation energy 21.0kJ mo1-1. Frankenberger and Poth (1987a) also de- monstrated the ability of a rhizobacterium, fluorescent pseudomonad isolated from a grass subfamily Festucoideae, to convert L-TRP into IAA in culture medium by ion suppression reverse-phase HPLC.

Auxins in soil can only influence plant growth if subject to direct uptake by the plant and not metabolized by other soil microorganisms. A precursor-inocuum interaction was recently established by Frankenberger and Poth (1987b) who conducted a greenhouse experiment to test the response of Douglas fir to inoculation of an ectomycorrhizae, Pisolithus tinctorius and the I A A precursor, TRP. Prior to initiating this study, they confirmed the ability of this ectomycorrhizae to derive IAA from L-TRP in culture medium by thin-layer chromatography, HPLC, enzyme-linked immunosorbent (mono- clonal antibody) assay (ELISA) and unequivocal identification by gas chromatography mass spectrometry. After one growing season, P. tinctorius stimulated the growth of potted seedlings of

Micro-organisms and plant hormones 3

NH2 cH2CHCOOH

H

TRP 'X~

OH

H

ILA

U 0 |

• I~--'-N'~C I"¢2 (;O~H

H

IPyA /

~ CH2CH2NH2

H

TAM

~ CH2C -- N

H

IAN

~ CH2CHEOH

H

TOL

tl ~C~CHO

H IAAId

.~ ~CHICOOH

H IAA

TAP, I~plophan IPyA. Indole-3.pyruvlc acid IAAId. Indole-3-1celaldehyde

--~CHcDCONH 2 ILA, Indole-3-1acl|c acid TAM, IWpla~nlne

H TOL, Iryplophol IAN, Indole-3-lcelonll rile

ZAM IAM, Indole-3-acetarnlde

Fig. 1. Proposed pathways for the biosynthesis of indole-3-acetic acid in soil (IAA) (Frankenberger and Brunner, 1983).

Douglas fir only when supplied with low concentrations of TRP (10 -8 to 10 -6 M, 0.34 to 34/xg kg -~ soil). Analyses of variance for seedling height, stem diameter, shoot-root dry weight, and root/shoot ratios showed significant differ- ences among the treatments (Table 1). There was basically no difference in growth between the inoculated and uninoculated treatments; however, the addition of a dilute solution of TRP (10 -8 to 10 -6 M, 0.34 to 34/xg kg -~ soil) with P. tinctorius promoted a dramatic increase, indicating a physiological rather than a nutrition- al effect. In regards to the biomass of shoots and roots, a similar trend was observed with enhanced yield among the 10 -7 and 10-SM TRP treatments.

A higher concentration of TRP applied at 10-3M inhibited the dry weights of the seedlings. The most dramatic influence among the interaction of P. tinctorius and TRP was the seedling root length, reflecting a phytohormone effect, with elongation being promoted upon 10 -8 to 10 6 M TRP applications. Root examina- tion revealed that the mycelial inoculum of P.

tinctorius was highly effective in establishing col- onization but it did not influence plant growth, unless TRP was provided.

Cytokinins

Cytokinins are found in both microbial and plant tissues. Investigations on microbial production of cytokinins have been focused primarily on plant pathogens. The isolation and quantification of cytokinins in nonpathogenic soil bacteria, as well as in soil, have received very little attention with usually cytokinin-like bioactivity via bioassays being reported (Nieto and Frankenberger, 1990). Among the biosynthetic pathways which have been proposed, adenine (ADE) is considered the common precursor for cytokinin production in plant and microbial tissues. Van Andel and Fuchs (1972) had reported the presence of cytokinins in 11 bacteria and 4 fungi. Since Van Andel's and Fuch's report, cytokinins have been detected in the following bacterial cultures, Agrobacterium tumefaciens, Azotobac-


Table 1. Influence of Pisolithus tinctorius (Pt) and tryptophan (TRP) on various growth parameters of Douglas fir (Franken- berger and Poth, 1987b)

Treatments Shoot Root Shoot Root Root / shoot Stem height length weight weight ratio diameter (cm) (cm) (g) (g) (ram)

Control 2.3 a* 42.6 ab 3.6 b 3.5 ab 0.97 0.79 abc Pt (inoculation) 2.3 a 40.5 ab 3.6 b 3.6 ab 1.00 0.80 abc Pt + 10 -3 M TRP 2.6 ab 32.2 a 2.8 a 2.9 a 0.98 0.50 a Pt + 10 4 M TRP 2.5 ab 38.8 ab 3.6 b 3.8 ab 1.02 0.75 abc Pt + 10 5 M TRP 4,5 bc 40.6 ab 3.8 b 4.0 b 1.01 0.82 abc Pt + 10 6 M TRP 3.6 abc 53.0 c 4.3 bcd 5.0 c 1.13 1.04 bc Pt + 10 -v M TRP 4.1 abc 54.6 c 4.7 cd 5.4 cd 1.12 1.07 bc P t+ 10 8 M T R P 5.1c 54.3c 5.1d 5.9d 1.14 1.12c

* Treatments followed by the same letter are not significantly different at the 0.05 level according to the Duncan multiple range test.

ter chroococcum, Bacillus cereus mycoides, and Lactobacillus acidophilus, and the fungi Taph- rina spp., Taphrina cerasi and Rhizopogon och- raceorubens, but there is little available evidence on the effect of these exogenously produced cytokinins on plant growth (Lynch, 1985).

Azotobacter chroococcum (ATCC 9043), A. vinelandii (ATCC 12981), A. beijerinckii (ATCC 12837), and two Pseudomonads (P. fluorescens, P. putida) species, enriched from the rhizosphere of a common grass, Festuca, were tested for their ability to produce cytokinins in culture media (Nieto and Frankenberger, 1989a). Among several purine ring constituents and isoprenoid com- pounds tested for their ability to stimulate bacterial cytokinin production, ADE and isopentyl alcohol (IA) enhanced cytokinin bioactivity to the greatest degree in the culture filtrates of all the bacteria tested. A. chroococcum was the most prolific producer of cytokinins. A treatment of 10-SM ADE and 10-3M IA, 200mg g-1 sucrose, 10-SM KNO3, pH 6.5, 32°C and shaken aerated conditions were found to be op- timum. Nieto and Frankenberger (1988) utilized HPLC/UV spectrometry for the identification and quantification of cytokinins produced by A. chroococcum (Fig. 2).

The isolation, quantification and bioactivity of cytokinins in soil have received very little attention. One of the major difficulties impeding soil analysis of plant growth regulators is that of sensitive and selective chemical detection. Nieto and Frankenberger (1989b) detected cytokinins in an Arlington soil. Various concentrations of ADE were applied as soil treatments, separately

and in combination with 10-3M IA and an inoculum of Azotobacter chroococcum, over a period of 7 days. Samples were filtered and cytokinins were detected by HPLC/UV spectrometry and the radish cotyledon bioassay. Only two cytokinins, t-zeatin (t-io6Ade) and zeatin riboside (i06Ado), were detected in any appreciable amounts. A treatment of 10 -6 M ADE enhanced cytokinin levels in soil to the greatest degree at 5 days post-treatment, whether applied alone or in combination with IA or an inoculum of A. chroococcum. Results indicated that both the indigenous microflora and A. chroococcum were capable of producing io6Ado; however, A. chroococ- cure was more effective in its production.

Ethylene

Ethylene (C2H4) is well known for its profound effects on plant growth. The biological production of C 2 H 4 is not limited to higher plants, since both pathogenic and nonpathogenic microorganisms are capable of synthesizing C 2 H 4.

Methionine (MET) is the sole precursor for C2H 4 biosynthesis in plants (Adams and Yang, 1979). In combination with glucose as an energy source, MET serves as an effective substrate for the microbial production of C 2 H 4 in a wide diverse group of fungi, yeast and bacteria (Ar- shad and Frankenberger, 1989; Dasilva et al., 1974; Lynch, 1972; Lynch and Harper, 1974a,b; Primrose, 1976; Primrose and Dilworth, 1976; Swanson et al., 1979; Thomas and Spencer, 1977). Arshad and Frankenberger (1989) ob-

Micro-organ&ms and plant hormones 5

I. i o6Ado t - i o 6 A d e c- io6Ade i 6 A d e

S F A N D A R D

. . . . . . . . St MPLE

,-I," 2 0 0 2 0 0 .

~*-'" i' X(nm) ~ / X (nrn) m

i ~ I I / I//~ , SAMPLE: BUTANOL I I I 1 1 / l i A EXTRACT OF CULTURE

~ J~.[~, . . - ~ L J SUPERNATANT FROM , I~" U ~ ' ~ d q ~ - w ~ ' ~ A z o f o b o c f e r chroococcum

~ L/"- tlJ

I I I I I 0 5 I0 15 20

ELUTION TIME (rain) Fig. 2. HPLC chromatogram of a culture supernatant of dihydrozeatin riboside; t-io6Ade, t-zeatin; c-i06Ade, c-zeatin;

I 1 I 25 30 35

Azotobacter chroococcum, io~Ado, zeatin riboside; H2-ioOAdo, i6Ade, isopentenyl adenine (Nieto and Frankenberger, 1988).

served that corn rhizosphere contained an appreciable number of microflora capable of deriv- ing C2H 4 from L-MET. Among the soil fungal isolates, Acremonium falciforme was an efficient C2H 4 producer. However, it followed a different pathway of C2H 4 biosynthesis from that eluci- dated in higher plants. We found that amino acids, organic acids and carbohydrates typically found in root exudates stimulate C z H 4 biosynthesis in soil (Arshad and Frankenberger, 1990). This indicates that an excellent site for microbial biosynthesis of C2H 4 is the rhizosphere, where substrate levels and microflora populations are comparatively very high.

Ethylene has been identified as a common component of the soil atmosphere and under certain conditions has been shown to reach a concentration high enough to influence plant growth and development (Smith, 1976). Ethylene concentrations as low as 10 ppb can

evoke plant responses and concentrations of 25 ppb result in decreased fruit and flower development (Primrose, 1979). Jackson and Campbell (1975) were the first to demonstrate rapid movement of radiolabeled [~4C]-C2H4 applied to the roots of tomato plants producing a specific epi- nastic response. However, little direct evidence is available in the existing literature on the microbial influence of CzH 4 production on plant growth. Arshad and Frankenberger (1988) de- monstrated the influence of microbial produced L-MET derived C z H 4 on etiolated pea seedlings by observing the classical 'triple' response which includes reduction in elongation, swelling of the hypocotyl, and a change in the direction of growth (horizontal) (Fig. 3). Etiolated pea seedling responded to C2H 4 derived from L-MET by the inoculum, A. falciforme (Table 2) and the soil indigenous microflora (Table 3). A similar response was observed when seedlings were ex-


Fig. 3. Response of etiolated pea seedlings to exogenous L-methionine-derived C,H, produced by A. falciforme in the absence of AgNO, (Arshad and Frankenberger, 1988).

posed to C2H, gas. The specific anti-C,H, prop- erties of Ag(1) confirmed that C,H, was the fungal metabolite responsible for the observed effects. Moreover, the culture was incubated outside of the root zone and thus C,H, derived from L-MET in the medium affected the growth of pea seedlings.

Conclusions

The studies reported here reveal the ability of specific inocula as well as the soil indigenous microflora to derive plant hormones from precursors provided in pure culture and in soil. Inoculation with specific organisms may prove

Table 2. Influence of L-methionine-derived C,H, produced by A. falciforme on etiolated pea seedlings grown in autoclaved soil (Arshad and Frankenberger, 1988)

Treatment* Seedling Seedling length (cm) diameter (mm)

Control 6.23 b*** 1.91 ab AgNO, (240 mg L-l)** 1.51 b 1.87a Inoculated (A. falciforme) 6.67 b 2.07 b L-Methionine (10 mM) 7.22 b 2.10 b L-Methionine (10 mM) + inoculation 2.58 a 2.44~ L-Methionine (10 mM) + AgNO, (240 mg L-l) 6.11 b 1.93 ab L-Methionine (10 mM) + inoculation

+ AgNO, (240 mg L-r) 5.77 b 2.07 b

* Samples that did not receive AgNO, received NaNO, (240mgL-‘) ** AgNO, is an inhibitor of C,H, action. *** Values followed by the same letter were not significantly different at the 0.05 level according to the Duncan multiple range

test.

Micro-organisms and plant hormones 7

Table 3. Influence of L-methionine-derived CzH 4 produced by soil indigenous microflora on etiolated pea seedlings (Arshad and Frankenberger, 1988)

Treatment* Seedling Seedling length (cm) diameter (mm)

Control AgNO 3 (240 mg L -i) L-Methionine (5 raM) L-Methionine (5 mM) + AgNO~ (240 mg L ') L-Methionine (10 raM) L-Methionine (10 mM) + AgNO 3 (240 mg L- ~ )

6.56 b** 1.87 a 13.50 d 1.93 ab 5.14 ab 2.49 c

l l .10c 2.06 ab 3.90 a 2.75 d

10.10c 2.11 b

* Samples that did not receive AgNO 3 received NaNO 3 (240 mg L ~) ** Values followed by the same letter were not significantly different at the 0.05 level according to the Duncan multiple range

test.

beneficial in affecting plant growth in some cases but results strongly support our hypothesis that inoculation in the presence of a specific physiological precursor is more effective than inoculation alone. Similarly, precursor-inoculum interactions are often much better in creating plant responses than the application of precursors alone, indicating the superiority of particular inocula over the soil indigenous microflora in synthesizing PGRS. Plant responses may be con- trolled by varying the amount of precursors applied since this response is concentration dependent. Rhizosphere microflora should be exten- sively screened for their ability to synthesize plant hormones from a given substrate.

References

Adams D O and Yang S F 1979 Ethylene biosynthesis: identification of l-aminocylopropane-l-carboxylic acid as an intermediate in the conversion of methionine to ethylene. Proc. Natl. Acad. Sci. USA 76, 170-174.

Andel O M van and Fuchs A 1972 Interference with plant growth regulation by microbial metabolites. In Phytotoxins in Plant Disease. Eds. R K S Wood, A Ballic and A Graniti. pp 227-249. Academic Press, London.

Arshad M and Frankenberger W T Jr 1988 Influence of ethylene produced by soil microorganisms on etiolated pea seedings. Appl. Env. Microbiol. 54, 2728-2732.

Arshad M and Frankenberger W T Jr 1989 Biosynthesis of ethylene by Acremonium falciforme. Soil Biol. Biochem. 21, 633-638.

Arshad M and Frankenberger W T Jr 1990 Ethylene accumu- lation in soil in response to organic amendments. Soil Sci. Soc. Am. J. (in press).

Barea J M and Brown M E 1974 Effects on plant growth produced by Azotobacter paspali related to synthesis of plant growth regulating substances. J. Appl. Bacteriol. 37, 583-593.

Brown M E and Burlingham S K 1968 Production of plant growth substances by Azotobacter chroococcum. J. Gen. Microbiol. 53, 135-144.

Dasilva E J, Henriksson E and Henriksson L A 1974 Ethylene production by fungi. Plant Sci. Len. 2, 63-66.

D6bereiner J, Marriel I E and Nery M 1976 Ecological distribution of Spirillum lipoferum. Beyerinck. Can. J. Microbiol. 22, 1464-1473.

Frankenberger W T Jr and Brunner W 1983 Methods of detection of auxin-indole-3-acetic acid in soils by high performance liquid chromatography. Soil Sci. Soc. Am. J. 47,237-241.

Frankenberger W T Jr and Poth M 1987a Determination of substituted indole derivatives by ion suppression-reverse- phase high-performance liquid chromatography. Anal. Biochem. 165, 300-308.

Frankenberger W T Jr and Poth M 1987b Biosynthesis of indole-3-acetic acid by the pine ectomycorrhizal fungus Pisolithus tinctorius. Appl. Environ. Microbiol. 53, 2908- 2913.

Frankenberger W T Jr and Poth M 1988 L-tryptophan transaminase of a bacterium isolated from the rhizosphere of Fustuca octoflora (Graminae). Soil Biol. Biochem. 2ll, 299-3(14.

Gamborg O I and Wetter L R 1963 An aromatic amino acid transaminase from mung bean. Can. J. Biochem. Physiol. 41, 1733-1740.

Gruen H E 1959 Auxins and fungi. Annu. Rev. Plant Physiol. 10, 405-440.

Hubbell D H, Tien T M, Gaskin M H and Lee J 1979 Physiological interaction in the Azospirillum- grass root association. In CRC Associative Symbiosis. Eds. P B Vose and A P Ruschel. 1, 1-6.

Hussain A, Arshad M, Hussain A and Hussain F 1987 Response of maize (Zea mays) to Azotobacter inoculation under fertilized and unfertilized conditions. Biol. Fertil. Soils 4, 73-77.

Jackson M B and Campbell D J 1975 Movement of ethylene from roots to shoots, a factor in the responses of tomato plants to waterlogged conditions. New Phytol. 74, 397- 406.

Jagnow G 1987 Inoculation of cereal crops and forage grasses with nitrogen-fixing rhizosphere bacteria: Possible causes

8 Micro-organisms and p lan t h o r m o n e s

of success and failure with regard to yield response- A review. Z. Pflanzenernaehr. Bodenkd. 150, 361-368.

Kaper J M and Veldstra H 1958 Metaboism of tryptophan by Agrobacterium tumefaciens. Biochim. Biophys. Acta 30, 401-420.

Lee M, Breckenridge C and Knowles R 1970 Effect of some culture conditions on the production of indole-3-acetic acid and a gibberellin-like substance by Azotobacter vinelandii. Can. J. Microbiol. 16, 1325-1330.

Libbert E and Brunn K 1961 Nachweis von indol-3-brenz- trauben saure und indol-3-athanol (tryptophol) bei der enzymatischen auxin bildung aus tryptophan in vitro. Naturwissenschaft 48, 741.

Lynch J M 1972 Identification of substrates and isolation of microorganisms responsible for ethylene production in the soil. Nature 240, 45-46. London.

Lynch J M 1985 Origin, nature and biological activity of aliphatic substances and growth hormones found in soil. In Soil Organic Matter and Biological Activity. Eds. D Vaughan and R E Malcolm. pp 151-174. Martinus Nijhoff/ Dr W Junk Publishers. Dordrecht/Boston/Lancaster.

Lynch J M and Harper S H T 1974a Formation of ethylene by a soil fungus. J. General Microbiol. 80, 187-195.

Lynch J M and Harper S H T 1974b Fungal growth rate and the formation of ethylene in soil. J. Gen. Microbiol. 85, 91-96.

Nieto K F and Frankenberger W T Jr 1988 Determination of cytokinins by ion suppression-reverse phase high performance liquid chromatography. J. Liq. Chrom. 11, 2907- 2925.

Nieto K F and Frankenberger W T Jr 1989a Biosynthesis of cytokinins produced by Azotobacter chroococcurn. Soil Biol. Biochem. 21, 967-972.

Nieto K F and Frankenberger W T Jr 1989b Biosynthesis of cytokinins in soil. Soil Sci. Soc. Am. J. 53, 735-740.

Nieto K F and Frankenberger W T Jr 1990 Microbial production of cytokinins. In Soil Biochemistry. Vol. 6. pp 191- 248. Eds. J M Bollag and G Stotzky. Marcel Dekker, New York.

Perley J E and Stowe B B 1966a The production of tryp- tamine from tryptophan by Bacillus cereus (KVT). Bio- chem. J. 100, 169-174.

Perley J E and Stowe B B 1966b On the ability of Taphrina deformans to produce indole acetic acid from tryptophan by way of triptomine. Plant Physiol. 41,234-237.

Phillips D A and Torrey J G 1970 Cytokinin production by Rhizobium japonicum. Physiol. Plant. 23, 1057-1063.

Primrose S B 1976 Formation of ethylene by Escherichia coli. J. Gen. Microbiol. 95, 159-165.

Primrose S B 1979 A review, ethylene and agriculture: the role of the microbes. J. Appl. Bacteriol. 46, 1-25.

Primrose S B and Dilworth M J 1976 Ethylene production by bacteria. J. Gen. Microbiol. 93, 177-181.

Puppo A and Rigaud J 1978 Cytokinins and morphological aspects of French-bean roots in the presence of Rhizobium. Physiol. Plant. 42, 205-206.

Sembdner G, Gross D, Liebisch H W and Schneider G 1980 Biosynthesis and metabolism of plant hormones. In Hor- monal Regulation of Development. I. Molecular Aspects of Plant Hormones. Ed. J MacMillan. pp 281-444. Spring- er, New York.

Smith A M 1976 Ethylene in soil biology. Annu. Rev. Phytopathol. 14, 53-73.

Swanson B T, Wilkins H F and Kennedy B 1979 Factors affecting ethylene production by some plant pathogenic bacteria. Plant and Soil 51, 19-26.

Thomas K C and Spencer M 1977 L-methionine as an ethylene precursor in Saccharomyces cerevisiae. Can. J. Microbiol. 23, 1669-1674.

Tien T M, Gaskins M H and Hubbell D H 1979 Plant growth substances produced by Azospirillum bransilense and their effect on the growth of pearl millet (Pennisetum americanum L.). Appl. Env. Microbiol. 37, 1016-1024.

Truelsen T A 1972 Indole-3-pyruvic acid as an intermediate in the conversion of tryptophan to indole-3-acetic acid. I. Some characteristics of tryptophan transaminase from mung bean seedling. Physiol. Plant. 26, 289-295.

Documents

Microbial production of plant hormones