Interactions of Azospirillum brasilense and Bacillus megaterium var. phosphaticum and Glomus

fasciculatum in the rhizosphere of tomato plants were studied. There was no significant difference in plant

growth parameters between YAM-inoculated treatments. Plants inoculated with the phosphobacteria had

significantly higher shoot length, which was equivalent to VAM and other VAM + phosphobacteria combinations.

Azospirullum or phosphobacteria alone increased plant biomass compared with the uninoculated control. VAM

index was significantly reduced with the addition of phosphobacteria. There was no difference in tissue nutrient

concentrations between treatments

K., Udaiyan

S., Manian

Thomson T., Edathil

Universiti Putra Malaysia Institutional Repository

PertanikaJ. Trop. Agric. Sci. 18(3): 187-191 (1995) ISSN: 0126-6128© Universiti Pertanian Malaysia PressEffect of Interactions of Three Growth-promoting Microorganisms on VAMColonization, Spore Density, Plant Growth and Nutrient Accumulation inTomato (Lycopersicon esculentum) SeedlingsTHOMSON T. EDATHIL, S. MANIAN and K. UDAIYANMicrobiology Unit, Dept. of BotanyBharathiar UniversityCoimbatore 641 046 Tamil Nadu, IndiaKeywords: vesicular-arbuscular mycorrhiza, Azospirillum brasilense, Bacillus megaterium var. phosphaticum,Lycopersicon esculentumABSTRAKKajian dibuat terhadap interaksi Azospirillum brasilense dan Bacillus megaterium var. phospharicumdan Glomus fasciculatum dalam rizosfera tanaman tomato. Tidak terdapat perbezaan yang signifikan dalamparameter pertumbuhan tanaman antara rawatan-rawatan inokulat - VAM. Tanaman-tanaman yang dinokulatdengq,n fosfobakteria jelas mempunyai tunas yang l.ebih panjang yang mana bersamaan dengan VAM dancantuman VAM + fosfobakteria yang lain. Cuma Azospirullum atau fostobakteria sahaja menambahkanbiojisim tanaman dibandingkan dengan kawalan tak inokulat. Indeks VAM jelas menurun dengan penambahanfosfobakteria. Pemekatan nutrien tisu tidak berbeza antara rawatan.ABSTRACTInteractions of Azospirillum brasilense and Bacillus megaterium var. phosphaticum and Glomusfasciculatum in the rhizosphere of tomato plants were studied. There was no significant difference in plantgrowth parameters between YAM-inoculated treatments. Plants inoculated with the phosphobacteria hadsignificantly higher shoot length, which was equivalent to VAM and other VAM + phosphobacteria combinations.Azospirullum or phosphobacteria alone increased plant biomass compared with the uninoculated control. VAMindex was significantly reduced with the addition ofphosphobacteria. There was no difference in tissue nutrientconcentrations between treatments.INTRODUCTIONInteractions of growth-promoting microbialpopulations in the rhizosphere of VA-mycorrhizal plants have been studied by manyworkers (Barea et al. 1983; Pacovsky and Fuller1985; Linderman 1988; Baas 1990). Subba Raoet al. (1985) reported that the synergisticinteractions of VAM and Azospirillum brasilensesignificantly increased dry matter productionand grain yield of barley. Response of plants tocolonization by mycorrhizas depends on manybiotic and environmental factors. Plant-availableP is considered to influence the degree ofmycorrhizal symbiosis (Bethlenfalvay et al. 1982).Among the many soil microorganisms knownto solubilize unavailable forms of P,phosphobacteria have been used as bacterialfertilizer (Bagyaraj 1984). These bacteria survivefor a longer period in the rhizosphere ofmycorrhizal roots (Linderman 1988). Hencethis trial aimed to study the interactions ofVAM fungus with Azospirillum andphosphobacteria in rhizosphere soils of tomatoseedlings and their effect on plant growth,tissue nutrient concentration, VAM colonizationand spore density.MATERIALS AND METHODSThe soil used was a nutrient deficient (N 225, P22.5, K 780, Zn 0.20 and Cu 0.78 kg/ha·1) alluvialdeposit of sandy loam with pH 7.2 and EC 0.2milli S/cm·1 from the Bharathiar UniversityTHOMSON T. EDATHIL, S. MANIAN AND K. UDAIYANCampus, Coimbatore. A mixture of equal partsof soil and sand autoclaved at 121°C and lSlb/inch2 (1 h each on three consecutive days,followed by 1 week incubation at roomtemperature) was used to fill 30 x 12 cmpolyethylene bags (about 3 kg per bag). Asbacteria require an organic substratum for initialestablishment in the soil (Lynch 1983; SubbaRao 1993), SO g of autoclaved, (l21°C, IS lb/inch2) dried cowdung was added to the topsoilin each bag.A stock culture of Glomus fasciculatum wasused as VAM inoculum, since it is known thatthis species is most effective in enhancinggrowth and P uptake (Sulochana et al. 1989;Sivaprasad et al. 1992). It was maintained in apot culture of 90-day-old maize. Fresh culturesof Azospirillum brasilense and phosphobacteria,Bacillus megatenum var. phosphaticum (obtainedfrom the Tamil Nadu Agricultural University,Coimbatore) were used as bacterial inocula.Ten grams of VAM inoculum soil, containingapprox. 644 spores (64 spores/g dry soil) alongwith lyphae and infected root fragments/l0 gcharcoal base containing about 109 bacterialcells, (1 g charcoal base containing 108 bacterialcells) were placed as a thin layer about 2 cmbelow the soil surface in the bags. The controlbags received autoclaved inocula. Thetreatments used were: (i) YAM-free (control),(ii) Azospirillum, (iii) phosphobacteria, (iv) VAM,(v) VAM + Azospirillum, (vii) VAM+ Azospirillum+ phosphobacteria. Seeds of tomato (Lywpersiconesculentum Mill.) cv. Co 1 were sown in all thebags at the rate of 10 seeds/barfl. The bagswere kept in a greenhouse, watered regularlyand the seedlings were thinned on the Sth dayafter emergence (DAE) to maintain oneseedling per bag. Each treatment was replicatedfour times.At 60 DAE, the plants were harvested andgrowth parameters such as shoot and root length,leaf area, biomass, tissue nutrient (N, P, K, Znand eu) concentrations, VAM colonization index(VAM!) and spore density were determined.Leaf area was measured using a leaf area meter.Plant biomass was recorded after drying at 60°Cfor 12 h. Determination ofVAMI was done afterstaining the root samples following the methodof Phillips and Hayman (1970) and using thescoring method of Edathil et al. (1994). Sporedensity was assessed using the modified wet-sieving and decanting method (Gerdemann andNicolson 1963) and expressed as the number ofspores per gram of dry soil. Tissue nutrientconcentration was determined following thestandard methods of Jackson (1973).RESULTSThere was no significant difference in the growthparameters of tomato seedlings between VAMtreatments. Phosphobacteria-inoculated seedlingsexhibited the highest shoot length (73 cm),which was equivalent to VAM (67 cm) and VAM+ phosphobacteria (64 cm) combinations.Seedlings inoculated with Azospirillum orphosphobacteria alone had higher biomass thanthe uninoculated control. Leaf area and rootlength were more or less equal in all treatments(Table 1).TABLE 1Effect of interactions of microorganisms in the rhizosphere on the growth of tomato plantsTreatment Leaf area Shoot length Root length Biomass(cm2) (cm) (cm) (g)YAM-free (control) 23.0S' 61.00bc 46.7S'b 2.48dAzospirillum 17.77' 60.2Sbc 33.2Sb 4.23'Phosphobacteria 22.22' 73.00' 43.7S'b 4.12'VAM 20.81" 67.00'b 42.00'b 3.87'bVAM + Azospirillum 19.32' 60.75bc 40.2Sbc 3.01 bdVAM + Phosphobacteria 18.20' 64.7S'b 46.7S'b 3.0SbcdVAM + Azospirillum+ Phosphobacteria 23.1S' 64.S0'b S8.7S' 3.39'bdValues are mean of four replications.Values with the same letter are not significantly different P> O.OS according to Duncan's new multiplerange test.188 PERTANlKAJ. TROP. ACRIC. SCI. VOL. 18 NO.3, 1995EFFECf OF INTERACTIONS OF THREE-GROWfH PROMOTING MICROORGANISMSThe VAM and VAM + Azospirillum-inoculatedplants registered higher VAMI thanphosphobacteria-inoculated treatments. However,spore density was equal in all VAM treatments(Table 2). The accumulation ofN, K, Zn and Cuin plant tissue was equal in all treatments. In thecase of P accumulation, there was no regulartrend (Table 3).DISCUSSIONThe enhancement of plant growth with theaddition of VAM fungi (Nicolson 1960; Koske etal. 1975; Tinker 1975, 1978; Menge et aL 1978;Koske" 1981; Abbott and Robson 1982),Azospirillum (Barea et al. 1983; Pacovsky andFuller 1985; Palanisami 1985; Subba Rao et ai.1985) and phosphobacterium (Graeves andTABLE 2Effect of interactions of microorganisms in the rhizosphere of tomato plants onVAM colonization and spore densityTreatment VAMI(%)Spore Density(individuals/g-I dry soil)YAM-free (control)AzospirillumPhosphobacteriaVAMVAM + AzospirillumVAM + PhosphobacteriaVAM + Azospirillum+ Phosphobacteria62.25- 12.61'69.67- 11.34-49.10bc 11.16-42.51'b 9.23-Values are means of four replicationsValues with the same letter are not significantly different at P>0.05 according toDuncan's new multiple range test.TABLE 3Effect of interactions of microorganisms in the rhizosphere on tissue nutrientconcentrations in tomato plantsTreatment N P K Zn Cu(%) (%) (%) (%) (%)YAM-free (control) 1.82- O.14b 4.3- 0.011' 0.001'Azospirillum 1.99- 0.15-b 4.2- 0.01' 0.0013-Phosphobacteria 2.10- 0.16- 3.6- 0.01' 0.0012-VAM 1.67- O.14b 4.1' 0.01' 0.001'VAM + Azospirillum 1.74- 0.16- 4.4- 0.01' 0.0011'VAM + Phosphobacteria 1.81' 0.12< 4.1' 0.01' 0.0011'VAM + Azospirillum+ Phosphobacteria 2.01' 0.17- 4.2- 0.01' 0.0014-Values are means of four replicationsValues with the same letter are not significantly different at P>0.05 according to Duncan's newmultiple range test.PERTANlKAJ. TROP. AGRIC. SCI. VOL. 18 NO.3, 1995 189THOMSON T. EDATHIL, S. MANIAN AND K. UDAIYANWebley 1965; Bagyaraj 1984; Meyer andLinderman 1986) has been well dccumented.Abbott and Robson (1982) reported that VAMfungi would manifest their performance to agreater extent in low nutrient soils. As sufficientnutrients are available in soil, the test plantscould directly absorb the nutrients rather thandepending on VAM fungi or other rhizospheremicroorganisms. Futhermore, the addition oforganic manure will result in increased soilmicrobial population releasing organic acids suchas lactic acid, all of which have chelatingproperties which will ultimately promote Psolubilization (Sperber 1958a, 1958b; Louw andWebley 1959; Duff et at. 1963; Banik and Dey1982). If nutrient availability, especiallyphosporus, is high, the host plant may show anegative growth response to VAM fungi andVAM colonization can be reduced (Hayman etal. 1975; Johnson 1976; Sparling and Tinker1978; Koide 1991). Data in Table 2 substantiatethis observation because addition ofphosphobacteria significantly reduced VAMindex in tomato plants. The VAMI wassignificantly higher in VAM and VAM +Azospirillum treatments.Higher shoot length was observed in thetreatment with phosphobacteria or itscombination with VAM fungus (Table 1). Shootelongation may be a function of the excretion ofcertain growth promoting substances by thebacteria, because researchers have proved thatthe growth promotion by B. megaterium is mainlydue to their excretion of growth-promotinghormones and vitamins (Banik and Dey 1982;Meyer and Linderman 1986).Tien et al. (1979) reported the productionof plant hormones by Azospirillum. The planthormones present in bacterial cultures mayimprove the formation and development of VAmycorrhiza (Azcon et al. 1978).The unaffected nature of VAMF sporedensity in the rhizosphere of tomato seedlingsco-inoculated with either Azospirillum orphosphobacteria probably indicates the positiveinteractions among these growth-promotingmicr~rganisms (Table 2).Since there is no difference in the tissueconcentrations between the treatments (Table3), it may not serve as an indication of thebeneficial microbial interactions in a nutrient-rich (organic manure amended) soil with regardto nutrient accumulation.REFERENCESABBOTr, L.K. and A.D. ROBSON. 1982. Infectivity ofvesicular-arbuscular mycorrhizal fungi inagricultural soils. Australian Journal ofAgricultural Ressearch 33: 1049-1059.AzCON, R., C. AzCON, G. DE AGUILAR andJ.M. BAREA.1978. Effects of plant hormones present inbacterial cultures on the formation andresponse to VA endomycorrhiza. New Phytology80: 359-364.BAAS, R. 1990. 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(Ag.) thesis, Tamil Nadu Agric. Univ.Coimbatore, India.PHILLIPS, J.M. and D.S. HAYMAN. 1970. Improvedprocedures for clearing roots and staining para-sitic and vesicular-arbuscular mycorrhizal fungifor rapid assessment of infection. Transactionsof the British Mycological Society 55: 158-161.SIVAPRASAD, P., K..K.. SULOCHANA, B. GEORGE and MA.SALA. 1992. Growth and phosphorus uptake ofcashew (Anacardium occidentale L.) as influ-enced by inoculation with VA mycorrhizae.Cashew 6: 16-18.SPARLING, G.D. and P.B. TINKER. 1978. Mycorrizas inPennine grassland. In Endomycorrhizas, ed. F.E.Sanders, B. Mosse and P.B. Tinker, p. 545-560.ew York: Academic Press.SPERBER, J.I. 1958a. The influence of apatidesolubilizing orgnisms in the rhizosphere andsoil. Australian Journal of Agricultural Research91: 778-781.SPERBER,J.I. 1958b. Solution of apatide by soil micro-organisms producing organic acids. AustralianJournal of Agricultural Research 91: 782-787.SUBBA RAO, N.S., K..V.B.R TILAK and C.S. SINGH.1985. Effect of combined inoculation of VAMand Azospirillum brasilense on pearl millet(Pennisetum americanum). Plant Soil 84: 283-286.SUBBA RAo, N.S. 1993. Biofmilizers in Agriculture andForestry. New Delhi: Oxford and IBH Publish-ing, p. 84-101.SULOCHANA, T., C. MANOCHARACHARY and P.R RAo.1989. Growth response and root colonizationin cultivars of sesame to VAM fungi. CurrentScience 58: 519-520.TIEN, T.M., M.H. GASKINS and D.H. HUBBEL. 1979.Plant growth substances by Azospirillumbrasilense and their effect on the growth ofpearl millet (Pennisetum americanum L.). AppliedEnvironmental Microbiology 37: 1016-1024.TINKER, P.B. 1975. Effects of vesicular-arbuscularmycorrhizas on higher plants. Symposia of theSociety for Experimental Biology 29: 325-329.TINKER, P.B. 1978. Effects of vesicular-arbuscularmycorrhizas on plant nutrition and plantgrowth. Physiology of Vegetation 16: 743-761.(Received 28 March 1995)(Accepted 28 February 1996)PERTANIKAJ. TROP. AGRIC. SCI. VOL. 18 NO.3, 1995 191

Effect of interactions of three growth-promoting microorganisms on VAM colonization, spore density, plant growth and nutrient accumulation in tomato (lycopersicon esculentum) seedlings

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Effect of interactions of three growth-promoting microorganisms on VAM colonization, spore density, plant growth and nutrient accumulation in tomato (lycopersicon esculentum) seedlings

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