EFFECT OF ABIOTIC FACTORS ON SPORE ATTACHMENT OF PASTEURIA PENETRANS TO JUVENILES (J2) OF MELOIDOGYNE INCOGNITA

Indian J. Nemato/. 24(2): 195-199 (1994) EFFECT OF ABIOTIC FACTORS ON SPORE ATTACHMENT OF PASTEURIA PENETRANS TO JUVENILES (J2) OF MELOIDOGYNE INCOGNITA RAMESH CHAND AND J.S. GIll. Division of Nematology, Indian Agricultural Rp.search Institute, New Delhi-110012 Abstract: Experimento; were carried out to study the effect of three important abiotic factors viz., temperature, moisture and ph on spore attachment of Pasteuria penetrans. Maximum adherence of spores witj1isi1 average of 25.0 spores/ J2 was observed at 26 C in comparlo;onto 9.68 and 13.38 spores/j2 at 23 and 29 C, respectively. Out of four moisture levels tested for spore attachment, nematode recovery and spore attachment were maximum at a moisture level of 50% (23.03)followed by 13:77, 9.06 and 3.95 at 75, 100 and 25 per cent moisture level respectively. The per cent encumberance at 50 per cent treabnent was higher. However, lowest J2 was recorvered (32.13%) from soil with 25% moisture content and highest recovery (55.80) at 75% moisture treabnent. With regard to ph more spore attachment (28.84 spores/ J2) was observed at 7.6 ph, which decreao;ed to 11.91 and 8.31 sporp.s/j2 in 6.4 and 5.2 ph, respectively. Keywords: Posteu"o penetrons. Meloidogyne incognito. temperature, moisture, ph, spore attachment.. Root-knot nematode (Meloidogyne spp.) is the most common, widely prevalent nematode causing serious problem on large number of crops and infliciting serious yield losses. Although nematicides decrease nematode population quickly, their field application has been almost negligible due to various reasons., the prominent among them being the prohibitive cost and possible environmental pollution. Accordingly, a need was feltto exploit biocontrol agents. Pas teuria penetrans, a mycelial endospore forming bacterial parasite has recently been projected as a biocontrol agent as an alternative to chemical control for managing nematodes (Mankau & Prasad, 1972; Sayre, 1984; Stirling, 1984; Brown and Smart 1984 Maheswari et al., 1988). In India, oq:urrence of strains of P. penetrahs has been reported which differed primarily in respect of pathogenicity (Singh & Dhawan, 1990). One of the first and foremost requirement for the use of P. penetrans as biocontrol agent is to determine its host specificity on the basis of spore adherence to the specific nematode body cuticle. Therefore, the present investigations were undertaken to study the effect of abiotic factors like temperature, mositure and ph on the spore attachment of P. penetrans specific to juveniles (J2) of Meloidogyne incognita. MATERIALS AND METHODS The strain of P. penetrans used in this investigation is specific and pathogenic to the root-knot nematode M. incognita. Two hundred freshly hatched J2 of M. incognta in 4 ml of water were mixed with 20 g of soil in small petridishes (5 cm dia). containing 1.5 x 105 bacterial spore density of P. penetrans. All the treatments were replicated 4 times. Petridishes were incubated at three different temperatures for studying the temperature effect but for studying moisture and ph incubation temperature was kept 26 C as maximum spore adherence was recorded at this temperature. In order to study the effect of soil moisture on spore attachment, 25, 50, 75 and 100

196 RAMESH CHAND AND J.S. GILL per cent of field capacity moisture levels were maintained in petridishes containing bacterial spores and M. incognita J2 in desert soil. These were incubated at 26 C for 72 hours, washed and finally observed for spore attachment. The effect of different soil ph on spore attachment was studied using desert soil. Four different ph levels viz., 4.5, 6.0, 7.5 and 9.0 were set initially and also tested at the end of the experiment. RESULTS AND DISCUSSION Temperature: The spore attachment occurred at all three temperatures with minimum being at 23 C and maximum at 26 C. However, further increase in temperature to 29 C. spore attachment was significantly reduced. All the treatments were significantly different from each other. The average number of spores attached per J2 significantly increased from 9.68 to 25.05 with the increase in t2mperature from 23 to 26 C. The per cent increase of spore attachment over 23 C treatment was 158.7 and 38.22 at 26 C and 29 C, respectively. Further, comparing the spore attachment at 29 C with 26 C, 53.10 per cent reduction was recorded. The maximum number of juveniles carrying bacterial spores was also at 26 C and minimum at 23 C, the latter differed significantly with the former but at par with 29 C treatment (Table 1). The results obtained were at variance from that of earlier workers, who reported maximum spore attachment between 22.5 and 30 C to J2 of M. jauanica (Stirling, 1981) or betweent 25 and 30 C for M. incognita (Maheswary et.al, 1988; Soma Sekhar and Gill 1990). However, the optimum temperah:re of 26 C observed in the present study isvery close to that of Singh and Dhawan (1992)who found that maximum spore attachment occurred at 25 C to Heterodera cajani. These variations in optimum tempeature requirement could be po~ible due to difference in nematode species or race as well as the bacterial strain under test. In present case, the optimum temperature for nematode activity is around 27 C (Taylor, 1968) which corresponds favourably with the spore attachment too. Thus, present findings are more realistic as the optimum temperature range for the spore adherence and also the nematode activity seems very close. TABLE 1. Effect of temperature on spore attachment of P. penetrans on J2 of M. incognita. (Mean of 4 replications) Temperature No. attachedlj2 25.05b 13.38< 0.62 9.68' ofno. spores 87.78' of 91:15'< 6.74 96.15b J2(%) en<:wnbered No. of J2 recovered from soil(%) 53.13 48.38 50.30 NS

ABIOTIC FACTORS INFLUENCE P. PENETRANS SPORE ATTACHMENT 197 Maximum nematode number from the soil was recovered at 23 C followed by 29 C and 26 C. However, the difference among the treatments was statisitically insignificant. Incidently, it was interesting to record that eventhough there was maximum spore attachment and maximum number of J2 getting encumbered at 26 C., the per cent nematode recovery in this treatment was comparatively lower than other treatments. This may be explained keeping in view the observations of Brown and Smart (1985) where the heavily encumbered J2 become sluggish and penetrate the root in lesser number. Furthp.r, it has also been reported that encumbered J2 often showed swarming phenomenon (Bhattacharya and Swarup ( 1988) which impaired their movement and also affected their extraction through the sieves and tissue paper. Moisture: The results presented in Table 2 showed that spore attachment occurred at allmoisture levels, maximum and minimum being at 50 and 25 per cent moisture levels. Further, all the treatments were significantly different from each other. These observations are in accordance with those of Guiran and Demeure (1978) who reported that nematodes are most active in soil with moisture level at 40 to 60 per cent of the field capacity and move through soil in water films. As the soil, either dry or increase in moisture, the nematode activity decreases. Further, the variations of moisture content either towards lower side Le. 25 per cent or higher side Le. 75 percent inhibited spore attachment. This could be explained that at higher moisture level, the nematode movement might be hindered because of water film thickness or pos...,ibly due to reduced activity rate for want of oxygen (Van Gundy, 1965; Wallace, 1966). It is apparent that the nematode activity seemed to be more at 50 per cent moisture level, and this explains for the higher number of bacterial spores getting adhered to juveniles. The number of juveniles encumbered with bacterial spores was also observed to be the highest in 50 per cent moisture treatment which even though was at par with other treatment., but differed significantly from the 25 per cent moisture treatment. The latter having the minimum spores on nematodes. Maximum nematode recovery was in 75 per cent moisture treatment which being at par with other treatment., but differed significantly from 25 per cent moisture level. TABLE 2. Effect of moisture on spore attachment of P. penetrans on second-stage juvenile of M. Incognita. Moisture 25 No. attached;j2 23.03b 13.77c 9.06d 3.95" 1.22 of No. spores 90.46b 82.92" of 6.01 93.56b 94.9Gb52.30b,/2(%) encumberedno. recovered 4.77 32.13" 55.80b 53.00bfromof J2 soil

198 RAMFSHCHANDAND J.S. Glli. TABlE 3. Effectof Fnal 6.4 7.6 8.4 5.2 phspores 28.84<92.77b 21.16d89.61a It.91b on 8.31a85.84a spore 45.63. sou(%) No. recovered 41.80 41.00 NS 87.81a 4.19 J2(%) No. encumbered of attachment No. of from of 2.33 of P. penetrans on second-stage juvenile of M. Incognlta. ph levels ph: The spore adherence increased with increase in ph level and was highest at 7.6 ph (fable 3). Further increase in ph from 7.6 to 8.4 resulted in significantly decrease in spore adherence. All the treatments significantly differed from each other. The per cent increase in spore attachment over the 5.2 ph level was 43.32, 247.05 and 154.63 in 6.4, 7.6, and 8.4 ph levels, respectively. The findings are in confoimity with those of Davies et al. (1988) who observed maximum spore attachment at ph 7;0 and much less at ph 4.0 or9.0. However, Kamra (1993) reported maxim",m spore attachment at ph 8.0 than at ph 4.0 and 6.0. Singh Dhawan (1992) reported more spore attachment in alkaline ph than acidic ones. The percentage. of juveniles encumbered was minimum at lowest ph, which was at par with other treatments, but differed significantly froin 7.6 ph level treatment. The highest percentage of nema10de recove!y was also decreased in ph level 7.6, but such difference among. the treatments was found. to be insignificant. CONCLUSION Inconclusion, it maybe soil temperature, soil moisture and ph that have profound influence on spore attachment of P. penetrans, specific to M. incogn ita juveniles. The optimum value of these abiotic parameters synchronises with the movement and activity of M. incognita J2 under natural conditions, thereby indicating the possible use of this biocontrol agent for root-knot nematode, M. incognita. ACKNOWLEDGEMENT The authors are greatly thankful to Head of the Division of Nematology, IARI, New Delhi for.providing facilities. REFERENCES BhattachaJya. D. and Swamp.G. (1988). Pasteuria penetrans, a pathogenofthe genus Heterodera, its effect on nematode biology and control. Indian J. Nematol. 18: 61-70. Brown, S.M. Bc Smart, G.C. Jr. (1984).AtuiclUnent of Bedll/us penetrans to Meloldogyne I~cognita. Nematroplca 14: 171-172. Brown, S.M. Bc Smart, G.C. Jr. (1985), Root penetrationby Meloldogyne Incognitajuveniles.irlfectedwith&cil/us penetratns. J. Nematol. 17 : 123-126. Davies, K.G., Kerry, B.K. Bc Flynn, C.A. (1988). Observationson the pathogenicityof Pasteuria penetrans, a parasite of root-knot nematode. Ann. appl. BioI.'112: 491-501.

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