I= USE OF GROWTH-PROMOTING BACTERIA IN THE CONTROL OF ROOT -KNOT NEMATODE AND ROOT-INFECTING FUNGI OF CROP PLANTS
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Title of Thesis
USE OF GROWTH-PROMOTING BACTERIA IN THE CONTROL OF ROOT -KNOT NEMATODE AND ROOT-INFECTING FUNGI OF CROP PLANTS

Author(s)
Imran Ali Siddiqui
Institute/University/Department Details
Department Of Botany/ University Of Karachi
Session
2000
Subject
Botany
Number of Pages
359
Keywords (Extracted from title, table of contents and abstract of thesis)
bacteria, root knot, nematode, fungi, crop plant, pseudomonas aeruginosa, bacillus subtilis, okra, sunflower, wheat, uridbean, fenugreek, lucerne, peanut, cotton, sesbania sesban, lettuce, soybean, tomato, corn, spinach, meloidogyne javanica, l. aeruginosa, b. subtilis, f. oxysporum. f. solani, r. solani

Abstract
Thirty two isolates of Pseudomonas aeruginosa and a strain of Bacillus subtilis were isolated from the rhizosphere of 4 wild plants such as Cynodon dactylon (L.) Peers., Setarria vertieellata (L.) P. Beauv., Cyprus rotundus L., Eriocloa procera (Retz.) C.E. Hubb., and 15 cultivated plants like okra (Abelmoschus esculentus (L.) Moench), sunflower (Helianthus annuus L.), wheat (Triticum aestivurn L.), uridbean (Vigna mungo (L.) Hepper), fenugreek (Trigonella foenum-graecum L.), lucerne (Medicago staiva L.), peanut (Arachis hypogea L.), cotton (Gossypillm arboreum L.), Sesbania sesban L., sorghum (Sorghum bicolor L.), lettuce (lactuca sativa L.), soybean [Glycine max L. (Mere)], tomato (Lycopersicon esculentum MilL), corn (Zea mays L.) and spinach (Spinacea oleracea L.) growing at the Karachi University Campus, Malir, Darsanochanoo, Kathore, Thatta and Tandojam.

In in vitro studies, cell-free culture filtrates of some isolates of l. aeruginosa and B. subtilis inhibited egg hatching and showed nematicidal activity against Meloidogyne javanica, root-knot nematode. The cell-free culture filtrate of P. aeruginusa strain 1E-6 and its ethyl acetate and hexane extracts showed significant mortality of M javanica larvae with maximum mortality when ethyl acetate extract at 1.0 mg/ml was used indicating that active compound responsible for nematicidal activity was soluble in ethyl acetate and hence may be intermediary in polarity. Active nematicidal compound was heat labile and hence may be pfoteinaceous or glycoproteinaceous in nature. Rhizobacterial strains beside suppressipn of root-knot nematode also inhibited radial growth of root-infecting fungi viz., Macrophomina phaseolina, Fusarium oxysporum, F '. ,solani and Rhizoctonia solani producing zones of inhibitions. Some isolates/strains of p'. aeruginosa also lysed the fungal hyphae. In dual culture plate assay, rhizobacterial strains showed variability in antagonistic activity against root-infecting fungi.

In the greenhouse and field experiments, seed treatment, bare-root-dip treatment and soil drench with strains/isolates of p'. aeruginosa and B. subtilis significantly controlled the infection of M. javancia .Phaseolina,. F. oxysporum. F. solani and R. solani on leguminous plants like mungbean [Vigna radiata (L.) Wilczek and uridbean [Vigna mungo (L.) Hepper] and tomato (lycopersicon esculentuml MilL) used as non-leguminous test crop. Strains of P. aeruginosa and B. subtilis also increased plant growth and yield. Numbers of nodules per root system in leguminous plants were also significantly higher in the presence of plant growth-promoting rhizobacterial (PGPR) strains. Of the P. aenrginosa strains used, strain IE6 was found as the most potent isolate in the suppression of root-knot and root-rot diseases and its efficacy was similar to that of Carbofuran an nematicide. PGPR showed better biocontrol and growth promoting effects in 15-day-old plants as compared to plants harvested after 30 and 45 days.

Activity of culture filtrate was greater compared to the dead cells of B. subtilis whereas viable and heat-killed cells of P. aeruginosa were found more effective then culture filtrate in reducing nematode penetration. Application of bacteria in the root zone showed better results in reducing nematode invasion into tomato roots as compared to their application on leaf or in stem. Aqueous cell suspension of P. aeruginosa was found more effective as compared to cell free culture filtrate in the control of root-infecting fungi and the root-knot nematode in tomatos. Root-rot and root-knot disease progressively decreased with an increase in filtrate concentration.

Highest inoculum level of P. aeruginosa (7.4x 108 cfu/ml) in the presence of lowest population density of M javanica (500 J2/plant) showed maxi/num inhibition on root-knot development in tomato. An inoculum level 2.5x 108 cfu/ml of the bacterium was found as optimum for the enhancement of tomato growth. P. aeruginosa was reisolated from rhizosphere and inner tissues of root and shoot. Bacterial population was much higher in the rhizosphere as compared to inner tissues of plant root and shoot. Rhizosphere colonization by P. aeruginosa seems to be governed by : i) initial inoculum size and ii) severity of the root-knot disease. Endoroot and endoshoot colonization was dependent on degree of root colonization by Fusarium oxysporum. A higher inoculum level of R. solani (3 ml/kg soil) and population density of M. javanica (2000 eggs/pot) resulted in 100% death of tomato seedlings in sterilized soil. Infected plants treated with P. aeruginoso showed slower disease progression and continued to grow until the harvest time. Biocontrol and growth promoting potential of the bacterium enhanced when kept at 50% or 75% moisture holding capacity where as 25% MHC reduced bacterial efficacy.

P. aerllginosa when used alone or in combination with fungal antagonists viz., Trichoderma spp., Verticillium chlamydopsorium, Paecilomyces Iilacinus, Talaromyces flavus, Slachybotrys atra and Bradyrhizobia the root-nodule bacteria showed sih'T1ificant suppression of infection caused by M. phaseolina, F. solani and R. solani, the root-infecting fungi and M javanica, the root-knot nematode m mungbean, uridbean, tomato and chili. In dual culture growth of V. chlamydosporium, P. lilacinus, T. flavus and T. harzianum was inhibited by P. aeruginosa whereas S. atra was not inhibited by the bacterium. Combined use of P. aeruginosa and microbial antagonists showed better control of root-rot and root-knot diseases then their separate use. P. aeruginosa used with either V. chlamydosporium or P. lilacinus reduced population buildup of M. javanica in tomato under field conditions. Co-inoculation of P. aeruginosa with fresh isolate of rhizobia showed better results in the control of root rot-root knot disease complex and produced greater number of nodules with enhancement in plant growth as compared to isolates obtained from culture collection. Fresh isolate of Bradyrhizobium spp., showed slight reduction in population of P. aeruginosa in the rhizosphere but not to an extant that could reduce biocontrol and growth promoting effects of the bacterium.

P.aerugnosa strains (IE-6 and Pa-7) used with 3 fungicides viz., Captan, Benlate and Bevistin significantly suppressed root-rot and root-kpot infection in chickpea (Cicer arietinum L.). Fungicide Bavistin was found highly toxic to one strain of P. aeruginosa as compared to other pesticides used. Captan and Bavistin had no additive effect on P. aerl/ginosa in reducing root-knot infection whereas Benlate and Carbofuran, a nematicide enhanced the activity P. aeruginosa in the control of root-knot nematode.

Ethanolic extract of neem cake and brown seaweeds viz., Stoechospermum marginatum and Sargassum tenenrrimum caused significant mortality of M. .javanica larvae in vitro. Soil amendment with neem cake, Datura fastuosa or brown seaweeds alone or in combination with PGPR strains significantly suppressed root-rot and rootknot infection in uridbean and root-knot infestation and nematode soil population in tomato. No adverse effect of organic amendments was observed on bacterial survival in the rhizosphere. Use of P. aeruginosa alone or in combination with urea at 0. I 5 g/kg soil and or potash at 0.1 g/kg significantly suppressed root-rot and root-knot infection on mungbean roots. Highest reduction in gall formation was recorded in treatment where P. aeruginosa strain IE-6 was used in combination with urea and potash. Potash used alone increased root-knot infection induced by M javanica.

P. aeruginosa was found to survive for 244 days on talc powder. Ta1c-based inoculum of the bacterium when incorporated in soil at rates of 3% w/w significantly controlled root-rot and root-knot diseases with improvement in growth of tomato under greenhouse and field conditions.

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1 0 Contents
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2 1 Introduction 1
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3 3 Experimental Results
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4 4 Discussion
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5 5 References
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  5.1 Appendices Acknowledgments
  5.2 List Of Publications Selected Published Papers