x Wavelength PSM Phosphate solubilizing microorganism

Transcription

1 LIST OF ABBREVIATIONS Da Daltons EPS Exopolysaccharide ACC 1-aminocyclopropane-1- by Base pairs carboxylic acid EDTA Ethylenediaminotetraacetic acid oc Degree Celsius HPLC High-performance liquid chromatography Fig Figure IAA Indole-3-acetic acid g Gram IAM Indole-3-acetamide h Hour (s) SAM S-adenosylmethionine ul Microliter TRP Trytophan um Micromolar PGPR Plant growth-promoting mg Milligram rhizobacteria CFU Colony forming units ml Milliliter DNA Deoxyribonucleic acid mm Millimeter UV Ultraviolet mins Minute (s) w/v Weight per volume ns Nonsignificant ANOVA Analysis of variance rpm Revolution per minute Percent spp. Species (plural) Negative sp. Species (single) CAS Chrome S azurol PCR Polymerase chain reaction HCN Hydrogen cyanide N Nitrogen PHA Polyhydroxyalkanoates GC Gas chromatography PPYG Polypeptone yeast extract OD Optical density glucose agar MW Molecular weight PVK Pikovskaya medium ACCD ACC deaminase Fe Iron TLC Thin layer chromatography FMM Fiss glucose minimal medium DHBA 2,3 - dihydroxy benzoic acid SDS Sodium dodecyl sulphate DF Dworkin and Forster minimal medium PAGE Polyacrylamide gel electrophoresis NCBI National center of RNA Ribonucleic acid biotechnology and information PNPP p- nitrophenyl phosphate PNP p- nitrophenol KDa Kilodalton Rf Resolution factor x Wavelength PSM Phosphate solubilizing microorganism

2 LIST OF FIGURES AND TABLES LIST OF FIGURES Introduction Fig. 1 Beach grasses and shrubs growing on sandy dunes Fig. 2 Sand dune vegetation zones Fig. 3a Spinifex littoreus plants growing on the sand dunes on the Aswem Mandrem beach in North Goa Fig. 3b Ipomoea pes- caprae plants growing on the sand dunes on the Miramar beach in North Goa. Fig. 4 Levels of biological organization range from organism to biosphere in the coastal sand dune ecosystem. Chapter 1 Fig. 1.1 Effects of root exudates components on nutrient availability and uptake by plants and rhizosphere microbes Fig. 1.2 Possible plant-microbe interactions affecting plant growth Fig. 1.3 Model explaining PGPR stimulation of plant root elongation Fig. 1.4 The biosynthesis pathway leading to indole-3-acetic acid(iaa) Fig. 1.5 Siderophore mediated uptake of iron into the bacterial cell Chapter 2 Fig. 2.1 Fig. 2.2 Fig. 2.3 Fig. 2.4 Fig. 2.4 Fig. 2.5 Fig. 2.6 Fig. 2.7 Map showing the sampling sites Comparative activity of multiple enzyme production by rhizosphere and endophytic neutrophiles associated with Ipomoea pes- caprae and Spinifex littoreus Neutrophilic bacterial isolates showing production of enzymes Comparative activity of multiple enzyme production by rhizosphere and endophytic alkalophilic associated with Ipomoea pes- caprae and Spinifex littoreus Comparative activity of multiple enzyme production by rhizosphere and endophytic alkalophilic associated with Ipomoea pes- caprae and Spinifex littoreus Alkalophilic bacterial isolates showing production of enzymes Fluorescence exhibited by bacterial isolates grown on E2 medium plate on staining with Nile Blue A The yellow orange halo surrounding the bacterial colony is indicative of the production of an Fe binding compound such as siderophore, which removes Fe (III) from the Fe (III) CAS HDTMA complex in the plate and turns the blue dye to yellow color.

3 Fig. 2.8 Screening of neutrophilic rhizosphere and endophytic bacteria for production of siderophores Fig. 2.9 Screening of alkalophilic rhizosphere and endophytic bacteria for production of Siderophores Fig P solubilizing bacteria producing yellow halo/clear zone on Pikovskaya agar due to production of organic acids Fig Screening of P solubilizing neutrophilic rhizosphere and endophytic bacteria associated with Ipomoea pes- caprae and Spinifex littoreus Fig Screening of P solubilizing alkalophilic rhizosphere and endophytic bacteria associated with Ipomoea pes- caprae and Spinifex littoreus Chapter 3 Fig. 3.1 Fig. 3.2 Fig. 3.3 Fig. 3.4 Fig. 3.5 Cell location of polysaccharides produced by gram-positive and gram negative bacteria. Viscous exopolysaccharide produced by a) Sulphates stained with Alcian Blue (ph 2.5), b) Uronic acids stained with Alcian Blue ( ph 0.5) Scanning electron micrographs of exopolysaccharide formation by Growth curve and EPS production of when grown on glucose (1%) in PPYG Fig. 3.6 EPS production of the when grown on glucose (1%) in PPYG under shaker, shaker+static and static conditions Fig. 3.7 Fig. 3.8 Fig. 3.9 Effect of a) different carbon substrates and b) sucrose concentration on exopolysaccharide production by Effect of yeast extract concentration on EPS production by Effect of a) nitrogen concentration and b) glycine concentration on EPS production by Fig GC analysis profiles of a) Aldoses standard and b) Bacterial EPS Fig X-ray diffraction patterns of the exopolymers Fig Emulsification activity shown by M.arborescens Fig Agarose Gel Electrophoresis of Plasmid DNA from Microbacterium arborescens Fig Aggregation of soils by Fig Standard particle size distribution curves Fig Particle size distribution curves of uninoculated and inoculated sandy soil with Microbacterium arborescen Fig Particle size distribution curves of uninoculated and inoculated field soil with Fig Particle size distribution curves of uninoculated and inoculated mine reject soil with

4 Chapter 4 Fig S rrna base sequence of the selected isolates Fig. 4.2 Unrooted tree showing the phylogenetic relationships of B.subtilis (DQ287964) and members of the genus Bacillus based on 16S rdna sequences. Fig. 4.3 Unrooted tree showing the phylogenetic relationships of Kocuria rosea (DQ287963) and members of the genus Kocuria based on 16S rdna sequences. Fig. 4.4 Unrooted tree showing the phylogenetic relationships of Bacillus sp. MF-A4 (DQ287962) and members of the genus Bacillus based on 16S rdna sequences Fig. 4.5 Unrooted tree showing the phylogenetic relationships of (DQ287961) and members of the genus Microbacterium based on 16S rdna sequences. Fig. 4.6 CAS plate assay showing production of orange halos around the colonies of the isolates indicating the presence of siderophore production Fig. 4.7 Reddish colouration obtained in Arnows test in the supernatant of B.subtilis Fig. 4.8 Growth curve and siderophore production by B.subtilis Fig. 4.9 Effect of increasing iron concentration on siderophore production by B.subtilis Fig SDS-PAGE of possible receptor proteins of B. subtilus. Fig Plate assays of P solubilizers Fig B. subtilus shown hydrolyzing PNPP(p-nitrophenyl phosphate) to PNP(p-nitro phenol) Fig solubilizes inorganic phosphate, colour change in phenol red dye from pink to yellow, indicating a drop in ph due to release of organic acids into the medium. Fig P solubilization and lowering in ph of broth due to phosphate solubilizing activity of B. subtilus Fig P solubilization by A - M. arborescens, B- M.roseus & C- Bacillus sp. MF-A4 Fig 4.16 Production of hydrogen cyanide (HCN) (orange-red colour) by the sand dune bacterial isolates Fig Orange pigment produced by Microbacterium arboresecens on PPYG agar medium and in broth Fig Absorption spectrum of pigment from alkalophilic Microbacterium arborescens in acetone Fig HPLC chromatogram showing carotenoid fractions separated from the crude pigment extract of. Fig HPLC profile of the pigment of

5 Chapter 5 Fig. 5.1 Fig. 5.2 Fig. 5.3a Fig. 5.3b Fig. 5.4 In vitro inhibition of growth of Sclerotium sp. by antagonistic B. subtilis on a) PDA medium and b) PDA + 2% chitin Germination of Eggplant seedlings by Roll towel tube method Growth promoting effects of sand dune bacteria in non sterilized soil on eggplant seedlings Growth promoting effects of sand dune bacteria in sterilized soil on eggplant seedlings Growth promoting effects of sand dune bacteria in normal soil(ph 7) on eggplant seedlings

6 LIST OF TABLES Chapter 2 Table 2.1 Table 2.2 Table 2.3 Table 2.4a Table 2.4b Table 2.5 Table 2.6 Table 2.7a Table 2.7b Table 2.8a Table 2.8b Table 2.9a Table 2.9b Table 2.10a Table 2.10b Table 2.11 Total viable counts of bacterial groups associated with sand dune vegetation, Ipomoea pes- caprae and Spinifex littoreus in the three seasons. Total viable counts of bacterial groups associated with unvegetated areas of sand dunes. Neutrophiles and alkaliphiles in rhizosphere and endosphere Characterization of neutrophilic bacteria Characterization of alkaliphilic bacteria Percentage distribution of facultative and alkalitolerant rhizosphere and endophytic bacteria in three seasons Percentage of neutrophilic PHA producers Distribution of representative neutrophilic bacterial taxa in the rhizosphere and endophyte of sand dune plants Distribution of representative alkaliphilic bacterial taxa in the rhizosphere and endophytes of sand dune plants Neutrophilic isolates showing production of enzymes, siderophores and P solubilization Alkalophilic isolates showing production of enzymes, siderophores and Phosphate solubilization Percentage growth of neutrophiles associated with Ipomoea pescaprae and Spinifex littoreus on hydrocarbons Percentage distribution of solvent tolerant rhizosphere and endophytic neutrophilic bacteria in premonsoon, monsoon and postmonsoon period Percentage growth of alkalophiles associated with Ipomoea pescaprae and Spinifex littoreus on hydrocarbons Percentage distribution of solvent tolerant rhizosphere and endophytic alkalophilic bacteria in premonsoon, monsoon and postmonsoon period Screening of alkaliphilic bacteria for EPS production Chapter 3 Table 3.1 Table 3.2 Table 3.3 Table 3.4 Table 3.5 Alcian blue adsorption assay for exopolymer Solubility of the polymers isolated from Microbacterium arborescens in different solvents UV-VIS absorption maxima of the polymers Chemical composition of the polymers isolated from Monosaccharide composition (%) of the polymers produced by

7 Table 3.6a Table 3.6b Table 3.7a Table 3.7b Table 3.8a Table 3.8b Sieve analysis of uninoculated sandy soil Sieve analysis of inoculated Sandy soil Sieve analysis of uninoculated agricultural/field soil Sieve analysis of inoculated agricultural/field soil Sieve analysis of uninoculated mine reject soil Sieve analysis of inoculated mine reject soil Chapter 4 Table 4.1 Table 4.2 Table 4.3 Table 4.4 Table 4.5 Table 4.6 Table 4.7 Table 4.8 Table 4.9 Percentage 16S rrna sequence similarities between Bacillus subtilis and some closely related Bacillus spp. Percentage 16S rrna sequence similarities between Kocuria rosea and some closely related Kocuria spp. Percentage 16S rrna sequence similarities between Bacillus sp MF-A4 and some closely related Bacillus spp. Percentage 16S rrna sequence similarities between (DQ287961) and some closely related Microbacterium spp. Nature of the siderophores produced by the sand dune bacterial cultures Phosphate solubilization efficiency(se) of the bacterial cultures Phosphatase enzyme in the bacterial isolates Paper chromatography of organic acids produced by cultures Absorption characteristics of pigment in various solvents Chapter 5 Table 5.1 Table 5.2 Table 5.3 Table 5.4 Table 5.5 Table 5.6 Table 5.7 Effect of sand dune bacteria on eggplant seedling growth Growth promoting effects of sand dune bacteria in sterilized and nonsterilized soil on eggplant seedlings Growth promoting effects of sand dune bacteria in normal soil on eggplant seedling Change in total viable count of sand dune bacteria inoculated under nursery pot conditions in sterilized and nonsterilized soil conditions Change in total viable count of sand dune bacteria inoculated under nursery pot conditions in normal field soil Change in the soil properties of sterilized and non sterilized soils inoculated with sand dune bacteria Change in soil properties due to treatments with SDB grown at neutral ph.