Rangasamy Anandham

Characterization of Plant Growth-Promoting Traits of Bacteria Isolated from Larval Guts of Diamondback Moth Plutella xylostella (Lepidoptera: Plutellidae)

Eight bacterial isolates from the larval guts of Diamondback moths (Plutella xylostella) were tested for their plant growth–promoting (PGP) traits and effects on early plant growth. All of the strains tested positive for nitrogen fixation and indole 3-acetic acid (IAA) and salicylic acid production but negative for hydrogen cyanide and pectinase production. In addition, five of the isolates exhibited significant levels of tricalcium phosphate and zinc oxide solubilization; six isolates were able to oxidize sulfur in growth media; and four isolates tested positive for chitinase and β-1,3-glucanase activities. Based on their IAA production, six strains including four that were 1-aminocyclopropane-1-carboxylate (ACC) deaminase positive and two that were ACC deaminase negative were tested for PGP activity on the early growth of canola and tomato seeds under gnotobiotic conditions. Acinetobacter sp. PSGB04 significantly increased root length (41%), seedling vigor, and dry biomass (30%) of the canola test plants, whereas Pseudomonas sp. PRGB06 inhibited the mycelial growth of Botrytiscinerea, Colletotrichumcoccodes, C.gleospoiroides, Rhizoctoniasolani, and Sclerotiasclerotiorum under inxa0vitro conditions. A significant increase, greater than that of the control, was also noted for growth parameters of the tomato test plants when the seeds were treated with PRGB06. Therefore, the results of the present study suggest that bacteria associated with insect larval guts possess PGP traits and positively influence plant growth. Therefore, insect gut bacteria as effective PGP agents represent an unexplored niche and may broaden the spectrum of beneficial bacteria available for crop production.

Chemolithoautotrophic oxidation of thiosulfate and phylogenetic distribution of sulfur oxidation gene (soxB) in rhizobacteria isolated from crop plants

Twenty-one thiosulfate-oxidizing bacteria were isolated from rhizosphere soils and 16S rRNA analysis revealed that the isolates were affiliated with seven different phylogenetic groups within the Beta and Gamma subclasses of Proteobacteria and Actinobacteria. Among these, five genera, including Dyella, Burkholderia, Alcaligenes, Microbacterium and Leifsonia sp., represented new sulfur oxidizers in rhizosphere soils. The thiosulfate-oxidizing Dyella, Burkholderia, Alcaligenes, Microbacterium, Leifsonia and Pandoraea were able to grow chemolithotrophically with a medium containing thiosulfate and exhibited growth coupled with thiosulfate oxidation. They accumulated intermediate products such as sulfur, sulfite and trithionate in the spent medium during the time course of thiosulfate oxidation, and these products were finally oxidized into sulfate. Furthermore, they possessed thiosulfate-metabolizing enzymes such as rhodanese, thiosulfate oxidase, sulfite oxidase and trithionate hydrolase, suggesting that these bacteria use the S4 intermediate (S4I) pathway for thiosulfate oxidation. Phylogenetic analysis of the soxB gene revealed that Pandoraea sp. and Pandoraea pnomenusa strains formed a separate lineage within Betaproteobacteria.

Massilia jejuensis sp. nov. and Naxibacter suwonensis sp. nov., isolated from air samples

Two Gram-negative, motile, rod-shaped bacteria (strains 5317J-18T and 5414S-25T) were isolated from air samples collected in the Jeju Island and Suwon region of Korea, respectively. Phylogenetically, strain 5317J-18T was grouped with the genus Massilia with Massilia brevitalea byr23-80T as the closest relative (98.8% sequence similarity). Strain 5414S-25T was affiliated with the genus Naxibacter with Naxibacter haematophilus CCUG 38318T as the closest relative (98.8% sequence similarity). The mean DNA-DNA relatedness values between strain 5317J-18T and M. brevitalea DSM 18925T and Massilia aurea DSM 18055T were 43 and 36%, respectively. The mean DNA-DNA hybridization values between strain 5414S-25T and N. haematophilus KACC 13771T, M. brevitalea DSM 18925T, Massilia timonae DSM 16850T, Naxibacter varians KACC 13770T, M. aurea DSM 18055T, Massilia lutea DSM 17473T and Massilia albidiflava DSM 17472T ranged from 33 to 42%. Both novel strains had ubiquinone Q-8 as the predominant isoprenoid quinone and summed feature 3 (comprising iso-C15:0 2-OH and/or C16:1 omega7c) and C16:0 as the major fatty acids. Both strains also showed similar polar lipid profiles with phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol as the major polar lipids. The DNA G+C contents of strains 5317J-18T and 5414S-25T were 66.1 and 67.8%, respectively. On the basis of their phenotypic, chemotaxonomic and genotypic characteristics, the new strains represent novel species in the genera Massilia and Naxibacter. Strain 5317J-18T (=KACC 12634T=DSM 21309T) is proposed as the type strain of Massilia jejuensis sp. nov. and strain 5414S-25T (=KACC 12635T=DSM 21311T) is proposed as the type strain of Naxibacter suwonensis sp. nov.

Potential plant growth promoting traits and bioacidulation of rock phosphate by thiosulfate oxidizing bacteria isolated from crop plants

Thiosulfate oxidizing bacteria isolated from crop plants were tested for their traits related to plant growth promotion and their ability to solubilize Morocco rock phosphate (RP) through oxidation of thiosulfate to sulfuric acid. All the tested strains grew in Nfb medium (except Dyella ginsengisoli) and possessed β‐1,3 glucanase activity (except Burkholderia kururiensis). Of the fourteen tested strains, 2 were found positive for siderophore production, 3 each for phytohormones (IAA), and salicylic acid production. Based on qualitative and quantitative assays, 5 strains were found to efficiently solubilize tri‐calcium phosphate in Pikovskayas medium. Nine strains exhibited ACC (1‐aminocyclopropane‐1‐carboxylate) deaminase activity. In gnotobiotic experiments, Pandoraea sputorum ATSB28 which possessed the lowest ACC deaminase (0.44 nM of α‐Keto butyrate formed min–1 mg of protein–1) activity increased the primary root length of canola by 166%. Inoculation of Pandoraea sp. strain ATSB30 in mixture containing RP and thiosulfate significantly enhanced the water extractable‐P (1147 μg P g RP–1) and bicarbonate extractable‐P (1144 μg P g RP–1) on day 45. Glucose amendment resulted in increased RP solubilization as compared to glucose unamended treatments. Thiosulfate oxidizing bacteria tested in this study possessed at least one or more plant growth promoting traits apart from thiosulfate oxidation and solubilized the RP. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Cohnella yongneupensis sp. nov. and Cohnella ginsengisoli sp. nov., Isolated from Two Different Soils

Two aerobic, Gram-positive, rod-shaped bacterial strains, 5YN10-14(T) and GR21-5(T), were isolated from the Yongneup wetland and ginseng soil in Korea, respectively. The two strains formed ellipsoidal or oval spores positioned centrally or paracentrally in swollen sporangia. On the basis of 16S rRNA gene sequence analysis, these strains were related to members of the genus Cohnella. 16S rRNA gene sequence similarity between strains 5YN10-14(T) and GR21-5(T) was 95.9 %. Strains 5YN10-14(T) and GR21-5(T) showed, respectively, 94.3 and 95.2 % 16S rRNA gene sequence similarity to Cohnella thermotolerans CCUG 47242(T), 94.6 and 94.4 % to Cohnella hongkongensis HKU3(T), 94.7 and 94.7 % to Cohnella laeviribosi RI-39(T), and 95.4 and 94.8 % to Cohnella phaseoli GSPC1(T). The major fatty acids of strain 5YN10-14(T) were anteiso-C(15 : 0) (51.1 %), iso-C(16 : 0) (18.5 %) and C(16 : 0) (13.2 %), and the major fatty acids of strain GR21-5(T) were anteiso-C(15 : 0 ) (48.9 %), iso-C(16 : 0) (15.0 %) and iso-C(15 : 0) (12.2 %). The two strains contained menaquinone with seven isoprene units (MK-7) as the predominant quinone, and diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine as major polar lipids; however, strain 5YN10-14(T) also contained lysylphosphatidylglycerol as a major polar lipid, whereas strain GR21-5(T) had an unknown aminophospholipid as another major polar lipid. The DNA G+C contents of strains 5YN10-14(T) and GR21-5(T) were 58.8 and 61.3 mol%, respectively. Based on the results of the phylogenetic and phenotypic data presented, it was concluded that the two strains represent two novel species of the genus Cohnella , for which the names Cohnella yongneupensis sp. nov. (type strain 5YN10-14(T)=KACC 11768(T)=DSM 18998(T)) and Cohnella ginsengisoli sp. nov. (type strain GR21-5(T)=KACC 11771(T)=DSM 18997(T)) are proposed.

Pandoraea thiooxydans sp. nov., a facultatively chemolithotrophic, thiosulfate-oxidizing bacterium isolated from rhizosphere soils of sesame (Sesamum indicum L.)

A facultatively chemolithoautotrophic, thiosulfate-oxidizing, Gram-negative, aerobic, motile, rod-shaped bacterial strain, designated ATSB16(T), was isolated from rhizosphere soils of sesame (Sesamum indicum L.). 16S rRNA gene sequence analysis demonstrated that this strain was closely related to Pandoraea pnomenusa LMG 18087(T) (96.7 % similarity), P. pulmonicola LMG 18016(T) (96.5 %), P. apista LMG 16407(T) (96.2 %), P. norimbergensis LMG 18379(T) (96.1 %) and P. sputorum LMG 18819(T) (96.0 %). Strain ATSB16(T) shared 96.0-96.4 % sequence similarity with four unnamed genomospecies of Pandoraea. The major cellular fatty acids of the strain ATSB16(T) were C(17 : 0) cyclo (33.0 %) and C(16 : 0) (30.6 %). Q-8 was the predominant respiratory quinone. The major polar lipids were phosphatidylmethylethanolamine, diphosphatidylglycerol, phosphatidylethanolamine and two unidentified aminophospholipids. Hydroxyputrescine and putrescine were the predominant polyamines. The genomic DNA G+C content of the strain was 64.0 mol%. On the basis of the results obtained from this study, strain ATSB16(T) represents a novel species of the genus Pandoraea, for which the name Pandoraea thiooxydans sp. nov. is proposed. The type strain is ATSB16(T) (=KACC 12757(T) =LMG 24779(T)).

Thiosulfate oxidation and mixotrophic growth of Methylobacterium oryzae

Thiosulfate oxidation and mixotrophic growth with succinate or methanol plus thiosulfate was examined in nutrient-limited mixotrophic condition for Methylobacterium oryzae CBMB20, which was recently characterized and reported as a novel species isolated from rice. Methylobacterium oryzae was able to utilize thiosulfate in the presence of sulfate. Thiosulfate oxidation increased the protein yield by 25% in mixotrophic medium containing 18.5 mmol.L-1 of sodium succinate and 20 mmol.L-1 of sodium thiosulfate on day 5. The respirometric study revealed that thiosulfate was the most preferable reduced inorganic sulfur source, followed by sulfur and sulfite. Thiosulfate was predominantly oxidized to sulfate and intermediate products of thiosulfate oxidation, such as tetrathionate, trithionate, polythionate, and sulfur, were not detected in spent medium. It indicated that bacterium use the non-S4 intermediate sulfur oxidation pathway for thiosulfate oxidation. Thiosulfate oxidation enzymes, such as rhodanese and sulfite oxidase activities appeared to be constitutively expressed, but activity increased during growth on thiosulfate. No thiosulfate oxidase (tetrathionate synthase) activity was detected.

Rudaea cellulosilytica gen. nov., sp. nov., isolated from soil.

A yellow-pigmented, Gram-negative, aerobic, rod-shaped bacterium, strain KIS3-4T, was isolated from soil collected on Daechung Island in the West Sea of Korea. Phylogenetic analysis based on the 16S rRNA gene sequence placed strain KIS3-4T in a distinct lineage in the family Xanthomonadaceae. Strain KIS3-4T shared 87.3-93.7% sequence similarity with members of the family Xanthomonadaceae, and was related most closely to the genera Dyella and Dokdonella. In its biochemical characteristics, strain KIS3-4T was clearly separable from other genera within the family Xanthomonadaceae on the basis of the hydrolysis of cellulose and urea, high G+C content (64 mol%) and fatty acid profile. Major fatty acids (>10% of the total fatty acids) were iso-C17:1omega9c (32.8%), iso-C17:0 (18.0%) and iso-C16:0 (12.7%). Q-8 was the predominant respiratory quinone. Phosphatidylethanolamine and several unidentified aminophospholipids and phospholipids were present. Based on its unique phenotypic, genotypic and phylogenetic features, strain KIS3-4T represents a novel genus and species, for which the name Rudaea cellulosilytica gen. nov., sp. nov. is proposed. The type strain of Rudaea cellulosilytica is KIS3-4T (=KACC 12734T=JCM 15422T).

Leucobacter denitrificans sp. nov., isolated from cow dung

The bacterial strain M1T8B10T was isolated from cow dung in Suwon, Republic of Korea. The strain was a Gram stain-positive rod, nonmotile, and non-spore-forming. According to 16S rRNA gene sequence analysis, the strain fell within the clade of the genus Leucobacter, showing the highest sequence similarities with Leucobacter aridicollis L-9T (98.7%), Leucobacter iarius 40T (98.4%), and Leucobacter komagatae JCM 9414T (98.2%). Cell-wall peptidoglycan contained the diagnostic diamino acid 2,4-diaminobutyric acid of the genus Leucobacter, showing B-type cross-linked peptidoglycans. The major fatty acids were anteiso-C15:0, iso-C16:0, and anteiso-C17:0. The quinone system consisted of the menaquinones MK-11 (78%) and MK-10 (22%). The polar lipid profiles contained diphosphatidylglycerol, phosphatidylglycerol, and an unidentified glycolipid. Differences in several physiological features including nitrate reduction enabled the isolate to be differentiated from all recognized Leucobacter species. Based on these phylogenetic, chemotaxonomic, and phenotypic results, the isolate represents a novel species, for which the name Leucobacter denitrificans sp. nov. is proposed. The type strain is M1T8B10T (=KACC 14055T =NBRC 106309T).

Parapedobacter luteus sp. nov. and Parapedobacter composti sp. nov., isolated from cotton waste compost

Two aerobic, Gram-reaction-negative, non-spore-forming bacterial strains, 4M29T and 4M40T, were isolated from cotton composts. The two strains grew in the presence of 0-5% (w/v) NaCl (optimum growth in the absence of NaCl), at pH 6.0-8.0 (optimum, pH 7.0) and at 15-45 degrees C (optimum, 30 degrees C). The strains shared 97.1% 16S rRNA gene sequence similarity. Strains 4M29T and 4M40T showed the next highest levels of 16S rRNA gene sequence similarity to Parapedobacter koreensis Jip14T (95.6 and 94.4%, respectively) and Parapedobacter soli DCY14T (95.2 and 93.8%). The level of DNA-DNA relatedness between strains 4M29T and 4M40T was 38%. The two strains contained iso-C15:0, summed feature 3 (comprising iso-C15:0 2-OH and/or C16:1omega7c) and iso-C17:0 3-OH as major fatty acids, MK-7 as the major respiratory quinone, homospermidine as the only polyamine and phosphatidylethanolamine as the major polar lipid. The DNA G+C contents of strains 4M29T and 4M40T were 47.6 and 48.6 mol%, respectively. On the basis of phylogenetic and phenotypic data, strains 4M29T and 4M40T are considered to represent two novel species of the genus Parapedobacter, for which the names Parapedobacter luteus sp. nov. (type strain 4M29T =KACC 10955T =JCM 15977T) and Parapedobacter composti sp. nov. (type strain 4M40T =KACC 10972T =JCM 15978T) are proposed.