Facultative Methylotrophic Bacteria Research Articles

Genomic profiling and characteristics of a C1 degrading heterotrophic fresh-water bacterium Paracoccus sp. strain DMF.

Paracoccus species are metabolically versatile gram-negative, aerobic facultative methylotrophic bacteria showing enormous promise for environmental and bioremediation studies. Here we report, the complete genome analysis of Paracoccus sp. strain DMF (P. DMF) that was isolated from a domestic wastewater treatment plant in Kanpur, India (26.4287°N, 80.3891°E) based on its ability to degrade a recalcitrant organic solvent N, N-dimethylformamide (DMF). The results reveal a genome size of 4,202,269 base pairs (bp) with a G + C content of 67.9%. The assembled genome comprises 4141 coding sequences (CDS), 46 RNA sequences, and 2 CRISPRs. Interestingly, catabolic operons related to the conventional marine-based methylated amines (MAs) degradation pathway were functionally annotated within the genome of an obligated aerobic heterotroph that is P. DMF. The genomic data-based characterization presented here for the novel heterotroph P. DMF aims to improve the understanding of the phenotypic gene products, enzymes, and pathways involved with greater emphasis on facultative methylotrophic motility-based latent pathogenicity.

Molecular Identification of Methylorubrum extorquens using PCR-Amplified MxaF Gene Fragments as A Molecular Marker

Methylotrophs bacteria are ubiquitous, and they have the ability to consume single carbon (C1) which makes them biological conversion machines. It is the first study to find facultative methylotrophic bacteria in contaminated soils in Iraq. Conventional PCR was employed to amplify MxaF that encodes methanol dehydrogenase enzyme. DNA templates were extracted from bacteria isolated from five contaminated sites in Basra. The gene specific PCR detected Methylorubrum extorquens as the most dominant species in these environments. The ability of M. extorquens to degrade aliphatic hydrocarbons compound was tested at the laboratory. Within 7 days, gas chromatographic (GC) studies of remaining utilized crude oil revealed that 61.14 % of the initial content had been degraded, and GC fingerprinting of the utilized aliphatic compounds revealed significant reductions in C12, C13, C14, and C15. Globally this is the first time found a new strain of M. extorquens has the ability to degrade aliphatic hydrocarbons compound. Conventional PCR and gene sequencing revealed the presence of the facilitative methylotrophic bacteria in polluted areas in Basra. M. extorquens was dominant and showed a substantial ability to degrade crude oil which makes them an important tool to be employed in bioremediation.

Pink Pigmented Facultative Methylotrophs (PPFMs): Potential Bioinoculants for Sustainable Crop Production

Bacteria belonging to the genus Methylobacterium, popularly known as pink pigmented facultative methylotrophic (PPFM) bacteria, are well known for their distinct ability to use single-carbon compounds like methanol, formate and formaldehyde, and also a variety of multi-carbon substrates lacking carbon-carbon bonds. These bacterial groups are ubiquitously distributed, especially in phyllosphere and rhizosphere, and their occurrence have been reported in more than 100 species of plants so far. PPFMs have profound influence on soil fertility, crop growth and yield. The ability for phosphate acquisition, nitrogen fixation, iron chelation and phytohormone production indicate the possibility of developing them as promising biofertilizer candidates. In addition, many of them possess biocontrol activity against several phytopathogens. PPFMs induce several physiological changes in plants, making the plants more resistant to biotic and abiotic stress. They can therefore be promising alternatives to conventional chemical inputs in sustainable agricultural systems.

Natural Occurrence of Pink Pigmented Facultative Methylotrophic (PPFM)Bacteria on Phyllosphere of Mulberry (Morus Indica)

Natural Occurrence of Pink Pigmented Facultative Methylotrophic (PPFM)Bacteria on Phyllosphere of Mulberry (Morus Indica)

Cytobacillus pseudoceanisediminis sp. nov., A Novel Facultative Methylotrophic Bacterium with High Heavy Metal Resistance Isolated from the Deep Underground Saline Spring

In this study, we present the characterization of the BNO1T bacterial strain isolated from the deep subsurface saline spring at the Baksan Neutrino Observatory INR RAS (Kabardino-Balkaria, Russia). The complete genome sequence of the strain BNO1T is 5,347,902bp, with a GC content 41 and 49%. The cell wall peptidoglycan contains meso-diaminopimelic acid. The major isoprenoid quinone is MK-7 and the polar lipids are diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylethanolamine. The major fatty acids are anteiso-C15:0 (23.34%), iso-C15:0 (20.10%), C16:0 (11.96%), iso-C16:0 (10.88%), and anteiso-C17:0 (10.79%). The 16S rRNA gene sequence clearly demarcated the strain as belonging to Cytobacillus genera. Based on the phylogenetic analysis, ANI (average nucleotide identity) and dDDH (digital DNA-DNA hybridization) assessments we propose to assign the strain BNO1T and other related strains to new species and to name it Cytobacillus pseudoceanisediminis sp. nov. (The values of ANI and dDDH between BNO1T and Cytobacillus oceanisediminis CGMCC 1.10115T are 80.65% and 24.7%, respectively; values of ANI and dDDH between BNO1T and Cytobacillus firmus NCTC 10335T are 89% and 38%, respectively). Genomic analysis of strain BNO1T revealed pathways for C1 compounds oxidation and two pathways for C1 compounds assimilation: serine and ribulose monophosphate pathways. In addition, strain BNO1T contains a plasmid (342,541bp) coding multiple genes involved in heavy metal ion balance. Moreover, heavy metal toxicity testing confirmed the high potential of the strain BNO1T as a source of metal resistance genes and enzymes. The type strain is BNO1T (= BIM B-1921T = VKM B-3664T).

Impact of foliar fertilization of nutrients and biofertilizer on the growth and yield of hybrid cotton (Gossypium hirsutum L.)

The newly released high yielding cotton cultivars are higher boll retention rate than conventional cultivars that have higher overall nutrient demand during the flowering and boll development stages. Therefore, this field experiment was conducted at during summer, 2020 at Annamalai University experimental farm, Department of Agronomy, Faculty of Agriculture, Annamalai Nagar, Tamil Nadu, India, 608 002 to ascertain the impact of foliar application of essential nutrients and biofertilizers on growth and yield of cotton. The experiment was laid out in randomized block design with twelve treatments of foliar application macro and micro-nutrients with and without Pink pigmented facultative methylotrophic bacteria (PPFM) and replicated thrice. Foliar application of macro, micronutrients and PPFM registered the increased values of growth characters, yield attributes and yield of hybrid cotton. Among the foliar application practice tested, foliar application of 2% urea + 1% PPFM at flowering and boll development stages significantly resulted in the enhances values of growth characters viz., plant height (157.40 cm), Dry matter production (6003 kg/ha), number of monopodial branches (3.28/plant), growth analysis like Leaf area index (4.30), Crop growth rate (7.01 g/m2/day) and Relative growth rate (0.0144 g/g/day), yield attributes viz., number of sympodial branches (21.77/plant), number of squares (83.77/plant), number of bolls (45.04/plant), boll weight (5.58 g) and seed cotton yield (3185 kg/ha). This was statistically on par with the foliar application of 2% MgSO4 + 1% PPFM at flowering and boll development stages. The lower values of growth characters, yield attributes and yield were recorded under water spray treatment. Based on the results of field investigation it can be concluded that foliar application of 2% urea + 1% PPFM at flowering and boll development stages can possibly enhance the growth and yield of hybrid cotton grown under dryland condition.

Role of EPS in mitigation of plant abiotic stress: The case of Methylobacterium extorquens PA1

Methylobacterium extorquens is a facultative methylotrophic Gram-negative bacterium, often associated with plants, that exhibits a unique ability to grow in the presence of high methanol concentrations, which serves as a single carbon energy source. We found that M. extorquens strain PA1 secretes a mixture of different exopolysaccharides (EPSs) when grown in reference medium or in presence of methanol, that induces the secretion of a peculiar and heterogenous mixture of EPSs, with different structure, composition, repeating units, bulk and a variable degree of methylation. These factors influenced 3D structure and supramolecular assets, diffusion properties and hydrodynamic radius, and likely contribute to increase methanol tolerance and cell stability. No direct methanol involvement in the EPSs solvation shell was detected, indicating that the polymer exposure to methanol is water mediated. The presence of methanol induces no changes in size and shape of the polymer chains, highlighting how water-methanol mixtures are a good solvent for refEPS and metEPS.

Siderophore for Lanthanide and Iron Uptake for Methylotrophy and Plant Growth Promotion in Methylobacterium aquaticum Strain 22A.

Methylobacterium and Methylorubrum species are facultative methylotrophic bacteria that are abundant in the plant phyllosphere. They have two methanol dehydrogenases, MxaF and XoxF, which are dependent on either calcium or lanthanides (Lns), respectively. Lns exist as insoluble minerals in nature, and their solubilization and uptake require a siderophore-like substance (lanthanophore). Methylobacterium species have also been identified as plant growth-promoting bacteria although the actual mechanism has not been well-investigated. This study aimed to reveal the roles of siderophore in Methylobacterium aquaticum strain 22A in Ln uptake, bacterial physiology, and plant growth promotion. The strain 22A genome contains an eight-gene cluster encoding the staphyloferrin B-like (sbn) siderophore. We demonstrate that the sbn siderophore gene cluster is necessary for growth under low iron conditions and was complemented by supplementation with citrate or spent medium of the wild type or other strains of the genera. The siderophore exhibited adaptive features, including tolerance to oxidative and nitrosative stress, biofilm formation, and heavy metal sequestration. The contribution of the siderophore to plant growth was shown by the repressive growth of duckweed treated with siderophore mutant under iron-limited conditions; however, the siderophore was dispensable for strain 22A to colonize the phyllosphere. Importantly, the siderophore mutant could not grow on methanol, but the siderophore could solubilize insoluble Ln oxide, suggesting its critical role in methylotrophy. We also identified TonB-dependent receptors (TBDRs) for the siderophore–iron complex, iron citrate, and Ln, among 12 TBDRs in strain 22A. Analysis of the siderophore synthesis gene clusters and TBDR genes in Methylobacterium genomes revealed the existence of diverse types of siderophores and TBDRs. Methylorubrum species have an exclusive TBDR for Ln uptake that has been identified as LutH. Collectively, the results of this study provide insight into the importance of the sbn siderophore in Ln chelation, bacterial physiology, and the diversity of siderophore and TBDRs in Methylobacterium species.

Development of deficit irrigation program for improving the water use efficiency, growth, and yield of groundnut in the semiarid tropical region of India

In this study, we assessed the efficiency of a deficit irrigation program (DIP) on groundnut (<i>Arachis hypogaea</i> L.) yield and economic profits. The DIP was designed to combine frequency and volume of irrigation to develop an appropriate irrigation water management (IWM) strategy for groundnut. Two field experiments were conducted at the Agricultural College and Research Institute, Madurai, Tamil Nadu, India, with a bunch variety of groundnut, VRI 2, under a randomized block design (RBD) with three replications. The treatments included DIP in combination with a foliar application of pink-pigmented facultative methylotrophic bacteria (PPFM). The irrigation frequencies adopted for the first set of treatments were high frequency (5/5), medium frequency (7/5), and lower frequency (7/7) starting from 14 days after sowing (DAS). The effects of early water stress on groundnut pod yield and water use efficiency were investigated in a second experiment by starting irrigation at 21 DAS. The number of irrigation frequencies and irrigation volume was fixed to develop an easily readable irrigation chart for end-users. The number of irrigation frequencies was fixed as three, and the volume of irrigation was fixed as a depth of 3 cm. In the first experiment, a mini mobile pumping unit was used for scheduling irrigation frequencies; in the second experiment, the surface drip method was used. In both experiments, we also assessed the effects of foliar application of PPFM, widely used as a growth stimulant for several crops, especially under drought conditions. Overall, we observed that DIP led to up to 25.6% water saving compared to surface irrigation. Use of the DIP at the medium frequency with PPFM spray had a significantly positive influence on water use efficiency (WUE) and irrigation water use efficiency (IWUE) compared to conventional drip irrigation (CDI). Compared to surface irrigation (SI), DIP had a positive impact on growth parameters, yield attributes, and economic profitability with a higher benefit-cost ratio. Additionally, there were only minor differences between deficit irrigation (DI) results starting from 21 DAS and 14 DAS. Irrespective of the irrigation techniques, PPFM application had a positive effect on the yield attributes and WUE. Our results indicated that DI at a medium frequency (7/5) with PPFM spray, starting from 21 DAS under drip fertigation, is the best irrigation management innovation (IMI) strategy for optimum utilization of irrigation water without significant reduction in pod yield. This management had a positive influence on WUE and IWUE of summer groundnut in sandy clay loam soil. However, the exact scheduling of the irrigation may differ in different environments as well as soil types. Nevertheless, we propose that the application and further optimization of the DIP devised in this study would help the farmers attain a similar yield using a lower amount of irrigation water and lead to significant economic profits for the farmers.

Pink Pigmented Facultative Methylotrophic Bacteria Isolated from Fermented Philippine Shrimp Paste.

Pink pigmented facultative methylotrophic (PPFM) bacteria are ecologically distributed microorganisms. They have been isolated in many types of ecosystems like soil, water, air, in association with plants and even as pathogens in humans. However, a yet unexplored area for PPFM bacteria research is in food. Hence, the objective of this study was to establish the presence of PPFM bacteria in Philippine fermented food, in particular shrimp paste, and characterise them phenotypically and genotypically. A total of 13 PPFM bacteria were obtained from Philippine shrimp paste. Sequencing of the 16S rRNA gene revealed that the PPFM bacterial isolates belong to the genus Methylobacterium. A total of 35 phenotypic characterisations were performed that included morphological, biochemical and physiological tests. Phylogenetic tree was constructed to establish the genetic relatedness of the isolates. Morphological test results showed that all 13 isolates were consistent with the established phenotypic characters of the genus such as pink colony colour, Gram negative and rod-shaped. Biochemically, the use of API® 20 NE tests showed heterogeneity of results and physiological tests exhibited that the isolates are primarily mesophilic and halotolerant, being able to grow at 2% salt. Phylogenetic analysis showed that the isolates are Methylobacterium populi, M. dankookense, M. lusitanum, M. radiotolerans and M. zatmanii. This study confirmed the presence and diversity of PPFM bacteria in Philippine shrimp paste. Further studies are needed to show the functional activity of Methylobacterium in Philippine shrimp paste production.

Potentials, Utilization, and Bioengineering of Plant Growth-Promoting Methylobacterium for Sustainable Agriculture

Plant growth-promoting bacteria (PGPB) have great potential to provide economical and sustainable solutions to current agricultural challenges. The Methylobacteria which are frequently present in the phyllosphere can promote plant growth and development. The Methylobacterium genus is composed mostly of pink-pigmented facultative methylotrophic bacteria, utilizing organic one-carbon compounds as the sole carbon and energy source for growth. Methylobacterium spp. have been isolated from diverse environments, especially from the surface of plants, because they can oxidize and assimilate methanol released by plant leaves as a byproduct of pectin formation during cell wall synthesis. Members of the Methylobacterium genus are good candidates as PGPB due to their positive impact on plant health and growth; they provide nutrients to plants, modulate phytohormone levels, and protect plants against pathogens. In this paper, interactions between Methylobacterium spp. and plants and how the bacteria promote crop growth is reviewed. Moreover, the following examples of how to engineer microbiomes of plants using plant-growth-promoting Methylobacterium are discussed in the present review: introducing external Methylobacterium spp. to plants, introducing functional genes or clusters to resident Methylobacterium spp. of crops, and enhancing the abilities of Methylobacterium spp. to promote plant growth by random mutation, acclimation, and engineering.

IMPACT OF PINK PIGMENTED FACULTATIVE METHYLOTROPHIC BACTERIA AS A BIO-STIMULANT ON GROWTH, HERB YIELD AND VOLATILE OIL ATTRIBUTES OF ROSEMARY (ROSMARINUS OFFICINALIS L.) PLANT

The effect of Pink Pigmented Facultative Methylotrophic (PPFM) bacteria as a bio-stimulant on vegetative growth, macro-nutrient elements ( N,P and K ) contents , chemical constituents and volatile oil percentage and components in rosemary plant (Rosmarinus officinalis L.) were studied during 2018 and 2019 seasons. Plant height (cm), number of lateral branches/plant, herb fresh & dry weights and root fresh & dry weights of rosemary plant were all greatly affected with all applied treatments of PPFM when compared with the control. The hugest effect was recorded by 1% PPFM bacteria spraying in both studied seasons. However, no significant effect was recorded due to PPFM applied treatments on P%. Highest values of total carbohydrates were recorded by both 1 and 3%. PPFM bacteria spraying in both studied seasons. Whereas, the highest values of C/N ratios were obtained with 3% PPFM in first season and 1% PPFM in the second one. A substantial effect from PPFM bacteria spraying was observed through its effect on reducing the level of NO3 and NO2 in rosemary herb when compared with the control. In any case, 3% of PPFM bacteria spraying was more effective than other treatments in recording the least values of NO3 and NO2 in first season compared to the control. Volatile oil % was enormously increased with 3% PPFM bacteria spraying in both studied seasons than other treatments or control. The main components of volatile oil of rosemary plant were Camphor, Eucalyptol, Verbenone, Borneol, Pinene, Pinanone, Bornyl acetate and Camphene. The treatments of PPFM bacteria spraying immensely stimulated the component % in volatile oil of rosemary herb than in the control. It could be concluded that the application of PPFM bacteria as foliar spraying in rosemary plant production had many advantages especially reducing NO3 and NO2 levels and increasing volatile oil % and volatile oil fractions not to mention it increased the percent of active ingredients.

Transaldolase in Bacillus methanolicus: biochemical characterization and biological role in ribulose monophosphate cycle

BackgroundThe Gram-positive facultative methylotrophic bacterium Bacillus methanolicus uses the sedoheptulose-1,7-bisphosphatase (SBPase) variant of the ribulose monophosphate (RuMP) cycle for growth on the C1 carbon source methanol. Previous genome sequencing of the physiologically different B. methanolicus wild-type strains MGA3 and PB1 has unraveled all putative RuMP cycle genes and later, several of the RuMP cycle enzymes of MGA3 have been biochemically characterized. In this study, the focus was on the characterization of the transaldolase (Ta) and its possible role in the RuMP cycle in B. methanolicus.ResultsThe Ta genes of B. methanolicus MGA3 and PB1 were recombinantly expressed in Escherichia coli, and the gene products were purified and characterized. The PB1 Ta protein was found to be active as a homodimer with a molecular weight of 54 kDa and displayed KM of 0.74 mM and Vmax of 16.3 U/mg using Fructose-6 phosphate as the substrate. In contrast, the MGA3 Ta gene, which encodes a truncated Ta protein lacking 80 amino acids at the N-terminus, showed no Ta activity. Seven different mutant genes expressing various full-length MGA3 Ta proteins were constructed and all gene products displayed Ta activities. Moreover, MGA3 cells displayed Ta activities similar as PB1 cells in crude extracts.ConclusionsWhile it is well established that B. methanolicus can use the SBPase variant of the RuMP cycle this study indicates that B. methanolicus possesses Ta activity and may also operate the Ta variant of the RuMP.

NEW BACTERIAL AGGREGATE-FORMING PRODUCER OF POLY-3-HYDROXYBUTYRATE

The growth and accumulation of carotenoid pigment and poly-3-hydroxybutyrate by a new strain of pink-colored facultative methylotrophic bacterium Methylorubrum extorquens LP in suspension culture and in cell aggregates were studied. The parameters that promote cell aggregation and the formation of poly-3-hydroxybutyrate have been determined.

English

This work highlighted a putative link between the physiological activity and genetic diversity of Methylobacterium species and the association with sugarcane roots and rhizoplane. In total, 40 isolates previously described as pink-pigmented facultative methylotrophic bacteria (PPFMs), were evaluated for their ability to fix nitrogen and solubilize inorganic phosphate, amylase and pectinase activity. This in vitro potential was positively correlated with the community isolated from the root tissues than those from the rhizoplane. Regarding the genomic fingerprinting, the (BOX-PCR) approach revealed a low similarity among the isolates, occurring sole 7 haplotypes harboring more than 70% of similarity among band patterns. These results revealed that the genomic fingerprinting of the isolates recovery from roots is different from the rhizoplane. Besides that, these haplotypes occurred on both sugarcane varieties. Using a phylogenetic sequencing approach based on the 16S rRNA gene, we observed a high abundance of sequences similar to Methylobacterium radiotolerans colonizing both plant tissue and sugarcane varieties were observed. Hence, it was suggested that the plant should select those Methylobacterium spp. with a high biotechnological potential to promote plant growth. Therefore, the bioprospection of specific endophytic bacterial groups comprise an important source of biotechnological potential to improve sugarcane growth and production. Key words: Pink-pigmented facultative methylotrophic bacteria, plant growth promotion, BOX-PCR, 16S rDNA, Methylobacterium radiotolerans.

Polyphasic characterization of Delftia acidovorans ESM-1, a facultative methylotrophic bacterium isolated from rhizosphere of Eruca sativa.

In this study, one bacterial strain, ESM-1, was isolated from rhizosphere of Eruca sativa, growing in Al Hofouf, Saudia Arabia, after enrichment with methanol as a sole carbon and energy source in a batch culture. ESM-1 was characterized by a polyphasic approach. The strain was identified as Delftia acidovorans at similarity level of 99.9% of the 16S rRNA gene sequences. Results of the Biolog Gen III MicroPlate test system showed that strain ESM-1 reacted positively to 47 (50%) including the one-carbon compound formic acid, and partially positive to 6 (∼6.4%) out of the 94 different the traits examined. The total cellular fatty acids composition of the strain ESM-1 was (C16:1ω7c/C16:1ω6c) and C16:0) and matched that of Delftia acidovorans at a similarity index of 0.9, providing a robustness to the ESM-1 identification. Furthermore, ESM-1 displayed a complex polar lipid profile consisting of phosphatidylethanolamine, phosphatidylglycerol, glycolipid, aminolipid, in addition to uncharacterized lipids. The DNA G+C content of the strain was 66.6 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that the strain ESM1-1 was clearly clustered within the Delftia clade and constructed a monophyletic subcluster with Delftia acidovorans NBRC14950. The results addressed that ESM-1 is a facultative methylotrophic bacterium indigenous to Al Hofouf region and opens the door for potential biotechnological applications (e.g., bioremediation) of this strain, in future. Additionally, these findings assure that the total cellular fatty acid analysis and 16S rRNA gene are reliable tool for bacterial characterization and identification.

A novel Delftia plant symbiont capable of autotrophic methylotrophy

A facultative methylotrophic bacterium, strain Lp-1, which was isolated from root nodules of lupine (Lupinus polyphyllus L.) on the medium with methanol as a carbon and energy source, exhibited high similarity of the 16S rRNA gene sequences to Delftia strains (94‒99.9%). The cells of Delftia sp. Lp-1 were motile gram-negative rods dividing by binary fission. Predominant fatty acids were C16:0 (34.2%), C16:1ω9 (14.5%), and C18:1ω7c (17.3%). Phosphatidylethanolamine, phosphatidylcholine, and phosphatidylglycerol were the dominant phospholipids. Q8 was the major ubiquinone. Optimal growth occurred at 24‒26°C and pH 7.1‒7.3; growth was inhibited by 1% NaCl. The organism oxidized methanol with the classical methanol dehydrogenase and used the ribulose bisphosphate pathway of C1 metabolism. Analysis of translated amino acid sequence of the large subunit of the MxaF methanol dehydrogenase revealed 85.5‒94% similarity to the sequences of such autotrophic methylotrophs of the class Alphaproteobacteria as Angulomicrobium, Starkeya, and Ancylobacter, indicating the possible acquisition of the mxaF gene via horizontal gene transfer. Delftia sp. Lp-1 (VKM B-3039, DSM 24446), the first methylotrophic member of the genus Delftia, was shown to be a plant symbiont, stimulating plant growth and morphogenesis, increasing the level of photosynthetic pigments and specific leaf weight. It possesses the nifH gene of nitrogen fixation, is capable of phosphate solubilization, synthesis of auxins and siderophores, and is antagonistic to plant pathogenic fungi and bacilli.

Control of Wilt and Rot Pathogens of Tomato by Antagonistic Pink Pigmented Facultative Methylotrophic Delftia lacustris and Bacillus spp.

The studies on the biocontrol potential of pink pigmented facultative methylotrophic (PPFM) bacteria other than the genus Methylobacterium are scarce. In the present study, we report three facultative methylotrophic isolates; PPO-1, PPT-1, and PPB-1, respectively, identified as Delftia lacustris, Bacillus subtilis, and Bacillus cereus by 16S rRNA gene sequence analysis. Hemolytic activity was tested to investigate the potential pathogenicity of isolates to plants and humans, the results indicates that the isolates PPO-1, PPT-1, and PPB-1 are not pathogenic strains. Under in vitro conditions, D. lacustris PPO-1, B. subtilis PPT-1, and B. cereus PPB-1 showed direct antagonistic effect by inhibiting the mycelial growth of fungal pathogens; Fusarium oxysporum f. sp. lycopersici (2.15, 2.05, and 1.95 cm), Sclerotium rolfsii (2.14, 2.04, and 1.94 cm), Pythium ultimum (2.12, 2.02, and 1.92 cm), and Rhizoctonia solani (2.18, 2.08, and 1.98 cm) and also produced volatile inhibitory compounds. Under plant growth chamber condition methylotrophic bacterial isolates; D. lacustris PPO-1, B. subtilis PPT-1, and B. cereus PPB-1 significantly reduced the disease incidence of tomato. Under greenhouse condition, D. lacustris PPO-1, B. subtilis PPT-1, and B. cereus PPB-1 inoculated tomato plants, when challenged with F. oxysporum f. sp. lycopersici, S. rolfsii, P. ultimum, and R. solani, increased the pathogenesis related proteins (β-1,3-glucanase and chitinase) and defense enzymes (phenylalanine ammonia lyase, peroxidase, polyphenol oxidase, and catalase) on day 5 after inoculation. In the current study, we first report the facultative methylotrophy in pink pigmented D. lacustris, B. subtilis, and B. cereus and their antagonistic potential against fungal pathogens. Direct antagonistic and ISR effects of these isolates against fungal pathogens of tomato evidenced their possible use as a biocontrol agent.

Evaluation of Biosynthetic Pathways of 2Н- and 13С-Labeled Amino Acids by an Obligate Methylotrophic Bacterium Methylobacillus Flagellatum and a Facultative Methylotrophic Bacterium Brevibacterium Methylicum

Evaluation of Biosynthetic Pathways of 2Н- and 13С-Labeled Amino Acids by an Obligate Methylotrophic Bacterium Methylobacillus Flagellatum and a Facultative Methylotrophic Bacterium Brevibacterium Methylicum

Methylobacterium indicum sp. nov., a facultative methylotrophic bacterium isolated from rice seed

Two pink pigmented, Gram-negative, motile, aerobic, rod shaped endophytic bacteria designated as SE2.11T and SE3.6 were isolated in different experiments from surface sterilized rice seeds. Both strains grew optimally at 28°C temperature. They were positive for catalase and nitrate reduction. The 16S rRNA gene sequence of the strains SE2.11T and SE3.6 displayed between 98.1% and 97.2% similarities with the validly published species of the genus Methylobacterium. The major cellular fatty acid was C18:1 ω7c in both the strains, a characteristic feature observed for members of the genus Methylobacterium. The predominant polar lipids were phospholipids including phosphatidylglycerol (PG), phosphatidylethanolamine (PE) and diphosphatidylglycerol (DPG). Phylogenetic analysis of 16S rRNA gene sequences resulted in the formation of a coherent cluster of strains SE2.11T and SE3.6 with closest relative Methylobacterium platani JCM 14648T. However, digital DNA–DNA hybridization (dDDH) of strains SE2.11T and SE3.6 with the closest type strain M. platani JCM 14648T revealed similarity of 35.5% and 35.4%, respectively. Further, the ANI analysis of strains SE2.11T and SE3.6 genomes revealed only 87.9% identity with M. platani JCM 14648T. Based on differences in biochemical, chemotaxonomic characteristics along with low identity at whole genome level we conclude that both strains represent a novel species of the genus Methylobacterium, for which the name Methylobacterium indicum sp. nov., is proposed. The type strain Methylobacterium indicum is SE2.11T (=MTCC 12298T=JCM 30761T) and SE3.6 (=MTCC 12299=JCM 30762) is another strain.