George V. Thomas

Cold active microbial lipases : Some hot issues and recent developments

Lipases are glycerol ester hydrolases that catalyze the hydrolysis of triglycerides to free fatty acids and glycerol. Lipases catalyze esterification, interesterification, acidolysis, alcoholysis and aminolysis in addition to the hydrolytic activity on triglycerides. The temperature stability of lipases has regarded as the most important characteristic for use in industry. Psychrophilic lipases have lately attracted attention because of their increasing use in the organic synthesis of chiral intermediates due to their low optimum temperature and high activity at very low temperatures, which are favorable properties for the production of relatively frail compounds. In addition, these enzymes have an advantage under low water conditions due to their inherent greater flexibility, wherein the activity of mesophilic and thermophilic enzymes are severely impaired by an excess of rigidity. Cold-adapted microorganisms are potential source of cold-active lipases and they have been isolated from cold regions and studied. Compared to other lipases, relatively smaller numbers of cold active bacterial lipases were well studied. Lipases isolated from different sources have a wide range of properties depending on their sources with respect to positional specificity, fatty acid specificity, thermostability, pH optimum, etc. Use of industrial enzymes allows the technologist to develop processes that closely approach the gentle, efficient processes in nature. Some of these processes using cold active lipase from C. antarctica have been patented by pharmaceutical, chemical and food industries. Cold active lipases cover a broad spectrum of biotechnological applications like additives in detergents, additives in food industries, environmental bioremediations, biotransformation, molecular biology applications and heterologous gene expression in psychrophilic hosts to prevent formation of inclusion bodies. Cold active enzymes from psychrotrophic microorganisms showing high catalytic activity at low temperatures can be highly expressed in such recombinant strains. Thus, cold active lipases are today the enzymes of choice for organic chemists, pharmacists, biophysicists, biochemical and process engineers, biotechnologists, microbiologists and biochemists.

Multifarious beneficial traits and plant growth promoting potential of Serratia marcescens KiSII and Enterobacter sp. RNF 267 isolated from the rhizosphere of coconut palms (Cocos nucifera L.).

Two plant growth promoting bacteria designated as KiSII and RNF 267 isolated from the rhizosphere of coconut palms were identified as Serratia marcescens and Enterobacter sp. based on their phenotypic features, BIOLOG studies and 16S rRNA gene sequence analysis. Both bacteria exhibited phosphate solubilization, ammonification, and production of indole acetic acid, β-1, 3 glucanase activities and 1-aminocyclopropane-1-carboxylate-deaminase activity. They could also tolerate a range of pH conditions, low temperature and salinity (NaCl). In addition, S. marcescens KiSII exhibited N- fixation potential, chitinase activity, siderophore production and antibiotics production. Seed bacterization with these bacteria increased the growth parameters of test plants such as paddy and cowpea over uninoculated control in green house assay. In coconut seedlings, significant increase in growth and nutrient uptake accompanied with higher populations of plant beneficial microorganisms in their rhizospheres were recorded on inoculation with both the PGPRs. The present study clearly revealed that PGPRs can aid in production of healthy and vigorous seedlings of coconut palm which are hardy perennial crops. They offer a scope to be developed into novel PGPR based bioinoculants for production of elite seedlings that can benefit the coconut farming community and the coconut based ecology.

Occurrence and activity of phosphate-solubilizing fungi from coconut plantation soils

SummaryThe occurrence of phosphate-solubilizing fungi in coconut plantation soil types was investigated. The laterite, alluvial and clayey soils harboured more of the P-solubilizing fungi than the sandy soils. The isolated P-solubilizing fungi solubilized 26 to 74 per cent of the tricalcium phosphate in 5 to 15 days. The competitive saprophytic ability of the active P-solubilizing fungi in soil varied between the isolates. Eight fungi with high P-solubilizing capacity and high competitive saprophytic ability were recognised. They have better capacity to survive in soil and express their role in P-solubilization.

Bespoke microbiome therapy to manage plant diseases

Information gathered with advanced nucleotide sequencing technologies, small molecule detection systems and computational biology is revealing that a community of microbes and their genes, now termed “the microbiome,” located in gut and rhizosphere, is responsible for maintaining the health of human beings and plants, respectively. Within the complete microbiome a “core-microbiome” exists that plays the pivotal role in well being of humans and plants. Recent studies in medicine have shown that an artificial mixture of bacteria representing the core gut microbiome of healthy person when transferred into gut of diseased person results in re-establishment of normal microflora in the latter leading to alleviation from diseased condition. In agriculture, though not exactly in similar manner as in medicine, success in plant disease management has been achieved through transfer of microbiome by mixing disease suppressive soils with disease conducive soils. A study more similar to artificial gut microbiome transfer in medical field has been recently reported in agriculture, in which transfer of microbiome via soil solutions (filtered and unfiltered) has shown ability to alleviate drought stress in Arabidopsis thaliana. However, the exact practice of transferring artificially cultivated core-microbiome as in medicine has not thus far been attempted in plant disease management. Nonetheless, as the gut and rhizosphere microbiome are known to share many common traits, there exists a good scope for accomplishing similar studies in agriculture. Based upon the information drawn from all recent works in microbiome studies of gut and rhizosphere, we propose that tailor-made core-microbiome transfer therapy can be a success in agriculture too and it could become a viable strategy for management of plant diseases in future.

Sequencing and analysis of a South Asian-Indian personal genome

BackgroundWith over 1.3 billion people, India is estimated to contain three times more genetic diversity than does Europe. Next-generation sequencing technologies have facilitated the understanding of diversity by enabling whole genome sequencing at greater speed and lower cost. While genomes from people of European and Asian descent have been sequenced, only recently has a single male genome from the Indian subcontinent been published at sufficient depth and coverage. In this study we have sequenced and analyzed the genome of a South Asian Indian female (SAIF) from the Indian state of Kerala.ResultsWe identified over 3.4 million SNPs in this genome including over 89,873 private variations. Comparison of the SAIF genome with several published personal genomes revealed that this individual shared ~50% of the SNPs with each of these genomes. Analysis of the SAIF mitochondrial genome showed that it was closely related to the U1 haplogroup which has been previously observed in Kerala. We assessed the SAIF genome for SNPs with health and disease consequences and found that the individual was at a higher risk for multiple sclerosis and a few other diseases. In analyzing SNPs that modulate drug response, we found a variation that predicts a favorable response to metformin, a drug used to treat diabetes. SNPs predictive of adverse reaction to warfarin indicated that the SAIF individual is not at risk for bleeding if treated with typical doses of warfarin. In addition, we report the presence of several additional SNPs of medical relevance.ConclusionsThis is the first study to report the complete whole genome sequence of a female from the state of Kerala in India. The availability of this complete genome and variants will further aid studies aimed at understanding genetic diversity, identifying clinically relevant changes and assessing disease burden in the Indian population.

Whole Genome Sequencing and Analysis of Plant Growth Promoting Bacteria Isolated from the Rhizosphere of Plantation Crops Coconut, Cocoa and Arecanut

Coconut, cocoa and arecanut are commercial plantation crops that play a vital role in the Indian economy while sustaining the livelihood of more than 10 million Indians. According to 2012 Food and Agricultural organizations report, India is the third largest producer of coconut and it dominates the production of arecanut worldwide. In this study, three Plant Growth Promoting Rhizobacteria (PGPR) from coconut (CPCRI-1), cocoa (CPCRI-2) and arecanut (CPCRI-3) characterized for the PGP activities have been sequenced. The draft genome sizes were 4.7 Mb (56% GC), 5.9 Mb (63.6% GC) and 5.1 Mb (54.8% GB) for CPCRI-1, CPCRI-2, CPCRI-3, respectively. These genomes encoded 4056 (CPCRI-1), 4637 (CPCRI-2) and 4286 (CPCRI-3) protein-coding genes. Phylogenetic analysis revealed that both CPCRI-1 and CPCRI-3 belonged to Enterobacteriaceae family, while, CPCRI-2 was a Pseudomonadaceae family member. Functional annotation of the genes predicted that all three bacteria encoded genes needed for mineral phosphate solubilization, siderophores, acetoin, butanediol, 1-aminocyclopropane-1-carboxylate (ACC) deaminase, chitinase, phenazine, 4-hydroxybenzoate, trehalose and quorum sensing molecules supportive of the plant growth promoting traits observed in the course of their isolation and characterization. Additionally, in all the three CPCRI PGPRs, we identified genes involved in synthesis of hydrogen sulfide (H2S), which recently has been proposed to aid plant growth. The PGPRs also carried genes for central carbohydrate metabolism indicating that the bacteria can efficiently utilize the root exudates and other organic materials as energy source. Genes for production of peroxidases, catalases and superoxide dismutases that confer resistance to oxidative stresses in plants were identified. Besides these, genes for heat shock tolerance, cold shock tolerance and glycine-betaine production that enable bacteria to survive abiotic stress were also identified.

Occurrence ofAzospirillum spp. in coconut-based farming systems

Occurrence of Azospirillum was investigated in coconut-based farming systems, such as high-density multispecies cropping (15 crops), multi-storeyed cropping (3 crops), mixed cropping with tea and coffee (2 crops), intercropping with tropical tubers (5 crops), mixed farming with grasses (3 crops) and in 3 crops, arecanut,Mimosa invisa and sugarcane from other plots. A total of 26 plantation crops and intercrops were included in the study. Incidence of Azospirillum was determined by 2,3,5-triphenyl-tetrazolium chloride reduction and by culturing root fragments in N-free semisolid malate medium. Root samples from guava, mango and mimosa did not show any tetrazolium reduction or sub-surface pellicular growth. The extent of occurrence of Azospirillum seemed to depend upon the crop combinations. In a mixed farming system where guinea grass was one of the component crops, more root fragments of coconut and pepper demonstrated tetrazolium reduction activity than when guinea grass was absent.Azospirillum lipoferum andA. brasilense constituted 42% and 45% of the isolates, respectively, in the coconut-based cropping systems. Isolates from guinea grass, sugarcane and jackfruit exhibited higher nitrogenase (C2H2 reduction) than those isolated from plantation crops, tuber crops and spices. The large variation in the extent of association and nitrogenase activity of isolates from different crops indicated the need for inoculation with efficient cultures in a number of crops in coconut-based cropping systems.

In situ cultivation and incorporation of green manure legumes in coconut basins. An approach to improve soil fertility and microbial activity.

SummaryNine species of green manure legumes were tested for their ability to grow and establish in basins under coconut.Pueraria phaseoloides, Mimosa invisa andCalopogonium mucunoides were suitable yielding 19.43, 17.00 and 14.71 kg of green matter per basin.P. phaseoloides andM. invisa were effectively nodulated with native soil rhizobia as compared toC. mucunoides. Green manuring at 20 kg per palm induced a high level of zymogenic response by microorganisms in the coconut rhizosphere. Enzymatic activity in coconut rhizosphere also increased due to green manure addition.

Distribution of Arbuscular Mycorrhizae Associated with Coconut and Arecanut Based Cropping Systems

Mycorrhizal fungi are widespread in agricultural systems and are especially relevant for organic agriculture. A study was conducted to assess diversity in the arbuscular mycorrhizal (AM) fungi associated with coconut and arecanut intercropping systems of Kasaragod and Thiruvananthapuram districts of Kerala. Mycorrhizal parameters like spore density, root colonization, species richness, and relative occurrence of species were recorded. Coconut cropping system was found to be densely vegetated with diverse AM fungi, Glomus spp., Gigaspora spp., and Acaulospora spp. The diversity of fungal species was found to be maximum in the high density multiple species cropping system plot of CPCRI, Kasaragod with coconut as the main crop and banana, pepper as intercrops and least in farmers’ plot of Mogral Puthur, Kasaragod with arecanut as main crop and banana as the intercrop. A total of fourteen AM fungi were identified from coconut and arecanut intercropping systems. Glomusaggregatum, Funneliformis mosseae, and Rhizophagus fasiculatum were observed in both the cropping systems studied. Among the six edaphic factors analysed, an inverse relationship of mycorrhizal population with soil pH and soil phosphorous was observed. An abundance of P-solubilizers, fluorescent Pseudomonas spp., N-fixers, Bacillus spp., and Trichoderma spp. were observed in the rhizosphere, with Bacillus spp. showing the maximum association with AM fungi. The higher AMF colonization in coconut cropping system imply that the AM fungi colonization is primarily dependent on the host plant rather than the climatic/edaphic components of an area.

Diversity, richness and degree of colonization of arbuscular mycorrhizal fungi in coconut cultivated along with intercrops in high productive zone of Kerala, India

Degree of arbuscular mycorrhizal (AM) association and its diversity were investigated in the coconut palm (Cocos nucifera L.) cultivated in crop mixed system under rain-fed condition in a highly productive humid tropical zone in Malappuram district of Kerala, India. Forty AM species belonging to ten genera viz. Acaulospora, Claroideoglomus, Dentiscutata, Diversispora, Funneliformis, Gigaspora, Glomus, Redeckera, Scutellospora and Septoglomus were recorded indicating high level of AM richness in coconut rhizosphere. Of these, eighteen AM fungal species are being reported for first time from this palm. Claroideoglomus, Glomus and Gigaspora were the most commonly present genera and Claroideoglomus etunicatum and Glomus aggregatum the common species. The diversity indices (Shannon indices values) of AM fungi associated with coconut palm in a crop mixed system and mono crop varied significantly. The Shannon index, Simpson’s index and evenness ranged from 1.40 ± 0.10 to 2.70 ± 0.01, 0.65 ± 0.033 to 0.90 ± 0.004, 0.51 ± 0.01 to 0.82 ± 0.01, respectively. Correlation between soil physio chemical characters and mycorrhizal parameters were worked out. Soil pH had negative correlation with spore count, root colonization, species richness, Shannon H and Simpson indices but had positive correlation with species evenness. Electrical conductivity showed positive correlation with root mycorrhizal colonization which indicated active AMF mediated uptake of nutrients.