Anand B. Patel

Metagenome of Mehsani Buffalo Rumen Microbiota: An Assessment of Variation in Feed-Dependent Phylogenetic and Functional Classification

Aim: To gain a greater understanding of the ecology and metabolic potential of the rumen microbiome with the changes in the animal diet. Methods: Diet composed of varying proportion of green and dry roughages along with grains was given to 8 Mehsani buffaloes, and rumen metagenome was sketched using shotgun semiconductor sequencing. Results: In the present study, the Bacteroidetes were found to be dominant at the phyla level and Prevotella at the genus level. The ratio of Firmicutes to Bacteroidetes was found to be higher in the solid fraction as compared to the liquid fraction. In the solid fraction of the dry roughage group, the significant increment (p < 0.05) in Bacteroidetes abundance was observed with increment of roughage concentration. At the genus level, Clostridium significantly increased with the increment in roughage concentration. A comparison of glycoside hydrolase and cellulosome functional genes revealed more glycoside hydrolase 3 encoding genes with higher fiber diet and significant difference in carbohydrate-active enzymes family composition between green and dry roughage groups of the liquid fraction. Conclusion: The present study provides a base to understand the modulating behavior of microbiota which can be manipulated to improve livestock nutrient utilization efficiency and for targeting the efficient catabolism of complex carbohydrate molecules as well.

High Potential Source for Biomass Degradation Enzyme Discovery and Environmental Aspects Revealed through Metagenomics of Indian Buffalo Rumen

The complex microbiomes of the rumen functions as an effective system for plant cell wall degradation, and biomass utilization provide genetic resource for degrading microbial enzymes that could be used in the production of biofuel. Therefore the buffalo rumen microbiota was surveyed using shot gun sequencing. This metagenomic sequencing generated 3.9 GB of sequences and data were assembled into 137270 contiguous sequences (contigs). We identified potential 2614 contigs encoding biomass degrading enzymes including glycoside hydrolases (GH: 1943 contigs), carbohydrate binding module (CBM: 23 contigs), glycosyl transferase (GT: 373 contigs), carbohydrate esterases (CE: 259 contigs), and polysaccharide lyases (PE: 16 contigs). The hierarchical clustering of buffalo metagenomes demonstrated the similarities and dissimilarity in microbial community structures and functional capacity. This demonstrates that buffalo rumen microbiome was considerably enriched in functional genes involved in polysaccharide degradation with great prospects to obtain new molecules that may be applied in the biofuel industry.

Metagenomic analysis of buffalo rumen microbiome: Effect of roughage diet on Dormancy and Sporulation genes.

Buffalo rumen microbiome experiences a variety of diet stress and represents reservoir of Dormancy and Sporulation genes. However, the information on genomic responses to such conditions is very limited. The Ion Torrent PGM next generation sequencing technology was used to characterize general microbial diversity and the repertoire of microbial genes present, including genes associated with Dormancy and Sporulation in Mehsani buffalo rumen metagenome. The research findings revealed the abundance of bacteria at the domain level and presence of Dormancy and Sporulation genes which were predominantly associated with the Clostridia and Bacilli taxa belonging to the phyla Firmicutes. Genes associated with Sporulation cluster and Sporulation orphans were increased from 50% to 100% roughage treatment, thereby promoting sporulation all along the treatments. The spore germination is observed to be the highest in the 75% roughage treatment both in the liquid and solid rumen fraction samples with respect to the decrease in the values of the genes associated with spore core dehydration, thereby facilitating spore core hydration which is necessary for spore germination.

Transcriptomic dissection of myogenic differentiation signature in caprine by RNA-Seq.

Muscle growth and development from the embryonic to the adult stage of an organism consists of a series of exquisitely regulated and orchestrated changes in expression of genes leading to muscle maturation. In this study, we performed whole transcriptome profiling of adult caprine skeletal muscle derived myoblast and fused myotubes. Using Ion Torrent PGM sequencing platform, a total of 948,776 and 799,976 reads were generated in myoblasts and fused myotubes, respectively. The sequence reads were analyzed on CLC Genomics Workbench using Bos taurus RNA database to study the gene expression in both stages to study different genes responsible for muscle development and regeneration. The up and down-regulated genes were analyzed for gene ontology (GO) and KEGG pathways by Database for Annotation, Visualization and Integrated Discovery (DAVID) database. We found many genes exclusive to multinuclear fused myotubes and contractile nature of skeletal muscle, whereas up-regulated genes in myoblast stage were related to cell division and transcriptional regulation. Out of 27 genes selected for expression validation by RT-qPCR (reverse transcriptase-quantitative polymerase chain reaction), 19 genes showed the expression pattern comparable with CLC Genomics Workbench findings. Further, mRNA originated muscle specific microRNAs (miRNA-1 and miRNA-133b) were also observed in the fused myotubes along with other miRNAs with possible importance in muscle development. This study highlights important genes responsible for muscle development and differentiation in adult skeletal muscle system.

Transcriptome analysis and SNP Identification in SCC of Horn in (Bos indicus) Indian cattle

Single Nucleotide Polymorphisms (SNPs) have become the marker of choice for genome wide association studies. In order to provide the best genome coverage for the analysis of disease, production and performance traits, a large number of relatively evenly distributed SNPs are needed. The main objective of present work was to identify large numbers of gene-associated SNPs using high-throughput sequencing in squamous cell carcinoma of horn. RNA-seq analysis was conducted on 2 tissues viz. Horn Cancer (HC) and Horn Normal (HN) in Kankrej breed of cattle. A total of 909,362 reads with average read length of 405 bp for HC and 583,491 reads with average read length of 411 bp for HN were obtained. We found 9532 and 7065 SNPs as well as 1771 and 1172 Indels in HC and HN, respectively, from which, 7889 SNPs and 1736 Indels were uniquely present in HC, 5886 SNPs and 1146 Indels were uniquely present in HN and reported first time in Bos indicus, whereas the rest are already reported in Bos taurus dbSNP database. The gene-associated SNPs and Indels were high in upregulated genes of HC as compared to HN. Analysis of differentially expressed genes was identified, these genes are involved in regulation of cell proliferation, apoptosis, gene transcription, cell survival and metabolism through various metabolic pathways. The result of transcriptome expression profiling was validated using Real Time quantitative PCR in nine randomly selected genes. We identified numbers aberrant signaling pathways responsible for carcinogenesis in HC which are also commonly altered in squamous cell carcinoma (SCC) of lung in human being. We conclude that a large number of altered genes and dysfunction of multiple pathways are involved in the development of Horn Cancer. The present findings contribute to theoretical information for further screening of genes and identification of markers for early diagnosis of HC as well as SNPs identified in this report provide a much needed resource for genetic studies in B. indicus and shall contribute to the development of a high density SNP array. Validation and testing of these SNPs using SNP arrays will form the material basis for gene associated SNPs in HC.

Functional gene profiling through metaRNAseq approach reveals diet-dependent variation in rumen microbiota of buffalo (Bubalus bubalis)

Recent advances in next generation sequencing technology have enabled analysis of complex microbial community from genome to transcriptome level. In the present study, metatranscriptomic approach was applied to elucidate functionally active bacteria and their biological processes in rumen of buffalo (Bubalus bubalis) adapted to different dietary treatments. Buffaloes were adapted to a diet containing 50:50, 75:25 and 100:0 forage to concentrate ratio, each for 6 weeks, before ruminal content sample collection. Metatranscriptomes from rumen fiber adherent and fiber-free active bacteria were sequenced using Ion Torrent PGM platform followed by annotation using MG-RAST server and CAZYmes (Carbohydrate active enzymes) analysis toolkit. In all the samples Bacteroidetes was the most abundant phylum followed by Firmicutes. Functional analysis using KEGG Orthology database revealed Metabolism as the most abundant category at level 1 within which Carbohydrate metabolism was dominating. Diet treatments also exerted significant differences in proportion of enzymes involved in metabolic pathways for VFA production. Carbohydrate Active Enzyme(CAZy) analysis revealed the abundance of genes encoding glycoside hydrolases with the highest representation of GH13 CAZy family in all the samples. The findings provide an overview of the activities occurring in the rumen as well as active bacterial population and the changes occurring through different dietary treatments.

Complete Genome Sequence of Escherichia Phage ADB-2 Isolated from a Fecal Sample of Poultry

ABSTRACT Escherichia phage ADB-2 was isolated from a chicken fecal sample. It is a virulent phage and shows effective inhibition of Escherichia coli strains. Here we announce the completely sequenced genome of Escherichia phage ADB-2, and major findings from its annotation are described.

Draft Genome Sequence of Paenibacillus sp. Strain DMB20, Isolated from Alang Ship-Breaking Yard, Which Harbors Genes for Xenobiotic Degradation

ABSTRACT Paenibacillus sp. strain DMB20, in cometabolism with other Proteobacteria and Firmicutes, exhibits azoreduction of textile dyes. Here, we report the draft genome sequence of this bacterium, consisting of 6,647,181 bp with 7,668 coding sequences (CDSs). The data presented highlight multiple sets of functional genes associated with xenobiotic compound degradation.

Comparative analysis of SNP candidates in disparate milk yielding river buffaloes using targeted sequencing

River buffalo (Bubalus bubalis) milk plays an important role in economy and nutritious diet in several developing countries. However, reliable milk-yield genomic markers and their functional insights remain unexposed. Here, we have used a target capture sequencing approach in three economically important buffalo breeds namely: Banni, Jafrabadi and Mehsani, belonging to either high or low milk-yield group. Blood samples were collected from the milk-yield/breed balanced group of 12 buffaloes, and whole exome sequencing was performed using Roche 454 GS-FLX Titanium sequencer. Using an innovative approach namely, MultiCom; we have identified high-quality SNPs specific for high and low-milk yield buffaloes. Almost 70% of the reported genes in QTL regions of milk-yield and milk-fat in cattle were present among the buffalo milk-yield gene candidates. Functional analysis highlighted transcriptional regulation category in the low milk-yield group, and several new pathways in the two groups. Further, the discovered SNP candidates may account for more than half of mammary transcriptome changes in high versus low-milk yielding cattle. Thus, starting from the design of a reliable strategy, we identified reliable genomic markers specific for high and low-milk yield buffalo breeds and addressed possible downstream effects.

Draft Genome Sequence of Achromobacter sp. Strain DMS1, Capable of Degrading Polyaromatic Hydrocarbons Isolated from the Industrially Perturbed Environment of Amlakhadi Canal, India

ABSTRACT Here, we report the draft genome sequence of Achromobacter sp. strain DMS1, which is 4.9 Mbp and has 3,727 coding sequences (CDSs), and is capable of degrading xenobiotic compounds and harboring genes for aromatic hydrocarbon metabolism. Its genome will unravel the basic mechanism involved in bioremediation of anthropogens.