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The narrow zone of soil around the plant roots with maximum microbial activity termed as rhizosphere. Rhizospheric bacteria promote the plant growth directly or indirectly by providing the nutrients and producing antimicrobial compounds. In this study, the rhizospheric microbiota of peanut plants was characterized from different farms using an Illumina-based partial 16S rRNA gene sequencing to evaluate microbial diversity and identify the core microbiome through culture-independent (CI) approach. Further, all rhizospheric bacteria that could grow on various nutrient media were identified, and the diversity of those microbes through culture-dependent method (CD) was then directly compared with their CI counterparts. The microbial population profiles showed a significant correlation with organic carbon and concentration of phosphate, manganese, and potassium in the rhizospheric soil. Genera like Sphingomicrobium, Actinoplanes, Aureimonas _A, Chryseobacterium, members from Sphingomonadaceae, Burkholderiaceae, Pseudomonadaceae, Enterobacteriaceae family, and Bacilli class were found in the core microbiome of peanut plants. As expected, the current study demonstrated more bacterial diversity in the CI method. However, a higher number of sequence variants were exclusively present in the CD approach compared to the number of sequence variants shared between both approaches. These CD-exclusive variants belonged to organisms that are more typically found in soil. Overall, this study portrayed the changes in the rhizospheric microbiota of peanuts in different rhizospheric soil and environmental conditions and gave an idea about core microbiome of peanut plant and comparative bacterial diversity identified through both approaches.

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This study is designed to investigate Escherichia coli for the antibiotic resistance genes (ARGs) and integrons from healthy as well as diarrhoeic/diseased animals/birds' faecal samples. A total of eight samples were selected for the study; from each animal, two samples were taken, one from healthy animals/birds and one from diarrhoeic/diseased animals/birds. Antibiotic sensitivity testing (AST) and whole genome sequencing (WGS) was performed for selected isolates. The E. coli isolates showed resistance to moxifloxacin, followed by erythromycin, ciprofloxacin, pefloxacin, tetracycline, levofloxacin, ampicillin, amoxicillin, and sulfadiazine (4/8, 50.00% each). The E. coli isolates were 100% sensitive to amikacin, followed by chloramphenicol, cefixime, cefoperazone, and cephalothin. A total of 47 ARGs from 12 different antibiotic classes were detected among the eight isolates by WGS. The different classes of antibiotics included aminoglycoside, sulphonamide, tetracycline, trimethoprim, quinolone, fosfomycin, phenicol, macrolide, colistin, fosmidomycin, and multidrug efflux. The class 1 integrons were detected in 6/8 (75.00%) isolates with 14 different gene cassettes.

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Textile wastewater contains dyes mixed with other contaminants in various concentrations. Bacteria-mediated decolorization and degradation of azo dyes have achieved momentum as a method of treatment attributed to their inexpensive, eco-friendly, and application to a wide range of azo dyes. However, a single species of bacteria is inefficient in decolorizing diverse groups of dyes which is one of the most significant challenges for environmental technologists working in bioremediation. In the present study, an aerobic bacterial consortium AUJ consisting of six different bacterial strains (Pseudomonas stutzeri AK1, Pseudomonas stutzeri AK2, Pseudomonas stutzeri AK3, Bacillus spp. AK4, Pseudomonas stutzeri AK5, and Pseudomonas stutzeri AK6) removed the individual azo dyes in the 24-94% range when used in more than 200 ppm concentration within 72-96 h. In addition, the consortium was able to decolorize 52.19% mixed dyes (100 ppm) and 44.55% Acid blue 113 when used at a concentration as high as 1100 ppm within 96 h. Optimization of various nutritional and environmental parameters revealed that glucose and yeast extract were the preferred carbon and nitrogen source, respectively, and analysis of treated dye products using high-performance liquid chromatography (HPLC), Fourier-transform infrared spectroscopy (FTIR), and gas chromatography-mass spectrometry (GC-MS) confirmed the breakdown of dye. In all, we present a bacterial consortium with a good ability of dye decolorization that can be used for degrading a wide variety of azo dyes.

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BACKGROUND: Sequencing driven metagenomics studies have been instrumental in various aspects of microbiology including identification of newer taxa. While this culture-independent approach has its own merits and demerits, several studies have focussed on comparing it with traditional culture-dependent (CD) approach. However, most of these comparative studies rely on Sanger sequencing of complete 16S rRNA gene from pure culture colonies to determine the culturable bacterial diversity. This approach undercounts culturable diversity as only fewer isolates are selected, sequenced, and identified. METHODS: In this study, we have used an Illumina based partial 16S sequencing to identify all the microbes growing on the media and directly comparing with its culture-independent (CI) counterpart. Eight different media were used to target different organisms from soil. Diversity on these media were compared with their CI counterpart. The NGS data was analysed using DADA2 to provide more resolution to the data. RESULTS: In line with studies of similar nature, current study presented higher bacterial diversity in CI approach. However, the current study reflected that a greater number of sequence variants were missed out in CI approach as compared to number of sequence variants shared with CD approach. We observed around 322 (5.98%) ASVs (Amplicon Sequence Variants) exclusively present in CD samples while, 234 (4.35%) ASVs were shared between both approaches. Most of these 322 CD exclusive ASVs were classified as Enterobacteriaceae family and Bacillus genus, with several ASVs annotated at the species level as well, and these organisms are more commonly observed in soil and were also detected in CI approach. Furthermore, 22 genera were exclusively detected in CD samples, most of which were reported from soil and water.

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The rhizosphere, a narrow zone of soil near plant roots, is a hot spot for microbial activity. Rhizosphere microbiota directly or indirectly benefit plants by supplementing nutrients, producing beneficial chemicals, or suppressing pathogens. Plants attract and modulate bacteria within the rhizosphere by releasing exudates. Plants also tend to select the rhizosphere microbiota based on their needs; a phenomenon termed as "rhizosphere effect". In this study, we characterized the rhizosphere microbiota of peanut plants across the crop development cycle from pre-sowing of seeds to post-harvest of crop under field conditions. The rhizosphere and bulk soil samples from different crop developmental stages were also compared. The composition of bulk soil microbiota resembled microbiota of pre-sowing and post-harvest soil and was markedly different from rhizosphere soil samples. Rhizosphere samples were enriched with multiple organisms mostly from the Proteobacteria, Firmicutes and Bacteroidota phyla. Differences in diversity were observed among the rhizosphere samples but not in bulk soil across different crop development stages. Pseudomonas_M indica was highly enriched during the germination of seeds. Furthermore, Plant Growth Promoting (PGP) bacteria like Bacillus were enriched during the middle stages of crop development but there was a decline in PGP organisms in the matured crop stage. We also observed a significant association of pH and Electrical Conductivity (EC) with the profiles of microbial community. Overall, this study portrayed the changes in rhizosphere microbiota of peanut during different developmental stages of crop and may help to design stage specific bio-strategies such as bio-fertilizer to improve crop yield.

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In dromedary camels, which are pseudo-ruminants, rumen or C1 section of stomach is the main compartment involved in fiber degradation, as in true ruminants. However, as camels are adapted to the harsh and scarce grazing conditions of desert, their ruminal microbiota makes an interesting target of study. The present study was undertaken to generate the rumen microbial profile of Indian camel using 16S rRNA amplicon and shotgun metagenomics. The camels were fed three diets differing in the source of roughage. The comparative metagenomic analysis revealed greater proportions of significant differences between two fractions of rumen content followed by diet associated differences. Significant differences were also observed in the rumen microbiota collected at different time-points of the feeding trial. However, fraction related differences were more highlighted as compared to diet dependent changes in microbial profile from shotgun metagenomics data. Further, 16 genera were identified as part of the core rumen microbiome of Indian camels. Moreover, glycoside hydrolases were observed to be the most abundant among all Carbohydrate-Active enzymes and were dominated by GH2, GH3, GH13 and GH43. In all, this study describes the camel rumen microbiota under different dietary conditions with focus on taxonomic, functional, and Carbohydrate-Active enzymes profiles.

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India is considered as the home tract of some of the best buffalo breeds. However, the genetic structure of the Indian river buffalo is poorly understood. Hence, there is a need to characterize the populations and understand the genetic structure of various buffalo breeds for selection and to design breeding strategies. In this study, we have analyzed genetic variability and population structure of seven buffalo breeds from their respective geographical regions using Axiom Buffalo Genotyping Array. Diversity, as measured by expected heterozygosity, ranged from 0.364 in Surti to 0.384 in Murrah breed, and pair-wise F ST values revealed the lowest genetic distance between Murrah and Nili-Ravi (0.0022), while the highest between Surti and Pandharpuri (0.030). Principal component analysis and structure analysis unveiled the differentiation of Surti, Pandharpuri, and Jaffarabadi in first two principal components and at K = 4, respectively, while remaining breeds were grouped together as a separate single cluster and admixed. Murrah and Mehsana showed early linkage disequilibrium (LD) decay, while Surti breed showed late decay. In LD blocks to quantitative trait locis (QTLs) concordance analysis, 4.65% of concordance was observed with 873 LD blocks overlapped with 2,330 QTLs. Overall, total 4,090 markers were identified from all LD blocks for six types of traits. Results of this study indicated that these single-nucleotide polymorphism (SNP) markers could differentiate phenotypically distinct breeds like Surti, Pandharpuri, and Jaffarabadi but not others. So, there is a need to develop SNP chip based on SNP markers identified by sequence information of local breeds.

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The walking catfish Clarias magur (Hamilton, 1822) (magur) is an important catfish species inhabiting the Indian subcontinent. It is considered as a highly nutritious food fish and has the capability to walk to some distance, and survive a considerable period without water. Assembly, scaffolding and several rounds of iterations resulted in 3,484 scaffolds covering ∼94% of estimated genome with 9.88 Mb largest scaffold, and N50 1.31 Mb. The genome possessed 23,748 predicted protein encoding genes with annotation of 19,279 orthologous genes. A total of 166 orthologous groups represented by 222 genes were found to be unique for this species. The Computational Analysis of gene Family Evolution (CAFE) analysis revealed expansion of 207 gene families and 100 gene families have rapidly evolved. Genes specific to important environmental and terrestrial adaptation, viz. urea cycle, vision, locomotion, olfactory and vomeronasal receptors, immune system, anti-microbial properties, mucus, thermoregulation, osmoregulation, air-breathing, detoxification, etc. were identified and critically analysed. The analysis clearly indicated that C. magur genome possessed several unique and duplicate genes similar to that of terrestrial or amphibians' counterparts in comparison to other teleostean species. The genome information will be useful in conservation genetics, not only for this species but will also be very helpful in such studies in other catfishes.

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BACKGROUND: Squamous Cell Carcinoma of horn, also known as horn cancer, is a prevailing type of cancer in cattles especially Bos indicus. It is one of the most prevalent disease in Indian bullocks often resulting in death and huge economic losses to farmers. Here, we have reported the use of targeted exome sequencing to identify variants present in horn cancer affected horn mucosa tissue and blood of the same animal to identify some of the prevalent markers of horn cancer. RESULTS: We have observed higher number of variants present in tissue as compared to blood as well as among cancer samples compared to samples from normal animals. Eighty six and 1437 cancer-specific variants were identified among the predicted variants in blood and tissue samples, respectively. Total 25 missense variants were observed distributed over 18 genes. KRT8 gene coding for Keratin8, one of the key constituents of horn, displayed 5 missense variants. Additionally, three other genes involved in apoptosis pathway and two genes involved in antigen presentation and processing also contained variants. CONCLUSIONS: Several genes involved in various apoptotic pathways were found to contain non-synonymous mutations. Keratin8 coding for Keratin, a chief constituent of horn was observed to have the highest number of mutations. In all, we present a preliminary report of mutations observed in horn cancer.

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The present study describes rumen microbiota composition and their functional profiles in Indian Surti buffaloes by metagenomic (MG) and metatranscriptomic (MT) approaches. The study compares samples from buffaloes fed three different proportion of roughages; green and dry type of roughage; and different rumen liquor fractions. Irrespective of sample, Bacteroidetes and Firmicutes were the most predominant bacterial phyla, followed by Proteobacteria, Fibrobacteres and Actinobacteria while, Prevotella, Bacteroides, Ruminococcus and Clostridium were the most abundant genera. Different proportions of taxa were observed in both MG and MT approaches indicating the differences in organisms present and organisms active in the rumen. Higher proportions of fungal taxa were observed in MT while important organisms like Fibrobacter and Butyrivibrio and abundant organisms like Bacteroides and Prevotella were underrepresented in MT data. Functionally, higher proportions of genes involved in Carbohydrate metabolism, Amino acid metabolism and Translation were observed in both data. Genes involved in Metabolism were observed to be underrepresented in MT data while, those involved in Genetic information processing were overrepresented in MT data. Further, genes involved in Carbohydrate metabolism were overexpressed compared to genes involved in Amino acid metabolism in MT data compared to MG data which had higher proportion of genes involved in Amino acid metabolism than Carbohydrate metabolism. In all significant differences were observed between both approaches, different fractions of rumen liquor (liquid and solid) and different proportions of roughage in diet.

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Textile industry is one of the anthropogenic activities that consume a large amount of water and pollute water bodies. It uses a massive amount of dyes, which is one of the main constituents of polluting textile effluent. In the present research, biodegradation of Acid Blue 113 dye, a commonly used textile di-azo dye, has been studied exploiting Pseudomonas stutzeri, strain AK6. The dye (300 ppm) was decolorized up to 86.2% within 96 h. The metabolites of Acid Blue 113 obtained after biodegradation were identified by various analytical techniques viz. HPLC (high-performance liquid chromatography) and GC-MS (gas chromatography-mass spectrometry). Genome analysis of isolate AK6 using IMG/M (Integrated Microbial Genomes and Microbiomes) system supported the role of azoreductase and laccase for the decolorization and degradation of azo dye. The ability of P. stutzeri AK6 to tolerate high amount of dye makes it a potential candidate for bioremediation and pre-processing to remove dyes from textile effluents.

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Members of the genus Aspergillus are extensively studied ascomycetes because of their ability to synthesize high value-added compounds and enzymes of industrial interest. Precise whole genome assembly and gene annotation are significant for gene functional analyses. Here, we report the draft genome sequencing, assembly and whole genome analysis of Aspergillus terreus P14_T3, isolated from rumen sample of cattle fed with coconut-coir. A total of 13,340 protein-coding genes were predicted, among them 493 are involved in degradation of complex carbohydrate polysaccharides. Further, it was found that 29 genes, encoding ß-glucosidase belong to Glycosyl hydrolase (GH) family 1 (3 gene), 3 (17 gene), 5 (4 gene), 17 (3 gene), 132 (2 gene). The tertiary structure of all the ß-glucosidases was designed by homology modeling; modeled structure AtBgl1.3 (GH1), AtBgl3.1 (GH3), AtBgl5.4 (GH5), AtBgl17.1 (GH17) show classical (α/ß) TIM-like barrel motif. Molecular docking of different ß-glucosidases with cellobiose revealed that conserved amino acids i.e. Glu, Trp, Arg, His, Tyr and Asp are taking part in substrate hydrolysis. Moreover, some other amino acids i.e. Ser, Phe, Gln and Asn are found to be involved in hydrogen bond formation and catalysis. These findings may provide valuable insights in designing ß-glucosidases with higher cellulose-hydrolyzing efficiency.

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Microbial colonisation in the forestomach of a ruminant is one of the most crucial factors in determining many of its physiological developments and digestive capabilities. The present study attempts to identify establishment pattern of microbes in relation to food, age and rumen development in the buffalo calves at every fortnight interval from birth to 6 months of age, followed by every month till animals became 1 year of age. Diversity study based on 16S rRNA gene sequencing identified rapidly changing bacterial population during initial 60 days of life, which got assemblage as rumen became physiologically mature with increasing age of animals. A lactate fermenting aerobic to facultative anaerobic genera found during initial 30 days of life were expeditiously replaced by strict anaerobic cellulolytic bacterial population with increasing age. The study confirms that initial colonisation mainly depends on the oral cavity and skin of the mother, followed by the surrounding environment and feed offered, which is reversed in order once animal gets older. Some of the well-described genera based on culture-dependent studies like Ruminococcus spp. were found to be in lesser proportion suggesting an additional role of other microbes or niche in cellulose degradation. We report the presence of Porphyromonas spp. and Mannheimia glucosidal for the first time in bovine infants.

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Eimeria species parasites can cause the enteric disease coccidiosis, most notably in chickens where the economic and welfare implications are significant. Seven Eimeria species are recognized to infect chickens, although understanding of their regional occurrence, abundance, and population structure remains limited. Reports of Eimeria circulating in chickens across much of the southern hemisphere with cryptic genotypes and the capacity to escape current anticoccidial vaccines have revealed unexpected levels of complexity. Consequently, it is important to supplement validated species-specific molecular diagnostics with new genus-level tools. Here, we report the application of Illumina MiSeq deep sequencing to partial 18S rDNA amplicons generated using Eimeria genus-specific primers from chicken caecal contents collected in India. Commercial Cobb400 broiler and indigenous Kadaknath type chickens were sampled under field conditions after co-rearing (mixed type farms, n = 150 chickens for each) or separate rearing (single type farms, n = 150 each). Comparison of MiSeq results with established Internal Transcribed Spacer (ITS) and Sequence Characterised Amplified Region (SCAR) quantitative PCR assays suggest greater sensitivity for the MiSeq approach. The caecal-dwelling Eimeria tenella and E. necatrix dominated each sample set, although all seven species which infect chickens were detected. Two of the three cryptic Eimeria genotypes were detected including OTU-X and OTU-Y, the most northern report for the latter to date. Low levels of DNA representing other Eimeria species were detected, possibly representing farm-level contamination with non-replicating oocysts or Eimeria DNA, or false positives, indicating a requirement for additional validation. Next generation deep amplicon sequencing offers a valuable resource for future Eimeria studies.

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BACKGROUND: The caecal microbiota plays a key role in chicken health and performance, influencing digestion and absorption of nutrients, and contributing to defence against colonisation by invading pathogens. Measures of productivity and resistance to pathogen colonisation are directly influenced by chicken genotype, but host driven variation in microbiome structure is also likely to exert a considerable indirect influence. METHODS: Here, we define the caecal microbiome of indigenous Indian Aseel and Kadaknath chicken breeds and compare them with the global commercial broiler Cobb400 and Ross 308 lines using 16S rDNA V3-V4 hypervariable amplicon sequencing. RESULTS: Each caecal microbiome was dominated by the genera Bacteroides, unclassified bacteria, unclassified Clostridiales, Clostridium, Alistipes, Faecalibacterium, Eubacterium and Blautia. Geographic location (a measure recognised to include variation in environmental and climatic factors, but also likely to feature varied management practices) and chicken line/breed were both found to exert significant impacts (p < 0.05) on caecal microbiome composition. Linear discriminant analysis effect size (LEfSe) revealed 42 breed-specific biomarkers in the chicken lines reared under controlled conditions at two different locations. CONCLUSION: Chicken breed-specific variation in bacterial occurrence, correlation between genera and clustering of operational taxonomic units indicate scope for quantitative genetic analysis and the possibility of selective breeding of chickens for defined enteric microbiota.

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The incidence and severity of respiratory diseases in commercial broiler chicken flocks have increased recently in India because of intensification of the broiler industry. Viral population are predominant in respiratory tract infections and they pose continuous economic burden to poultry industry by causing severe economic losses through decreased productivity [1], [2]. To understand viral metagenome of poultry associated with respiratory infections, we performed DNA virome sequencing and data analysis of broilers from 8 districts of Gujarat State in India. We report high quality sequencing reads and highly abundant DNA viral population present in the infected broiler birds. The raw sequencing data used to perform metagenomic analysis is available in the Sequence Read Archive (SRA) under the BioProject No. PRJNA322592 and Accession No. MAUZ00000000, MAVA00000000, MAVB00000000, MAVC00000000, MAVD00000000, MAVE00000000, MAVF00000000, MAVG00000000 (https://www.ncbi.nlm.nih.gov/bioproject/?term=PRJNA322592).

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Zebu (Bos indicus) is a domestic cattle species originating from the Indian subcontinent and now widely domesticated on several continents. In this study, we were particularly interested in understanding the functionally active rumen microbiota of an important Zebu breed, the Gir, under different dietary regimes. Metagenomic and metatranscriptomic data were compared at various taxonomic levels to elucidate the differential microbial population and its functional dynamics in Gir cattle rumen under different roughage dietary regimes. Different proportions of roughage rather than the type of roughage (dry or green) modulated microbiome composition and the expression of its gene pool. Fibre degrading bacteria (i.e. Clostridium, Ruminococcus, Eubacterium, Butyrivibrio, Bacillus and Roseburia) were higher in the solid fraction of rumen (P<0.01) compared to the liquid fraction, whereas bacteria considered to be utilizers of the degraded product (i.e. Prevotella, Bacteroides, Parabacteroides, Paludibacter and Victivallis) were dominant in the liquid fraction (P<0.05). Likewise, expression of fibre degrading enzymes and related carbohydrate binding modules (CBMs) occurred in the solid fraction. When metagenomic and metatranscriptomic data were compared, it was found that some genera and species were transcriptionally more active, although they were in low abundance, making an important contribution to fibre degradation and its further metabolism in the rumen. This study also identified some of the transcriptionally active genera, such as Caldicellulosiruptor and Paludibacter, whose potential has been less-explored in rumen. Overall, the comparison of metagenomic shotgun and metatranscriptomic sequencing appeared to be a much richer source of information compared to conventional metagenomic analysis.

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Cellulase hydrolyses the cellulose by cleaving the ß-1,4-linkages to produce mono-, oligo- and shorter polysaccharide units. These enzymes have applications in various industries such as pulp and paper, laundry, food and feed, textile, brewing industry and in biofuel production. In the present study we have cloned acid-cellulase gene (Cel-1) from the fosmid library of buffalo rumen metagenomic DNA and functionally expressed it in Escherichia coli. The ORF encoding cellulase consisted of 1176-bp, corresponding to protein of 391 amino acid and has catalytic domain belonging to glycosyl hydrolase family 5. The purified protein has a molecular weight of 43-kDa on SDS-PAGE and its expression was confirmed by western blotting. The tertiary structure of the cellulase (Cel-1) showed a classical (α/ß) TIM-like barrel motif. Model surface charge of Cel-1 predicted that surface near active site was mostly negative which might be responsible for the stability of enzyme at lower pH. The pH and temperature for maximum enzyme activity were 4.5 and 45°C respectively. Various metal ions enhanced the enzyme activity and in presence of Mn+2 activity was significantly increased. Cel-1 hydrolyzed pre-treated wheat straw and released reducing sugars (62.60%). These desirable properties of Cel-1 make it attractive for the bioconversion of biomass.

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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.

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