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Bacterial diversity associated with different primer pairs on different diets in the rumen microbiome of Kankrej cattle
A comprehensive analysis of the bacterial diversity in the rumen of Kankrej cattle was investigated in this study. Two groups of four animals were assigned to two diets of Dry and Green roughage respectively. In each dietary group, four animals (replicates) were group fed one of three dietary treatments for six weeks each; dietary treatments were K1 (50% concentrate: 50% dry/green roughage), K2 (25% concentrate: 75% dry/green roughage) and K3 (100% dry/green roughage). The rumen samples were collected at the end of six weeks period and separated to solid and liquid fractions. The genomic DNA was extracted from each sample and PCR-amplified for V1-V3, V4-V5 and V6-V8 hypervariable regions, sequenced on 454 Roche platform and analyzed for bacterial diversity using QIIME. A total of 600, 851 pyrotags were analysed in this study. Differences in community composition were based on UniFrac distance metric calculated by primer, diet, treatment, fraction and animal and analysed by Permanova test. Different primers had a significant (P<0.001) effect on community compositions. There was no difference between diets but the inclusion of concentrate had an effect on community composition (P<0.01). Also community compositions between fiber and liquid fractions were different (P<0.01). Phylogenetic analysis revealed significant differences in the rumen microbiome mediated by primer (P<0.001). In primer pair 1 associated bacterial communities, the predominant phyla were Bacteroidetes and Firmicutes which together constituted >90% of abundance. These two phyla were influenced by dietary treatment (P<0.001) and fraction (P<0.05). Among communities associated with primer pairs 2 and 3, Firmicutes was predominant and contributed up to 90% of the fiber fraction. Dietary treatment had an influence on the abundance of Bacteroidetes (P<0.001) and Firmicutes (P<0.001) with K1 showing higher (P<0.001) Bacteroidetes while K2 and K3 treatments had higher (P<0.001) abundance of Firmicutes. Other bacterial phyla such as Proteobacteria and Fibrobacter together contributed up to 6% of the bacterial abundance which was influenced by dietary treatment (P<0.001) and fraction (P<0.001) across all three primer pairs. The identified repertoire of bacterial populations was dependent on the primer pair, as targeting the V1-V3 region resulted in greater diversity profiles of the rumen microbiome in this study. Within each primer pair, there were no differences between dry and green roughages. However, inclusion of concentrate in the diet altered the community composition with noticeable shifts at the phylum level.
Keywords: rumen microbiome; Kankrej cattle; QIIME, concentrate, roughage