330 Identification and genomic characterization of two novel strains of Prevotella albensis as starch utilizers in the rumen of beef cows

Abstract

Abstract In cattle fed concentrate diets, rumen amylolytic bacteria digest starch into glucose, which is metabolized for growth. Since metagenomics studies have revealed that uncharacterized ruminal amylolytic bacteria far outnumber known starch utilizers, we have been pursing the identification of novel ruminal amylolytic bacteria. The same Operational Taxonomic Unit (OTU) was enriched independently from the rumen fluid of two beef cows after culturing with starch. Since this identification was performed using the V1–V3 region of the 16S rRNA gene, a metagenomic analysis was conducted to determine whether this OTU represented the same strain or different strains of the same species. A total of 9.25 and 9.16 million sequence reads were respectively generated from a select enriched starch culture from each cow, which had a relative abundance for this OTU of 67.9% and 74.0%, respectively. Contigs were assembled using the publicly available software ABySS, with contigs of at least 2kb in length used for further analysis. Of the enzymes identified by gene annotation of these contigs (using a combination of the online tools RAST and BLASTp), the presence of genes encoding α-amylase and lactate dehydrogenase enzymes further supported this OTU as corresponding to a starch utilizer. The alpha-amylase isoforms from the two rumens differed in amino acid length (538 vs 625) and sequence, with their respective closest affiliation being to an uncultured species of Lachnospiraceae (51% amino acid identity) and to Prevotella albensis (95% amino acid identity), respectively. The lactate dehydrogenase isoforms were also found to be different in length (348 aa vs 335 aa) and sequence (100% amino acid identity to Lactobacillus mucosae and 99% to an uncultured species of the genus Olsonella, respectively). These and other gene comparisons together suggest that two strains of the same starch-utilizing OTU have been identified in the rumen of beef cows.