Genomic diversity of Streptococcus uberis isolated from clinical mastitis of cattle in selected areas of Bangladesh

Streptococci are the major etiology in mastitis, a cause of huge economic losses in the dairy industries. Streptococcus (S.) agalactiae, S. dysagalactiae and S. uberis are mostly encountered in bovine mastitis; however, data on the diversity and characteristics of Streptococcus in clinical mastitis of cattle in Bangladesh is lacking. Thus, the present study was aimed to determine the diversity and antimicrobial resistance pattern of Streptococcus spp. isolated from clinical mastitis of cattle reared in Bangladesh. A total of 105 milk samples comprising eighty (80) from cattle with clinical mastitis (CCM) and twenty-five (25) from apparently healthy cattle (AHC) in four prominent dairy farms and one dairy community were purposively collected and examined in this study. Milk samples were enriched in Luria Bertani broth (LB) and Streptococcus spp. was isolated on Modified Edwards Medium and identified by 16S rRNA gene sequencing. Among eighty (80) clinical samples, eighteen (18) were positive for Streptococcus spp. while none of the milk from AHC revealed Streptococcus by cultural and molecular examination. Sequencing and phylogenetic analysis identified 55.6%, 33.3%, 5.6% and 5.6% of the Streptococcus isolates as S. uberis, S. agalactiae, S. hyovaginalis and S. urinalis, respectively. Antibiotic sensitivity testing with antimicrobials commonly used to treat clinical mastitis revealed 100%, 100%, and 30% of the S. agalactiae, S. hyovaginalis, and S. uberis as multidrug-resistant, respectively. Molecular characterization through whole genome sequencing of five (5) S. uberis isolates identified at least two novel ST types of S. uberis circulating in the study areas with one ST (4/5 isolates) clustered with the isolates from China, India and Thailand, and the other (1/5) with UK, Ireland and Australia. Pan-genome analysis and phylogeny of the core genome sequences also clustered the isolates into two sub-clusters, indicating the presence of at least two different subtypes of S. uberis in the study area. On virulence profiling, all the isolates of this study were found to harbor at least 35 virulence and putative virulence genes probably associated with intramammary infection (IMI) indicating all the S. uberis isolated in this study as potential pathogen. From the overall findings it was evident that Strepococcus occurring in bovine mastitis are diverse and S. uberis genome carries an array of putative virulence factors which need to be investigated genotypically and phenotypically to identify a specific trait or determinant governing the virulence and fitness of this bacterium. Moreover, Streptococcus isolated in this study carried multidrug resistance which needs careful consideration during the selection of a treatment regimen for mastitis.