Whole-Genome Sequence Analysis of Antimicrobial Resistance Genes in Streptococcus uberis and Streptococcus dysgalactiae Isolates from Canadian Dairy Herds

Front Vet Sci. 2017 May 22;4:63. doi: 10.3389/fvets.2017.00063. eCollection 2017.

Abstract

The objectives of this study are to determine the occurrence of antimicrobial resistance (AMR) genes using whole-genome sequence (WGS) of Streptococcus uberis (S. uberis) and Streptococcus dysgalactiae (S. dysgalactiae) isolates, recovered from dairy cows in the Canadian Maritime Provinces. A secondary objective included the exploration of the association between phenotypic AMR and the genomic characteristics (genome size, guanine-cytosine content, and occurrence of unique gene sequences). Initially, 91 isolates were sequenced, and of these isolates, 89 were assembled. Furthermore, 16 isolates were excluded due to larger than expected genomic sizes (>2.3 bp × 1,000 bp). In the final analysis, 73 were used with complete WGS and minimum inhibitory concentration records, which were part of the previous phenotypic AMR study, representing 18 dairy herds from the Maritime region of Canada (1). A total of 23 unique AMR gene sequences were found in the bacterial genomes, with a mean number of 8.1 (minimum: 5; maximum: 13) per genome. Overall, there were 10 AMR genes [ANT(6), TEM-127, TEM-163, TEM-89, TEM-95, Linb, Lnub, Ermb, Ermc, and TetS] present only in S. uberis genomes and 2 genes unique (EF-TU and TEM-71) to the S. dysgalactiae genomes; 11 AMR genes [APH(3'), TEM-1, TEM-136, TEM-157, TEM-47, TetM, bl2b, gyrA, parE, phoP, and rpoB] were found in both bacterial species. Two-way tabulations showed association between the phenotypic susceptibility to lincosamides and the presence of linB (P = 0.002) and lnuB (P < 0.001) genes and the between the presence of tetM (P = 0.015) and tetS (P = 0.064) genes and phenotypic resistance to tetracyclines only for the S. uberis isolates. The logistic model showed that the odds of resistance (to any of the phenotypically tested antimicrobials) was 4.35 times higher when there were >11 AMR genes present in the genome, compared with <7 AMR genes (P < 0.001). The odds of resistance was lower for S. dysgalactiae than S. uberis (P = 0.031). When the within-herd somatic cell count was >250,000 cells/mL, a trend toward higher odds of resistance compared with the baseline category of <150,000 cells/mL was observed. When the isolate corresponded to a post-mastitis sample, there were lower odds of resistance when compared with non-clinical isolates (P = 0.01). The results of this study showed the strength of associations between phenotypic AMR resistance of both mastitis pathogens and their genotypic resistome and other epidemiological characteristics.

Keywords: Streptococcus dysgalactiae; Streptococcus uberis; antimicrobial resistance; bovine mastitis; whole-genome sequence.