Genetic diversity and molecular epidemiology of Streptococcus uberis in high-prevalence mastitis herds

Streptococcus uberis is one of the most frequently isolated bacterial species from mastitis cows and presents significant control challenges due to its high genetic diversity. This study aimed to determine genetic variations of Strep. uberis causing mastitis across 3 high-prevalence farms (farm A, B, and C). Isolates were obtained from different longitudinal studies and were used to study relationships between virulence factors (VF), antimicrobial resistance (AMR), and strain diversity. Data on farm management and the mastitis control policy were collected during longitudinal collection of milk samples and revealed different policies among the 3 farms. Among all available Strep. uberis isolates, strains were selected based on the characteristics of infection episodes, choosing a maximum of 2 isolates per episode. Selected isolates were confirmed using MALDI-TOF and subsequently used for whole-genome sequencing (WGS). All VF and AMR genes were categorized into the core genes, as present in all isolates; the accessory gene, as present in more than 50% of all isolates; and the unique genes for the genes that were present in less than 50% of all isolates. Accessory genes and unique genes were used to determine the relationships between VF and AMR using the Fisher exact chi-squared tests. The WGS results from a total of 138 Strep. uberis isolates, obtained from 92 episodes, revealed 7 distinct phylogroups (I to VII) and 32 gene patterns. Farm B, the only farm with long-time dairy experience and managed solely by experienced dairy farmers, exhibited the highest genetic diversity. In contrast, the other farms, dominated by persistent Strep. uberis IMI, showed fewer dominant patterns and lower diversity. A lower AMR prevalence in farm C (10.7%, 6/56) was associated with lower antibiotic use, as antimicrobial usage must be authorized by a local veterinarian, but the farmers of farms A and B designed their own use. In addition, both phylogenetic and farm factors revealed a significant association between virulence and AMR . From all identified 35 VF genes and 16 AMR genes, the core pattern included 21 core genes and 8 accessory genes from the VF genes, whereas the unique genes contained 6 VF and all 16 AMR genes. The results revealed that an increase in the number of AMR genes was associated with greater virulence diversity. Significant correlations were observed with overall VF gene presence or absence (r = 0.437), as well as with the absence of VF genes from the core pattern (R = 0.523). No AMR genes were detected in patterns without VF variations from the core pattern. Furthermore, a significant correlation was demonstrated between AMR genes and specific VF genes, notably with the presence of cfu and the absence of hasA. Thus, these findings highlight the complex relationship between virulence factors and resistance genes, which are potentially influenced by factors such as farm management practices and bacterial traits. This underscores the need for further studies that should be specific to each mastitis scenario, enabling the development of effective control strategies to reduce the impact of Strep. uberis mastitis.