Insights into the Virulence and Antimicrobial Resistance of Staphylococcus hyicus Isolates from Spanish Swine Farms

Abstract

Staphylococcus hyicus is a significant pathogen in swine, primarily causing exudative epidermitis. Addressing S. hyicus infections requires both the characterization of virulence and antimicrobial resistance (AMR) in farm-recovered isolates. This study aimed to characterize the virulence, AMR, and biofilm formation of S. hyicus isolates from Spanish swine farms. A total of 49 isolates were analyzed, originating from animals with cutaneous, reproductive, and systemic clinical signs. Half of the isolates (49.0%) were positive for at least one virulence factor (VF) gene, with SHETA being the most frequent (28.6%). A high frequency of multidrug resistant (MDR) isolates was observed (83.7%), with significant resistance to commonly used antimicrobials, including lincosamides (83.7%), pleuromutilins (81.6%), penicillins (75.5%), and tetracyclines (73.5%). All isolates exhibited robust in vitro biofilm formation capacity (DC = 15.6 ± 7.0). Significant associations were found between VFs, biofilm formation, and AMR patterns, highlighting the link between the resistance to lincosamides and pleuromutilins (p < 0.001; Φ = 0.57) and macrolides (p < 0.001; Φ = 0.48), and the association of AMR with the ExhC and ExhD VF genes. These findings underscore the need for targeted diagnostics to improve management and therapeutic strategies to mitigate the impact of S. hyicus on swine production.

Keywords: Staphylococcus hyicus; antimicrobial resistance; biofilm formation; exfoliative toxins; exudative epidermitis; virulence.

Figure 2

Antimicrobial resistance (AMR) characterization at the class level of 49 Staphylococcus hyicus isolates from Spanish swine farms. (A) AMR phenotype clustering based on antimicrobial classes, using the unweighted pair group method with arithmetic mean (UPGMA) as the hierarchical clustering method. (B) Principal component analysis (PCA) of the AMR patterns, showing grouping based on the type of sample of each S. hyicus isolate.

Figure 1

Virulence factor (VF) characterization of 49 Staphylococcus hyicus isolates from Spanish swine farms. (A) Frequency (%) of each VF gene and (B) Principal component analysis (PCA) of the six evaluated VF genes, showing grouping based on aminoglycoside antimicrobial resistance (AMR) of each S. hyicus isolate. WT: wild type, susceptible; NWT: non-wild type, resistant.

Figure 3

Network associations between virulence factor (VF) genes and antimicrobial resistance (AMR) at the class level in 49 Staphylococcus hyicus isolates from Spanish swine farms. Node size is determined by the percentage occurrence of the VF gene or AMR class. Edge size is proportional to the magnitude of the association based on the Φ coefficient. The network was constructed using significant associations (p < 0.05).

Figure 4

Biofilm formation of Staphylococcus hyicus isolates from Spanish swine farms. Boxplots illustrating the quantitative biofilm formation of S. hyicus, comparing the type of sample and the clinical signs of the animals, the pig breed, the antimicrobial resistance to penicillins and lincosamides, or the presence of the ExhB virulence factor (VF) gene. Quantification was performed as the difference from the negative control (DC) in absorbance (OD₅₉₅). Each S. hyicus isolate is represented by a dot with a horizontal jitter for visibility. The horizontal box lines represent the first quartile, the median, and the third quartile. Whiskers extend to 1.5 times the interquartile range. Differences between groups were evaluated using the Wilcoxon rank-sum test. The level of statistical significance was represented with asterisks: four asterisks (****) indicated a p-value less than 0.0001; two asterisks (**) indicated a p-value between 0.001 and 0.01; one asterisk (*) indicated a p-value between 0.01 and 0.05; non-significance (ns) indicated a p-value higher than 0.05.