Staphylococcus aureus is a well-known commensal pathogen of many animal species, including humans, that causes community and nosocomial infections (Li et al., 2018). It is regarded as one of the world’s leading causes of food consumption-related disease outbreaks (Abdalrahman et al., 2015). Previous studies have revealed that S. aureus has often been isolated from raw milk and retail meat (Johler et al., 2018; Sahibzada et al., 2018).

S. aureus-associated food poisoning in humans and similar intramammary infections in animals are produced by strains with the capacity to produce a wide range of virulence factors, including enterotoxins (se); toxic shock syndrome toxin-1 (tst); exfoliative toxins (eta and etb); leukocidins (lukD/E/FS); and haemolysins (hla, hld, hlg, and hlgv; Le Marechal et al., 2011; Wilson et al., 2011). The role of virulence factors in disease causation is not as definitive as foodborne illness, and a plethora of factors are likely to be involved in disease pathogenesis (Fijalkowski et al., 2013).

Most S. aureus isolates are host-specific, and the potential for animals to act as a source of S. aureus infections for humans has been shown for some clonal lineages, such as ST398 (Mediavilla et al., 2012; Yan et al., 2014). Methicillin-resistant S. aureus (MRSA) ST398 can colonize animals and humans and can be transmitted between them (Li et al., 2015). MRSA ST398 has been isolated in Australia (Mitchell et al., 2014), New Zealand (Deborah et al., 2014), and Spain (Lozano et al., 2012). MSSA ST398 has been recovered from pigs in Germany and northeast China (Kadlec and Schwarz, 2010; Yan et al., 2014), humans in France and Taiwan, China (Rasigade et al., 2010; Huang and Chen, 2020), and retail meat in USA and Shanxi, China (Waters et al., 2011; Li et al., 2015). Recently, infections caused by MSSA ST398 have been described in humans (Valentin-Domelier et al., 2011; Mediavilla et al., 2012; David et al., 2013), and MSSA ST398 has been associated with efficient transmission between people, with greater capacity for adhesion to human skin (Uhlemann et al., 2012).

The food safety challenge we have faced during the current COVID-19 pandemic has increased concerns about the increasing numbers of outbreaks of staphylococcal food-borne intoxication. As a commensal bacterium in livestock, there has been little focus on MSSA ST398 contamination of retail meat and BTM in China. Therefore, the main objective of this study was to investigate the population structure of MSSA ST398 in retail pork and BTM by using whole-genome sequencing.

In this study, we carried out whole-genome sequencing to investigate the phylogeny and characterization of MSSA ST398 from retail pork and BTM in China. To our knowledge, this is the first time that such type of MSSA ST398 isolates have been whole-genome sequenced and made publicly available. The prevalence of S. aureus in retail pork products from other studies has ranged from 12.0 to 59.7% (Hanson et al., 2011; Kelman et al., 2011; Waters et al., 2011). The results from this study were similar to those in other published reports, since S. aureus was found in 33.5% of retail pork, and they also coincide with other reports on poultry products that the prevalence of S. aureus in poultry products that has been reported worldwide has ranged from 23.5 to 50.5% (Ou et al., 2017). The isolates from the retail pork samples may originate from S. aureus-positive animals, the surrounding environment, humans, and other sources during processing and commercialization in meat counters at supermarkets and retail stores. Our results indicated that these meat items may serve as reservoirs of S. aureus, as with other retail meat samples.

Due to the excessive use of antibiotics, the prevalence of drug-resistant bacteria is increasing. In this study, we observed high resistance rates. The resistance rates to PEN (97.0%), ERY (86.6%), and FFC (83.4%) that were detected in this study have commonly been reported among S. aureus isolates from meat samples (Buyukcangaz et al., 2013; Ge et al., 2017). These high resistance rates may be related to the use of antimicrobials for treating disease and as growth promoters or feed additives in livestock. In this study, the MDR isolate rate of S. aureus (92.5%) from retail pork was consistent with the report that indicated that 90.0% of the strains were isolated from ready-to-eat meat sandwiches in Egypt (Mahmoud et al., 2021). However, this rate was higher than those reported for BTM (55.4%) and retail ready-to-eat foods (75.8%) in China (Zhao et al., 2021; Yang et al., 2016). Retail pork that is contaminated by MDR S. aureus is potentially hazardous, and the food chain may be the key site where resistance is transmitted between the environment and humans. In this regard, it was estimated that 2.8 million patients in the USA will be treated each year for resistant bacteria, and more than 35,000 will die as a result (CDC, 2019).

Several molecular typing methods were used to characterize the isolates, including MLST and spa typing. MLST is a DNA sequencing technology that uses sequence analyses of housekeeping genes to discriminate between isolates. MLST also offers the advantage that it is highly reproducible, which makes it an excellent tool for global comparisons of population structures (Enright et al., 2000). spa typing is specific to staphylococci and analyses the polymorphisms in the protein A gene (Frenay et al., 1996). In this study, eight STs were identified, and ST15 was the predominant type, which was different from the results of other studies. Previous studies have reported that ST5 has been found in retail pork in the United States, while ST9 was predominantly obtained from Asian countries, including China (Bhargava et al., 2011; Li et al., 2017). Importantly, it has been reported that MSSA ST398 isolates collected from retail pork are primarily associated with community- and hospital-acquired MSSA infections in humans (Chen and Huang, 2014). Two spa types were identified among the MSSA ST398 isolates from retail pork, and the predominant type was t034, which is a common and dominant type in Europe and North America (Pantosti, 2012).

The phylogenetic analysis showed that MSSA ST398 from retail pork clustered together with MSSA ST398 from BTM, which suggested that ST398 has a wide range of hosts and is considered to be adapted to the colonization of nonhuman mammals (Golding et al., 2010). In this study, we found that all MSSA ST398 isolates carried DHA-1, which belongs to class C of the Ambler classification and to group 1 of the functional classification of Bush, Jacoby, and Medeiros. These results were different from those of another study in which, among the 56 S. aureus isolates, only one in Iran carried DHA-1 (Shahnaz et al., 2016). In our study, most of the MSSA ST398 isolates harboured blaZ (26/29), which was consistent with the result that most isolates were resistant to PEN. This observation is in line with a previous report that the blaZ gene confers PEN resistance (Johler et al., 2018). According to the results of this study, we suggested that MSSA ST398 could be a reservoir for the resistance genes. With regard to the risk of pathogenicity, the presence of virulence genes among the MSSA ST398 isolates was assessed in this study. All isolates carried eta, hla, and hlb, which coincided with the results of another study (Sun et al., 2019). SEs in particular are involved in human food poisoning (Hennekinne et al., 2012). In this study, we only detected SEs in BTM but none in retail pork, which can be attributed to the differences in geographical regions, sample sources and environments and needs to be further monitored. The detection of virulence genes in MSSA ST398 isolates reveals the lurking threat of retail pork and BTM, suggesting the need for implementing surveillance programs and prevention strategies.

Our findings suggested that the retail pork examined was highly contaminated with S. aureus. In addition, the MSSA ST398 harboured multiple virulence and exhibited multiple antimicrobial resistance. To our knowledge, this is the first report of the detection of MSSA ST398 in retail pork and BTM in China. The whole genome sequencing studies are crucial to survey the global epidemiology of infectious agents, including MSSA ST398, and provided a deeper knowledge of the epidemiology of this bacterium and may help in understanding how to prevent and treat infections without boosting antibiotic resistance.

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/Supplementary Material.

Ethical review and approval was not required for the animal study because The Ethical Statement is not applicable because sample collection from animals has been gathered.

XZ and YLi designed the work. XZ, MH, YZ, QZ, LL, and YLu collected samples. XZ and YLi analyzed and interpreted data. XZ drafted the article. XZ and YLi critically reviewed the article. XZ and CZ revised the manuscript. All authors contributed to the article and approved the submitted version.

This work was supported by the National Key Research and Development Project (2019YFA0904004); Shandong Agricultural Major Applied Technology Innovation Program (SD2019XM009); Major Scientific and Technological Innovation Project in Shandong Province (2019JZZY010719); and The High-Level Talents and Innovative Team Recruitment Program of the Shandong Academy of Agricultural Sciences, China (CXGC2018E10).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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