Genomic characterization of coagulase-negative staphylococci including methicillin-resistant Staphylococcus sciuri causing bovine mastitis
Introduction
Staphylococci are responsible for numerous infections in humans and animals including cellulitis, bacteraemia, endocarditis, pneumonia, and mastitis. Staphylococcus aureus is the most pathogenic species in the genus (Weese and van Duijkeren, 2010), and one of the most important aetiological agents of bovine mastitis (Keefe, 2012). In the past decade, the less pathogenic coagulase-negative staphylococci (CoNS), including S. sciuri, have increasingly been recognised as important bovine mastitis pathogens (Levison et al., 2016). S. sciuri is frequently isolated from the skin of humans and animals, the environment (Couto et al., 2003; Nemeghaire et al., 2014; Piessens et al., 2011), and in addition to causing bovine mastitis, is an opportunistic human pathogen (Hu et al., 2015; Taponen et al., 2016).
Worldwide, bacterial mastitis is a significant disease in dairy production systems resulting in reduced production and quality of milk, and significant economic losses (Petrovski et al., 2006). Without effective mastitis control programmes, affected herds in Australasia may have mastitis-related morbidity as high as 35% (Petrovski et al., 2009). The empirical use of topical and parenterally administered antimicrobials for the treatment of bovine mastitis is common, and consequently their use may be associated with the development of antimicrobial resistance in targeted and commensal bacteria (Oliver and Murinda, 2012).
Previously, antibacterial resistance in bovine mastitis staphylococci was negligible. However, CoNS associated with β-lactam, aminoglycoside and tetracycline resistance have become prominent causes of mastitis in dairy cows (Frey et al., 2013). Furthermore methicillin-resistant S. aureus (MRSA) and methicillin-resistant CoNS (MRCoNS) exhibiting multidrug-resistant profiles have been isolated from mastitic cows’ milk samples (Frey et al., 2013). This is a potential public health concern due to the possibility staphylococci containing the SCCmec element may act as reservoirs of antimicrobial resistance determinants which can be transferred via direct transmission of resistant pathogens between different hosts and/or lateral transfer of resistance genes through genetic recombination (Couto et al., 2003; Juuti et al., 2005).
Very few intramammary antimicrobials are approved for the treatment of bovine mastitis. In Australia, the most frequently used intramammary antimicrobials for bovine mastitis belong to the β-lactams class. However as the majority of β-lactam derivatives are ineffective against methicillin-resistant staphylococci, the occurrence and spread of MRSA and MRCoNS infections in dairy herds is a major issue facing the industry. The main objective of the present study was to genetically and phenotypically characterise, for the first time in Australia, a group of MRCoNS isolated from bovine mastitis cases at a single dairy farm.
Section snippets
Milk samples
A total of 320 milk samples (from 87 dairy cows) from a single dairy farm in south-east South Australia with a high incidence of mastitis were submitted frozen to the School of Animal and Veterinary Sciences, University of Adelaide, Roseworthy, South Australia for microbiological testing. Cows ranged from 4 to 10 years of age and were identified by farm staff as having clinical mastitis in one or more quarters based on signs of changes in milk (e.g. clots, wateriness and changed consistency)
Isolation and identification of staphylococci
Fifty-nine of the 320 milk samples (18.4%) grew a single bacterial isolate considered to be the pathogen. Thirty seven of the isolates were identified as CoNS, including S. chromogenes (n = 8), S. fleurettii (4), S. haemolyticus (5), S. sciuri (10), S. simulans (4), S. succinus (3) and S. xylosus (3).
Antimicrobial susceptibility profile and resistance scores
The antimicrobial MICs of the 37 CoNS are presented in Table 1. Nineteen CoNS (51.3%) were resistant to at least one antimicrobial with 15 (37.8%) resistant to two or more antimicrobials. Fourteen
Discussion
In our study, and to the best of our knowledge, we have identified and characterised for the first time in Australia, MRCoNS isolated from the milk of cows with bovine mastitis. PCR and whole genome sequencing confirmed the presence of mecA in five oxacillin resistant CoNS spp. (single isolates from S. haemolyticus, S. fleurettii, S. sciuri and two S. succinus isolates). In addition, whole genome sequencing identified the presence of a mecA homologue in four oxacillin resistant S. sciuri
Funding
This study was supported in part by ARC Linkage project LP130100736 with Zoetis and Luoda Pharma as the main industry partners.
Transparency declarations
None to declare.
Authors contribution
MK participated in study design, carried out culture, analysed culture results, and wrote the preliminary manuscript, GWC, DJT, HV and ADO contributed to writing and discussion, AAA and FH carried out the PCR, RRA carried out culture and tested susceptibility of the isolates, AH contributed to sample collection and provided farm information, MO, SA, and SP, carried out whole genome sequencing and analysis, editing and contributed to discussion, SA contributing to study design, KRP conceived the
Acknowledgements
The authors would like to thank Professor Stefan Schwarz, Amanda Ruggero, Hongfei Pi, Ali Khazandi, Benjamin Linn, Budoor Ayoob, Chelsea Smith, David Oswald, Gabe Laucke, Hannah Cohen and Rhyanna Tedesco for their technical support during this study.
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These authors contributed equally.