Vancomycin-intermediate livestock-associated methicillin-resistant Staphylococcus aureus ST398/t9538 from swine in Brazil

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) has been mainly related with pig farming, in Europe and North America, with the ST398 as the most commonly identified type of LA-MRSA. Here we present the draft genome of the first vancomycin-intermediate MRSA ST398/t9538 isolated from a swine presenting exudative epidermitis in Brazil.

Methicillin-resistant Staphylococcus aureus (MRSA) animal infection has been reported since the 1970s and is now referred to as livestock-associated MRSA (LA-MRSA). It has only been since 2005, with the studies of MRSA pigassociated strains from sequence type 398, that LA-MRSA started to have greater importance to the medical-scientific community (Leonard & Markey 2008). In Europe and North America, the ST398 remains the most commonly identified type of LA-MRSA (Smith 2015); nevertheless, clinical infections by LA-MRSA ST398 are still rare.
In South America, LA-MRSA ST398 has only been associated with porcine carriage in Peru (Arriola et al. 2011) and milk contamination in Brazil (Silva et al. 2014). The epidemiology and public health impact of LA-MRSA in South America remains poorly addressed. Here we present the isolation, phenotypic and genomic characterisation of MRSA ST398/t9538 from a swine presenting exudative epidermitis in Brazil.
The strain SA7112 was isolated in 2012, from a skin swab collected from a 45-day-old swine presenting skin exfoliation with sebaceous exudation and crust formation in Rio Grande do Sul state, Brazil. The skin swab was plated on sheep blood agar (5%) and incubated for 24 h at 37ºC. The hemolytic white colonies were identified as S. aureus by polymerase chain reaction (PCR) as described by Kearns et al. (1999). A single colony was used for: antimicrobial susceptibility profiling, research of mecA gene by PCR (Kearns et al. 1999) and further genome sequencing.
Whole genome sequencing was performed through Illumina ® Miseq platform with paired-end library. The de novo assembly was performed with CLC Main Workbench 7.5.1 (CLC Bio, Denmark) and Geneious 8.0.5 (Biomatters Ltd, Auckland, New Zealand) and resulted in 22 scaffolds with an N 50 of 466,156. Mapping and ordering of obtained scaffolds with reference strain S0385, an MRSA ST398 (NC_017333), was performed with CLC Microbial Genomics Module (CLC Bio, Denmark) and Mauve multiple genome aligner (Darling et al. 2010) and demonstrated the existence of two plasmids pSA7112-1 (KX011076) and pSA7112-2 (KX011077) ( Table II). The SA7112 draft genome (LNTF00000000.1) comprises ~2.8 Mbp, with an overall G+C content of 32.89%. Automatic genome annotation was performed with NCBI Prokaryotic Genome Annotation Pipeline.
Chromosomal analysis enabled SA7112 typing as SC-Cmec V(5C2&5) subtype c, spa t9538 and ST398. The ST398 has only been reported in one human methicillinsusceptible S. aureus (MSSA) Brazilian strain (Gales et al. 2015) that was further typed as spa t034. Even though the identified spa t9538 has not been associated with LA-MRSA, it is closely related to t034 that is characterised as a livestock-associated spa type. While this is the first report of LA-MRSA ST398 carrying a SCCmec type V(5C2&5) subtype c in Brazil, it has already been established as the predominant LA-MRSA in Europe (Li et al. 2011).
The SA7112 genome presents the genes encoding aureolysin (aur), beta-hemolysin (hlb) and staphylococ-cal gamma-hemolysins (hlgA, hlgB and hlgC), which are related to bacteria escaping from the host immune system. The lukM-lukF-PV (Panton-Valentine bi-component leukotoxin) and the exfoliative toxin genes were not identified.   With regard to the resistance profile, only the fexA, norA, tetM and tetK resistance genes were detected in SA7112 chromosome while both identified plasmids appear to be mostly responsible for the SA7112 resistance phenotype.