Detection and Molecular characterisation of Virulence Genes in Antibiotic Resistant Staphylococcus aureus from milk in the North West Province, South Africa
Abstract
Staphylococcus aureus (S. aureus) is a common microorganism present on the skin and
mucosal surfaces of humans and animals. However, a variety of infections, ranging from mild
skin infections to severe infections such as bacteraemia, osteomylitis, and toxin-mediated
diseases can be caused by this bacterium. These infections become more severe if strain
harbours antibiotic resistance genes together with virulence genes. The aim of this study was to
investigate the occurrence, antibiotic susceptibilities, virulence genes profiles and genetic
relationships of S. aureus in milk obtained from the North-West Province. To achieve this, 200
samples of raw, tank and pasteurised milk were obtained randomly from supermarkets, shops
and some farms in the North-West Province during the period of May 2012 to April 2013. S.
aureus was isolated and positively identified using morphological, biochemical tests, protein
profile analysis (MALDI-TOF mass spectrometry) and molecular (PCR) methods. The isolates
were characterised by determining their antimicrobial resistance profiles, detection of genes
encoding enterotoxins, exfoliative toxins and collagen adhesins. Moreover, the relationships of
the isolates from different stations and milk types were compared using their antibiotic
inhibition zone diameter data, RAPD-PCR and ERIC-PCR fmgerprinting data.Among all the
samples examined, 30 of 40 raw milk samples (75%), 25 of 85 of tank milk samples (29%) and
10 of 75 pasteurised milk samples (13%) were positive for S. aureus. One hundred and fifty-six
PCR-confirmed S. aureus isolates were obtained from the 75 contaminated milk samples. A
large proportion (60-1 00%) of the isolates was resistant to penicillin G, ampicillin, oxacillin,
vancomycin, teicoplanin and erythromycin. On the contrary, low level resistance (8.3 to 40%)
was observed for gentamycin, kanamycin and sulphamethoxazole. Methicillin resistance was
detected in 59% of the multidrug resistant isolates. However, a small proportion (20.6%) of
these isolates possessed the PBP2a which codes for methicillin resistance in S. aureus. In
addition, 32.7% of isolates possessed the sec gene whereas the sea, seb sed, see, cna, eta,etb
genes were not detected. A total of 18 RAPD patterns were generated from 74 randomly
selected milk isolates while 9 banding patterns was obtained with ERIC-PCR indicating a high
genetic diversity among the isolates. However, the presence of toxin genes was not associated
with any particular genotype. The findings of this study indicate that raw, tank and pasteurised
milk in the North-West Province were contaminated with toxigenic and multi-drug resistant S.
aureus strains. Moreover, even if some of these S .aureus isolates were genetically diverse, in
some situations isolates from different types of milk were very similar based on phenotypic and
genotypic typing assays. This emphasise the need for the implementation of better control
measures to reduce contamination as well as the spread of virulent S. aureus strains. This may
greatly reduce human suffering due to foodborne infections that may occur through the
consumption of contaminated food products.