Evaluation of isolation procedures and chromogenic agar media for the detection of MRSA in nasal swabs from pigs and veal calves

Abstract

Since the emergence of MRSA in livestock, screening of animals for the detection of MRSA is widely practised. Different procedures are published for animal samples but a systematic comparison of methods has not been performed. The objective of this study was to compare three available commonly used procedures and three chromogenic agars for detecting MRSA in nasal swabs from pigs (n = 70) and veal calves (n = 100). Procedures 1 and 2 used a pre-enrichment comprising Mueller Hinton broth with 6.5% NaCl followed by selective enrichment with 4 μg/ml oxacillin + 75 μg/ml aztreonam (procedure 1) and 5 μg/ml ceftizoxime + 75 μg/ml aztreonam (procedure 2) respectively. Procedure 3 used a selective enrichment broth only, containing 4% NaCl, 5 μg/ml ceftizoxime + 50 μg/ml aztreonam. After selective enrichment, media were streaked on to three different chromogenic agars. Significantly more MRSA were found for pig as well as for veal calf samples with procedures 1 and 2. No significant differences were found between procedures 1 and 2. For nasal swabs from pigs significantly more MRSA-positive samples were found when MRSA Screen (Oxoid) or MRSASelect™ (Bio-Rad) agars were used compared to MSRA ID (bioMérieux). For calf samples no significant differences between the different agars were found.

In conclusion, the results of this study show that procedures 1 and 2, both using additional high salt pre-enrichment are superior and should be recommended for MRSA detection in nasal swabs from pigs and veal calves. The preferred choice of chromogenic agar depends on the sample matrix.

Introduction

The prevalence of Methicillin resistant Staphylococcus aureus (MRSA) is increasing world-wide, especially since the emergence of community-acquired and animal-related MRSA (Khanna et al., 2008, Nahimana et al., 2006, Tiemersma et al., 2004). Recently, a specific MRSA clone has been reported at unexpected high prevalence among pig farmers and veterinarians in different geographical areas (Voss et al., 2005, Weese and van Duijkeren, 2009). Strains belonging to this clone are resistant to SmaI macrorestriction and therefore referred to as non-typable (NT-MRSA). They all belong to Multi Locus Sequence Type 398 (ST398) and show closely related spa-types (mainly t011, t108 and t1254) (De Neeling et al., 2007). A case control study showed that pig and cattle farmers have an increased risk for being positive for ST398 (Van Loo et al., 2007). The source of these human infections can be found in the pig population and veal calves.

Screening for MRSA among various human populations with increased risk has become important for control of nosocomial infections. In human health care settings, studies have shown that different procedures employed for the detection of MRSA from clinical specimens have varying results depending on the isolation methods used (Brown et al., 2005). For animal samples less is known about differences between MRSA detection procedures, in particular on the detection of ST398 in pig and veal calf samples.

Three existing commonly used procedures are applied for MRSA screening in pig samples (De Neeling et al., 2007 (procedure 1)) and human samples (Wertheim et al., 2001; with additional pre-enrichment (procedure 2)) (Van Duijkeren et al., 2008 (procedure 3)). To ascertain the performance of these MRSA detection methods, we conducted a study to compare three different procedures for the isolation of ST398 and the usefulness of three different chromogenic agar media. Nasal swabs of pigs and veal calves were used as matrix.