Materials and Methods

Antibiotic resistance profiles of Staphylococcus spp. from white button mushrooms and handlers

A total of 432 white button mushroom samples were collected as outlined 12 in Rossouw and Korsten. Sampling sites included two large-scale commercial mushroom farms located in Gauteng Province, South Africa, which follow similar production practices, operating under Global-G.A.P. Integrated Farm Assurance Standards V5.1. Mushrooms that were void of defects and at the ready-to-harvest stage were randomly sampled based on uniformity of size, shape and maturity. Mushrooms were aseptically harvested by researchers.12 Packed mushrooms handled by pickers and packers prior to punnet sealing were also collected. Sampling of hands was done according to standard hand swab procedures.13 Hand swabs were collected using the Copan Venturi Transystem (Copan, Italy) from five pickers’ and five packers’ dominant hands on a weekly basis for a period of 15 weeks, resulting in 150 hand samples. Samples were placed in a cooler box and transported to the laboratory for analysis within 24 h. At the time of the study, ethical clearance for non-invasive swabbing of the hands was not mandatory and was therefore waived by the Ethics Committee of the Faculty of Natural and Agricultural Sciences, University of Pretoria. Commercial and personal permission to conduct non-invasive hand swabbing was granted by the commercial entity as well as by the persons whose hands were sampled. Confidentiality was maintained throughout the process.

A sample (250 g) was aseptically obtained from each mushroom sample and homogenised using a handheld blender (Russell Hobbs, Johannesburg, South Africa). Ten grams of the homogenised sample was added to 90 mL tryptone soy broth (Merck, Johannesburg) in a sterile homogeniser bag, macerated for 5 min and incubated for 24 h at 37 °C. Contents were subsequently plated onto Baird-Parker agar (Merck) and incubated for 24 h at 37 °C.

Staphylococcus spp. resistance in mushrooms Hand swabs were placed into 9 mL buffered peptone water (Merck), and incubated for 24 h at 37 °C and plated onto Baird-Parker agar plates, which were incubated for 24 h at 37 °C. Presumptive Staphylococcus spp. were selected from the Baird-Parker agar, based on a dark grey to black colony morphology which was surrounded by a clear zone. Isolates were purified on Baird-Parker agar and the purified isolate identity was determined using matrix-assisted laser desorption ionisation time- of-flight mass spectrometry in combination with the Bruker Biotyper

Article TitleAntibiotic resistance profiles of Staphylococcus spp. from white button mushrooms and handlers

Abstract

The presence of Staphylococcus spp. has increasingly been reported in food products and poses a public health threat. The aim of this study was to determine the diversity of Staphylococcus spp. and the antibiotic resistance profiles of isolates obtained from freshly harvested and packed ready-to-eat mushrooms (n=432) and handlers’ hands (n=150). A total of 56 Staphylococcus isolates 46.4% (n=26) from hands and 53.6% (n=30) from mushrooms were recovered belonging to 10 species. Staphylococcus succinus isolates (n=21) were the most prevalent, of which 52.4% came from mushrooms and 47.6% from hands. This was followed by S. equorum isolates n=12; 91.7% (n=11) from mushrooms and 8.3% (n=1) from hands and S. saprophyticus n=9; 66.7% (n=6) from mushrooms and 33.3% (n=3) from hands. Six isolates that were characterised as multidrug resistant were isolated from hands of handlers. Most (83.9%; n=47) of the 56 isolates were resistant to penicillin 53.2% (n=25) from mushrooms and 46.8% (n=22) from hands and 14.3% (n=8) were resistant to cephalosporin classes 25% (n=2) from mushrooms and 75% (n=6) from hands, both of which are used to treat staphylococcal infections. Antibiotic resistance genes blaZ 25.0% (n=14) of all isolates of which 71.4% (n=10) were from hands and 28.57% (n=4) from mushrooms, tetL and tetK both 1.8% (n=1) from hands, mecA 5.4% (n=3) from hands and ermA 1.8% (n=1) from mushrooms were detected from the 56 isolates. Only two (25.0%) of the eight methicillin-resistant staphylococci harboured the mecA gene, while only 11 (23%) of the 47 penicillin-resistant isolates harboured the blaZ gene 36.4% (n=4) from mushrooms and 63.6% (n=7) from hands. Our results demonstrate that food handlers and harvested and packed ready-to-eat mushrooms could be a source of diverse Staphylococcus spp. that exhibit antimicrobial resistance. Clinically relevant S. aureus was only detected on one handler’s hand; however, the isolate was not multidrug resistant. The presence of diverse Staphylococcus spp. on mushrooms and the hands of handlers is a potential public health concern due to their potential to cause opportunistic infections.

staphylococci, antibiotic genes, antibiotic resistance


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