High throughput multiple locus variable number of tandem repeat analysis (MLVA) of Staphylococcus aureus from human, animal and food sources

Staphylococcus aureus is a major human pathogen, a relevant pathogen in veterinary medicine, and a major cause of food poisoning. Epidemiological investigation tools are needed to establish surveillance of S. aureus strains in humans, animals and food. In this study, we investigated 145 S. aureus isolates recovered from various animal species, disease conditions, food products and food poisoning events. Multiple Locus Variable Number of Tandem Repeat (VNTR) analysis (MLVA), known to be highly efficient for the genotyping of human S. aureus isolates, was used and shown to be equally well suited for the typing of animal S. aureus isolates. MLVA was improved by using sixteen VNTR loci amplified in two multiplex PCRs and analyzed by capillary electrophoresis ensuring a high throughput and high discriminatory power. The isolates were assigned to twelve known clonal complexes (CCs) and -a few singletons. Half of the test collection belonged to four CCs (CC9, CC97, CC133, CC398) previously described as mostly associated with animals. The remaining eight CCs (CC1, CC5, CC8, CC15, CC25, CC30, CC45, CC51), representing 46% of the animal isolates, are common in humans. Interestingly, isolates responsible for food poisoning show a CC distribution signature typical of human isolates and strikingly different from animal isolates, suggesting a predominantly human origin

Foodborne viruses: Detection, risk assessment, and control options in food processing

In a recent report by risk assessment experts on the identification of food safety priorities using the Delphi technique, foodborne viruses were recognized among the top rated food safety priorities and have become a greater concern to the food industry over the past few years. Food safety experts agreed that control measures for viruses throughout the food chain are required. However, much still needs to be understood with regard to the effectiveness of these controls and how to properly validate their performance, whether it is personal hygiene of food handlers or the effects of processing of at risk foods or the interpretation and action required on positive virus test result. This manuscript provides a description of foodborne viruses and their characteristics, their responses to stress and technologies developed for viral detection and control. In addition, the gaps in knowledge and understanding, and future perspectives on the application of viral detection and control strategies for the food industry, along with suggestions on how the food industry could implement effective control strategies for viruses in foods. The current state of the science on epidemiology, public health burden, risk assessment and management options for viruses in food processing environments will be highlighted in this review

Origin of the animal and food isolates.

a<p>the number of MRSA isolates is indicated in brackets.</p>b<p>the origin of one of the 13 food poisoning associated isolates is unknown.</p>c<p>NA : not applicable.</p

Electrophoregrams showing multiplex PCR amplicons resolved by capillary electrophoresis

<p><b>a. PCR1</b> ten dye-colored coamplified VNTR loci. <b>b. PCR2</b> six dye-colored coamplified VNTR loci.</p

Dendrogram of the HARMONY collection using MLVA-16_Orsay.

<p>Color coding is according to MLST clonal complex assignment whereas clustering is done according to the displayed MLVA data. Strain Id, clonal complex, sequence type, spa type, spa code and geographic origin are indicated. MLVA cluster bootstrap values are shown for the main clusters.</p

VNTRs and associated oligonucleotide primers.

<p>(*HGDI calculated on the 90 isolates from the HARMONY collection).</p>a<p>Multiplex PCR reaction.</p>b<p>Expected amplicon size for strain Mu50 RefSeq NC_002758 (by convention, corresponding number of repeated units for strain Mu50).</p>c<p>Observed allele size range: amplicons length (number of repeated units).</p

Minimum spanning tree of the 251 <i>S. aureus</i> isolates using MLVA-16_Orsay.

<p>Minimum spanning tree of the 251 <i>S. aureus</i> isolates (106 human-associated isolates, 98 animal-associated isolates and 47 isolates from food products among which 13 were related with food-poisoning) using MLVA-16_Orsay. Each circle represents a MLVA genotype. The size of each circle indicates the number of isolates within this MLVA genotype. The different clusters are annotated. The host origin is indicated with a specific color. Isolates involved in food poisoning events are represented by black circles. Human and food isolates are highlighted with two different hatch patterns.</p

Constitution of the different MLVA schemes.

a<p>the VNTR locus name reflects genome localisation (in kb) in strain Mu50 refseq NC_002758.</p>b<p>alternative names given in the literature.</p>c<p>from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033967#pone.0033967-Hardy1" target="_blank">[52]</a>.</p>d<p>from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033967#pone.0033967-Schouls1" target="_blank">[23]</a>.</p>e<p>from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033967#pone.0033967-Gilbert1" target="_blank">[53]</a>.</p>f<p>from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033967#pone.0033967-Sabat1" target="_blank">[54]</a>.</p>g<p>MLVA14_Orsay corresponds to the 14 panel 1-panel 2 loci in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033967#pone.0033967-Pourcel1" target="_blank">[24]</a>.</p>h<p>MLVA10_Orsay corresponds to the 10 panel 1 loci in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033967#pone.0033967-Pourcel1" target="_blank">[24]</a>.</p>i<p>MLVA8_Bilthoven corresponds to the 8 loci used by <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033967#pone.0033967-Schouls1" target="_blank">[23]</a>.</p