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Staphylococcal food poisoning (SFP) is a major foodborne illness caused by staphylococcal enterotoxins (SEs). It is a well known fact that foodborne outbreak investigations are solely characterized by commercially available immunoassay kits. However, these assays encompass only few enterotoxins such as SEA-SEE which are renowned as “classical” enterotoxins and unable to detect any other novel enterotoxins even though their involvement is predicted. In this context, the present study involved development of a sandwich ELISA immunoassay for the specific detection of “non-classical” enterotoxin G (SEG). The toxin belongs to enterotoxin gene cluster ( egc ) which comprises a bunch of five toxin genes that are known to co-express. Thus, the developed assay might indirectly speculate the presence of other toxins in the cluster. The efficiency of ELISA was compared with PCR analysis where all strains possessing seg were found positive for toxin production. Additionally, analogous to other studies which reported the co-occurrence of seg and sei , the PCR analysis accomplished in the study evinced the same. The sandwich format allowed sensitive detection with a detection limit of 1 ng/mL. High specificity was achieved in presence of non-target protein as well as bacteria. Likewise, staphylococcal protein A (SpA) interference that is inevitably associated with immunoassays was eliminated by implementation of anti-SEG IgY in our study. Consequently, chicken IgY were used to capture target antigen in developed sandwich ELISA. Further, spiking studies and analysis on natural samples emphasized the robustness as well as applicability of developed method. Altogether, the established assay could be a reliable detection tool for the routine investigation of SEG as well as to predict other egc toxins in samples from food and clinical sources. [ABSTRACT FROM AUTHOR]

Abstract: Genes encoding staphylococcal enterotoxins (SEs) are carried by mobile genetic elements, and enterotoxin gene clusters (egc) are pathogenicity island-borne structures comprising several SE genes, which are frequently found among clinical Staphylococcus aureus isolates. In the present study, we investigated the distribution and the genetic variability of egc loci in S. aureus strains isolated from raw milk and soft cheese in Minas Gerais, Brazil. Ninety-two isolates were submitted to PCR detection of individual egc-borne SE genes (seg, sei, sem, sen, seo, seu), and egc loci were typed using PCR-RFLP. PCR products of egc positive isolates were sequenced. Ninety-one isolates harbored at least one SE gene, which generated 14 different genotypes. The sei gene was the most widely distributed (97.8%), and was found in combination with seg in 49 isolates (53.3%). Altogether, a complete set of individual egc genes was detected in 37 isolates (40%). However, egc loci were detected by PCR-RFLP in only 4 isolates, and classified as egc1 (n=2), egc3 (n=1), and egc4 (n=1). This investigation demonstrated the low occurrence of the egc in S. aureus isolated from dairy products. However, the frequency of complete sets of individual egc-borne genes reflects either the presence of these SE genes outside egc or the existence of new egc types in these strains. [Copyright &y& Elsevier]

Abstract: A multiplex PCR assay that allows for the rapid screening of the 19 genes that encode staphylococcal enterotoxins (SEs) (sea to see, and seg to sei), SE-like (SEl) toxins (sej to ser, and seu), and toxic shock syndrome toxin-1 (TSST-1) (tst) was developed in this study. These toxins are included in the pyrogenic toxin superantigen (PTSAg) family and are responsible for many diseases such as staphylococcal food poisoning (SFP) and TSS. The primers were designed based on dual priming oligonucleotide (DPO) technology to detect all of the 19 SAg genes in three sets of PCR. The developed multiplex PCR was applied to 143 Staphylococcus aureus strains isolated from pork and chicken meat in Korea. Almost 50% of the strains possessed at least one of the 19 SAg genes. The most frequently found genes were seg, sei, sem, and sen (53 isolates, 37%), which were often found simultaneously in the same isolate. In those isolates, the seo (39 isolates, 27%) or seu (6 isolates, 4%) genes were frequently found together and this combination (seg, sei, sem, sen, and seo or seu) was considered to be a part of the enterotoxin gene cluster (egc). The sea gene (10 isolates, 7%) was the gene most frequently detected out of all the classical SE genes (sea to see). Although these classical SEs are considered to be major etiological factors in SFP, newly described SE or SEl genes (seg to ser, and seu) were more frequently detected than the classical SE genes in this study. There was no isolate detected containing the seb, sec, sek, sel, or seq genes. S. aureus possessing mobile genetic elements known to encode these SAg genes, such as egc, were presumed to be widely distributed among pork and chicken meats in Korea. The multiplex PCR developed in this study could be applied to the investigation of SAg genes in S. aureus strains isolated from various sources. [Copyright &y& Elsevier]