RIDASCREEN® Verotoxin is an enzyme immunoassay for the detection of pathogenic E. coli (EHEC/STEC/VTEC) by using verotoxin 1 and 2 (SLT-I and SLT-II) which are formed during enrichment as indicator/detectable molecule for the pathogen.
Enterobacteriacea of the species Escherichia coli (E. coli) are part of the intestinal flora of healthy humans, as well as many farm animals. Therefore, they are indicators of fecal contamination of water and food. Due to different pathogenicity factors, often plasmid-coded or transmitted by phages, E. coli are considered as facultative pathogenic. Since 1977, E. coli are known which are able to produce two cytotoxins, verotoxin 1 and 2. Due to the similarities of the verotoxins to the shiga toxin of Shigella dysenteriae, they have also been labeled as the shiga-like toxin 1 and 2 (SLT-I and SLT-II).
The ”prototype” of enterohemorrhagic E. coli (EHEC) was first described in 1982. The clinical symptoms which are caused by EHEC range from light to severe gastroenteritis to hemorrhagic colitis which occurs in 10 – 20 % of all infections. Also life-threatening and post-infectious complications in 5 – 10 % of the infections can occur. Especially infants and small children, but also old patients or patients with a weakened immune system are affected. Due to its high environmental resistance and relative acid tolerance, the infection level for EHEC is only around 10² microbes. Main sources of infection are products from cattle, sheep and goats, especially raw or insufficiently cooked meat and poorly stored meat products and raw milk. Chains of infection from human to human, especially in public institutions such as child care centers, senior citizen homes or hospitals, as well as direct contact to animals are of importance.
Today’s recommended procedure for food analysis consists of a pre-enrichment of the food sample for 6 h, inoculation of an aliquot of the pre-enrichment into a fresh broth and subsequently incubation overnight (16 – 18 h). After that the bacteria should be isolated. At the same time, as the safest pathogenicity factor, the ability of enriched bacteria to produce verotoxin should be carried out using the enrichment broth. This can be done by means of PCR or immunologically by ELISA. In comparison to PCR, ELISA techniques such as the RIDASCREEN® Verotoxin assay, are simple methods which are practical for any laboratory. An enrichment of the bacteria is required for both methods since the amount of cells of pathogenic E. coli as well as the toxin content in food can be very low.
|Test Format||Microtiter plate (12 strips with 8 removable wells each)|
|Standard Range||Positive control (inactivated verotoxins)|
|Sample Preparation||Primary enrichment in mTSB (6 h).
Secondary enrichment in mTSB containing Mitomycin C (16 – 18 h).
After centrifugation the supernatant of the culture is used directly in the assay.
|Incubation Time||1 h 45 min|
|Detection Limit||Enrichment for 16 – 18 hours is sufficient to detect the toxins in the supernatant of the culture. Relevant data show that already 1 cfu of a verotoxin producing E. coli strain in the original sample is detectable with the RIDASCREEN® Verotoxin test under suitable conditions.|
|Cross Reactivity||Cross-reactions: no cross-reactions detected beside the specific detection of SLT-I and SLT-II|