We use cookies on our website. Some are necessary for the operation of the website. You can also allow cookies for statistical purposes. You can adjust the data protection settings or agree to all cookies directly.
Data Protection Declaration
Imprint
Investigation on contamination of raw consumed food of vegetable origin with VTEC / STEC / EHEC using serological and molecular detection methods
Project
Project code: BfR-BIOS-08-1322-403
Contract period: 01.04.2009
- 31.12.2010
Purpose of research: Applied research
Vegetables and their products intended for raw human consumption constitute an important, previously underestimated source of infection for STEC / EHEC. Outbreaks of EHEC infections after ingestion of contaminated vegetables and salad were reported from several countries. The largest outbreak of EHEC infection in over 6000 patients in Japan was due to EHEC contaminated radish sprouts.
In Germany, vegetables are usually not examined for STEC / EHEC, and only little data are available on this subject. No data on human patients who have become infected with EHEC after consumption of vegetables are reported from the Robert Koch-Institut (RKI 2009)
In the research project 40 mixed salad / sprout samples were examined in 2009 for their microbial load and the load of (pathogenic) E. coli. Almost all of the products contained high numbers of bacteria. The aerobic colony count was 106 to 107 per gram and Enterobacteriaceae were largely contributing to this. On VRBD agar for detection of Enterobacteriaceae bacterial counts were determined between 105 to larger than 106 cfu per gram. The benchmark for E. coli (1x102 CFU / g) was exceeded in 5 samples, but not the warning value of 1x103 CFU / g.
E. coli could be isolated after enrichment from 23 (57.5%) of the samples. Using real-time PCR the genes for Shiga toxins (stx genes) were detected in 2 (5.0%) and for other virulence markers of EHEC, such as intimin (eae), in one (2.5%) sample. The gene for the heat-stable enterotoxin (EAST) was found in 17 (42.5%) of the samples. Four EAST-positive E. coli and one STEC could be isolated from the salads. Examination and serological typing of E. coli isolates from the lettuce samples revealed strains belonging to human pathogenic groups (O91: H21, O146:H21).
Regardless of the high amount of other bacteria that were present in the samples real-time PCR revealed highly reliable findings on pathogenic E. coli. In spiking experiments right down to three CFU of EHEC O157:H7 / 25g salad could still be detected after overnight en-richment by real-time PCR. Due to the high burden of other Enterobacteriaceae, however, the isolation of pathogenic E. coli from the samples proved to be difficult. Due to the high numbers of other bacteria that are initially present, even after enrichment 1,000 fold higher amounts of other Enterobacteriaceae are found, compared to E. coli.
In the continuation of the project, procedures shall be developed that allow a more specific enrichment and isolation of pathogenic E. coli from salad samples purchased for raw consumption. With the application of these procedures further 120 samples shall be analyzed for pathogenic E. coli including isolation of the strains in order to perform a risk analysis for this particular type of food in regard to pathogenic E. coli.
31 samples of pre-cut mixed salad and 9 samples of sprouts, which are retailed as products ready for raw consumption for humans were examined for their microbiological status. All samples were purchased within the sell-by date. Salads were washed according to the manufacturer. All samples contained strong microbial loads, Enterobacteriaceae were found in counts between 105 and larger then 106 cfu / g. E. coli could be isolated from 23 of the 40 samples, the warning value for E. coli of 103 cfu / g) was not exceeded in any sample. For the detection of pathogenic E. coli Real Time PCR methods were used. Two samples were positive for shiga-toxin (stx) genes and 18 other contained eae or east genes as indicators of human pathogenic E. coli. The isolation of E. coli from the samples is problematic, since 1000-fold higher concentrations of other Enterobacteriaceae were found before and after enrichment of the samples. Therefore, specific methods for the selective enrichment and isolation of E. coli will be developed for vegetable foods. Our investigations could show that precut mixed salad constitute risk foods with respect to pathogenic E. coli. In the continuation of the research project, methods shall be developed to improve the examination of vegetable food samples for pathogenic E. coli.
Food poisoning with presumptive B. cereus is caused by various bacterial toxins: hemolysin BL (hbl), a non-hemolytic enterotoxin (nhe), a cytotoxin (cytk) and an emetic toxin (cer). The aim of this project was the evaluation of the BTEC-RPLA test and also the evaluation of PCR Primers and conditions suitable for the detection of the toxin genes hbl, nhe and cytk. To this end we tested 40 Bacillus spp. Strains, 20 strains belong to the B. cereus Group. With these two methods we were able to detect the main virulence factors of presumptive B. cereus.
Altogether both methods can be now used for the routine diagnosis due to this evaluation.
Section overview
Subjects
- Food microbiology
- Toxicology
