Production of small, acid-soluble spore proteins inClostridium perfringensnonfoodborne gastrointestinal disease isolates

The molecular basis for the differences in heat resistance between spores of Clostridium perfringens food-borne versus nonfoodborne isolates remains unknown. Since a recent study demonstrated the role of small, acid-soluble spore proteins (SASPs) in heat resistance of spores of food-borne isolates, in the current study, we evaluated the expression of SASP-encoding genes (ssp) and the production of SASPs in nonfoodborne isolates. Our results demonstrated the presence of all three ssp genes in five surveyed nonfoodborne isolates. A β-glucuronidase assay showed that these ssp genes are expressed specifically during sporulation. Furthermore, nonfoodborne isolate F4969 produced SASPs at a level similar to that of food-borne isolate SM101. Collectively, these results suggest that the difference in the levels of heat resistance between spores of food-borne and the nonfoodborne isolates is not the result of impaired expression of ssp genes and (or) decreased production of SASPs in nonfoodborne isolates.

Comparison of the β-Glucuronidase Assay and the Conventional Method for Identification of Escherichia coli on Eosin-Methylene Blue Agar

The purpose of this study was to compare the IMViC (indole, methyl red, Voges-Proskauer and citrate utilization) tests with the β-glucuronidase (GUD) assay for the identification of suspect Escherichia coli on Levine's eosin-methylene blue (EMB) agar. After testing 258 suspect E. coli colonies from raw meat and meat products, 163 and 44 were found to be E. coli and non-E. coli, respectively, by both methods. Nine isolates were IMViC positive (i.e., + + − − or − + − −) but GUD negative; among these isolates, six were confirmed to be E. coli by API 20E (bioMérieux, Marcy-I'Etoile, France) with the remaining three being non-E. coli. There were 42 isolates that were IMViC negative but GUD positive; among these isolates, seven were pure E. coli cultures, 33 were mixed cultures containing E. coli, and the remaining two were Proteus spp. The sensitivities for the identification of E. coli on EMB were 80.9% (169/209) and 97.1% (203/209), respectively, by the IMViC tests and GUD assay; whereas the specificities were 93.9% (46/49) and 95.9% (47/49), respectively, by the IMViC tests and GUD assay. It is proposed that the GUD assay can be an effective alternative to the conventional IMViC tests for the identification of suspect E. coli on EMB.

An Improved Membrane Filtration Method for Enumeration of Faecal Coliforms and E. Coli by a Fluorogenic β-Glucuronidase Assay

A fluorogenic β-glucuronidase assay comprising membrane filtration followed by selective enumeration on m-FC agar at 44.5°C and further confirmation using tlie 4-metliylumbelliferyl-β-D-glucuronide (MUG) containing medium was evaluated for the detection of Escherichia coli in water. A total of 200 typical blue and non-typical blue colonies were isolated from sea and fresh water samples using initial selective enumeration on m-FC agar. Pure cultures of the selected colonies were further tested using the MUG assay and identified using the API 20E method. Of the colonies tested which were shown to be positive using the MUG assay 99.4% were Escherichia coli. The results of this study indicate the combination of the m-FC method followed by the MUG assay to be highly efficient for the selection and confirmation of E. coli from a wide range of environmental waters.

Rapid Detection of Bifidobacterium dentium by Enzymatic Hydrolysis of β-Glucuronide Substrates

Enzyme profiles and fermentation patterns of bifidobacteria were studied to determine phenotypic characteristics that allow the rapid detection of Bifidobacterium dentium and its differentiation from Bifidobacterium adolescentis, Bifidobacterium angulatum, Bifidobacterium catenulatum, and Bifidobacterium pseudocatenulatum. Among 43 bifidobacterial strains tested, the production of β-glucuronidase was limited to six strains of B. dentium. The presence of B. dentium on a selective medium may be rapidly confirmed by the detection of β-glucuronidase activity. Columbia agar containing propionic acid was chosen to enumerate bifidobacteria previously cultivated in MRS medium. After 48 h of incubation, β-glucuronidase activity was determined by using a plate staining procedure. B. dentium strains gave positive results for β-glucuronidase activity after application of the overlay solution of β-glucuronide substrate. The β-glucuronidase assay is a rapid screening method for B. dentium. This method might be useful for selection of nonpathogenic strains or detection of fecal contamination from human origin.