Bacterial agglutination and polyacrylamide gel electrophoresis for typing Clostridium difficile.

Bacterial agglutination and polyacrylamide gel electrophoresis (PAGE) were methods evaluated for typing strains of Clostridium difficile. A panel of four antisera, obtained by immunizing rabbits with washed whole cells of different strains of C. difficile, produced distinctive patterns of agglutination. Ethylenediaminetetraacetate (EDTA) extracts subjected to PAGE also produced distinctive protein profiles. Excellent correlation between the two methods was observed when geographically distant isolates were typed without knowledge of their clinical origin. Both typing methods should receive further evaluation for their value as tools for epidemiological studies.

A rapid (4--6-hour) urine-culture system for direct identification and direct antimicrobial susceptibility testing.

This study evaluates a new direct rapid system for urine cultures, including detection and quantitation of positive specimens by Gram stain, direct identification by 4--6-hour incubation of sediment with reagent strips, and antibiotic susceptibility testing by direct (3--4-hour) disk-elution methods. Of 987 routine urine specimens, 121 had significant (less than or equal to 10(5) colony-forming units/ml) gram-negative bacilluria, of which 89% were detected by the Gram stain. Direct rapid identification was correct in 94%. Results of direct disk-elution antimicrobial tests showed overall agreement with results of standard disk diffusion of 93% of tests, and major discrepancies in 4%. For urine specimens with gram-negative bacilluria, this system permitted detection, quantitation, identification, and antimicrobial susceptibility testing in four to six hours with reasonable, though not complete, accuracy.

Factors influencing heat-labile Escherichia coli enterotoxin activity.

In this study, conditions for production, detection, and storage of heat-labile Escherichia coli enterotoxin (LT) in culture filtrates from E. coli H-10407 were defined by using the adrenal tumor cell assay system. An enriched medium containing 0.6% yeast extract, 2% Casamino Acids, and 0.25% glucose buffered at pH 8.5 produced the highest LT activity of the various test media. In E. coli strain H-10407, LT activity was markedly decreased if the initial pH of the culture media was reduced to pH 7.5 or less. In contrast to E. coli P-263, if strain H-10407 was grown in the presence of mitomycin C there was no increase in LT production. Crude-culture filtrates containing LT can be stored at 4 degrees C for several days without an appreciable loss of activity; however, for long-term storage lyophilization or freezing at -70 degrees C is recommended.

Toxigenic Escherichia coli and childhood diarrhea.

Stool specimens were examined from 40 children with diarrhea who were under three years of age to determine the incidence of enterotoxigenic Escherichia coli in endemic diarrhea. Heat-labile E. coli enterotoxin was assayed in the very sensitive and reproducible cultured adrenal tumor cell system. Toxigenic E. coli were isolated from only one stool specimen and in this case infection with Shigella dysenteriae was also present. None of the eight classic enteropathogenic E. coli isolates were positive in the adrenal assay. This study suggests that heat-labile enterotoxin-producing E. coli are not an important cause of endemic childhood diarrhea in Southern California.

Antibiotic susceptibility testing of gram-negative nonfermentative bacilli.

A study was undertaken to determine if current methods of antibiotic susceptibility testing could be successfully applied to the gram-negative nonfermentative bacilli. Using clinical isolates and reference strains, experiments were conducted on the inherent reliability of the Bauer-Kirby method, as well as the effect of certain modifications on the method such as elimination of the 2- to 5-h incubation in broth and use of different agar media. Results obtained using these modifications were compared to the results obtained by the standard method. It was shown that the two modifications investigated had a significant effect on the interpretation of zone diameters. It was further shown that the standard Bauer-Kirby method with some exceptions correlates with minimal inhibitory concentrations as determined by broth dilution methods. Results suggest that the Bauer-Kirby method may be a reliable technique for testing the antibiotic susceptibility of the nonfermentative bacilli.