Search

The morbidity and mortality associated with Vibrio -mediated waterborne diseases necessitates the development of sensitive detection technologies that are able to elucidate the identity, potential pathogenicity, susceptibility, and viability of contaminating bacteria in a timely manner. For this purpose, we have designed a single multiplex PCR assay to simultaneously amplify 95 diagnostic regions (encompassing species/serogroup-specific, antimicrobial resistance, and known toxin markers) and combined it with a long oligonucleotide microarray to create a platform capable of rapidly detecting and discriminating the major human pathogenic species from the genus Vibrio : V. cholerae, V. parahaemolyticus, V. vulnificus , and V. mimicus . We were able to validate this strategy by testing 100 geographically and temporally distributed isolates and observed an excellent concordance between species- and serotype-level microarray-based identification and traditional typing methods. In addition to accurate identification, the microarray simultaneously provided evidence of antibiotic resistance genes and mobile genetic elements, such as sulfamethoxazole-trimethoprim constins and class I integrons, and common toxin ( ctxAB, rtxA, hap, hlyA, tl, tdh, trh, vvhA, vlly , and vmhA ) and pathogenicity ( tcpA , type III secretion system) genes that are associated with pathogenic Vibrio . The versatility of this method was further underscored by its ability to detect the expression of known toxin and virulence genes from potentially harmful viable but nonculturable organisms. The results suggest that this molecular identification method provides rapid and definitive information that would be of value in epidemiological, environmental, and health risk assessment surveillance.

Diarrhea is a major cause of preventable illness in sub-Saharan Africa. Although most cases of bacterial gastroenteritis do not require antimicrobial treatment antimicrobial use is widespread. We examined the bacterial causes of diarrhea and monitored antimicrobial susceptibilities of isolates through clinic-based surveillance in a rural Kenyan community. From May 1997 through April 2003 diarrheal stool samples from persons presenting to 4 sentinel health centers were cultured by standard techniques for routine bacterial enteric pathogens for which antimicrobial susceptibilities were determined. A random subset of specimens was also evaluated for diarrheagenic Escherichia coli. Among stool specimens from 3445 persons 1092 (32%) yielded at least 1 bacterial pathogen. Shigella species was most commonly isolated (responsible for 16% of all illnesses; 54% of isolates were Shigella flexneri). Campylobacter species and diarrheagenic E. coli predominated among children aged < 5 years and were progressively replaced by Shigella species with increasing age. With the exception of Campylobacter species susceptibility to the antimicrobials used most widely in the community was low: < 40% for all isolates tested and < 25% for Shigella species. Most persons were treated with an antimicrobial to which their isolate was resistant. Susceptibility to specific antimicrobials was inversely proportional to the frequency with which they were prescribed. The utility of available antimicrobials for treating bacterial diarrhea in rural western Kenya is substantially limited by reduced susceptibility. More judicious use of appropriate antimicrobials is warranted. Efforts to prevent illness through provision of clean water improved hygiene and vaccine development should be strengthened. (authors)