C O Tacket

Role of the eaeA gene in experimental enteropathogenic Escherichia coli infection.

Enteropathogenic Escherichia coli (EPEC) infections are a leading cause of infant diarrhea in developing countries. Recently eaeA, a gene necessary for the characteristic intimate attachment of EPEC to epithelial cells in tissue culture, was described. We conducted a randomized, double-blind study to determine the role of the eaeA gene in human EPEC infection. 11 adult volunteers ingested 2 x 10(10) colony-forming units of O127:H6 EPEC strain E2348/69, and an equal number received the same dose of an isogenic eaeA deletion mutant constructed from E2348/69. Volunteers were monitored for the development of diarrhea, fever, and systemic and gastrointestinal complaints. Diarrhea developed in all 11 volunteers who received E2348/69 and in 4 of 11 who received the mutant (P = 0.002). Fever was more common in recipients of the wild-type strain (P = 0.024). Stool volumes were lower in recipients of the mutant. All volunteers seroconverted to E2348/69 LPS, but the geometric mean peak titers of serum IgG and IgA in recipients of the mutant were lower than those of recipients of the wild-type strain. IgA against LPS was detected in the jejunal fluid of six of six recipients of E2348/69 and 5/6 recipients of the mutant. This study unambiguously assigns a role for eaeA as an EPEC virulence gene, but the residual diarrhea seen in recipients of the mutant indicates that other factors are involved.

SAFETY, IMMUNOGENICITY, AND EFFICACY OF RECOMBINANT LIVE ORAL CHOLERA VACCINES, CVD 103 AND CVD 103-HgR

The genes encoding the A (toxic) subunit of cholera toxin were deleted from pathogenic Vibrio cholerae O1 strain 569B by recombinant techniques, leaving intact production of immunogenic, non-toxic B subunit. The resultant strain, CVD 103, evaluated for safety, immunogenicity, and efficacy as a live oral vaccine, was highly attenuated and elicited strong antibacterial and antitoxic immune responses; a single dose significantly protected volunteers against challenge with pathogenic V cholerae O1 of either serotype or biotype. A further derivative, CVD 103-HgR, which has an Hg++-resistance gene to differentiate it from wild-type vibrios, was also well-tolerated, immunogenic, and protective; moreover, faecal excretion of this derivative was significantly lower than that of CVD 103, which should minimise environmental spread of the vaccine. CVD 103-HgR is a candidate for expanded clinical trials in endemic areas.

Evaluation in volunteers of a candidate live oral attenuated Salmonella typhi vector vaccine.

Candidate vector vaccine strain CVD 906 (aroC- and aroD- derivative of virulent Salmonella typhi strain ISP1820) was evaluated in phase 1 clinical trials. The first nine volunteers ingested a single dose of 5 x 10(7) CVD 906 bacilli. At this dose CVD 906 stimulates remarkable systemic and mucosal immune responses, inasmuch as 89% of volunteers developed marked serum antibody levels to S. typhi antigens and high numbers of antigen-specific gut-derived antibody-secreting cells. Four (44%) volunteers developed asymptomatic vaccinemia 4-10 d after immunization and all volunteers excreted CVD 906 on at least one occasion. However, two volunteers developed febrile adverse reactions, one on the day of vaccination and the other on day 4. Of 11 volunteers who ingested a single dose of 5 x 10(3) CVD 906 bacilli, none displayed side effects but 27% developed significant serum responses to S. typhi LPS. In vitro, CVD 906 replicates for only nine generations in pooled human serum, indicating that CVD 906 growth is limited in this physiologically relevant medium. In phorbol myristate acetate-induced U937 human macrophage-like cells, CVD 906 replicates intracellularly to a lesser extent than parent strain ISP1820. Although, strain CVD 906 is attenuated and highly immunogenic, the occasional febrile reactions at high doses indicate that further attenuation of this strain is necessary.

Pediatric diarrhea: the challenge of prevention.

Conditions such as poverty, underdevelopment, and lack of education facilitate the widespread transmission of the pathogens that cause diarrheal disease, dysentery, and enteric fever in young children. Such infections produce high rates of morbidity, mortality, and adverse nutritional consequences in the first 2 years of life. Although rapid socioeconomic development has produced a precipitous decline in mortality due to diarrheal disease in the developed world, such a trend is not likely in developing countries unless alternative measures are pursued. Nonspecific interventions pursued have included oral rehydration therapy to prevent and treat dehydration, promotion of breast feeding, health education to teach maternal technology, and early realimentation to diminish the nutritional consequences of infant diarrhea. In addition, there is reason to be optimistic about the future development of various immunizing agents against the major enteric pathogens. Epidemiologic data support the conclusion that prior natural infection with enterotoxigenic E. coli, Shigella, rotavirus, and V. cholerae OL confers protective immunity. Among the divergent approaches being followed in the development of vaccines against rotavirus are: 1) use of animal rotavirus as possible attenuated strains; 2) attenuating human rotaviruses by passage in tissue culture; 3) development of hybrid reassortant strains by coinfecting tissue cultures with both an animal strain well adapted to tissue culture and a human strain and then selecting a hybrid virus that possesses the human virus neutralization antigen but grows to higher titre in tissue culture; 4) evaluation of rotaviruses isolated from asymptomatic infected neonates in nursery outbreaks for their safety, infectivity, and immunogenicity in older children; 5) cloning a DNA copy of the RNA genus responsible for the neutralization antigens of rotaviruses; and 6) preparation of a synthetic peptide of the critical epitope of the neutralization antigen.