Impact of lower challenge doses of enterotoxigenic Escherichia coli on clinical outcome, intestinal colonization and immune responses in adult volunteers

Abstract

A reliable and effective human challenge model is needed to help down-select the most promising ETEC vaccines currently under development. Such a model would need to reliably induce diarrhea in a high proportion of volunteers using the lowest possible inoculum to maximize safety and sensitivity. Previously we validated a challenge model that utilized a dose of 2x107 CFU of ETEC strain H10407 (LT+, ST+, CFA/I+ and O78+) to induce attack rates for moderate to severe diarrhea (MSD) of ~60–70%. Here we detail efforts to further refine the model in an attempt to determine if a lower challenge dose of H10407 can be used. Thirty subjects were randomized 1:1 to receive an oral administration of H10407 at doses of 106 or 105 CFU in bicarbonate buffer. After challenge, subjects were monitored for signs and symptoms of enteric illness and stool samples were collected to detect shedding of the challenge strain. Systemic and mucosal immune responses were measured using serum, antibody in lymphocyte supernatant and fecal samples. The attack rate was 13.3% (2/15) and 26.7% (4/15) for MSD in the 105 and 106 groups, respectively. Four MSD cases met criteria for early antibiotic treatment. All subjects but one shed the challenge strain in fecal samples. The frequency and magnitude of anti-LT toxin, CFA/I and LPS O78 immune responses were antigen, dose, severity of diarrhea and shedding levels dependent. Notably, although of lower magnitude, there were considerable immune responses in the subjects with no diarrhea. This may indicate that immune responses to asymptomatic infections of ETEC in children in the endemic countries may contribute to protection. Based on this and our prior studies, we conclude that a dose of 2x107 H10407 remains the lowest practical dose for use in future volunteer studies evaluating candidate vaccines and other preventive or therapeutic ETEC interventions.Trial registration: ClinicalTrials.gov NCT00844493.

Human Mucosal IgA Immune Responses against Enterotoxigenic Escherichia coli

“…After experimental infection with ETEC, most volunteers appeared to have a significant increase in strain-specific anti-O-antigen IgA levels, with 3 studies reporting a proportion of volunteers with elevated IgA levels, or response ratios, of more than 90% [ 17 , 32 , 53 ]. It also induces the strongest responses, as measured in ALS, compared to any of the other ETEC antigens [ 17 , 59 ]. Responses against ETEC O-antigens have been studied in many sample types, and both the strength and responder ratio seem to increase with increasing doses given to the volunteers, with a few exceptions [ 55 , 59 ].…”

“…It also induces the strongest responses, as measured in ALS, compared to any of the other ETEC antigens [ 17 , 59 ]. Responses against ETEC O-antigens have been studied in many sample types, and both the strength and responder ratio seem to increase with increasing doses given to the volunteers, with a few exceptions [ 55 , 59 ]. Chakraborty et al reported that O-antigen IgA responses were high compared to anti-LT and -CF responses, even when low doses of H10407 were given to the volunteers [ 59 ].…”

“…Responses against ETEC O-antigens have been studied in many sample types, and both the strength and responder ratio seem to increase with increasing doses given to the volunteers, with a few exceptions [ 55 , 59 ]. Chakraborty et al reported that O-antigen IgA responses were high compared to anti-LT and -CF responses, even when low doses of H10407 were given to the volunteers [ 59 ]. It can also be seen that the serum IgA response to O-antigens peaks relatively early, at around days 7–10, which is earlier than the response to other ETEC antigens like LT and colonization factors ( Table 1 ).…”

“…The most likely explanation for this increase is that it takes more than a single infection to optimize the immune response against this antigen [ 47 ]. Furthermore, anti-CFA/I IgA responders were few or absent when fecal samples from the infected volunteers were tested [ 17 , 59 ]. It should also be noted that increased anti-colonization factor IgA levels in serum after infection appeared to be more frequent in volunteers who did not develop diarrhea [ 17 , 60 , 61 , 62 ].…”

Enteropathogen seroepidemiology among children in low-resource settings

“…We defined incident seroconversions and seroreversions as a change in IgG across a pathogen's seropositivity cutoff. Vaccine immunogenicity and pathogen challenge studies among healthy adults often use a 4-fold increase in antibody levels (difference of +0.6 on the log 10 scale) as a criterion for seroconversion [76][77][78] . In a secondary analysis aimed to capture significant changes above a pathogen's seropositivity cutoff, we defined incident boosting episodes as a ≥4-fold increase in IgG to a final level above a seropositivity cutoff, and incident waning episodes as ≥4-fold decrease in IgG from an initial level above a seropositivity cutoff.…”