Intradermally administered enterotoxigenic E. coli vaccine candidate MecVax induces functional serum IgG antibodies against seven adhesins (CFA/I, CS1-CS6) and both toxins (STa, LT)

There are no vaccines licensed for enterotoxigenic Escherichia coli (ETEC), a leading bacterial cause of children’s diarrhea and travelers’ diarrhea. MecVax, a multivalent E. coli vaccine candidate composed of two epitope- and structure-based polyvalent proteins (toxoid fusion 3xSTa N12S -mnLT R192G/L211A and CFA/I/II/IV MEFA), is to induce broad anti-adhesin and antitoxin antibodies against heterogeneous ETEC pathovars. Administered intraperitoneally (IP) or intramuscularly (IM), MecVax was shown to induce antibodies against seven ETEC adhesins (CFA/I, CS1-CS6), which are produced by ETEC pathovars causing over 60% of ETEC-associated diarrheal cases and the moderate-to-severe cases, and both toxins (heat-labile toxin - LT and heat-stable toxin - STa) expressed by all ETEC strains. To further characterize immunogenicity of this protein-based injectable subunit vaccine candidate and to explore other parenteral administration routes for the product, in this study, we intradermally (ID) immunized mice with MecVax and measured antigen-specific antibody responses and further antibody functional activities against the adhesins and toxins targeted by the vaccine. Data showed that mice ID immunized with MecVax developed robust anti-CFA/I, -CS1, -CS2, -CS3, -CS4, -CS5, -CS6, -LT and anti-STa IgG responses. Furthermore, antibodies derived from MecVax via ID route inhibited adherence of ETEC or E. coli strains expressing any of the seven target adhesins (CFA/I, CS1-CS6) and neutralized enterotoxicity of LT and STa toxins. These results confirmed broad immunogenicity of MecVax and suggested that this multivalent ETEC subunit vaccine candidate can be effectively delivered via ID route. IMPORTANCE Enterotoxigenic Escherichia coli (ETEC) is a leading bacterial cause of diarrhea in children living in developing countries and international travelers. Developing an effective vaccine for ETEC diarrhea has been hampered because of challenges of virulence heterogeneity and difficulties of inducing neutralizing antibodies against the key STa toxin. MecVax, a subunit vaccine candidate carrying two polyvalent protein antigens for the first time induces functional antibodies against the most important ETEC adhesins which are associated with a majority of diarrheal cases and the moderate-to-severe cases but also against enterotoxicity of LT and more importantly STa toxin which plays a key role in children’s diarrhea and travelers’ diarrhea, potentially leading to development of a truly effective ETEC vaccine. Data from this study may also indicated that this ETEC subunit vaccine can be administered effectively via ID route, expanding clinical administration options for this vaccine product.

Enteropathogen Changes After Rotavirus Vaccine Scale-up

OBJECTIVES: To inform next steps in pediatric diarrhea burden reduction by understanding the shifting enteropathogen landscape after rotavirus vaccine implementation. METHODS: We conducted a case-control study of 1788 medically attended children younger than 5 years, with and without gastroenteritis, after universal rotavirus vaccine implementation in Peru. We tested case and control stools for 5 viruses, 19 bacteria, and parasites; calculated coinfection-adjusted attributable fractions (AFs) to determine pathogen-specific burdens; and evaluated pathogen-specific gastroenteritis severity using Clark and Vesikari scales. RESULTS: Six pathogens were independently positively associated with gastroenteritis: norovirus genogroup II (GII) (AF 29.1, 95% confidence interval [CI]: 28.0–32.3), rotavirus (AF 8.9, 95% CI: 6.8–9.7), sapovirus (AF 6.3, 95% CI: 4.3–7.4), astrovirus (AF 2.8, 95% CI: 0.0–4.0); enterotoxigenic Escherichia coli heat stable and/or heat labile and heat stable (AF 2.4, 95% CI: 0.6–3.1), and Shigella spp. (AF 2.0, 95% CI: 0.4–2.2). Among typeable rotavirus cases, we most frequently identified partially heterotypic strain G12P[8] (54 of 81, 67%). Mean severity was significantly higher for norovirus GII–positive cases relative to norovirus GII–negative cases (Vesikari [12.7 vs 11.8; P < .001] and Clark [11.7 vs 11.4; P = .016]), and cases in the 6- to 12-month age range relative to cases in other age groups (Vesikari [12.7 vs 12.0; P = .0002] and Clark [12.0 vs 11.4; P = .0016]). CONCLUSIONS: Norovirus is well recognized as the leading cause of pediatric gastroenteritis in settings with universal rotavirus vaccination. However, sapovirus is often overlooked. Both norovirus and sapovirus contribute significantly to the severe pediatric disease burden in this setting. Decision-makers should consider multivalent vaccine acquisition strategies to target multiple caliciviruses in similar countries after successful rotavirus vaccine implementation.

Molecular characterization of enterotoxigenic Escherichia coli toxins and colonization factors in children under five years with acute diarrhea attending Kisii Teaching and Referral Hospital, Kenya

Background Enterotoxigenic Escherichia coli (ETEC) is one of the leading causes of infectious diarrhea in children. There are no licensed vaccines against ETEC. This study aimed at characterizing Escherichia coli for ETEC enterotoxins and colonization factors from children < 5 years with acute diarrhea and had not taken antibiotics prior to seeking medical attention at the hospital. Methods A total of 225 randomly selected archived E. coli strains originally isolated from 225 children with acute diarrhea were cultured. DNA was extracted and screened by multiplex polymerase chain reaction (PCR) for three ETEC toxins. All positives were then screened for 11 colonization factors by PCR. Results Out of 225 E. coli strains tested, 23 (10.2%) were ETEC. Heat-stable toxin (ST) gene was detected in 16 (69.6%). ETEC isolates with heat-stable toxin of human origin (STh) and heat-stable toxin of porcine origin (STp) distributed as 11 (68.8%) and 5 (31.2%) respectively. Heat-labile toxin gene (LT) was detected in 5 (21.7%) of the ETEC isolates. Both ST and LT toxin genes were detected in 2 (8.7%) of the ETEC isolates. CF genes were detected in 14 (60.9%) ETEC strains with a majority having CS6 6 (42.9%) gene followed by a combination of CFA/I + CS21 gene detected in 3 (21.4%). CS14, CS3, CS7 and a combination of CS5 + CS6, CS2 + CS3 genes were detected equally in 1 (7.1%) ETEC isolate each. CFA/I, CS4, CS5, CS2, CS17/19, CS1/PCFO71 and CS21 genes tested were not detected. We did not detect CF genes in 9 (39.1%) ETEC isolates. More CFs were associated with ETEC strains with ST genes. Conclusion ETEC strains with ST genes were the most common and had the most associated CFs. A majority of ETEC strains had CS6 gene. In 9 (39.1%) of the evaluated ETEC isolates, we did not detect an identifiable CF.

Genomic Characteristics of Stx2e-Producing Escherichia coli Strains Derived from Humans, Animals, and Meats

Shiga toxin (Stx) can be classified into two types, Stx1 and Stx2, and different subtypes. Stx2e is a subtype commonly causing porcine edema disease and rarely reported in humans. The purpose of this study was to analyze the prevalence and genetic characteristics of Stx2e-producing Escherichia coli (Stx2e-STEC) strains from humans compared to strains from animals and meats in China. Stx2e-STEC strains were screened from our STEC collection, and whole-genome sequencing was performed to characterize their genetic features. Our study showed a wide distribution of Stx2e-STEC among diverse hosts and a higher proportion of Stx2e-STEC among human STEC strains in China. Three human Stx2e-STEC isolates belonged to O100:H30, Onovel26:H30, and O8:H9 serotypes and varied in genetic features. Human Stx2e-STECs phylogenetically clustered with animal- and food-derived strains. Stx2e-STEC strains from animals and meat showed multidrug resistance, while human strains were only resistant to azithromycin and tetracycline. Of note, a high proportion (55.9%) of Stx2e-STEC strains, including one human strain, carried the heat-stable and heat-labile enterotoxin-encoding genes st and lt, exhibiting a STEC/enterotoxigenic E. coli (ETEC) hybrid pathotype. Given that no distinct genetic feature was found in Stx2e-STEC strains from different sources, animal- and food-derived strains may pose the risk of causing human disease.

Enterotoxigenic Escherichia coli degrades the host MUC2 mucin barrier to facilitate critical pathogen-enterocyte interactions in human small intestine.

Enterotoxigenic Escherichia coli (ETEC) are a genetically diverse pathologic variant of E. coli defined by the production of heat-labile (LT) and/or heat-stable (ST) toxins. ETEC are estimated to cause hundreds of millions of cases of diarrheal illness annually. However, it is not clear that all strains are equally equipped to cause disease and asymptomatic colonization with ETEC is common in low-middle income regions lacking basic sanitation and clean water where ETEC are ubiquitous. Recent molecular epidemiology studies have revealed a significant association between strains which produce EatA, a secreted autotransporter protein, and the development of symptomatic infection. Here, we demonstrate that LT stimulates production of MUC2 mucin by goblet cells in human small intestine, enhancing the protective barrier between pathogens and enterocytes. In contrast, using explants of human small intestine as well as small intestinal enteroids, we show that EatA counters this host defense by engaging and degrading the MUC2 mucin barrier to promote bacterial access to target enterocytes and ultimately toxin delivery suggesting that EatA plays a crucial role in the molecular pathogenesis of ETEC. These findings may inform novel approaches to prevention of the acute diarrheal illness as well as the sequelae associated with ETEC and other pathogens that rely on EatA and similar proteases for efficient interaction with their human hosts.

Epidemiologic potentials and correlational analysis of Vibrio species and virulence toxins from water sources in greater Bushenyi districts, Uganda

Adequate water supply is one of the public health issues among the population living in low-income settings. Vibriosis remain a significant health challenge drawing the attention of both healthcare planners and researchers in South West districts of Uganda. Intending to clamp down the disease cases in the safest water deprive locality, we investigated the virulent toxins as contaminants and epidemiologic potentials of Vibrio species recovered from surface waters in greater Bushenyi districts, Uganda. Surface water sources within 46 villages located in the study districts were obtained between June and October 2018. Standard microbiological and molecular methods were used to analyse samples. Our results showed that 981 presumptive isolates retrieved cell counts of 10–100 CFU/g, with, with (640) 65% confirmed as Vibrio genus using polymerase chain reaction, which is distributed as follows; V. vulnificus 46/640 (7.2%), V. fluvialis 30/594 (5.1), V. parahaemolyticus 21/564 (3.7), V. cholera 5/543 (0.9), V. alginolyticus 62/538 (11.5) and V. mimicus 20/476 (4.2). The virulence toxins observed were heat-stable enterotoxin (stn) 46 (82.10%), V. vulnificus virulence gene (vcgCPI) 40 (87.00%), extracellular haemolysin gene {vfh 21 (70.00)} and Heme utilization protein gene {hupO 5 (16.70)}. The cluster analysis depicts hupO (4.46% n = 112); vfh (18.75%, n = 112); vcgCPI and stn (35.71%, & 41.07%, n = 112). The principal component analysis revealed the toxins (hupO, vfh) were correlated with the isolate recovered from Bohole water (BW) source, while (vcgCPI, stn) toxins are correlated with natural raw water (NRW) and open springs (OS) water sources isolates. Such observation indicates that surface waters sources are highly contaminated with an odds ratio of 1.00, 95% CI (70.48–90.5), attributed risk of (aR = 64.29) and relative risk of (RR = 73.91). In addition, it also implies that the surface waters sources have > 1 risk of contamination with vfh and > six times of contamination with hupO (aR = 40, − 66). This is a call of utmost importance to the population, which depends on these water sources to undertake appropriate sanitation, personal hygienic practices and potential measures that ensure water quality.