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Controlled human infection models are important tools for the evaluation of vaccines against diseases where an appropriate correlate of protection has not been identified. Enterotoxigenic Escherichia coli (ETEC) strain LSN03-016011/A (LSN03) is an LT enterotoxin and CS17-expressing ETEC strain useful for evaluating vaccine candidates targeting LT-expressing strains. We sought to confirm the ability of the LSN03 strain to induce moderate-to-severe diarrhea in a healthy American adult population, as well as the impact of immunization with an investigational cholera/ETEC vaccine (VLA-1701) on disease outcomes. A randomized, double-blinded pilot study was conducted in which participants received two doses of VLA1701 or placebo orally, one week apart; eight days after the second vaccination, 30 participants (15 vaccinees and 15 placebo recipients) were challenged with approximately 5 × 109 colony-forming units of LSN03. The vaccine was well tolerated, with no significant adverse events. The vaccine also induced serum IgA and IgG responses to LT. After challenge, 11 of the placebo recipients (73.3%; 95%CI: 48.0–89.1) and 7 of the VLA1701 recipients (46.7%; 95%CI: 24.8–68.8) had moderate-to-severe diarrhea (p = 0.26), while 14 placebo recipients (93%) and 8 vaccine recipients (53.3%) experienced diarrhea of any severity, resulting in a protective efficacy of 42.9% (p = 0.035). In addition, the vaccine also appeared to provide protection against more severe diarrhea (p = 0.054). Vaccinees also tended to shed lower levels of the LSN03 challenge strain compared to placebo recipients (p = 0.056). In addition, the disease severity score was lower for the vaccinees than for the placebo recipients (p = 0.046). In summary, the LSN03 ETEC challenge strain induced moderate-to-severe diarrhea in 73.3% of placebo recipients. VLA1701 vaccination ameliorated disease severity, as observed by several parameters, including the percentage of participants experiencing diarrhea, as well as stool frequency and ETEC severity scores. These data highlight the potential value of LSN03 as a suitable ETEC challenge strain to evaluate LT-based vaccine targets (NCT03576183). [ABSTRACT FROM AUTHOR]

Background. Enterotoxigenic E. coli (ETEC) is a principal cause of diarrhea in travelers, deployed military personnel, and children living in low to middle-income countries. ETEC expresses a variety of virulence factors including colonization factors (CF) that facilitate adherence to the intestinal mucosa. We assessed the protective efficacy of a tip-localized subunit of CF antigen I (CFA/I), CfaE, delivered intradermally with the mutant E. coli heat-labile enterotoxin, LTR192G, in a controlled human infection model (CHIM). Methods. Three cohorts of healthy adult subjects were enrolled and given three doses of 25 μg CfaE + 100 ng LTR192G vaccine intradermally at 3-week intervals. Approximately 28 days after the last vaccination, vaccinated and unvaccinated subjects were admitted as inpatients and challenged with approximately 2 × 107 cfu of CFA/I+ ETEC strain H10407 following an overnight fast. Subjects were assessed for moderate-to-severe diarrhea for 5 days post-challenge. Results. A total of 52 volunteers received all three vaccinations; 41 vaccinated and 43 unvaccinated subjects were challenged and assessed for moderate-to-severe diarrhea. Naïve attack rates varied from 45.5% to 64.7% across the cohorts yielding an overall efficacy estimate of 27.8% (95% confidence intervals: −7.5–51.6%). In addition to reducing moderate–severe diarrhea rates, the vaccine significantly reduced loose stool output and overall ETEC disease severity. Conclusions. This is the first study to demonstrate protection against ETEC challenge after intradermal vaccination with an ETEC adhesin. Further examination of the challenge methodology is necessary to address the variability in naïve attack rate observed among the three cohorts in the present study. [ABSTRACT FROM AUTHOR]

Few studies have examined how avian life-history traits vary within populations as elevation increases and climate becomes more severe. We compared egg and clutch sizes of Mountain Bluebirds (Sialia currucoides) nesting at two elevations (1500 m and 2500 m above sea level) in the Bighorn Mountains of Wyoming over two years. Eggs laid by females at the high-elevation site were, on average, significantly (6%) smaller in volume than eggs laid by their lower-elevation counterparts. Across elevations, egg size showed a significant positive correlation with female body condition (weight relative to size), and high-elevation females had significantly lower indices of condition than low-elevation females. Temperatures during clutch formation were colder at the high-elevation site, and egg size was negatively related to temperature after controlling for the effects of female condition. Clutches of females at high elevations were, on average, marginally smaller (by 5%, ~0.3 eggs) than clutches of low-elevation females. Unlike egg size, clutch size was unrelated to either female condition or temperature during clutch formation. This suggests that, when under energetic or nutritional stress at high elevations, females sacrifice egg size before sacrificing clutch size. Key words: clutch size, egg size, elevation, female condition, Mountain Bluebird, Sialia currucoides, temperature. Pocos estudios han examinado como los caracteres de historia de vida cambian dentro de poblaciones a medida que la elevacion aumenta y el clima se torna mas severo. Comparamos el tamano de los huevos y de la nidada de individuos de Sialia currucoides que se encontraban nidificando a dos altitudes (1500 m y 2500 m sobre el nivel del mar) en las Montanas Bighorn de Wyoming durante dos anos. Los huevos puestos por hembras del sitio a mayor altitud fueron, en promedio, significativamente (6%) menores en volumen que los huevos puestos por hembras en los sitios de baja altitud. A traves del rango de elevacion, el tamano de los huevos mostro una relacion positiva y significativa con la condicion corporal (peso en relacion con el tamano). Las hembras del sitio de mayor altitud tuvieron indices de condicion significativamente menores que las hembras del sitio de baja altitud. Las temperaturas durante la formacion de la nidada en el sitio de mayor altitud fueron, en promedio, marginalmente menores (en un 5%, ~0.3 huevos) que las nidadas de las hembras del sitio de menor elevacion. De modo contrario al tamano del huevo, el tamano de la nidada no se relaciono con la condicion de las hembras ni con la temperatura durante la formacion de la nidada. Esto sugiere que cuando las hembras se encuentran bajo estres energetico o nutricional a altitudes altas, estas sacrifican el tamafio del huevo antes de sacrificar el tamano de la nidada.

Background: Human challenge models for enterotoxigenic Escherichia coli (ETEC) facilitate vaccine down-selection. The B7A (O148:H28 CS6+LT+ST+) strain is important for vaccine development. We sought to refine the B7A model by identifying a dose and fasting regimen consistently inducing moderate-severe diarrhea. Methods: An initial cohort of 28 subjects was randomized (1:1:1:1) to receive B7A following an overnight fast at doses of 108 or 109 colony forming units (cfu) or a 90-minute fast at doses of 109 or 1010 cfu. A second cohort included naïve and rechallenged subjects who had moderate-severe diarrhea and were given the target regimen. Immune responses to important ETEC antigens were assessed. Results: Among subjects receiving 108 cfu of B7A, overnight fast, or 109 cfu, 90-minute fast, 42.9% (3/7) had moderate-severe diarrhea. Higher attack rates (71.4%; 5/7) occurred in subjects receiving 109 cfu, overnight fast, or 1010 cfu, 90-minute fast. Upon rechallenge with 109 cfu of B7A, overnight fast, 5/11 (45.5%) had moderate-severe diarrhea; the attack rate among concurrently challenge naïve subjects was 57.9% (11/19). Anti-CS6, O148 LPS and LT responses were modest across all groups. Conclusions: An overnight fast enabled a reduction in the B7A inoculum dose; however, the attack rate was inconsistent and protection upon rechallenge was minimal. [ABSTRACT FROM AUTHOR]

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of bacterial diarrhea both among children in low and middle income countries and in travelers to these regions. Although there are several approaches to develop an effective vaccine for ETEC, no licensed vaccines are currently available. The most advanced ETEC vaccine candidates include multiple colonization factors along with the heat labile toxin B subunit. In the absence of known correlates of protection, and to understand the mechanism of protection, monitoring immune responses to a majority of the vaccine associated antigens using various types of samples is needed. Unfortunately, a traditional ELISA is time consuming, labor intensive and requires substantial amounts of antigens and sample volumes. To address these constraints, we developed and validated a novel high throughput electrochemiluminescent (ECL) - based multiplex immunoassay using Meso Scale Discovery (MSD) platform for analyzing immune responses to ETEC antigens. The ETEC multiplex ECL assay is an 8-plex assay which includes the ETEC colonization factor antigens (CFA/I, CS1, CS2, CS3, CS5 and CS6) along with the two subunits of heat labile toxin (LTA and LTB). Our data suggested that a single dilution of sample provides a quantifiable result for a wide range of sample titers. To compare ETEC multiplex ECL with ELISA, we carried out assays using the same antigens with the two immunoassay platforms using a common sample set of serum and ALS (antibodies in lymphocyte supernatant) specimens. The MSD platform achieved excellent correlations with ELISA for the antigens tested, consistently detecting comparable antibody levels in the samples. The ETEC multiplex ECL can serve as a fundamental platform in evaluating performances of candidate ETEC vaccines in future field trials. • A novel ETEC multiplex ECL assay was developed which achieved excellent correlations with conventional ELISA. • ETEC multiplex ECL assay would minimize volumes of samples and antigens as well as save time and labor. • The ETEC multiplex assay could serve as an improved platform for evaluation of candidate ETEC vaccines. [ABSTRACT FROM AUTHOR]

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 . [ABSTRACT FROM AUTHOR]

Background: Household contacts of cholera patients have a 100 times higher risk of developing a cholera infection than the general population. To compare the genetic relatedness of clinical and water source Vibrio cholerae isolates from cholera patients' households across three outbreaks, we analyzed these isolates using whole-genome-sequencing (WGS) and multilocus variable-number tandem-repeat analysis (MLVA). Results: The WGS analyses revealed that 80% of households had source water isolates that were more closely related to clinical isolates from the same household than to any other isolates. While in another 20% of households an isolate from a person was more closely related to clinical isolates from another household than to source water isolates from their own household. The mean pairwise differences in single nucleotide-variant (SNV) counts for isolates from the same household were significantly lower than those for different households (2.4 vs. 7.7 p < 0.0001), and isolates from the same outbreak had significantly fewer mean pairwise differences compared to isolates from different outbreaks (mean: 6.2 vs. 8.0, p < 0.0001). Based on MLVA in outbreak 1, we observed that the majority of households had clinical isolates with MLVA genotypes related to other clinical isolates and unrelated to water source isolates from the same household. While in outbreak 3, there were different MLVA genotypes between households, however within the majority of households, the clinical and water source isolates had the same MLVA genotypes. The beginning of outbreak 2 resembled outbreak 1 and the latter part resembled outbreak 3. We validated our use of MLVA by comparing it to WGS. Isolates with the identical MLVA genotype had significantly fewer mean pairwise SNV differences than those isolates with different MLVA genotypes (mean: 4.8 vs. 7.7, p < 0.0001). Furthermore, consistent with WGS results, the number of pairwise differences in the five MLVA loci for isolates within the same household was significantly lower than isolates from different households (mean: 1.6 vs. 3.0, p < 0.0001). Conclusion: These results suggest that transmission patterns for cholera are a combination of person-to-person and water-to-person cholera transmission with the proportions of the two modes varying within and between outbreaks. [ABSTRACT FROM AUTHOR]

The article presents research which assessed the evidence for the heterozygote instability in house wrens microsatellite loci. The researchers analyzed 20 microsatellite loci of Troglodytes aedon also called as the songbird. They found the difference between the homologue and the allele affects the mutation rate of this species. The authors argue that instability in the bird's heterozygote is possible at microsatellite loci.

Female birds are hypothesized to mate outside the pairbond to secure alleles that enhance the fitness potential of their offspring. To test this, researchers typically compare fitness-related attributes of extra-pair (EP) and within-pair (WP) half-siblings. Often neglected, however, is the possibility that females may provide EP offspring with certain non-genetic advantages. For example, in species in which eggs hatch asynchronously, females may place EP offspring amongst earlier-laid eggs in the clutch. Because they tend to hatch first, chicks from earlier-laid eggs are often larger and more developed than their nestmates and thus have a competitive advantage. We tested for an association between offspring paternity and position in the laying/hatching sequence in a Wyoming population of the house wren Troglodytes aedon. Eggs in this population always hatch asynchronously, usually in the order laid, over 24–48 h, setting up a stair-step-like size hierarchy within broods early in the nestling stage. This suggested that we could thus use a chick's mass relative to that of its nestmates as an index of the chick's position in the laying sequence. We first confirmed this, showing that position in the laying sequence explains 70% of the variance in chick size relative to nestmates. We then compared masses of EP and WP offspring shortly after hatching in 27 broods with mixed paternity. On average, EP offspring weighed 14% more than their WP counterparts, a highly significant difference. Our results therefore suggest that EP offspring are more likely than WP offspring to occur in earlier-laid eggs, thus gaining a size-based competitive advantage. We recommend that, when comparing EP and WP half-siblings in species in which eggs hatch asynchronously, researchers test for this potential maternally derived effect on offspring performance. [ABSTRACT FROM AUTHOR]

In many bird species that conceal nests within cavities or burrows, only females incubate eggs, but both sexes feed young after hatching. How males in such species discover that eggs have hatched and start provisioning offspring is unknown. We video-taped 26 house wren (Troglodytes aedon) nests continuously before, during, and after hatching began to test four hypotheses as to how males might learn of hatching: (i) females signal hatching to males; (ii) males hear hatchlings vocalizing; (iii) males observe changes in their mate's behaviour (e.g., entering and exiting nests more frequently and/or carrying eggshells or food); and (iv) males enter the nest cavity and encounter hatchlings directly. We did not detect any unique visual or vocal displays by females immediately post-hatching and so could not support the first hypothesis. At only three of 26 nests did it appear that males might have been stimulated to begin feeding solely as a result of either hearing offspring from outside the nest or observing changes in their mate's behaviour. Rather, males at most nests did not begin delivering food until after they had entered nest cavities and presumably had direct visual and/or tactile contact with hatchlings, even though most of these males previously had the opportunity to hear offspring from outside nests and/or see their mates acting parentally. Our finding that most males do not start delivering food until they have had direct exposure to young is consistent with recent neurological studies in mammals and birds that suggest direct exposure to young triggers a cascade of gene expression in the brain that induces nurturing behaviour. [ABSTRACT FROM AUTHOR]

Indicator models of sexual selection suggest that costly ornaments signal reliable information regarding an individual's quality to potential mates. In species that produce altricial offspring, the amount of parental care provided by both males and females can impact reproductive success. The Good Parent Hypothesis proposes that ornamentation in biparental species can act as an honest signal of parental ability to potential mates. We tested this hypothesis using the mountain bluebird ( Sialia currucoides), a sexually dichromatic, socially monogamous species in which both sexes have structurally based ornamental plumage coloration. A male's plumage color predicted neither the rate at which it provisioned nestlings nor brood growth rate. The same was true for females. We also found no indication of assortative mating by color or body condition. Feeding rates within pairs were positively correlated, which we suggest may be due to pairs responding similarly to the perceived needs of nestlings or to local area prey availability. In sum, our results do not support the Good Parent Hypothesis as an explanation for the evolution of ornamental plumage color in mountain bluebirds. We suggest alternative hypotheses for the evolution of ornamental plumage color in this species. [ABSTRACT FROM AUTHOR]

Polymerase chain reaction (PCR)-based methods to determine the sex of birds are well established and have seen few modifications since they were first introduced in the 1990s. Although these methods allowed for sex determination in species that were previously difficult to analyse, they were not conducive to high-throughput analysis because of the laboriousness of DNA extraction and gel electrophoresis. We developed a high-throughput real-time PCR-based method for analysis of sex in birds, which uses noninvasive sample collection and avoids DNA extraction and gel electrophoresis. [ABSTRACT FROM AUTHOR]

Abstract Background There is no licensed vaccine against enterotoxigenic Escherichia coli (ETEC), a major cause of diarrhea-associated morbidity and mortality among infants and children in low-income countries and travelers. The results of this vaccination/challenge study demonstrate strong protection by an attenuated ETEC vaccine candidate, ACE527, when co-administered with a mucosal adjuvant, the double-mutant heat-labile toxin (dmLT) of ETEC. Methods Sixty healthy adults participated in a randomized, placebo-controlled, double-blind study with three doses of lyophilized ACE527 (∼3 × 109 of each strain per dose) administered orally with or without dmLT adjuvant (25 µg/dose). Six months later, 36 of these volunteers and a control group of 21 unvaccinated volunteers were challenged with virulent ETEC strain H10407. The primary outcome was severe diarrhea, defined as passing >800 g of unformed stools during the inpatient period following challenge. Findings The vaccine was well tolerated and induced robust immune responses to key antigens. The protective efficacy (PE) against the primary outcome of severe diarrhea was 65.9% (95% confidence interval [CI] 5.4–87.7, p = 0.003). Among subjects receiving the adjuvanted vaccine, the attack rate of severe diarrhea was 23.1, while in unimmunized controls it was 67.7%. The PE against diarrhea of any severity was 58.5% (95% CI 3.8– 82.1, p = 0.016). There was a strong inverse correlation between shedding of the vaccine strain after either of the first two doses and absence of severe diarrhea upon challenge (RR = 0.29, 95% CI 0.08–1.05, p = 0.041). Challenge strain shedding was 10-fold lower in those receiving the adjuvant than in those receiving vaccine alone. The unadjuvanted vaccine was not protective (PE = 23.1%). Interpretation The results of this study support further development of ACE527 + dmLT as a vaccine for children in endemic countries and travelers. This is the first clinical demonstration that dmLT can contribute significantly to vaccine efficacy and may warrant testing with other oral vaccines. (ClinicalTrials.gov registration: NCT01739231). [ABSTRACT FROM AUTHOR]

Shigella causes morbidity and mortality worldwide, primarily affecting young children living in low-resource settings. It is also of great concern due to increasing antibiotic resistance, and is a priority organism for the World Health Organization. A Shigella vaccine would decrease the morbidity and mortality associated with shigellosis, improve child health, and decrease the need for antibiotics. Controlled human infection models (CHIMs) are useful tools in vaccine evaluation for early up- or down-selection of vaccine candidates and potentially useful in support of licensure. Over time, the methods employed in these models have become more uniform across sites performing CHIM trials, although some differences in conduct persist. In November 2017, a Shigella CHIM workshop was convened in Washington, District of Columbia. Investigators met to discuss multiple aspects of these studies, including study procedures, clinical and immunological endpoints, and shared experiences. This article serves as a uniform procedure by which to conduct Shigella CHIM studies., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.)