Exploring the views of infection consultants in England on a novel delinked funding model for antimicrobials: the SMASH study.

Objectives: A novel 'subscription-type' funding model was launched in England in July 2022 for ceftazidime/avibactam and cefiderocol. We explored the views of infection consultants on important aspects of the delinked antimicrobial funding model. Methods: An online survey was sent to all infection consultants in NHS acute hospitals in England. Results: The response rate was 31.2% (235/753). Most consultants agreed the model is a welcome development (69.8%, 164/235), will improve treatment of drug-resistant infections (68.5%, 161/235) and will stimulate research and development of new antimicrobials (57.9%, 136/235). Consultants disagreed that the model would lead to reduced carbapenem use and reported increased use of cefiderocol post-implementation. The presence of an antimicrobial pharmacy team, requirement for preauthorization by infection specialists, antimicrobial stewardship ward rounds and education of infection specialists were considered the most effective antimicrobial stewardship interventions. Under the new model, 42.1% (99/235) of consultants would use these antimicrobials empirically, if risk factors for antimicrobial resistance were present (previous infection, colonization, treatment failure with carbapenems, ward outbreak, recent admission to a high-prevalence setting).Significantly higher insurance and diversity values were given to model antimicrobials compared with established treatments for carbapenem-resistant infections, while meropenem recorded the highest enablement value. Use of both 'subscription-type' model drugs for a wide range of infection sites was reported. Respondents prioritized ceftazidime/avibactam for infections by bacteria producing OXA-48 and KPC and cefiderocol for those producing MBLs and infections with Stenotrophomonas maltophilia, Acinetobacter spp. and Burkholderia cepacia. Conclusions: The 'subscription-type' model was viewed favourably by infection consultants in England.

Use of cidofovir in a patient with severe mpox and uncontrolled HIV infection.

A 48-year-old man with poorly controlled HIV presented with severe human monkeypox virus (hMPXV) infection, having completed 2 weeks of tecovirimat at another hospital. He had painful, ulcerating skin lesions on most of his body and oropharyngeal cavity, with subsequent Ludwig's angina requiring repeated surgical interventions. Despite commencing a second, prolonged course of tecovirimat, he did not objectively improve, and new lesions were still noted at day 24. Discussion at the UK National Health Service England High Consequence Infectious Diseases Network recommended the use of 3% topical and then intravenous cidofovir, which was given at 5 mg/kg; the patient made a noticeable improvement after the first intravenous dose. He received further intravenous doses at 7 days and 21 days after the dose and was discharged at day 52. Cidofovir is not licensed for use in treatment of hMPXV infection. Data for cidofovir use in hMPXV are restricted to studies in animals. Four other documented cases of cidofovir use against hMPXV have been reported in the USA in 2022, but we present its first use in the UK. The scarcity of studies into the use of cidofovir in this condition clearly shows the need for robust studies to assess efficacy, optimum dosage, timing, and route of administration.

Mucosal-Associated Invariant T cells exhibit distinct functional signatures associated with protection against typhoid fever.

We have previously demonstrated that Mucosal-Associated Invariant T (MAIT) cells secrete multiple cytokines after exposure to Salmonella enterica serovar Typhi (S. Typhi), the causative agent of typhoid fever in humans. However, whether cytokine secreting MAIT cells can enhance or attenuate the clinical severity of bacterial infections remain debatable. This study characterizes human MAIT cell functions in subjects participating in a wild-type S. Typhi human challenge model. Here, we found that MAIT cells exhibit distinct functional signatures associated with protection against typhoid fever. We also observed that the cytokine patterns of MAIT cell responses, rather than the average number of cytokines expressed, are more predictive of typhoid fever outcomes. These results might enable us to objectively, based on functional parameters, identify cytokine patterns that may serve as predictive biomarkers during natural infection and vaccination.

Immunogenicity of a Third Scheduled Dose of Rotarix in Australian Indigenous Infants: A Phase IV, Double-blind, Randomized, Placebo-Controlled Clinical Trial.

BACKGROUND: Rotarix (GlaxoSmithKline) oral rotavirus vaccine is licensed as 2 doses in the first 6 months of life. In settings with high child mortality rates, clinical protection conferred by 2 doses of Rotarix is reduced. We assessed vaccine immune response when an additional dose of Rotarix was given to Australian Aboriginal children 6 to <12 months old. METHODS: ORVAC is a 2-stage, double-blind, randomized, placebo-controlled trial. Australian Aboriginal children 6 to <12 months old who had received 1 or 2 prior doses of Rotarix rotavirus vaccine were randomized 1:1 to receive an additional dose of Rotarix or matched placebo. The primary immunological end point was seroresponse defined as an anti-rotavirus immunoglobulin A level ≥20 AU/mL, 28-56 days after the additional dose of Rotarix or placebo. RESULTS: Between March 2018 and August 2020, a total of 253 infants were enrolled. Of these, 178 infants (70%) had analyzable serological results after follow-up; 89 were randomized to receive Rotarix, and 89 to receive placebo. The proportion with seroresponse was 85% after Rotarix compared with 72% after placebo. There were no occurrences of intussusception or any serious adverse events. CONCLUSIONS: An additional dose of Rotarix administered to Australian Aboriginal infants 6 to <12 months old increased the proportion with a vaccine seroresponse. CLINICAL TRIALS REGISTRATION: NCT02941107.

Development and validation of a dynamic 48-hour in-hospital mortality risk stratification for COVID-19 in a UK teaching hospital: a retrospective cohort study.

OBJECTIVES: To develop a disease stratification model for COVID-19 that updates according to changes in a patient's condition while in hospital to facilitate patient management and resource allocation. DESIGN: In this retrospective cohort study, we adopted a landmarking approach to dynamic prediction of all-cause in-hospital mortality over the next 48 hours. We accounted for informative predictor missingness and selected predictors using penalised regression. SETTING: All data used in this study were obtained from a single UK teaching hospital. PARTICIPANTS: We developed the model using 473 consecutive patients with COVID-19 presenting to a UK hospital between 1 March 2020 and 12 September 2020; and temporally validated using data on 1119 patients presenting between 13 September 2020 and 17 March 2021. PRIMARY AND SECONDARY OUTCOME MEASURES: The primary outcome is all-cause in-hospital mortality within 48 hours of the prediction time. We accounted for the competing risks of discharge from hospital alive and transfer to a tertiary intensive care unit for extracorporeal membrane oxygenation. RESULTS: Our final model includes age, Clinical Frailty Scale score, heart rate, respiratory rate, oxygen saturation/fractional inspired oxygen ratio, white cell count, presence of acidosis (pH <7.35) and interleukin-6. Internal validation achieved an area under the receiver operating characteristic (AUROC) of 0.90 (95% CI 0.87 to 0.93) and temporal validation gave an AUROC of 0.86 (95% CI 0.83 to 0.88). CONCLUSIONS: Our model incorporates both static risk factors (eg, age) and evolving clinical and laboratory data, to provide a dynamic risk prediction model that adapts to both sudden and gradual changes in an individual patient's clinical condition. On successful external validation, the model has the potential to be a powerful clinical risk assessment tool. TRIAL REGISTRATION: The study is registered as 'researchregistry5464' on the Research Registry (www.researchregistry.com).

A controlled human infection model of Streptococcus pyogenes pharyngitis (CHIVAS-M75): an observational, dose-finding study.

BACKGROUND: Streptococcus pyogenes is a leading cause of infection-related morbidity and mortality. A reinvigorated vaccine development effort calls for new clinically relevant human S pyogenes experimental infection models to support proof of concept evaluation of candidate vaccines. We describe the initial Controlled Human Infection for Vaccination Against S pyogenes (CHIVAS-M75) study, in which we aimed to identify a dose of emm75 S pyogenes that causes acute pharyngitis in at least 60% of volunteers when applied to the pharynx by swab. METHODS: This observational, dose-finding study was done in a clinical trials facility in Melbourne (VIC, Australia). Groups of healthy volunteers aged 18-40 years, at low risk of complicated S pyogenes disease, and without high type-specific anti-emm75 IgG antibodies against the challenge strain were challenged and closely monitored as inpatients for up to 6 days, and then as outpatients for 6 months. Antibiotics were started upon diagnosis (clinical signs and symptoms of pharyngitis and a positive rapid molecular test) or after 5 days in those without pharyngitis. Rapid test results were confirmed by standard bacterial culture. After a sentinel participant, cohorts of five and then ten participants were challenged, with protocol-directed dose-escalation or de-escalation for subsequent cohorts. The primary outcome was the proportion of participants at each dose level with pharyngitis by day 5 after challenge. The study is registered with ClinicalTrials.gov, NCT03361163. FINDINGS: Between July 10, 2018, and Sept 23, 2019, 25 healthy adults were challenged with emm75 S pyogenes and included in analyses. Pharyngitis was diagnosed in 17 (85%; 95% CI 62-97) of 20 participants at the starting dose level (1-3 × 105 colony-forming units [CFU]/mL). This high proportion prompted dose de-escalation. At the lower dose level (1-3 × 104 CFU/mL), pharyngitis was diagnosed in one of five participants. Immunological, biochemical, and microbiological results supported the clinical picture, with acute symptomatic pharyngitis characterised by pharyngeal colonisation by S pyogenes accompanied by significantly elevated C-reactive protein and inflammatory cytokines (eg, interferon-γ and interleukin-6), and modest serological responses to streptolysin O and deoxyribonuclease B. There were no severe (grade 3) or serious adverse events related to challenge. INTERPRETATION: We have established a reliable pharyngitis human infection model with reassuring early safety findings to accelerate development of vaccines and other interventions to control disease due to S pyogenes. FUNDING: Australian National Health and Medical Research Council.

OPTIMUM study protocol: an adaptive randomised controlled trial of a mixed whole-cell/acellular pertussis vaccine schedule.

INTRODUCTION: Combination vaccines containing whole-cell pertussis antigens were phased out from the Australian national immunisation programme between 1997 and 1999 and replaced by the less reactogenic acellular pertussis (aP) antigens. In a large case-control study of Australian children born during the transition period, those with allergist diagnosed IgE-mediated food allergy were less likely to have received whole-cell vaccine in early infancy than matched population controls (OR: 0.77 (95% CI, 0.62 to 0.95)). We hypothesise that a single dose of whole-cell vaccine in early infancy is protective against IgE-mediated food allergy. METHODS AND ANALYSIS: This adaptive double-blind randomised controlled trial is investigating whether a mixed whole-cell/aP vaccine schedule prevents allergic disease in the first year of life. The primary outcome is IgE-mediated food allergy by 12 months of age. Secondary outcomes include new onset of atopic dermatitis by 6 or 12 months of age; sensitisation to at least one allergen by 12 months of age; seroconversion in anti-pertussis toxin IgG titres after vaccination with aP booster at 18 months of age; and solicited systemic and local adverse events following immunisation with pertussis-containing vaccines. Analyses will be performed using a Bayesian group sequential design. ETHICS AND DISSEMINATION: This study has been approved by the Child and Adolescent Health Service Human Research Ethics Committee, Perth, Western Australia (RGS 00019). The investigators will ensure that this trial is conducted in accordance with the principles of the Declaration of Helsinki and with the International Conference on Harmonisation Guidelines for Good Clinical Practice. Individual consent will be requested. Parents will be reimbursed reasonable travel and parking costs to attend the study visits. The dissemination of these research findings will follow the National Health and Medical Research Council of Australia Open Access Policy. TRIAL REGISTRATION NUMBER: ACTRN12617000065392p.

Human Salmonella Typhi exposure generates differential multifunctional cross-reactive T-cell memory responses against Salmonella Paratyphi and invasive nontyphoidal Salmonella.

OBJECTIVE: There are no vaccines for most of the major invasive Salmonella strains causing severe infection in humans. We evaluated the specificity of adaptive T memory cell responses generated after Salmonella Typhi exposure in humans against other major invasive Salmonella strains sharing capacity for dissemination. METHODS: T memory cells from eleven volunteers who underwent controlled oral challenge with wt S. Typhi were characterised by flow cytometry for cross-reactive cellular cytokine/chemokine effector responses or evidence of degranulation upon stimulation with autologous B-lymphoblastoid cells infected with either S. Typhi, Salmonella Paratyphi A (PA), S. Paratyphi B (PB) or an invasive nontyphoidal Salmonella strain of the S. Typhimurium serovar (iNTSTy). RESULTS: Blood T-cell effector memory (TEM) responses after exposure to S. Typhi in humans evolve late, peaking weeks after infection in most volunteers. Induced multifunctional CD4+ Th1 and CD8+ TEM cells elicited after S. Typhi challenge were cross-reactive with PA, PB and iNTSTy. The magnitude of multifunctional CD4+ TEM cell responses to S. Typhi correlated with induction of cross-reactive multifunctional CD8+ TEM cells against PA, PB and iNTSTy. Highly multifunctional subsets and T central memory and T effector memory cells that re-express CD45 (TEMRA) demonstrated less heterologous T-cell cross-reactivity, and multifunctional Th17 elicited after S. Typhi challenge was not cross-reactive against other invasive Salmonella. CONCLUSION: Gaps in cross-reactive immune effector functions in human T-cell memory compartments were highly dependent on invasive Salmonella strain, underscoring the importance of strain-dependent vaccination in the design of T-cell-based vaccines for invasive Salmonella.

Point of Care Nucleic Acid Testing for SARS-CoV-2 in Hospitalized Patients: A Clinical Validation Trial and Implementation Study.

There is an urgent need for rapid SARS-CoV-2 testing in hospitals to limit nosocomial spread. We report an evaluation of point of care (POC) nucleic acid amplification testing (NAAT) in 149 participants with parallel combined nasal and throat swabbing for POC versus standard lab RT-PCR testing. Median time to result is 2.6 (IQR 2.3-4.8) versus 26.4 h (IQR 21.4-31.4, p < 0.001), with 32 (21.5%) positive and 117 (78.5%) negative. Cohen's κ correlation between tests is 0.96 (95% CI 0.91-1.00). When comparing nearly 1,000 tests pre- and post-implementation, the median time to definitive bed placement from admission is 23.4 (8.6-41.9) versus 17.1 h (9.0-28.8), p = 0.02. Mean length of stay on COVID-19 "holding" wards is 58.5 versus 29.9 h (p < 0.001). POC testing increases isolation room availability, avoids bed closures, allows discharge to care homes, and expedites access to hospital procedures. POC testing could mitigate the impact of COVID-19 on hospital systems.

Whole-Cell Pertussis Vaccination and Decreased Risk of IgE-Mediated Food Allergy: A Nested Case-Control Study.

BACKGROUND: Rates of food allergy have increased markedly in Australia and other high- income countries in recent years. On the basis of ecological observations, and the known immunologic characteristics of whole-cell pertussis (wP) compared with acellular pertussis (aP) vaccines, we hypothesized that wP vaccination in infancy protects against the development of food allergy. OBJECTIVE: To determine whether infants who receive wP in infancy were less likely to develop IgE-mediated food allergy than those who received aP. METHODS: Retrospective cohort-nested case-control study of Australian children born in the period 1997 to 1999, the period of transition from using wP-containing to aP-containing vaccines. Children diagnosed with IgE-mediated food allergy were individually matched to 10 controls by date of birth, socioeconomic decile, and jurisdiction of birth. The odds ratio of vaccination with wP versus aP among cases and matched controls was calculated using conditional logistic regression. RESULTS: The odds ratio of receiving the first dose as wP (rather than aP) among cases of food allergy compared with controls was 0.77 (95% CI, 0.62-0.95). The results of secondary analyses (any dose as wP vs aP-only, and wP-only vs aP-only) were broadly similar. CONCLUSIONS: Australian infants who received wP vaccines were less likely to be diagnosed with food allergy in childhood than contemporaneous children who received aP vaccines. If a protective effect is confirmed in a randomized controlled trial, wP or mixed wP and aP vaccination schedules could form part of an effective strategy for combating the rise in food allergies.

The ORVAC trial protocol: a phase IV, double-blind, randomised, placebo-controlled clinical trial of a third scheduled dose of Rotarix rotavirus vaccine in Australian Indigenous infants to improve protection against gastroenteritis.

INTRODUCTION: Rotavirus vaccines were introduced into the Australian National Immunisation Program in 2007. Despite this, Northern Territory Indigenous children continue to be hospitalised with rotavirus at a rate more than 20 times higher than non-Indigenous children in other Australian jurisdictions, with evidence of waning protection in the second year of life. We hypothesised that scheduling an additional (third) dose of oral human rotavirus vaccine (Rotarix, GlaxoSmithKline) for children aged 6 to <12 months would improve protection against clinically significant all-cause gastroenteritis. METHODS AND ANALYSIS: This Bayesian adaptive clinical trial will investigate whether routinely scheduling an additional dose of Rotarix for Australian Indigenous children aged 6 to <12 months old confers significantly better protection against clinically important all-cause gastroenteritis than the current two-dose schedule at 2 and 4 months old. There are two coprimary endpoints: (1) seroconversion from baseline serum anti-rotavirus immunoglobulin A (IgA) titre <20 U/mL prior to an additional dose of Rotarix/placebo to serum anti-rotavirus IgA titre >20 U/mL following the administration of the additional dose of Rotarix/placebo and (2) time from randomisation to medical attendance (up to age 36 months old) for which the primary reason is acute gastroenteritis/diarrhoea. Secondary endpoints include the change in anti-rotavirus IgA log titre, time to hospitalisation for all-cause diarrhoea and for rotavirus-confirmed gastroenteritis/diarrhoea, and rotavirus notification. Analysis will be based on Bayesian inference with adaptive sample size. ETHICS, REGISTRATION AND DISSEMINATION: Ethics approval has been granted by Central Australian Human Research Ethics Committee (HREC-16-426) and Human Research Ethics Committee of the Northern Territory Department of Health and Menzies School of Health Research (HREC-2016-2658). Study investigators will ensure the trial is conducted in accordance with the principles of the Declaration of Helsinki and with the ICH Guidelines for Good Clinical Practice. Individual participant consent will be obtained. Results will be disseminated via peer-reviewed publication. The trial is registered with Clinicaltrials.gov (NCT02941107) and important modifications to this protocol will be updated. TRIAL REGISTRATION NUMBER: NCT02941107; Pre-results.

Identification of the determinants of incomplete vaccination in Australian children.

BACKGROUND AND AIMS: We aimed to understand the risk factors associated with incomplete vaccination, which may help to identify and prioritise opportunities to intervene. METHODS: Consenting parents of children <6 years old attending an outpatient clinic completed a questionnaire, which captured demographic information and their level of agreement with belief statements about vaccination using a 7-point Likert scale. Vaccination status was determined from the Australian Childhood Immunisation Register and deemed either "complete" (no doses overdue) or "incomplete" (1 or more doses overdue) at the time of questionnaire completion. RESULTS: Of 589 children of respondents, 116 (20%) had an incomplete vaccination status. Of these, nearly two-thirds (63%) of parents believed that their child was, in fact, fully-vaccinated. Compared to those with a complete vaccine status, children with an incomplete vaccine status were more likely to be born overseas (p < 0.001), have a larger family size (p = 0.02) and to have parents with lower educational attainment (p = 0.001). Parents of children with an incomplete status reported more doubt about the importance of vaccination and greater concern about vaccine safety, compared to parents of children with a complete status. CONCLUSION: Most parents are supportive of vaccination. Sociodemographic factors may contribute more to the risk of incomplete vaccination than attitudes or beliefs. Some parents are unaware of their child's vaccination status, suggesting that simple and modern reminders may assist parents to keep up to date.

Controlled human infection for vaccination against Streptococcus pyogenes (CHIVAS): Establishing a group A Streptococcus pharyngitis human infection study.

Group A Streptococcus (GAS) is a highly-adapted and human-restricted pathogen responsible for a high global burden of disease across a diverse clinical spectrum. Vaccine development has been impeded by scientific, regulatory, and commercial obstacles. Human infection studies (HIS) are increasingly contributing to drug, diagnostics, and vaccine development, reducing uncertainty at early stages, especially for pathogens with animal models that incompletely reproduce key elements of human disease. We review the small number of historical GAS HIS and present the study protocol for a dose-ranging inpatient study in healthy adults. The primary objective of the study is to establish a new GAS pharyngitis HIS with an attack rate of at least 60% as a safe and reliable platform for vaccine evaluation and pathogenesis research. According to an adaptive dose-ranging study design, emm75 GAS doses manufactured in keeping with principles of Good Manufacturing Practice will be directly applied by swab to the pharynx of carefully screened healthy adult volunteers at low risk of severe complicated GAS disease. Participants will remain as closely monitored inpatients for up to six days, observed for development of the primary outcome of acute symptomatic pharyngitis, as defined by clinical and microbiological criteria. All participants will be treated with antibiotics and followed as outpatients for six months. An intensive sampling schedule will facilitate extensive studies of host and organism dynamics during experimental pharyngitis. Ethics approval has been obtained and the study has been registered at ClinicalTrials.gov (NCT03361163).

Protocol for Pertussis Immunisation and Food Allergy (PIFA): a case-control study of the association between pertussis vaccination in infancy and the risk of IgE-mediated food allergy among Australian children.

INTRODUCTION: Atopic diseases, including food allergy, have become a predominant cause of chronic illness among children in developed countries. In Australia, a rise in hospitalisations among infants coded as anaphylaxis to foods coincided with the replacement of whole-cell pertussis (wP) vaccine with subunit acellular pertussis (aP) vaccine on the national immunisation schedule in the late 1990s. Atopy is characterised by a tendency to mount T helper type 2 (Th2) responses to otherwise innocuous environmental antigens. Compared with infants who receive aP as their first pertussis vaccine, those who receive wP appear less likely to mount Th2 immune responses to either vaccine or extraneous antigens. We therefore speculate that removal of wP from the vaccine schedule contributed to the observed rise in IgE-mediated food allergy among Australian infants. METHODS AND ANALYSIS: This is a retrospective individually matched case-control study among a cohort of Australian children born from 1997 to 1999, the period of transition from wP to aP vaccines; we include in the cohort children listed on Australia's comprehensive population-based immunisation register as having received a first dose of either pertussis vaccine by 16 weeks old. 500 cohort children diagnosed as having IgE-mediated food allergy at specialist allergy clinics will be included as cases. Controls matched to each case by date and jurisdiction of birth and regional socioeconomic index will be sampled from the immunisation register. Conditional logistic regression will be used to estimate OR (±95% CI) of receipt of wP (vs aP) as the first vaccine dose among cases compared with controls. ETHICS AND DISSEMINATION: The study is approved by all relevant human research ethics committees: Western Australia Child and Adolescent Health Services (2015052EP), Women's and Children's Hospital (HREC/15/WCHN/162), Royal Children's Hospital (35230A) and Sydney Children's Hospital Network (HREC/15/SCHN/405). Outcomes will be disseminated through publication and scientific presentation. TRIAL REGISTRATION NUMBER: NCT02490007.

The NICE-GUT trial protocol: a randomised, placebo controlled trial of oral nitazoxanide for the empiric treatment of acute gastroenteritis among Australian Aboriginal children.

INTRODUCTION: Diarrhoeal disease is the second leading cause of death in children under 5 years globally, killing 525 000 annually. Australian Aboriginal and Torres Strait Islander (hereafter Aboriginal) children suffer a high burden of disease. Randomised trials in other populations suggest nitazoxanide accelerates recovery for children with Giardia, amoebiasis, Cryptosporidium, Rotavirus and Norovirus gastroenteritis, as well as in cases where no enteropathogens are found. METHODS AND ANALYSIS: This double blind, 1:1 randomised, placebo controlled trial is investigating the impact of oral nitazoxanide on acute gastroenteritis in hospitalised Australian Aboriginal children aged 3 months to <5 years. Dosing is based on age-based dosing. The primary endpoint is the time to resolution of 'significant illness' defined as the time from randomisation to the time of clinical assessment as medically ready for discharge, or to the time of actual discharge from hospital, whichever occurs first. Secondary endpoints include duration of hospitalisation, symptom severity during the period of significant illness and following treatment, duration of rehydration and drug safety. Patients will be followed for medically significant events for 60 days. Analysis is based on Bayesian inference. Subgroup analysis will occur by pathogen type (bacteria, virus or parasite), rotavirus vaccination status, age and illness severity. ETHICS AND DISSEMINATION: Ethics approval has been granted by the Central Australian Human Research Ethics Committee (HREC-14-221) and the Human Research Ethics Committee of the Northern Territory Department of Health and Menzies School of Health Research (HREC2014-2172). Study investigators will ensure that the trial is conducted in accordance with the principles of the Declaration of Helsinki. Individual participant consent will be obtained. Results will be disseminated via peer-reviewed publication. TRIAL REGISTRATION NUMBER: ACTRN12614000381684.

Assessment and Translation of the Antibody-in-Lymphocyte Supernatant (ALS) Assay to Improve the Diagnosis of Enteric Fever in Two Controlled Human Infection Models and an Endemic Area of Nepal.

New diagnostic tests for enteric fever are urgently needed to assist with timely antimicrobial treatment of patients and to measure the efficacy of prevention measures such as vaccination. In a novel translational approach, here we use two recently developed controlled human infection models (CHIM) of enteric fever to evaluate an antibody-in-lymphocyte supernatant (ALS) assay, which can detect recent IgA antibody production by circulating B cells in ex vivo mononuclear cell culture. We calculated the discriminative ability of the ALS assay to distinguish diagnosed cases in the two CHIM studies in Oxford, prior to evaluating blood culture-confirmed diagnoses of patients presenting with fever to hospital in an endemic areas of Kathmandu, Nepal. Antibody responses to membrane preparations and lipopolysaccharide provided good sensitivity (>90%) for diagnosing systemic infection after oral challenge with Salmonella Typhi or S. Paratyphi A. Assay specificity was moderate (~60%) due to imperfect sensitivity of blood culture as the reference standard and likely unrecognized subclinical infection. These findings were augmented through the translation of the assay into the endemic setting in Nepal. Anti-MP IgA responses again exhibited good sensitivity (86%) but poor specificity (51%) for detecting blood culture-confirmed enteric fever cases (ROC AUC 0.79, 95%CI 0.70-0.88). Patients with anti-MP IgA ALS titers in the upper quartile exhibited a clinical syndrome synonymous with enteric fever. While better reference standards are need to assess enteric fever diagnostics, routine use of this ALS assay could be used to rule out infection and has the potential to double the laboratory detection rate of enteric fever in this setting over blood culture alone.

Challenge of Humans with Wild-type Salmonella enterica Serovar Typhi Elicits Changes in the Activation and Homing Characteristics of Mucosal-Associated Invariant T Cells.

Gastrointestinal infections by Salmonella enterica serovar Typhi (S. Typhi) are rare in industrialized countries. However, they remain a major public health problem in the developing world with an estimated 26.9 million new cases annually and significant mortality when untreated. Recently, we provided the first direct evidence that CD8+ MAIT cells are activated and have the potential to kill cells exposed to S. Typhi, and that these responses are dependent on bacterial load. However, MAIT cell kinetics and function during bacterial infections in humans remain largely unknown. In this study, we characterize the human CD8+ MAIT cell immune response to S. Typhi infection in subjects participating in a challenge clinical trial who received a low- or high dose of wild-type S. Typhi. We define the kinetics of CD8+ MAIT cells as well as their levels of activation, proliferation, exhaustion/apoptosis, and homing potential. Regardless of the dose, in volunteers resistant to infection (NoTD), the levels of CD8+ MAIT cells after S. Typhi challenge fluctuated around their baseline values (day 0). In contrast, volunteers susceptible to the development of typhoid disease (TD) exhibited a sharp decline in circulating MAIT cells during the development of typhoid fever. Interestingly, MAIT cells from low-dose TD volunteers had higher levels of CD38 coexpressing CCR9, CCR6, and Ki67 during the development of typhoid fever than high-dose TD volunteers. No substantial perturbations on the levels of these markers were observed in NoTD volunteers irrespective of the dose. In sum, we describe, for the first time, that exposure to an enteric bacterium, in this case S. Typhi, results in changes in MAIT cell activation, proliferation, and homing characteristics, suggesting that MAIT cells are an important component of the human host response to bacterial infection.

Importance of Salmonella Typhi-Responsive CD8+ T Cell Immunity in a Human Typhoid Fever Challenge Model.

Typhoid fever, caused by the human-restricted organism Salmonella enterica serovar Typhi (S. Typhi), constitutes a major global health problem. The development of improved attenuated vaccines is pressing, but delayed by the lack of appropriate preclinical models. Herein, we report that high levels of S. Typhi-responsive CD8+ T cells at baseline significantly correlate with an increased risk of disease in humans challenged with a high dose (~104 CFU) wild-type S. Typhi. Typhoid fever development was associated with higher multifunctional S. Typhi-responsive CD8+ T effector memory cells at baseline. Early decreases of these cells in circulation following challenge were observed in both S. Typhi-responsive integrin α4ß7- and integrin α4ß7+ CD8+ T effector memory (TEM) cells, suggesting their potential to home to both mucosal and extra-intestinal sites. Participants with higher baseline levels of S. Typhi-responsive CD8+ T memory cells had a higher risk of acquiring disease, but among those who acquired disease, those with a higher baseline responses took longer to develop disease. In contrast, protection against disease was associated with low or absent S. Typhi-responsive T cells at baseline and no changes in circulation following challenge. These data highlight the importance of pre-existing S. Typhi-responsive immunity in predicting clinical outcome following infection with wild-type S. Typhi and provide novel insights into the complex mechanisms involved in protective immunity to natural infection in a stringent human model with a high challenge dose. They also contribute important information on the immunological responses to be assessed in the appraisal and selection of new generation typhoid vaccines.

Evaluation of the Clinical and Microbiological Response to Salmonella Paratyphi A Infection in the First Paratyphoid Human Challenge Model.

Background: To expedite the evaluation of vaccines against paratyphoid fever, we aimed to develop the first human challenge model of Salmonella enterica serovar Paratyphi A infection. Methods: Two groups of 20 participants underwent oral challenge with S. Paratyphi A following sodium bicarbonate pretreatment at 1 of 2 dose levels (group 1: 1-5 × 103 colony-forming units [CFU] and group 2: 0.5-1 × 103 CFU). Participants were monitored in an outpatient setting with daily clinical review and collection of blood and stool cultures. Antibiotic treatment was started when prespecified diagnostic criteria were met (temperature ≥38°C for ≥12 hours and/or bacteremia) or at day 14 postchallenge. Results: The primary study objective was achieved following challenge with 1-5 × 103 CFU (group 1), which resulted in an attack rate of 12 of 20 (60%). Compared with typhoid challenge, paratyphoid was notable for high rates of subclinical bacteremia (at this dose, 11/20 [55%]). Despite limited symptoms, bacteremia persisted for up to 96 hours after antibiotic treatment (median duration of bacteremia, 53 hours [interquartile range, 24-85 hours]). Shedding of S. Paratyphi A in stool typically preceded onset of bacteremia. Conclusions: Challenge with S. Paratyphi A at a dose of 1-5 × 103 CFU was well tolerated and associated with an acceptable safety profile. The frequency and persistence of bacteremia in the absence of clinical symptoms was notable, and markedly different from that seen in previous typhoid challenge studies. We conclude that the paratyphoid challenge model is suitable for the assessment of vaccine efficacy using endpoints that include bacteremia and/or symptomatology. Clinical Trials Registration: NCT02100397.