Persistent Gastric Colonization with Burkholderia pseudomallei and Dissemination from the Gastrointestinal Tract following Mucosal Inoculation of Mice

Melioidosis is a disease of humans caused by opportunistic infection with the soil and water bacterium Burkholderia pseudomallei. Melioidosis can manifest as an acute, overwhelming infection or as a chronic, recurrent infection. At present, it is not clear where B. pseudomallei resides in the mammalian host during the chronic, recurrent phase of infection. To address this question, we developed a mouse low-dose mucosal challenge model of chronic B. pseudomallei infection and investigated sites of bacterial persistence over 60 days. Sensitive culture techniques and selective media were used to quantitate bacterial burden in major organs, including the gastrointestinal (GI) tract. We found that the GI tract was the primary site of bacterial persistence during the chronic infection phase, and was the only site from which the organism could be consistently cultured during a 60-day infection period. The organism could be repeatedly recovered from all levels of the GI tract, and chronic infection was accompanied by sustained low-level fecal shedding. The stomach was identified as the primary site of GI colonization as determined by fluorescent in situ hybridization. Organisms in the stomach were associated with the gastric mucosal surface, and the propensity to colonize the gastric mucosa was observed with 4 different B. pseudomallei isolates. In contrast, B. pseudomallei organisms were present at low numbers within luminal contents in the small and large intestine and cecum relative to the stomach. Notably, inflammatory lesions were not detected in any GI tissue examined in chronically-infected mice. Only low-dose oral or intranasal inoculation led to GI colonization and development of chronic infection of the spleen and liver. Thus, we concluded that in a mouse model of melioidosis B. pseudomallei preferentially colonizes the stomach following oral inoculation, and that the chronically colonized GI tract likely serves as a reservoir for dissemination of infection to extra-intestinal sites

Seroprevalence and prevention of hepatitis B, measles and rubella among healthcare workers in Dili, Timor-Leste

Introduction: The World Health Organisation recommends that healthcare workers (HCWs) are immune to measles and rubella, and those at risk of exposure are offered the hepatitis B vaccine. No formal programme for occupational assessment and provision of vaccinations to HCWs currently exists in Timor-Leste. Methods: A cross-sectional study was undertaken to determine the seroprevalence of hepatitis B, measles and rubella among HCWs in Dili, Timor-Leste. All patient-facing employees at three healthcare institutions during April–June 2021 were invited to participate. Epidemiological data were collected by interview-questionnaire and a serum sample was collected by phlebotomy and analysed at the National Health Laboratory. Participants were contacted to discuss their results. Relevant vaccines were offered to seronegative individuals and those with active hepatitis B infection were referred for further assessment and management in a hepatology clinic as per national guidelines. Results: Three-hundred-and-twenty-four HCWs were included (representing 51.3% of all eligible HCWs working at the three participating institutions). Sixteen (4.9%; 95% CI: 2.8–7.9%) had active hepatitis B infection, 121 (37.3%; 95% CI: 32.1–42.9%) had evidence of previous (cleared) hepatitis B infection, 134 (41.4%; 95% CI: 35.9–46.9%) were hepatitis B seronegative, and 53 (16.4%; 95% CI: 12.5–20.8%) had been vaccinated. Two-hundred-and-sixty-seven (82.4%; 95% CI: 77.8–86.4%) and 306 (94.4%; 95% CI: 91.4–96.7%) individuals exhibited antibodies to measles and rubella, respectively. Interpretation: There are significant immunity gaps and a high prevalence of hepatitis B infection among HCWs in Dili Municipality, Timor-Leste. Routine occupational assessment and targeted vaccination of this group would be beneficial and should include all types of HCWs. This study provided an opportunity to develop a programme for the occupational assessment and vaccination of HCWs and forms the template for a national guideline. Funding: This work was supported by the Department of Foreign Affairs and Trade, Australian Government [Complex Grant Agreement Number 75889].Full Tex

A population-representative serosurvey estimating vaccine-induced immunity against measles, rubella, hepatitis B and severe acute respiratory syndrome coronavirus 2 in Timor-Leste

Background Serosurveillance can be used to assess population immunity to vaccine-preventable diseases (VPDs). This study aimed to determine seroprevalence of four VPDs across Timor-Leste and identify immunity gaps. Methods A population-representative three-stage cluster random sample of census-enumerated households were visited between October 2021 and February 2023. Occupants aged above one year were tested for measles immunoglobulin G (IgG), rubella IgG, hepatitis B surface antibody (HBsAb), hepatitis B core antibody (HBcAb) and severe acute respiratory syndrome 2 (SARS-CoV-2) IgG, using serological assays with a priori determined cut-offs. Sample- and response-weighted mixed effects logistic regression models were used to estimate seroprevalence in relevant age-strata. Findings Of 2613 eligible households, 1908 (73.0%) participated. Of 8427 occupants, 4750 (56.4%) participated. Measles IgG seroprevalence was low among children, particularly those aged 10–14 (33.2%, 95% confidence interval (CI) 27.8–38.6%). Rubella IgG seroprevalence was high in all ages (93.2%, 95% CI 92.2–94.2%). SARS-CoV-2 IgG seroprevalence was high, including in young children not eligible for vaccination (74.0%, 95% CI 70.4–77.6%). HBsAb seroprevalence was lowest among adolescents aged 15–19 (12.1%, 95% CI 6.8–17.5%) but higher among younger children, who also had low HBcAb seroprevalence. Interpretation The pattern of measles immunity is consistent with low virus transmission and suboptimal childhood vaccine uptake. These data have informed supplementary immunisation activities. High rubella IgG seroprevalence suggests recent or ongoing virus transmission and a need for congenital rubella syndrome surveillance. Hepatitis B data provide evidence of recent improvements in vaccine-induced immunity and protection. This study demonstrates how serosurveillance can directly influence national vaccine strategies. Funding This study was funded by the Department for Foreign Affairs and Trade, Australian Government (Complex Grant Agreement Number 75889).Full Tex

Vaccine Preventable Disease Seroprevalence in a Nationwide Assessment of Timor-Leste (VASINA-TL): Study protocol for a population-representative cross-sectional serosurvey

Introduction Historic disruption in health infrastructure combined with data from a recent vaccine coverage survey suggests there are likely significant immunity gaps to vaccine preventable diseases and high risk of outbreaks in Timor-Leste. Community-based serological surveillance is an important tool to augment understanding of population-level immunity achieved through vaccine coverage and/or derived from prior infection. Methods and analysis This national population-representative serosurvey will take a three-stage cluster sample and aims to include 5600 individuals above 1 year of age. Serum samples will be collected by phlebotomy and analysed for measles IgG, rubella IgG, SARS-CoV-2 antispike protein IgG, hepatitis B surface antibody and hepatitis B core antigen using commercially available chemiluminescent immunoassays or ELISA. In addition to crude prevalence estimates and to account for differences in Timor-Leste's age structure, stratified age-standardised prevalence estimates will be calculated, using Asia in 2013 as the standard population. Additionally, this survey will derive a national asset of serum and dried blood spot samples which can be used for further investigation of infectious disease seroepidemiology and/or validation of existing and novel serological assays for infectious diseases. Ethics and dissemination Ethical approval has been obtained from the Research Ethics and Technical Committee of the Instituto Nacional da Saúde, Timor-Leste and the Human Research Ethics Committee of the Northern Territory Department of Health and Menzies School of Health Research, Australia. Co-designing this study with Timor-Leste's Ministry-of-Health and other relevant partner organisations will allow immediate translation of findings into public health policy, which may include changes to routine immunisation service delivery and/or plans for supplementary immunisation activities.Full Tex

Single-View Echocardiography by Nonexpert Practitioners to Detect Rheumatic Heart Disease A Prospective Study of Diagnostic Accuracy

BACKGROUND: Echocardiographic screening can detect asymptomatic cases of rheumatic heart disease (RHD), facilitating access to treatment. Barriers to implementation of echocardiographic screening include the requirement for expensive equipment and expert practitioners. We aimed to evaluate the diagnostic accuracy of an abbreviated echocardiographic screening protocol (single parasternal-long-axis view with a sweep of the heart) performed by briefly trained, nonexpert practitioners using handheld ultrasound devices. METHODS: Participants aged 5 to 20 years in Timor-Leste and the Northern Territory of Australia had 2 echocardiograms: one performed by an expert echocardiographer using a GE Vivid I or Vivid Q portable ultrasound device (reference test), and one performed by a nonexpert practitioner using a GE Vscan handheld ultrasound device (index test). The accuracy of the index test, compared with the reference test, for identifying cases with definite or borderline RHD was determined. RESULTS: There were 3111 enrolled participants; 2573 had both an index test and reference test. Median age was 12 years (interquartile range, 10-15); 58.2% were female. Proportion with definite or borderline RHD was 5.52% (95% CI, 4.70-6.47); proportion with definite RHD was 3.23% (95% CI, 2.61-3.98). Compared with the reference test, sensitivity of the index test for definite or borderline RHD was 70.4% (95% CI, 62.2-77.8), specificity was 78.1% (95% CI, 76.4-79.8). CONCLUSIONS: Nonexpert practitioners can be trained to perform single parasternal-long-axis view with a sweep of the heart echocardiography. However, the specificity and sensitivity are inadequate for echocardiographic screening. Improved training for nonexpert practitioners should be investigated