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Trachoma, caused by ocular Chlamydia trachomatis infection, is targeted for global elimination as a public health problem by 2030. To provide evidence for use of antibodies to monitor C. trachomatis transmission, we collated IgG responses to Pgp3 antigen, PCR positivity, and clinical observations from 19,811 children aged 1-9 years in 14 populations. We demonstrate that age-seroprevalence curves consistently shift along a gradient of transmission intensity: rising steeply in populations with high levels of infection and active trachoma and becoming flat in populations near elimination. Seroprevalence (range: 0-54%) and seroconversion rates (range: 0-15 per 100 person-years) correlate with PCR prevalence (r: 0.87, 95% CI: 0.57, 0.97). A seroprevalence threshold of 13.5% (seroconversion rate 2.75 per 100 person-years) identifies clusters with any PCR-identified infection at high sensitivity ( >90%) and moderate specificity (69-75%). Antibody responses in young children provide a robust, generalizable approach to monitor population progress toward and beyond trachoma elimination.

Trachoma is an infectious disease characterized by repeated exposures to Chlamydia trachomatis (Ct) that may ultimately lead to blindness. Efficient identification of communities with high infection burden could help target more intensive control efforts. We hypothesized that IgG seroprevalence in combination with geospatial layers, machine learning, and model-based geostatistics would be able to accurately predict future community-level ocular Ct infections detected by PCR. We used measurements from 40 communities in the hyperendemic Amhara region of Ethiopia to assess this hypothesis. Median Ct infection prevalence among children 0-5 years old increased from 6% at enrollment, in the context of recent mass drug administration (MDA), to 29% by month 36, following three years without MDA. At baseline, correlation between seroprevalence and Ct infection was stronger among children 0-5 years old (ρ = 0.77) than children 6-9 years old (ρ = 0.48), and stronger than the correlation between active trachoma and Ct infection (0-5y ρ = 0.56; 6-9y ρ = 0.40). Seroprevalence was the strongest concurrent predictor of infection prevalence at month 36 among children 0-5 years old (cross-validated R2 = 0.75, 95% CI: 0.58-0.85), though predictive performance declined substantially with increasing temporal lag between predictor and outcome measurements. Geospatial variables, a spatial Gaussian process, and stacked ensemble machine learning did not meaningfully improve predictions. Serological markers among children 0-5 years old may be an objective tool for identifying communities with high levels of ocular Ct infections, but accurate, future prediction in the context of changing transmission remains an open challenge.

The MORDOR trial in Niger, Malawi, and Tanzania found that biannual mass distribution of azithromycin to children younger than 5 years led to a 13.5% reduction in all-cause mortality (NCT02048007). To help elucidate the mechanism for mortality reduction, we report IgG responses to 11 malaria, bacterial, and protozoan pathogens using a multiplex bead assay in pre-specified substudy of 30 communities in the rural Niger placebo-controlled trial over a three-year period (n = 5642 blood specimens, n = 3814 children ages 1-59 months). Mass azithromycin reduces Campylobacter spp. force of infection by 29% (hazard ratio = 0.71, 95% CI: 0.56, 0.89; P = 0.004) but serological measures show no significant differences between groups for other pathogens against a backdrop of high transmission. Results align with a recent microbiome study in the communities. Given significant sequelae of Campylobacter infection among preschool aged children, our results support an important mechanism through which biannual mass distribution of azithromycin likely reduces mortality in Niger.

BackgroundWHO promotes the SAFE strategy for the elimination of trachoma as a public health programme, which promotes surgery for trichiasis (ie, the S component), antibiotics to clear the ocular strains of chlamydia that cause trachoma (the A component), facial cleanliness to prevent transmission of secretions (the F component), and environmental improvements to provide water for washing and sanitation facilities (the E component). However, little evidence is available from randomised trials to support the efficacy of interventions targeting the F and E components of the strategy. We aimed to determine whether an integrated water, sanitation, and hygiene (WASH) intervention prevents the transmission of trachoma.MethodsThe WASH Upgrades for Health in Amhara (WUHA) was a two-arm, parallel-group, cluster-randomised trial in 40 rural communities in Wag Hemra Zone (Amhara Region, Ethiopia) that had been treated with 7 years of annual mass azithromycin distributions. The randomisation unit was the school catchment area. All households within a 1·5 km radius of a potential water point within the catchment area (as determined by the investigators) were eligible for inclusion. Clusters were randomly assigned (at a 1:1 ratio) to receive a WASH intervention either immediately (intervention) or delayed until the conclusion of the trial (control), in the absence of concurrent antibiotic distributions. Given the nature of the intervention, participants and field workers could not be masked, but laboratory personnel were masked to treatment allocation. The WASH intervention consisted of both hygiene infrastructure improvements (namely, construction of a community water point) and hygiene promotion by government, school, and community leaders, which were implemented at the household, school, and community levels. Hygiene promotion focused on two simple messages: to use soap and water to wash your or your child's face, and to always use a latrine for defecation. The primary outcome was the cluster-level prevalence of ocular chlamydia, measured annually using conjunctival swabs in a random sample of children aged 0-5 years from each cluster at 12, 24, and 36 month timepoints. Analyses were done in an intention-to-treat manner. This trial is ongoing and is registered at ClinicalTrials.gov, NCT02754583.FindingsBetween Nov 9, 2015, and March 5, 2019, 40 of 44 clusters assessed for eligibility were enrolled and randomly allocated to the trial groups (20 clusters each, with 7636 people from 1751 households in the intervention group and 9821 people from 2211 households in the control group at baseline). At baseline, ocular chlamydia prevalence among children aged 0-5 years was 11% (95% CI 6 to 16) in the WASH group and 11% (5 to 18) in the control group. At month 36, ocular chlamydia prevalence had increased in both groups, to 32% (24 to 41) in the WASH group and 31% (21 to 41) in the control group (risk difference across three annual monitoring visits, after adjustment for prevalence at baseline: 3·7 percentage points; 95% CI -4·9 to 12·4; p=0·40). No adverse events were reported in either group.InterpretationAn integrated WASH intervention addressing the F and E components of the SAFE strategy did not prevent an increase in prevalence of ocular chlamydia following cessation of antibiotics in an area with hyperendemic trachoma. The impact of WASH in the presence of annual mass azithromycin distributions is currently being studied in a follow-up trial of the 40 study clusters. Continued antibiotic distributions will probably be important in areas with persistent trachoma.FundingNational Institutes of Health-National Eye Institute.TranslationFor the Amharic translation of the abstract see Supplementary Materials section.

BackgroundSerological surveys with multiplex bead assays can be used to assess seroprevalence to multiple pathogens simultaneously. However, multiple methods have been used to generate cut-off values for seropositivity and these may lead to inconsistent interpretation of results. A literature review was conducted to describe the methods used to determine cut-off values for data generated by multiplex bead assays.Methodology/principal findingsA search was conducted in PubMed that included articles published from January 2010 to January 2020, and 308 relevant articles were identified that included the terms "serology", "cut-offs", and "multiplex bead assays". After application of exclusion of articles not relevant to neglected tropical diseases (NTD), vaccine preventable diseases (VPD), or malaria, 55 articles were examined based on their relevance to NTD or VPD. The most frequently applied approaches to determine seropositivity included the use of presumed unexposed populations, mixture models, receiver operating curves (ROC), and international standards. Other methods included the use of quantiles, pre-exposed endemic cohorts, and visual inflection points.Conclusions/significanceFor disease control programmes, seropositivity is a practical and easily interpretable health metric but determining appropriate cut-offs for positivity can be challenging. Considerations for optimal cut-off approaches should include factors such as methods recommended by previous research, transmission dynamics, and the immunological backgrounds of the population. In the absence of international standards for estimating seropositivity in a population, the use of consistent methods that align with individual disease epidemiological data will improve comparability between settings and enable the assessment of changes over time.

The Trachoma Control Program in Amhara region, Ethiopia, scaled up the surgery, antibiotics, facial cleanliness, and environmental improvement (SAFE) strategy in all districts starting in 2007. Despite these efforts, many districts still require additional years of SAFE. In 2017, four districts were selected for the assessment of antibody responses against Chlamydia trachomatis antigens and C. trachomatis infection to better understand transmission. Districts with differing endemicity were chosen, whereby one had a previous trachomatous inflammation-follicular (TF) prevalence of ≥ 30% (Andabet), one had a prevalence between 10% and 29.9% (Dera), one had a prevalence between 5% and 10% (Woreta town), and one had a previous TF prevalence of < 5% (Alefa) and had not received antibiotic intervention for 2 years. Survey teams assessed trachoma clinical signs and took conjunctival swabs and dried blood spots (DBS) to measure infection and antibody responses. Trachomatous inflammation-follicular prevalence among children aged 1-9 years was 37.0% (95% CI: 31.1-43.3) for Andabet, 14.7% (95% CI: 10.0-20.5) for Dera, and < 5% for Woreta town and Alefa. Chlamydia trachomatis infection was only detected in Andabet (11.3%). Within these districts, 2,195 children provided DBS. The prevalence of antibody responses to the antigen Pgp3 was 36.9% (95% CI: 29.0-45.6%) for Andabet, 11.3% (95% CI: 5.9-20.6%) for Dera, and < 5% for Woreta town and Alefa. Seroconversion rate for Pgp3 in Andabet was 0.094 (95% CI: 0.069-0.128) events per year. In Andabet district, where SAFE implementation has occurred for 11 years, the antibody data support the finding of persistently high levels of trachoma transmission.

Multiplex bead assays (MBAs) for serologic testing have become more prevalent in public health surveys, but few studies have assessed their test performance. As part of a trachoma study conducted in a rural part of Ethiopia in 2016, dried blood spots (DBS) were collected from a random sample of 393 children aged 0 to 9 years, with at least two separate 6-mm DBS collected on a filter card. Samples eluted from DBS were processed using an MBA on the Luminex platform for antibodies against 13 antigens of nine infectious organisms: Chlamydia trachomatis, Vibrio cholera, enterotoxigenic Escherichia coli, Cryptosporidium parvum, Entamoeba histolytica, Camplyobacter jejuni, Salmonella typhimurium Group B, Salmonella enteritidis Group D, and Giardia lamblia. Two separate DBS from each child were processed. The first DBS was run a single time, with the MBA set to read 100 beads per well. The second DBS was run twice, first at 100 beads per well and then at 50 beads per well. Results were expressed as the median fluorescence intensity minus background (MFI-BG), and classified as seropositive or seronegative according to external standards. Agreement between the three runs was high, with intraclass correlation coefficients of > 0.85 for the two Salmonella antibody responses and > 0.95 for the other 11 antibody responses. Agreement was also high for the dichotomous seropositivity indicators, with Cohen's kappa statistics exceeding 0.87 for each antibody assay. These results suggest that serologic testing on the Luminex platform had strong test performance characteristics for analyzing antibodies using DBS.

BackgroundBiannual mass azithromycin administration to preschool children reduces all-cause mortality, but the mechanism for the effect is not understood. Azithromycin has activity against malaria parasites, and malaria is a leading cause of child mortality in the Sahel. The effect of biannual versus annual azithromycin distribution for trachoma control on serological response to merozoite surface protein 1 (MSP-119), a surrogate for malaria incidence, was evaluated among children in Niger.MethodsMarkers of malaria exposure were measured in two arms of a factorial randomized controlled trial designed to evaluate targeted biannual azithromycin distribution to children under 12 years of age compared to annual azithromycin to the entire community for trachoma control (N = 12 communities per arm). Communities were treated for 36 months (6 versus 3 distributions). Dried blood spots were collected at 36 months among children ages 1-5 years, and MSP-119 antibody levels were assessed using a bead-based multiplex assay to measure malaria seroprevalence.ResultsAntibody results were available for 991 children. MSP-119 seropositivity was 62.7% in the biannual distribution arm compared to 68.7% in the annual arm (prevalence ratio 0.91, 95% CI 0.83 to 1.00). Mean semi-quantitative antibody levels were lower in the biannual distribution arm compared to the annual arm (mean difference - 0.39, 95% CI - 0.05 to - 0.72).ConclusionsTargeted biannual azithromycin distribution was associated with lower malaria seroprevalence compared to that in a population that received annual distribution. Trial Registration Clinicaltrials.gov NCT00792922.

Little is known about enteropathogen seroepidemiology among children in low-resource settings. We measured serological IgG responses to eight enteropathogens (Giardia intestinalis, Cryptosporidium parvum, Entamoeba histolytica, Salmonella enterica, enterotoxigenic Escherichia coli, Vibrio cholerae, Campylobacter jejuni, norovirus) in cohorts from Haiti, Kenya, and Tanzania. We studied antibody dynamics and force of infection across pathogens and cohorts. Enteropathogens shared common seroepidemiologic features that enabled between-pathogen comparisons of transmission. Overall, exposure was intense: for most pathogens the window of primary infection was

BACKGROUND:Program decision-making for trachoma elimination currently relies on conjunctival clinical signs. Antibody tests may provide additional information on the epidemiology of trachoma, particularly in regions where it is disappearing or elimination targets have been met. METHODS:A cluster-randomized trial of mass azithromycin distribution strategies for trachoma elimination was conducted over three years in a mesoendemic region of Niger. Dried blood spots were collected from a random sample of children aged 1-5 years in each of 24 study communities at 36 months after initiation of the intervention. A multiplex bead assay was used to test for antibodies to two Chlamydia trachomatis antigens, Pgp3 and CT694. We compared seropositivity to either antigen to clinical signs of active trachoma (trachomatous inflammation-follicular [TF] and trachomatous inflammation-intense [TI]) at the individual and cluster level, and to ocular chlamydia prevalence at the community level. RESULTS:Of 988 children with antibody data, TF prevalence was 7.8% (95% CI 6.1 to 9.5) and TI prevalence was 1.6% (95% CI 0.9 to 2.6). The overall prevalence of antibody positivity to Pgp3 was 27.2% (95% CI 24.5 to 30), and to CT694 was 23.7% (95% CI 21 to 26.2). Ocular chlamydia infection prevalence was 5.2% (95% CI 2.8 to 7.6). Seropositivity to Pgp3 and/or CT694 was significantly associated with TF at the individual and community level and with ocular chlamydia infection and TI at the community level. Older children were more likely to be seropositive than younger children. CONCLUSION:Seropositivity to Pgp3 and CT694 correlates with clinical signs and ocular chlamydia infection in a mesoendemic region of Niger. TRIAL REGISTRATION:ClinicalTrials.gov NCT00792922.

Robust surveillance methods are needed for trachoma control and recrudescence monitoring, but existing methods have limitations. Here, we analyse data from nine trachoma-endemic populations and provide operational thresholds for interpretation of serological data in low-transmission and post-elimination settings. Analyses with sero-catalytic and antibody acquisition models provide insights into transmission history within each population. To accurately estimate sero-conversion rates (SCR) for trachoma in populations with high-seroprevalence in adults, the model accounts for secondary exposure to Chlamydia trachomatis due to urogenital infection. We estimate the population half-life of sero-reversion for anti-Pgp3 antibodies to be 26 (95% credible interval (CrI): 21-34) years. We show SCRs below 0.015 (95% confidence interval (CI): 0.0-0.049) per year correspond to a prevalence of trachomatous inflammation-follicular below 5%, the current threshold for elimination of active trachoma as a public health problem. As global trachoma prevalence declines, we may need cross-sectional serological survey data to inform programmatic decisions.

The target end date for the global elimination of trachoma as a public health problem is 2020. As countries begin the process for submitting their dossier for the validation of elimination of trachoma as a public health problem, strategies for post-validation surveillance must be considered. Seroprevalence of antibodies against antigens from the causative bacteria Chlamydia trachomatis (Ct) in young children has been shown to reflect trachomatous inflammation-follicular (TF) rates in both endemic and previously endemic settings. However, none of these studies has directly compared age seroprevalence in the same communities before and after mass drug administration (MDA) for trachoma. Here we report a marked shift in age seroprevalence curves in four villages in Kapilvastu District, Nepal, before and after MDA. Clinical examinations were performed and blood was taken before (N = 659) and 5 years after (N = 646) MDA. Rates of TF decreased from 17.6% in ≤ 9-year-olds before MDA (N = 52) to 0% in ≤ 9-year-olds (N = 73) after MDA. Positive antibody responses to Ct in the entire population decreased from 82.1% pre-MDA to 35.8% post-MDA, whereas those among ≤ 9-year-olds decreased from 59.6% to 4.1%. These data show that the postintervention decrease in TF was reflected in a drop in anti-Ct antibody responses, suggesting that antibody responses could be useful indicators for post-validation surveillance.

Monitoring trachoma transmission with antibody data requires characterization of decay in IgG to Chlamydia trachomatis antigens. In a 3-year longitudinal cohort in a high-transmission setting, we estimated a median IgG half-life of 3 years and a seroreversion rate of 2.5 per 100 person-years (95% confidence interval, 1.6–3.5). Clinical Trials Registration NCT02754583. [ABSTRACT FROM AUTHOR]

We describe here a lateral flow-based assay (LFA) for the detection of antibodies against immunodominant antigen Pgp3 from Chlamydia trachomatis, the causative agent of urogenital chlamydia infection and ocular trachoma. Optimal signal detection was achieved when the gold-conjugate and test line contained Pgp3, creating a dual sandwich capture assay. The LFA yielded positive signals with serum and whole blood but not with eluted dried blood spots. For serum, the agreement of the LFA with the non-reference multiplex assay was 96%, the specificity using nonendemic pediatric sera was 100%, and the inter-rater agreement was κ=0.961. For whole blood, the agreement of LFA with multiplex was 81.5%, the specificity was 100%, and the inter-rater agreement was κ=0.940. The LFA was tested in a field environment and yielded similar results to those from laboratory-based testing. These data show the successful development of a lateral flow assay for detection of antibodies against Pgp3 with reliable use in field settings, which would make antibody-based testing for trachoma surveillance highly practical, especially after cessation of trachoma elimination programs.

BackgroundThe WHO seeks to control trachoma as a public health problem in endemic areas. Achham District in western Nepal was found to have TF (trachoma follicular) above 20% in a 2006 government survey, triggering 3 annual mass drug administrations finishing in 2010. Here we assess the level of control that has been achieved using surveillance for clinical disease, ocular chlamydia trachomatis infection, and serology for antibodies against chlamydia trachomatis protein antigens.MethodsWe conducted a cross-sectional survey of children aged 1-9 years in communities in Achham District in early 2014 including clinical examination validated with photographs, conjunctival samples for Chlamydia trachomatis (Amplicor PCR), and serological testing for antibodies against chlamydia trachomatis protein antigens pgp3 and CT694 using the Luminex platform.FindingsIn 24 randomly selected communities, the prevalence of trachoma (TF and/or TI) in 1-9 year olds was 3/1124 (0.3%, 95% CI 0.1 to 0.8%), and the prevalence of ocular chlamydia trachomatis infection was 0/1124 (0%, 95% CI 0 to 0.3%). In 18 communities selected because they had the highest prevalence of trachoma in a previous survey, the prevalence of TF and/or TI was 7/716 (1.0%, 95% CI 0.4 to 2.0%) and the prevalence of ocular chlamydia trachomatis infection was 0/716 (0%, 95% CI 0 to 0.5%). In 3 communities selected for serological testing, the prevalence of trachoma was 0/68 (0%, 95% CI 0 to 5.3%), the prevalence of ocular chlamydia trachomatis infection was 0/68 (0%, 95% CI 0 to 0.5%), the prevalence of antibodies against chlamydia trachomatis protein antigen pgp3 was 1/68 (1.5%, 95% CI 0.04% to 7.9%), and the prevalence of antibodies against chlamydia trachomatis protein antigen CT694 was 0/68 (0%, 95% CI 0 to 5.3%).Conclusion/significanceThis previously highly endemic district in Nepal has little evidence of recent clinical disease, chlamydia trachomatis infection, or serological evidence of trachoma, suggesting that epidemiological control has been achieved.

Background: Trachoma recrudescence after elimination as a public health problem has been reached is a concern for control programs globally. Programs typically conduct district-level trachoma surveillance surveys (TSS) ≥ 2 years after the elimination threshold is achieved to determine whether the prevalence of trachomatous inflammation-follicular (TF) among children ages 1 to 9 years remains <5%. Many TSS are resulting in a TF prevalence ≥5%. Once a district returns to TF ≥5%, a program typically restarts costly mass drug administration (MDA) campaigns and surveys at least twice, for impact and another TSS. In Amhara, Ethiopia, most TSS which result in a TF ≥5% have a prevalence close to 5%, making it difficult to determine whether the result is due to true recrudescence or to statistical variability. This study's aim was to monitor recrudescence within Amhara by waiting to restart MDA within 2 districts with a TF prevalence ≥5% at TSS, Metema = 5.2% and Woreta Town = 5.1%. The districts were resurveyed 1 year later using traditional and alternative indicators, such as measures of infection and serology, a "wait and watch" approach. Methods/Principal findings: These post-surveillance surveys, conducted in 2021, were multi-stage cluster surveys whereby certified graders assessed trachoma signs. Children ages 1 to 9 years provided a dried blood spot and children ages 1 to 5 years provided a conjunctival swab. TF prevalence in Metema and Woreta Town were 3.6% (95% Confidence Interval [CI]:1.4–6.4) and 2.5% (95% CI:0.8–4.5) respectively. Infection prevalence was 1.2% in Woreta Town and 0% in Metema. Seroconversion rates to Pgp3 in Metema and Woreta Town were 0.4 (95% CI:0.2–0.7) seroconversions per 100 child-years and 0.9 (95% CI:0.6–1.5) respectively. Conclusions/Significance: Both study districts had a TF prevalence <5% with low levels of Chlamydia trachomatis infection and transmission, and thus MDA interventions are no longer warranted. The wait and watch approach represents a surveillance strategy which could lead to fewer MDA campaigns and surveys and thus cost savings with reduced antibiotic usage. Author summary: The return of trachoma transmission after elimination as a public health problem has been reached is a concern for control programs globally. Currently, many district-level trachoma surveillance surveys (conducted ≥2 years since elimination threshold has been reached) are resulting in a prevalence above the established threshold. Once a district returns above threshold, a program typically restarts costly mass drug administration campaigns and conducts more surveys. This study's aim was to monitor recrudescence through a "wait and watch" approach within Amhara, Ethiopia. This entailed waiting to restart mass drug administration within 2 districts with trachoma prevalence above but close to the threshold at surveillance survey, then surveying the districts 1 year later using traditional and alternative indicators. These post-surveillance surveys assessed traditional trachoma signs, and additionally, children provided a dried blood spot for serology outcomes and a conjunctival swab for infection. The results of the study demonstrated that both districts had a prevalence below threshold with low levels of infection and serological evidence of transmission, and thus mass drug administration interventions are no longer warranted. The wait and watch approach represents a surveillance strategy that could lead to fewer surveys and mass drug administration campaigns and thus savings in programmatic costs with reduced usage of antibiotics. [ABSTRACT FROM AUTHOR]

Baseline trachoma surveys in Côte d'Ivoire (2019) identified seven evaluation units (EUs) with a trachomatous inflammation–follicular (TF) prevalence ≥10%, but a trachomatous trichiasis (TT) prevalence in individuals ≥15 y of age below the elimination threshold (0.2%). Two of these EUs, Bondoukou 1 and Bangolo 2, were selected for a follow-up survey to understand the epidemiology of trachoma using additional indicators of Chlamydia trachomatis infection (DNA from conjunctival swabs) and exposure (anti-Pgp3 and Ct694 antibodies from dried blood spots [DBSs]). A two-stage cluster sampling methodology was used to select villages and households. All individuals 1–9 y of age from each selected household were recruited, graded for trachoma and had a conjunctival swab and DBS collected. Conjunctival swabs and DBSs were tested using Cepheid GeneXpert and a multiplex bead assay, respectively. The age-adjusted TF and infection prevalence in 1- to 9-year-olds was <1% and <0.3% in both EUs. Age-adjusted seroprevalence was 5.3% (95% confidence interval [CI] 1.5 to 15.6) in Bondoukou 1 and 8.2% (95% CI 4.3 to 13.7) in Bangolo 2. The seroconversion rate for Pgp3 was low, at 1.23 seroconversions/100 children/year (95% CI 0.78 to 1.75) in Bondoukou 1 and 1.91 (95% CI 1.58 to 2.24) in Bangolo 2. Similar results were seen for CT694. These infection, antibody and clinical data provide strong evidence that trachoma is not a public health problem in either EU. [ABSTRACT FROM AUTHOR]

Examined the development of seven infants with "missing milestones" in motor development. Found that three children had normal development, three developed global developmental delay, and one was diagnosed with multiple cavernous haemangiomata in the brain. Suggested that missing milestones can be a benign variation of normal motor development or the first sign of neurodevelopmental disorder. (Author/KB)