A general framework to support cost-efficient fecal egg count methods and study design choices for large-scale STH deworming programs-monitoring of therapeutic drug efficacy as a case study.

BACKGROUND: Soil-transmitted helminth (STH) control programs currently lack evidence-based recommendations for cost-efficient survey designs for monitoring and evaluation. Here, we present a framework to provide evidence-based recommendations, using a case study of therapeutic drug efficacy monitoring based on the examination of helminth eggs in stool. METHODS: We performed an in-depth analysis of the operational costs to process one stool sample for three diagnostic methods (Kato-Katz, Mini-FLOTAC and FECPAKG2). Next, we performed simulations to determine the probability of detecting a truly reduced therapeutic efficacy for different scenarios of STH species (Ascaris lumbricoides, Trichuris trichiura and hookworms), pre-treatment infection levels, survey design (screen and select (SS); screen, select and retest (SSR) and no selection (NS)) and number of subjects enrolled (100-5,000). Finally, we integrated the outcome of the cost assessment into the simulation study to estimate the total survey costs and determined the most cost-efficient survey design. PRINCIPAL FINDINGS: Kato-Katz allowed for both the highest sample throughput and the lowest cost per test, while FECPAKG2 required both the most laboratory time and was the most expensive. Counting of eggs accounted for 23% (FECPAKG2) or ≥80% (Kato-Katz and Mini-FLOTAC) of the total time-to-result. NS survey designs in combination with Kato-Katz were the most cost-efficient to assess therapeutic drug efficacy in all scenarios of STH species and endemicity. CONCLUSIONS/SIGNIFICANCE: We confirm that Kato-Katz is the fecal egg counting method of choice for monitoring therapeutic drug efficacy, but that the survey design currently recommended by WHO (SS) should be updated. Our generic framework, which captures laboratory time and material costs, can be used to further support cost-efficient choices for other important surveys informing STH control programs. In addition, it can be used to explore the value of alternative diagnostic techniques, like automated egg counting, which may further reduce operational costs. TRIAL REGISTRATION: ClinicalTrials.gov NCT03465488.

Potential drivers for schistosomiasis persistence: Population genetic analyses from a cluster-randomized urogenital schistosomiasis elimination trial across the Zanzibar islands.

The World Health Organization's revised NTD Roadmap and the newly launched Guidelines target elimination of schistosomiasis as a public health problem in all endemic areas by 2030. Key to meeting this goal is elucidating how selective pressures imposed by interventions shape parasite populations. Our aim was to identify any differential impact of a unique cluster-randomized tri-armed elimination intervention (biannual mass drug administration (MDA) applied alone or in association with either mollusciciding (snail control) or behavioural change interventions) across two Zanzibarian islands (Pemba and Unguja) on the population genetic composition of Schistosoma haematobium over space and time. Fifteen microsatellite loci were used to analyse individual miracidia collected from infected individuals across islands and intervention arms at the start (2012 baseline: 1,522 miracidia from 176 children; 303 from 43 adults; age-range 6-75, mean 12.7 years) and at year 5 (2016: 1,486 miracidia from 146 children; 214 from 25 adults; age-range 9-46, mean 12.4 years). Measures of genetic diversity included allelic richness (Ar), Expected (He) and Observed heterozygosity (Ho), inbreeding coefficient (FST), parentage analysis, estimated worm burden, worm fecundity, and genetic sub-structuring. There was little evidence of differential selective pressures on population genetic diversity, inbreeding or estimated worm burdens by treatment arm, with only the MDA+snail control arm within Unguja showing trends towards reduced diversity and altered inbreeding over time. The greatest differences overall, both in terms of parasite fecundity and genetic sub-structuring, were observed between the islands, consistent with Pemba's persistently higher mean infection intensities compared to neighbouring Unguja, and within islands in terms of infection hotspots (across three definitions). These findings highlight the important contribution of population genetic analyses to elucidate extensive genetic diversity and biological drivers, including potential gene-environmental factors, that may override short term selective pressures imposed by differential disease control strategies. Trial Registration: ClinicalTrials.gov ISRCTN48837681.

Population genomics of ancient and modern Trichuris trichiura.

The neglected tropical disease trichuriasis is caused by the whipworm Trichuris trichiura, a soil-transmitted helminth that has infected humans for millennia. Today, T. trichiura infects as many as 500 million people, predominantly in communities with poor sanitary infrastructure enabling sustained faecal-oral transmission. Using whole-genome sequencing of geographically distributed worms collected from human and other primate hosts, together with ancient samples preserved in archaeologically-defined latrines and deposits dated up to one thousand years old, we present the first population genomics study of T. trichiura. We describe the continent-scale genetic structure between whipworms infecting humans and baboons relative to those infecting other primates. Admixture and population demographic analyses support a stepwise distribution of genetic variation that is highest in Uganda, consistent with an African origin and subsequent translocation with human migration. Finally, genome-wide analyses between human samples and between human and non-human primate samples reveal local regions of genetic differentiation between geographically distinct populations. These data provide insight into zoonotic reservoirs of human-infective T. trichiura and will support future efforts toward the implementation of genomic epidemiology of this globally important helminth.

Prediction of gestational age using urinary metabolites in term and preterm pregnancies.

Assessment of gestational age (GA) is key to provide optimal care during pregnancy. However, its accurate determination remains challenging in low- and middle-income countries, where access to obstetric ultrasound is limited. Hence, there is an urgent need to develop clinical approaches that allow accurate and inexpensive estimations of GA. We investigated the ability of urinary metabolites to predict GA at time of collection in a diverse multi-site cohort of healthy and pathological pregnancies (n = 99) using a broad-spectrum liquid chromatography coupled with mass spectrometry (LC-MS) platform. Our approach detected a myriad of steroid hormones and their derivatives including estrogens, progesterones, corticosteroids, and androgens which were associated with pregnancy progression. We developed a restricted model that predicted GA with high accuracy using three metabolites (rho = 0.87, RMSE = 1.58 weeks) that was validated in an independent cohort (n = 20). The predictions were more robust in pregnancies that went to term in comparison to pregnancies that ended prematurely. Overall, we demonstrated the feasibility of implementing urine metabolomics analysis in large-scale multi-site studies and report a predictive model of GA with a potential clinical value.

RNA profiles reveal signatures of future health and disease in pregnancy.

Maternal morbidity and mortality continue to rise, and pre-eclampsia is a major driver of this burden1. Yet the ability to assess underlying pathophysiology before clinical presentation to enable identification of pregnancies at risk remains elusive. Here we demonstrate the ability of plasma cell-free RNA (cfRNA) to reveal patterns of normal pregnancy progression and determine the risk of developing pre-eclampsia months before clinical presentation. Our results centre on comprehensive transcriptome data from eight independent prospectively collected cohorts comprising 1,840 racially diverse pregnancies and retrospective analysis of 2,539 banked plasma samples. The pre-eclampsia data include 524 samples (72 cases and 452 non-cases) from two diverse independent cohorts collected 14.5 weeks (s.d., 4.5 weeks) before delivery. We show that cfRNA signatures from a single blood draw can track pregnancy progression at the placental, maternal and fetal levels and can robustly predict pre-eclampsia, with a sensitivity of 75% and a positive predictive value of 32.3% (s.d., 3%), which is superior to the state-of-the-art method2. cfRNA signatures of normal pregnancy progression and pre-eclampsia are independent of clinical factors, such as maternal age, body mass index and race, which cumulatively account for less than 1% of model variance. Further, the cfRNA signature for pre-eclampsia contains gene features linked to biological processes implicated in the underlying pathophysiology of pre-eclampsia.

Efficacy and safety of co-administered ivermectin and albendazole in school-aged children and adults infected with Trichuris trichiura in Côte d'Ivoire, Laos, and Pemba Island, Tanzania: a double-blind, parallel-group, phase 3, randomised controlled trial.

BACKGROUND: Preventive chemotherapy with albendazole or mebendazole remains one of the cornerstones of soil-transmitted helminth control. However, these drugs are less effective against Trichuris trichiura. Combined ivermectin-albendazole is a promising treatment alternative, yet robust evidence is lacking. We aimed to demonstrate superiority of co-administered ivermectin-albendazole over albendazole monotherapy in three distinct epidemiological settings. METHODS: We conducted a double-blind, parallel-group, phase 3, randomised controlled trial in community members aged 6-60 years infected with T trichiura in Côte d'Ivoire, Laos, and Pemba Island, Tanzania, between Sept 26, 2018, and June 29, 2020. Participants with at least 100 T trichiura eggs per g of stool at baseline were randomly assigned (1:1) using computer-generated randomisation sequences in varying blocks of four, six, and eight, stratified by baseline T trichiura infection intensity, to orally receive either a single dose of ivermectin (200 µg/kg) plus albendazole (400 mg) or albendazole (400 mg) plus placebo. Patients, field staff, and outcome assessors were masked to treatment assignment. The primary outcome was cure rate against T trichiura, defined as the proportion of participants with no eggs in their faeces 14-21 days after treatment, assessed by Kato-Katz thick smears, and analysed in the available-case population according to intention-to-treat principles. Safety was a secondary outcome and was assessed 3 h and 24 h after drug administration. The trial is registered at ClinicalTrials.gov, NCT03527732. FINDINGS: Between Sept 13 and Dec 18, 2019, Jan 12 and April 5, 2019, and Sept 26 and Nov 5, 2018, 3737, 3694, and 1435 community members were screened for trial eligibility in Côte d'Ivoire, Laos, and Pemba Island, respectively. In Côte d'Ivoire, Laos, and Pemba Island, 256, 274, and 305 participants, respectively, were randomly assigned to the albendazole group, and 255, 275, and 308, respectively, to the ivermectin-albendazole group. Primary outcome data were available for 722 participants treated with albendazole and 733 treated with ivermectin-albendazole. Ivermectin-albendazole showed significantly higher cure rates against T trichiura than albendazole in Laos (66% [140 of 213]vs 8% [16 of 194]; difference 58 percentage points, 95% CI 50 to 65, p<0·0001) and Pemba Island (49% [140 of 288]vs 6% [18 of 293], 43 percentage points, 36 to 49, p<0·0001) but had similar efficacy in Côte d'Ivoire (14% [32 of 232]vs 10% [24 of 235], 4 percentage points, -2 to 10, p=0·24). No serious adverse events were reported; observed events were mostly classified as mild (95% [266 of 279] in the albendazole group and 91% [288 of 317] in the ivermectin-albendazole group), and all were transient in nature. INTERPRETATION: Treatment with ivermectin-albendazole resulted in higher efficacy against trichuriasis than albendazole alone in Laos and Pemba Island but not in Côte d'Ivoire. We recommend implementation of this combination therapy for soil-transmitted helminth control in countries with high T trichiura prevalence and proven enhanced efficacy of this treatment, particularly where ivermectin is beneficial against other endemic helminthiases. FUNDING: Bill & Melinda Gates Foundation.

Individual responses to a single oral dose of albendazole indicate reduced efficacy against soil-transmitted helminths in an area with high drug pressure.

BACKGROUND: Albendazole (ALB) is administered annually to millions of children through global deworming programs targeting soil-transmitted helminths (STHs: Ascaris lumbricoides, Trichuris trichiura and hookworms, Necator americanus and Ancylostoma duodenale). However, due to the lack of large individual patient datasets collected using standardized protocols and the application of population-based statistical methods, little is known about factors that may affect individual responses to treatment. METHODOLOGY/PRINCIPAL FINDINGS: We re-analyzed 645 individual patient data from three standardized clinical trials designed to assess the efficacy of a single 400 mg oral dose of ALB against STHs in schoolchildren from different study sites, each with varying history of drug pressure based on duration of mass drug administration programs: Ethiopia, low; Lao People's Democratic Republic (PDR), moderate; Pemba Island (Tanzania), high. Using a Bayesian statistical modelling approach to estimate individual responses (individual egg reduction rates, ERRi), we found that efficacy was lower in Pemba Island, particularly for T. trichiura. For this STH, the proportion of participants with a satisfactory response (ERRi ≥50%), was 65% in Ethiopia, 61% in Lao PDR but only 29% in Pemba Island. There was a significant correlation between ERRi and infection intensity prior to drug administration (ERRi decreasing as a function of increasing infection intensity). Individual age and sex also affected the drug response, but these were of negligible clinical significance and not consistent across STHs and study sites. CONCLUSIONS/SIGNIFICANCE: We found decreased efficacy of ALB against all the STHs analyzed in Pemba Island (Tanzania), an area with high drug pressure. This does not indicate causality, as this association may also be partially explained by differences in infection intensity prior to drug administration. Notwithstanding, our results indicate that without alternative treatment regimens, program targets will not be achievable on Pemba Island because of inadequate efficacy of ALB. TRIAL REGISTRATION: The study was registered on Clinicaltrials.gov (ID: NCT03465488) on March 7, 2018.

Long-term outcomes of ivermectin-albendazole versus albendazole alone against soil-transmitted helminths: Results from randomized controlled trials in Lao PDR and Pemba Island, Tanzania.

BACKGROUND: Preventive chemotherapy is the cornerstone of soil-transmitted helminth (STH) control. Long-term outcomes and adequate treatment frequency of the recently recommended albendazole-ivermectin have not been studied to date. METHODOLOGY/PRINCIPAL FINDINGS: Double-blind randomized controlled trials were conducted in Lao PDR, Pemba Island, Tanzania and Côte d'Ivoire between 2018 and 2020 to evaluate the efficacy and safety of ivermectin-albendazole versus albendazole-placebo in Trichuris trichiura-infected individuals aged 6 to 60. In the framework of this study, in Lao PDR 466 and 413 participants and on Pemba Island, 558 and 515 participants were followed-up six and 12 months post-treatment, respectively. From each participant at least one stool sample was processed for Kato-Katz diagnosis and cure rates (CRs), egg reduction rates (ERRs) and apparent reinfection rates were calculated. If found helminth-positive at six months, participants were re-treated according to their allocated treatment. Long-term outcomes against T. trichiura based on CRs and ERRs of ivermectin-albendazole compared to albendazole were significantly higher at six months in Lao PDR (CR, 65.8 vs 13.4%, difference; 52.4; 95% CI 45.0-60.0; ERRs, 99.0 vs 79.6, difference 19.4; 95% CI 14.4-24.4) and Pemba Island (CR, 17.8 vs 1.4%, difference; 16.4; 95% CI 11.6-21.0; ERRs, 84.9 vs 21.2, difference 63.8; 95% CI 50.6-76.9) and also at 12 months in Lao PDR (CR, 74.0 vs 23.4%, difference; 50.6; 95% CI 42.6-61.0; ERRs, 99.6 vs 91.3, difference 8.3; 95% CI 5.7-10.8) and Pemba Island (CR, 19.5 vs 3.4%, difference; 16.1; 95% CI 10.7-21.5; ERRs, 92.9 vs 53.6, difference 39.3; 95% CI 31.2-47.4) respectively. Apparent reinfection rates with T. trichiura were considerably higher on Pemba Island (100.0%, 95% CI, 29.2-100.0) than in Lao PDR (10.0%, 95% CI, 0.2-44.5) at 12 months post-treatment for participants treated with albendazole alone. CONCLUSIONS/SIGNIFICANCE: The long-term outcomes against T. trichiura of ivermectin-albendazole are superior to albendazole in terms of CRs and ERRs and in reducing infection intensities. Our results will help to guide decisions on how to best use ivermectin-albendazole in the context of large-scale PC programs tailored to the local context to sustainably control STH infections. TRIAL REGISTRATION: ClinicalTrials.gov registered with clinicaltrials.gov, reference: NCT03527732, date assigned: 17 May 2018.

Evaluation of two communication tools, slideshow and theater, to improve participants' understanding of a clinical trial in the informed consent procedure on Pemba Island, Tanzania.

BACKGROUND: Clinical trial participants are required to sign an informed consent form (ICF). However, information is lacking on the most effective methods to convey trial relevant information prior to inviting participants to sign the ICF, being particularly pertinent in low-income countries. A previous study on Pemba Island, Tanzania, found that a verbal information session (IS) was significantly better than providing an ICF alone. However, knowledge gaps remained. Building on these findings, we investigated the effect of adding a slideshow or a theater to the IS in the informed consent procedure of an anthelminthic clinical trial. METHODOLOGY/PRINCIPAL FINDINGS: A total of 604 caregivers were randomized into the control group that only received an ICF (n = 150) or an ICF plus one of three intervention strategies: (i) verbal IS (n = 135), (ii) verbal IS with a slideshow (n = 174) or (iii) verbal IS followed by a theater (n = 145). All modes of information covered the same key messages. Participants' understanding was assessed using a semi-structured questionnaire. The mean score of caregivers in the control group (ICF only) was 4.41 (standard deviation = 1.47). Caregivers attending the IS alone were more knowledgeable than those in the control group (estimated difference in mean scores: 2.40, 95% confidence interval (CI) 1.95 to 2.86, p < 0.01). However, there was no evidence of an improvement compared to the IS only when participants attended a slideshow (0.09, 95% CI -0.53 to 0.35, p = 0.68) or a theater (0.28, 95% CI -0.27 to 0.82, p = 0.32). Three out of 10 key messages remained largely misunderstood, regardless of the mode of information group. CONCLUSIONS/SIGNIFICANCE: Our study confirmed that, in this setting, an ICF alone was not sufficient to convey clinical trial-related information. An IS was beneficial, however, additional theater and slideshows did not further improve understanding. Future research should explore methods to improve communication between study teams and participants for different key messages, study types and settings.

Assessment of fecal calprotectin and fecal occult blood as point-of-care markers for soil-transmitted helminth attributable intestinal morbidity in a case-control substudy conducted in Côte d'Ivoire, Lao PDR and Pemba Island, Tanzania.

BACKGROUND: Infections with soil-transmitted helminths (STHs) may result in chronic inflammatory disorders affecting the human host. The objective of this study was to evaluate Fecal Calprotectin (FC) and Fecal Occult Blood (FOB) in individuals infected and non-infected with STHs to identify potential intestinal morbidity markers. METHODS: Stool from participants diagnosed positive for Trichuris trichiura and concomitant STH infections from three countries was used to perform FC and FOB point-of-care assays. Simultaneously, identified STH negative participants underwent FC and FOB testing as controls. Potential associations between test results and determinants were analyzed using multivariable logistic regression. FINDINGS: In total, 1034 T. trichiura infected cases (mostly light infections) and 157 STH negative controls were tested for FC and FOB. Among all participants tested, 18·5% had ≥ 50 µg/g FC concentration, while 14 (1·2%) were positive for FOB. No statistically significant association was found between T. trichiura infection or Ascaris lumbricoides co-infection and FC concentration, while an inverse association (odds ratio (OR): 0·45, 95% credible intervals (CrI): 0·26, 0·75) was found between hookworm co-infection and FC concentration. In Lao PDR, the proportion of participants in the ≥ 50 µg/g FC category was significantly higher in the oldest age category compared to the 5-11 years group (OR: 3·31, 95% CrI: 1·62, 7·24). Too few participants were found positive for FOB to derive any conclusions. INTERPRETATION: Studies are needed to better understand the relationship between intestinal morbidity and STH infections. Suitable, standardized, low-cost markers of STH attributable morbidity to better monitor the impact of STH control interventions are necessary. FUNDING: BMGF (OPP1153928).

Insights gained from conducting a randomised controlled trial on Ivermectin-Albendazole against Trichuris trichiura in Côte d'Ivoire, Lao PDR and Pemba Island.

There is only limited scientific literature on trial methodology, trial procedures and mitigation strategies to overcome challenges faced during clinical research taking place in resource constrained healthcare environments. Organisational, cultural, infrastructural and ethical challenges may vary between settings although conduct of clinical trials for the same disease (in our case soil-transmitted helminth (STH) infections) share similar risks for implementation. We use the example of a phase III randomised controlled trial, conducted between 2018 and 2020 in Côte d'Ivoire, Lao PDR and Pemba Island (Tanzania), to share challenges faced and mitigation strategies to guide future planning of studies in similar settings. We describe the planning, screening, enrolment and implementation phases in each of the three settings. Our findings indicate that involvement of local staff and close collaboration are essential factors for successful trial preparation and implementation. A strategic plan adapted to each setting with a distinct focus on community engagement and workforce is crucial to proceed efficiently. Mutual trust between the trial population and the trial team is of utmost importance and allows for early reaction and adaption to emerging issues.

Dissection of the gut microbiota in mothers and children with chronic Trichuris trichiura infection in Pemba Island, Tanzania.

BACKGROUND: Soil-transmitted helminthiases are important neglected tropical diseases that result in a notably high number of disability-adjusted life years worldwide. Characterizing the interactions between the human intestinal microbiome and helminths is of interest in the development of alternative treatments that do not rely on chemotherapeutics and do not lead to drug resistance. METHODS: We recruited and obtained fecal samples from 32 pairs of mothers and children on Pemba Island and monitored their intestinal microbiota using 16S rRNA gene sequencing. RESULTS: We observed that microbial changes occur in the gut microbiota of infected mothers and children. Some short-chain fatty acid (SCFA)-producing bacteria and carbohydrate-degrading bacteria exhibited lower abundance in the infected individuals. Potentially pathogenic Campylobacter and proinflammatory Methanobrevibacter in infected mothers and opportunistic Enterococcus in infected children exhibited greater abundance. CONCLUSIONS: Our findings could reveal the microbiota profiling in T. trichiura-infected individuals, indicate the potential roles of key microbiota in the host and aid to the development of novel strategies to control T. trichiura infection.

Diagnosis of soil-transmitted helminths using the Kato-Katz technique: What is the influence of stirring, storage time and storage temperature on stool sample egg counts?

BACKGROUND: Soil-transmitted helminths infect about one fifth of the world's population and have a negative impact on health. The Kato-Katz technique is the recommended method to detect soil-transmitted helminth eggs in stool samples, particularly in programmatic settings. However, some questions in its procedure remain. Our study aimed to investigate the effect of storage time, storage temperature and stirring of stool samples on fecal egg counts (FECs). METHODOLOGY/PRINCIPAL FINDINGS: In the framework of a clinical trial on Pemba Island, United Republic of Tanzania, 488 stool samples were collected from schoolchildren. These samples were evaluated in three experiments. In the first experiment (n = 92), two Kato-Katz slides were prepared from the same stool sample, one was stored at room temperature, the other in a refrigerator for 50 hours, and each slide was analyzed at nine time points (20, 50, 80, 110, 140 minutes, 18, 26, 42 and 50 hours). In the second experiment (n = 340), whole stool samples were split into two, one part was stored at room temperature, and the other part was put in a refrigerator for 48 hours. From each part one Kato-Katz slide was prepared and analyzed at three time points over two days (0, 24 and 48 hours). In the third experiment (n = 56), whole stool samples where stirred for 15 seconds six times and at each time point a Kato-Katz slide was prepared and analyzed. Mean hookworm FECs of Kato-Katz slides stored at room temperature steadily decreased following slide preparation. After two hours, mean hookworm FECs decreased from 22 to 16, whereas no reduction was observed if Kato-Katz slides were stored in the refrigerator (19 vs 21). The time x storage interaction effect was statistically significant (coefficient 0.26, 95% CI: 0.17 to 0.35, p < 0.0001). After 24 hours mean hookworm FECs dropped close to zero, irrespective of the storage condition. Whole stool samples stored at room temperature for one day resulted in a mean hookworm FEC decrease of 23% (p < 0.0001), compared to a 13% reduction (p < 0.0001) if samples were stored in the refrigerator. Fecal egg counts of A. lumbricoides and T. trichiura remained stable over time regardless of storage temperature of whole stool samples. Finally, we found a significant reduction of the variation of hookworm and T. trichiura eggs with increasing rounds of stirring the sample, but not for A. lumbricoides. For hookworm we observed a simultaneous decrease in mean FECs, making it difficult to draw recommendations on stirring samples. CONCLUSIONS/SIGNIFICANCE: Our findings suggest that stool samples (i) should be analyzed on the day of collection and (ii) should be analyzed between 20-30 minutes after slide preparation; if that is not possible, Kato-Katz slides can be stored in a refrigerator for a maximum of 110 minutes.

The detection of Schistosoma bovis in livestock on Pemba Island, Zanzibar: A preliminary study.

Schistosoma bovis is a parasitic trematode of ungulates transmitted by freshwater snails in Sub-Saharan Africa causing bovine intestinal schistosomiasis that leads to chronic morbidity and significant agricultural economic losses. The recently reported occurrence of Bulinus globosus infected with S. bovis for the first time on Pemba Island (Zanzibar, United Republic of Tanzania) is a cause of concern for livestock/wildlife health and complicates the surveillance of Schistosoma haematobium. To confirm that local cattle are infected with S. bovis, fresh faecal samples were collected from six adult cows surrounding two schistosomiasis transmission sites in Kinyasini, Pemba Island. Schistosome eggs were concentrated, egg hatching stimulated and miracidia were individually captured and identified by analysis of the partial mitochondrial cytochrome c oxidase subunit 1 (cox1) and the partial nuclear internal transcribed spacer region (ITS1+5.8S+ITS2). Two S. bovis miracidia were collected from one faecal sample with two cox1 haplotypes, one matching cox1 data obtained from S. bovis cercariae, collected previously at the same site in Pemba, the other matching S. bovis cox1 data originating from coastal Tanzania. The findings conclude that S. bovis transmission has been established on Pemba Island and is likely to have been imported through livestock trade with East Africa. Increasing the sensitivity of non-invasive diagnostics for bovine schistosomiasis, together with wider sampling, will enable a better assessment on the epidemiology of S. bovis on Pemba Island.

Efficacy, safety and acceptability of a new chewable formulation versus the solid tablet of mebendazole against hookworm infections in children: An open-label, randomized controlled trial.

BACKGROUND: Soil-transmitted helminths (STHs) infect almost 1·5 billion people worldwide. The control of STH infections is based on preventive chemotherapy using either albendazole or mebendazole. Before being widely used, a sufficient body of evidence on efficacy, safety and acceptability is warranted for the new chewable child-friendly formulation of mebendazole that was recently developed. METHODS: We conducted a randomised controlled superiority trial in four primary schools and kindergartens on Pemba Island, Tanzania. We considered eligible children aged 3 to 12 years with a hookworm infection intensity of at least 50 eggs per gram (EPG) of stool and no chronic diseases. Participants were allocated to treatment arms (ratio 1:1) using a computer generated random sequence. Our primary outcome was geometric mean based egg reduction rate (ERR) against hookworm assessed 14-21 days post-treatment. This trial complete and is registered with ClinicalTrials.gov, number NCT03995680 (June 24, 2019). FINDINGS: 397 children were eligible and randomised into the solid (198) or chewable (199) tablet arms, of whom 393 were analysed. We found no significant difference between both formulations in terms of ERR (solid 70·8% versus chewable 68·5%, difference in ERRgeometric mean 2·3%-points, 95% CI -7·8 to 12·6, p = 0.65) and CR (11·2 versus 12·7%, 95% CI -4·9 to 7·9, p = 0.65) against hookworm infections. Adverse events were mild in both treatment arms. INTERPRETATIONS: Though we could not demonstrate superiority in terms of efficacy of the new formulation, the difference between arms was small and therefore, the chewable formulation could be safely used as an alternative to swallowable tablets, in particular in young children who may have swallowing difficulties. This might help increase compliance and, consequently, enhance the effect of preventive chemotherapy.

Analytical and Clinical Assessment of a Portable, Isothermal Recombinase Polymerase Amplification (RPA) Assay for the Molecular Diagnosis of Urogenital Schistosomiasis.

Accurate diagnosis of urogenital schistosomiasis is crucial for disease surveillance and control. Routine diagnostic methods, however, lack sensitivity when assessing patients with low levels of infection still able to maintain pathogen transmission. Therefore, there is a need for highly sensitive diagnostic tools that can be used at the point-of-care in endemic areas. Recombinase polymerase amplification (RPA) is a rapid and sensitive diagnostic tool that has been used to diagnose several pathogens at the point-of-care. Here, the analytical performance of a previously developed RPA assay (RT-ShDra1-RPA) targeting the Schistosoma haematobium Dra1 genomic region was assessed using commercially synthesised S. haematobium Dra1 copies and laboratory-prepared samples spiked with S. haematobium eggs. Clinical performance was also assessed by comparing diagnostic outcomes with that of a reference diagnostic standard, urine-egg microscopy. The RT-ShDra1-RPA was able to detect 1 × 101 copies of commercially synthesised Dra1 DNA as well as one S. haematobium egg within laboratory-spiked ddH2O samples. When compared with urine-egg microscopy, the overall sensitivity and specificity of the RT-ShDra1-RPA assay was 93.7% (±88.7-96.9) and 100% (±69.1-100), respectively. Positive and negative predictive values were 100% (±97.5-100) and 50% (±27.2-72.8), respectively. The RT-ShDra1-RPA therefore shows promise as a rapid and highly sensitive diagnostic tool able to diagnose urogenital schistosomiasis at the point-of-care.

Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar.

BACKGROUND: Efforts to control and eliminate schistosomiasis have accelerated over the past decade. As parasite burden, associated morbidity and egg excretion decrease, diagnosis with standard parasitological methods becomes harder. We assessed the robustness and performance of a real-time PCR (qPCR) approach in comparison with urine filtration microscopy and reagent strip testing for the diagnosis of Schistosoma haematobium infections of different intensities. METHODS: The robustness of DNA isolation and qPCR was validated in eight laboratories from Europe and Africa. Subsequently, 792 urine samples collected during cross-sectional surveys of the Zanzibar Elimination of Schistosomiasis Transmission (ZEST) project in 2012-2017 were examined with qPCR in 2018. Diagnostic sensitivity of the qPCR was calculated at different infection intensity categories, using urine filtration microscopy as reference test. Spearman's rank correlation between Ct-values and S. haematobium egg counts was assessed and Ct-value percentiles for infection intensity categories determined. RESULTS: S. haematobium Dra1 DNA-positive samples were identified correctly in all eight laboratories. Examination of urine samples from Zanzibar revealed Dra1 DNA in 26.8% (212/792) by qPCR, S. haematobium eggs in 13.3% (105/792) by urine filtration, and microhaematuria in 13.8% (109/792) by reagent strips. Sensitivity of the qPCR increased with augmenting egg counts: 80.6% (29/36) for counts between 1 and 4 eggs, 83.3% (15/18) for counts between 5 and 9 eggs, 100% (23/23) for counts between 10 and 49 eggs, and 96.4% (27/28) for counts of 50+ eggs. There was a significant negative correlation between Ct-values and egg counts (Spearman's rho = - 0.49, P < 0.001). Seventy-five percent of the Ct-values were ≥ 33 in the egg-negative category, < 31 in the light intensity category, and < 24 in the heavy intensity category. CONCLUSIONS: While the sensitivity of the qPCR was ~ 80% for very light intensity infections (egg counts < 10), in general, the Dra1 based qPCR assay detected twice as many S. haematobium infections compared with classical parasitological tests. The qPCR is hence a sensitive, urine-based approach for S. haematobium diagnosis that can be used for impact assessment of schistosomiasis elimination programmes, individual diagnosis, and in improved format also for verification and certification of elimination. TRIAL REGISTRATION: ISRCTN, ISRCTN48837681 . Registered 05 September 2012 - Retrospectively registered.

Snail-Related Contributions from the Schistosomiasis Consortium for Operational Research and Evaluation Program Including Xenomonitoring, Focal Mollusciciding, Biological Control, and Modeling.

The Schistosomiasis Consortium for Operational Research and Evaluation (SCORE) was created in 2008 to answer questions of importance to program managers working to reduce the burden of schistosomiasis in Africa. In the past, intermediate host snail monitoring and control was an important part of integrated schistosomiasis control. However, in Africa, efforts to control snails have declined dramatically over the last 30 years. A resurgence of interest in the control of snails has been prompted by the realization, backed by a World Health Assembly resolution (WHA65.21), that mass drug administration alone may be insufficient to achieve schistosomiasis elimination. SCORE has supported work on snail identification and mapping and investigated how xenomonitoring techniques can aid in the identification of infected snails and thereby identify potential transmission areas. Focal mollusciciding with niclosamide was undertaken in Zanzibar and Côte d'Ivoire as a part of elimination studies. Two studies involving biological control of snails were conducted: one explored the association of freshwater riverine prawns and snail hosts in Côte d'Ivoire and the other assessed the current distribution of Procambarus clarkii, the invasive Louisiana red swamp crayfish, in Kenya and its association with snail hosts and schistosomiasis transmission. SCORE also supported modeling studies on the importance of snail control in achieving elimination and a meta-analysis of the impact of molluscicide-based snail control programs on human schistosomiasis prevalence and incidence. SCORE's snail control studies contributed to increased investment in building capacity, and specimens collected during SCORE research deposited in the Schistosomiasis Collections at the Natural History Museum (SCAN) will provide a valuable resource for the years to come.

SCORE Operational Research on Moving toward Interruption of Schistosomiasis Transmission.

As part of its diverse portfolio, the Schistosomiasis Consortium for Operational Research and Evaluation (SCORE) included two cluster-randomized trials evaluating interventions that could potentially lead to interruption of schistosomiasis transmission (elimination) in areas of Africa with low prevalence and intensity of infection. These studies, conducted in Zanzibar and Côte d'Ivoire, demonstrated that multiyear mass drug administration (MDA) with praziquantel failed to interrupt the transmission of urogenital schistosomiasis, even when provided biannually and/or supplemented by small-scale implementation of additional interventions. Other SCORE activities related to elimination included a feasibility and acceptability assessment of test-treat-track-test-treat (T5) strategies and mathematical modeling. Future evaluations of interventions to eliminate schistosomiasis should recognize the difficulties inherent in conducting randomized controlled trials on elimination and in measuring small changes where baseline prevalence is low. Highly sensitive and specific diagnostic tests for use in very low-prevalence areas for schistosomiasis are not routinely available, which complicates accurate measurement of infection rates and assessment of changes resulting from interventions in these settings. Although not encountered in these two studies, as prevalence and intensity decrease, political and community commitment to population-wide MDA may decrease. Because of this potential problem, SCORE developed and funded the T5 strategy implemented in Egypt, Kenya, and Tanzania. It is likely that focal MDA campaigns, along with more targeted approaches, including a T5 strategy and snail control, will need to be supplemented with the provision of clean water and sanitation and behavior change communications to achieve interruption of schistosome transmission.