Antibodies to Helminth Defense Molecule-1 are associated with inflammation, organomegaly, and decreased nutritional status in schistosomiasis japonica.

Immunomodulation enhances parasite fitness by reducing inflammation-induced morbidity in the mammalian host, as well as by attenuating parasite-targeting immune responses. Using a whole proteome differential screening method, we identified Schistosoma japonicum Helminth Defense Molecule (SjHDM-1) as a target of antibodies expressed by S. japonicum resistant, but not susceptible, individuals. In a longitudinal cohort study (N=644) conducted in a S. japonicum endemic region of the Philippines, antibody levels to SjHDM-1 did not predict resistance to reinfection but were associated with increased measures of inflammation. Individuals with high levels of anti-SjHDM-1 IgG had higher levels of C-reactive protein compared to individuals with low anti-SjHDM-1. High anti-SjHDM-1 IgG responses were also associated with reduced biomarkers of nutritional status (albumin), as well as decreased anthropometric measures of nutritional status (WAZ and HAZ) and increased measures of hepatomegaly. Our results suggest that anti-SjHDM-1 responses inhibit the immunomodulatory function of SjHDM-1, resulting in increased morbidity.

Population Pharmacokinetics of Praziquantel in Pregnant and Lactating Filipino Women Infected with Schistosoma japonicum.

An estimated 40 million women of reproductive age are infected with one of three species of the waterborne parasite Schistosoma spp. Treatment with praziquantel (PZQ) via mass drug administration (MDA) campaigns is the mainstay of schistosomiasis control for populations living in areas of endemicity. The World Health Organization recommends that pregnant and lactating women be included in schistosomiasis MDA programs, and several recent studies have evaluated the safety and efficacy of PZQ use during pregnancy. To date, there are no data describing PZQ pharmacokinetics (PK) during pregnancy or among lactating postpartum women. As part of a randomized controlled trial investigating the safety and efficacy of PZQ during human pregnancy, we examined the PK of this therapeutic drug among three distinct cohorts of women infected with S. japonicum in Leyte, Philippines. Specifically, we studied the PK properties of PZQ among early- and late-gestation pregnant women (n = 15 each) and lactating postpartum women (n = 15) with schistosomiasis. We found that women in early pregnancy had increased apparent clearance and lower area-under-the-curve (AUC0-24) values that may be related to physiological changes in drug clearance and/or changes in oral bioavailability. There was no relationship between body weight and apparent clearance. The mean ± standard deviation partition ratio of plasma to breast milk was 0.36. ± 0.13. The estimated median infant PZQ daily dose would be 0.037 mg/kg of body weight ingested from breast milk, which is significantly lower than the dosage required for antischistosomal activity and not known to be harmful to the infant. Our PK data do not support the suggestion to delay breastfeeding 72 h after taking PZQ. Results can help inform future drug efficacy studies in pregnant and lactating women with schistosomiasis.

Parasite-derived circulating microRNAs as biomarkers for the detection of human Schistosoma japonicum infection.

Novel tools for early diagnosis and monitoring of schistosomiasis are urgently needed. This study aimed to validate parasite-derived miRNAs as potential novel biomarkers for the detection of human Schistosoma japonicum infection. A total of 21 miRNAs were initially validated by real-time-polymerase chain reaction (RT-PCR) using serum samples of S. japonicum-infected BALB/c mice. Of these, 6 miRNAs were further validated with a human cohort of individuals from a schistosomiasis-endemic area of the Philippines. RT-PCR analysis showed that two parasite-derived miRNAs (sja-miR-2b-5p and sja-miR-2c-5p) could detect infected individuals with low infection intensity with moderate sensitivity/specificity values of 66%/68% and 55%/80%, respectively. Analysis of the combined data for the two parasite miRNAs revealed a specificity of 77.4% and a sensitivity of 60.0% with an area under the curve (AUC) value of 0.6906 (P = 0.0069); however, a duplex RT-PCR targeting both sja-miR-2b-5p and sja-miR-2c-5p did not result in an increased diagnostic performance compared with the singleplex assays. Furthermore, the serum level of sja-miR-2c-5p correlated significantly with faecal egg counts, whereas the other five miRNAs did not. Targeting S. japonicum-derived miRNAs in serum resulted in a moderate diagnostic performance when applied to a low schistosome infection intensity setting.

Serum Exosomal miRNAs for Grading Hepatic Fibrosis Due to Schistosomiasis.

Chronic infection with Schistosoma japonicum or Schistosoma mansoni results in hepatic fibrosis of the human host. The staging of fibrosis is crucial for prognosis and to determine the need for treatment of patients with schistosomiasis. This study aimed to determine whether there is a correlation between the levels of serum exosomal micro-ribonucleic acids (miRNAs) (exomiRs) and fibrosis progression in schistosomiasis. Reference gene (RG) validation was initially carried out for the analysis of serum exomiRs expression in staging liver fibrosis caused by schistosome infection. The expression levels of liver fibrosis-associated exomiRs in serum were determined in a murine schistosomiasis model and in a cohort of Filipino schistosomiasis japonica patients (n = 104) with different liver fibrosis grades. Of twelve RG candidates validated, miR-103a-3p and miR-425-5p were determined to be the most stable genes in the murine schistosomiasis model and subjects from the schistosomiasis-endemic area, respectively. The temporal expression profiles of nine fibrosis-associated serum exomiRs, as well as their correlations with the liver pathologies, were determined in C57BL/6 mice during S. japonicum infection. The serum levels of three exomiRs (miR-92a-3p, miR-146a-5p and miR-532-5p) were able to distinguish subjects with fibrosis grades I-III from those with no fibrosis, but only the serum level of exosomal miR-146a-5p showed potential for distinguishing patients with mild (grades 0-I) versus severe fibrosis (grades II-III). The current data imply that serum exomiRs can be a supplementary tool for grading liver fibrosis in hepatosplenic schistosomiasis with moderate accuracy.

Maternal anemia type during pregnancy is associated with anemia risk among offspring during infancy.

BACKGROUND: We evaluated the association between etiology of maternal anemia and iron status throughout infancy. METHODS: Samples from a study designed to examine Praziquantel treatment during pregnancy were used (n = 359). All women were infected with schistosomiasis and randomized to Praziquantel or placebo at 16 ± 2 weeks' gestation. Hemoglobin, serum ferritin (SF), soluble transferrin receptor (sTfR), hepcidin, C-reactive protein, and interleukin-6 were measured in maternal and infant blood. The relationship between both maternal Praziquantel treatment and etiology of anemia and infant iron status was evaluated. RESULTS: Maternal iron-deficiency anemia was associated with increased risk of infant anemia at 6 months of age. Infants of mothers with the lowest levels of circulating hepcidin during gestation, likely a marker for iron deficiency, had higher sTfR:SF levels and lower hemoglobin levels, particularly at 12 months of age. Maternal non-iron-deficiency anemia (NIDA) did not impact infant anemia risk or iron status. Maternal treatment for schistosomiasis had no effect on infant hematologic status. CONCLUSIONS: Maternal iron deficiency anemia was associated with an increased risk for anemia or iron deficiency during late infancy. We did not observe an association between maternal NIDA and increased risk for iron deficiency during infancy.

Praziquantel for the treatment of schistosomiasis during human pregnancy.

In 2014, an estimated 40 million women of reproductive age were infected with Schistosoma haematobium, S. japonicum and/or S. mansoni. In both 2003 and 2006, the World Health Organization (WHO) recommended that all schistosome-infected pregnant and breastfeeding women be offered treatment, with praziquantel, either individually or during treatment campaigns. In 2006, WHO also stated the need for randomized controlled trials to assess the safety and efficacy of such treatment. Some countries have yet to follow the recommendation on treatment and many programme managers and pregnant women in other countries remain reluctant to follow the recommended approach. Since 2006, two randomized controlled trials on the use of praziquantel during pregnancy have been conducted: one against S. mansoni in Uganda and the other against S. japonicum in the Philippines. In these trials, praziquantel treatment of pregnant women had no significant effect on birth weight, appeared safe and caused minimal side-effects that were similar to those seen in treated non-pregnant subjects. Having summarized the encouraging data, on efficacy, pharmacokinetics and safety, from these two trials and reviewed the safety data from non-interventional human studies, we recommend that all countries include pregnant women in praziquantel treatment campaigns. We identify the barriers to the treatment of pregnant women, in countries that already include such women in individual treatments and mass drug administration campaigns, and discuss ways to address these barriers.

En 2014, on estimait que 40 millions de femmes en âge de procréer étaient infectées par Schistosoma haematobium, S. japonicum et/ou S. mansoni. En 2003 et 2006, l'Organisation mondiale de la Santé (OMS) a recommandé qu'un traitement au praziquantel soit offert, individuellement ou dans le cadre de campagnes de traitement, à toutes les femmes enceintes et allaitantes infectées par le schistosome. En 2006, l'OMS a également affirmé la nécessité d'essais contrôlés randomisés pour évaluer l'innocuité et l'efficacité de ce traitement. Néanmoins, certains pays ne suivent toujours pas la recommandation relative au traitement et dans d'autres pays, bon nombre de gestionnaires de programme et de femmes enceintes demeurent réticents à suivre l'approche recommandée. Depuis 2006, deux essais contrôlés randomisés sur l'utilisation du praziquantel au cours de la grossesse ont été menés: l'un sur S. mansoni en Ouganda et l'autre sur S. japonicum aux Philippines. Dans le cadre de ces essais, le traitement au praziquantel des femmes enceintes n'a pas eu d'effet notable sur le poids à la naissance, s'est révélé sans danger et a provoqué des effets secondaires minimes, similaires à ceux constatés chez les femmes traitées qui n'étaient pas enceintes. Ayant résumé les données encourageantes sur l'efficacité, la pharmacocinétique et l'innocuité tirées de ces deux essais et examiné les données de sécurité provenant d'études non interventionnelles sur l'homme, nous recommandons que tous les pays incluent les femmes enceintes dans des campagnes de traitement au praziquantel. Nous mettons en évidence les obstacles qui empêchent le traitement des femmes enceintes ­ dans des pays les incluant déjà dans des traitements individuels et des campagnes d'administration massive de médicaments ­ et décrivons des moyens permettant de surmonter ces obstacles.

En 2014, se estima que 40 millones de mujeres en edad reproductiva estaban infectadas con Schistosoma haematobium, S. japonicum y/o S. mansoni. Tanto en 2003 como en 2006, la Organización Mundial de la Salud (OMS) recomendó que todas las mujeres embarazadas y lactantes infectadas con esquistosoma recibieran tratamiento, con praziquantel, ya fuera individualmente o durante las campañas de tratamiento. En 2006, la OMS también informó de la necesidad de ensayos aleatorizados controlados para evaluar la seguridad y la eficacia de dicho tratamiento. Algunos países todavía tienen que seguir la recomendación sobre el tratamiento y muchos gestores de programas y mujeres embarazadas en otros países siguen siendo reacios a seguir el enfoque recomendado. Desde 2006, se han llevado a cabo dos ensayos aleatorizados controlados sobre el uso de praziquantel durante el embarazo: uno contra el S. mansoni en Uganda y el otro contra el S. japonicum en Filipinas. En estos ensayos, el tratamiento con praziquantel en mujeres embarazadas no tuvo un efecto significativo sobre el peso en el momento del nacimiento, pareció seguro y causó efectos secundarios mínimos, similares a los observados en sujetos no embarazadas tratadas. Después de resumir los alentadores datos sobre la eficacia, la farmacocinética y la seguridad de estos dos ensayos y revisar los datos de seguridad de los estudios observacionales en humanos, recomendamos que todos los países incluyan a mujeres embarazadas en las campañas de tratamiento con praziquantel. Identificamos las barreras para el tratamiento de mujeres embarazadas, en países que ya incluyen a mujeres en los tratamientos individuales y en las campañas masivas de administración de medicamentos, y analizamos las formas de abordar estas barreras.

Anemia of Inflammation during Human Pregnancy Does Not Affect Newborn Iron Endowment.

Background: To our knowledge, no studies have addressed whether maternal anemia of inflammation (AI) affects newborn iron status, and few have addressed risk factors for specific etiologies of maternal anemia. Objectives: The study aims were to evaluate 1) the contribution of AI and iron deficiency anemia (IDA) to newborn iron endowment, 2) hepcidin as a biomarker to distinguish AI from IDA among pregnant women, and 3) risk factors for specific etiologies of maternal anemia. Methods: We measured hematologic biomarkers in maternal blood at 12 and 32 wk of gestation and in cord blood from a randomized trial of praziquantel in 358 pregnant women with Schistosoma japonicum in The Philippines. IDA was defined as anemia with serum ferritin <30 ng/mL and non-IDA (NIDA), largely due to AI, as anemia with ferritin ≥30 ng/mL. We identified cutoffs for biomarkers to distinguish IDA from NIDA by using area under the curve (AUC) analyses and examined the impact of different causes of anemia on newborn iron status (primary outcome) by using multivariate regression modeling. Results: Of the 358 mothers, 38% (n = 136) had IDA and 9% (n = 32) had NIDA at 32 wk of gestation. At 32 wk of gestation, serum hepcidin performed better than soluble transferrin receptor (sTfR) in identifying women with NIDA compared with the rest of the cohort (AUCs: 0.75 and 0.70, respectively) and in identifying women with NIDA among women with anemia (0.73 and 0.72, respectively). The cutoff that optimally distinguished women with NIDA from women with IDA in our cohort was 6.1 µg/L. Maternal IDA, but not NIDA, was associated with significantly lower newborn ferritin (114.4 ng/mL compared with 148.4 µg/L; P = 0.042). Conclusions: Hepcidin performed better than sTfR in identifying pregnant women with NIDA, but its cost may limit its use. Maternal IDA, but not NIDA, is associated with decreased newborn iron stores, emphasizing the need to identify this cause and provide iron therapy. This trial was registered at www.clinicaltrials.gov as NCT00486863.

Droplet Digital PCR Diagnosis of Human Schistosomiasis: Parasite Cell-Free DNA Detection in Diverse Clinical Samples.

Background: Schistosomiasis japonica remains a major public health and socioeconomic concern in Southeast Asia. Sensitive and accurate diagnostics can play a pivotal role in achieving disease elimination goals. Methods: We previously reported a novel droplet digital polymerase chain reaction (ddPCR) assay targeting the mitochondrial gene nad1 to diagnose schistosomiasis japonica. The tool identified both prepatent and patent infections using Schistosoma japonicum DNA isolated from serum, urine, salivary glands, and feces in a murine model. The assay was validated here using clinical samples collected from 412 subjects resident in an area moderately endemic for schistosomiasis in the Philippines. Results: S. japonicum DNA present in human stool, serum, urine, and saliva was detected quantitatively with high sensitivity. The capability to diagnose cases of human schistosomiasis using noninvasively collected clinical samples, the higher level of sensitivity obtained compared with the microscopy-based Kato-Katz test, and the capacity to quantify infection intensity have important public health implications for schistosomiasis control and programs targeting other neglected tropical diseases. Conclusions: This verified ddPCR method represents a valuable new tool for the diagnosis and surveillance of schistosomiasis, particularly in low-prevalence and low-intensity areas approaching elimination and in monitoring where disease emergence or re-emergence is a concern.

National Influenza Surveillance in the Philippines from 2006 to 2012: seasonality and circulating strains.

BACKGROUND: The results of routine influenza surveillance in 13 regions in the Philippines from 2006 to 2012 are presented, describing the annual seasonal epidemics of confirmed influenza virus infection, seasonal and alert thresholds, epidemic curve, and circulating influenza strains. METHODS: Retrospective analysis of Philippine influenza surveillance data from 2006 to 2012 was conducted to determine seasonality with the use of weekly influenza positivity rates and calculating epidemic curves and seasonal and alert thresholds using the World Health Organization (WHO) global epidemiological surveillance standards for influenza. RESULTS: Increased weekly influenza positive rates were observed from June to November, coinciding with the rainy season and school opening. Two or more peaks of influenza activity were observed with different dominant influenza types associated with each peak. A-H1N1, A-H3N2, and two types of B viruses circulated during the influenza season in varying proportions every year. Increased influenza activity for 2012 occurred 8 weeks late in week 29, rather than the expected week of rise of cases in week 21 as depicted in the established average epidemic curve and seasonal threshold. The intensity was severe going above the alert threshold but of short duration. Southern Hemisphere vaccine strains matched circulating influenza virus for more surveillance years than Northern Hemisphere vaccine strains. CONCLUSIONS: Influenza seasonality in the Philippines is from June to November. The ideal time to administer Southern Hemisphere influenza vaccine should be from April to May. With two lineages of influenza B circulating annually, quadrivalent vaccine might have more impact on influenza control than trivalent vaccine. Establishment of thresholds and average epidemic curve provide a tool for policy-makers to assess the intensity or severity of the current influenza epidemic even early in its course, to help plan more precisely resources necessary to control the outbreak. Influenza surveillance activities should be continued in the Philippines and funding for such activities should already be incorporated into the Philippine health budget.

Can mass drug administration lead to the sustainable control of schistosomiasis?

BACKGROUND: In the Philippines, the current national control strategy for schistosomiasis is annual mass drug administration (MDA) with 40 mg/kg of praziquantel in all schistosomiasis-endemic villages with a prevalence ≥10%. METHODS: A cross-sectional survey of schistosomiasis was conducted in 2012 on 18 221 individuals residing in 22 schistosomiasis-endemic villages in the province of Northern Samar. The prevalence of schistosomiasis, intensity of Schistosoma infection, and morbidity of disease were assessed. RESULTS: Despite an active schistosomiasis-control program in Northern Samar for >30 years, which included a MDA campaign in the last 5 years, the mean prevalence of schistosomiasis among 10 435 evaluated subjects was 27.1% (95% confidence interval [CI], 26.3%-28.0%), and the geometric mean intensity of infection among 2832 evaluated subjects was 17.2 eggs per gram of feces (95% CI, 16.4-18.1). Ultrasonography revealed high levels of schistosomiasis-induced morbidity in the schistosomiasis-endemic communities. Left lobe liver enlargement (≥70 mm) was evident in 89.3% of subjects. Twenty-five percent of the study population had grade II/III liver parenchyma fibrosis, and 13.3% had splenomegaly (≥100 mm). CONCLUSIONS: MDA on its own was insufficient to control the prevalence of schistosomiasis, intensity of Schistosoma infection, or morbidity of the disease. Alternative control measures will be needed to complement the existing national MDA program.

Childhood malnutrition and parasitic helminth interactions.

BACKGROUND: There is evidence to support that nutritional deficiency can reduce the body's immune function, thereby decreasing resistance to disease and increasing susceptibility to intestinal parasites. METHODS: A cross-sectional survey was carried out on 693 school-aged children from 5 schistosomiasis-endemic villages in Northern Samar, the Philippines. Data on dietary intake, nutritional status, and intestinal parasitic infection were collected. RESULTS: The prevalence of stunting, thinness, and wasting was 49.2%, 27.8%, and 59.7% of all children. The proportion of children infected with Schistosoma japonicum (15.6%, P = .03) and hookworm (22.0%, P = .05) were significantly lower among children who met the recommended energy and nutrient intake (RENI) for total calories. The percentage of children infected with Trichuris trichiura was highest among children who did not meet the RENI for energy (74.1%, P = .04), iron (73.4%, P = .01), thiamine (74.0%, P = .00), and riboflavin (73.3%, P = .01). Susceptibility to having 1 or more parasitic infections was significantly associated with poor intake of energy (P = .04), thiamine (P = .02), and riboflavin (P = .01).The proportion of stunted children was significantly higher among children who did not meet the RENI for energy (68.9%, P = .002), protein (54.0%, P = .004), or niacin (30.8%, P = .02) and for those infected with hookworm (31.8%, P = .0002). After adjusting for potential confounders, protein intake less than the RENI (odds ratio [OR], 1.48; 95% confidence interval [CI], 1.03-2.14), and hookworm infection (OR, 1.77; 95% CI, 1.22-2.55) were the major predictors of stunting. CONCLUSIONS: The results support the hypothesis that poor nutrient intake may increase susceptibility to parasitic diseases and together they negatively affect childhood nutritional status.

A review of health education activities targeting schoolchildren for the control of soil-transmitted helminthiasis in Southeast Asia, with emphasis upon the Magic Glasses approach.

Soil-transmitted helminths continue to be a serious problem causing disease and morbidity globally. Children, mostly school-aged, are more at risk of these infections. The main strategy for control remains to be the mass drug administration (MDA) of antihelminthic drugs. With the limitation of MDA to prevent re-infection, the need for additional approaches such as hygiene education and improvements in water, sanitation and hygiene (WASH) infrastructure are required. Although the importance of health education as a crucial component of an integrated approaches to STH control is highlighted, this component has often been disregarded because the other more complex solutions have been the focus of most studies and programmes. We performed literature searches from four bibliographic databases - Scopus, PubMed, Web of Science and Cochrane Library - to determine availability of studies on the impact of health education interventions targeting STH infections on schoolchildren in Southeast Asia. Our review found only three studies that evaluated health education interventions targeting children. The current lack of evidence in this area suggests the need for more studies assessing the impact of health education intervention for STH control. A successful health education programme for STH called "The Magic Glasses" has been developed targeting schoolchildren in China and the Philippines. This public health intervention displayed significant impact in terms of improving knowledge, attitude and practices, reducing prevalence of STH infections in schoolchildren and encouraging compliance to MDA. This article details the successes and benefits of the Magic Glasses programme as a promising control tool for STH in the Southeast Asian region.

Development and assessment of a novel gold immunochromatographic assay for the diagnosis of schistosomiasis japonica.

Background: The neglected zoonosis, schistosomiasis japonica, remains a major public health problem in the Philippines. The current study aims to develop a novel gold immunochromatographic assay (GICA) and evaluate its performance in the detection of Schistosoma japonicum infection. Methods: A GICA strip incorporating a S. japonicum saposin protein, SjSAP4 was developed. For each GICA strip test, diluted serum sample (50 µl) was loaded and strips were scanned after 10 min to convert the results into images. ImageJ was used to calculate an R value, which was defined as the signal intensity of the test line divided by the signal intensity of the control line within the cassette. After determination of optimal serum dilution and diluent, the GICA assay was evaluated with sera collected from non-endemic controls (n = 20) and individuals living in schistosomiasis-endemic areas of the Philippines (n = 60), including 40 Kato Katz (KK)-positive participants and 20 subjects confirmed as KK-negative and faecal droplet digital PCR assay (F_ddPCR)-negative at a dilution of 1:20. An ELISA assay evaluating IgG levels against SjSAP4 was also performed on the same panel of sera. Results: Phosphate-buffered saline (PBS) and 0.9% NaCl were determined as optimal dilution buffer for the GICA assay. The strips tested with serial dilutions of a pooled serum sample from KK-positive individuals (n = 3) suggested that a relatively wide range of dilutions (from 1:10 to 1:320) can be applied for the test. Using the non-endemic donors as controls, the GICA strip showed a sensitivity of 95.0% and absolute specificity; while using the KK-negative and F_ddPCR-negative subjects as controls, the immunochromatographic assay had a sensitivity of 85.0% and a specificity of 80.0%. The SjSAP4-incorperated GICA displayed a high concordance with the SjSAP4-ELISA assay. Conclusions: The developed GICA assay exhibited a similar diagnostic performance with that of the SjSAP4-ELISA assay, yet the former can be performed by local personnel with minimal training with no requirement for specialised equipment. The GICA assay established here represents a rapid, easy-to-use, accurate and field-friendly diagnostic tool for the on-site surveillance/screening of S. japonicum infection.

Comparative assessment of the SjSAP4-incorporated gold immunochromatographic assay for the diagnosis of human schistosomiasis japonica.

Background: Schistosomiasis, a disease caused by parasites of the genus Schistosoma, remains a global public health threat. This study aimed to validate the diagnostic performance of a recently developed gold immunochromatographic assay (GICA) for the detection of S. japonicum infection in a rural endemic area of the Philippines. Methods: Human clinical samples were collected from 412 subjects living in Laoang and Palapag municipalities, Northern Samar, the Philippines. The presence of Schistosoma-specific antibodies in serum samples was tested with the SjSAP4-incorporated GICA strips and the results were converted to fully quantitative data by introducing an R value. The performance of the established GICA was further compared with other diagnostic tools, including the Kato-Katz (KK) technique, point-of-care circulating cathodic antigen (POC-CCA), droplet digital (dd) PCR, and enzyme-linked immunosorbent assays (ELISAs). Results: The developed GICA strip was able to detect KK positive individuals with a sensitivity of 83.3% and absolute specificity. When calibrated with the highly sensitive faecal ddPCR assay, the immunochromatographic assay displayed an accuracy of 60.7%. Globally, the GICA assay showed a high concordance with the SjSAP4-ELISA assay. The schistosomiasis positivity rate determined by the GICA test was similar to those obtained with the SjSAP4-ELISA assay and the ddPCR assay performed on serum samples (SR_ddPCR), and was 2.3 times higher than obtained with the KK method. Conclusion: The study further confirms that the developed GICA is a valuable diagnostic tool for detecting light S. japonicum infections and implies that this point-of-care assay is a viable solution for surveying endemic areas of low-intensity schistosomiasis and identifying high-priority endemic areas for targeted interventions.

Development of CRISPR/Cas13a-based assays for the diagnosis of Schistosomiasis.

BACKGROUND: Schistosomiasis is a disease that significantly impacts human health in the developing world. Effective diagnostics are urgently needed for improved control of this disease. CRISPR-based technology has rapidly accelerated the development of a revolutionary and powerful diagnostics platform, resulting in the advancement of a class of ultrasensitive, specific, cost-effective and portable diagnostics, typified by applications in COVID-19/cancer diagnosis. METHODS: We developed CRISPR-based diagnostic platform SHERLOCK (Specific High-sensitivity Enzymatic Reporter unLOCKing) for the detection of Schistosoma japonicum and S. mansoni by combining recombinase polymerase amplification (RPA) with CRISPR-Cas13a detection, measured via fluorescent or colorimetric readouts. We evaluated SHERLOCK assays by using 150 faecal/serum samples collected from Schistosoma-infected ARC Swiss mice (female), and 189 human faecal/serum samples obtained from a S. japonicum-endemic area in the Philippines and a S. mansoni-endemic area in Uganda. FINDINGS: The S. japonicum SHERLOCK assay achieved 93-100% concordance with gold-standard qPCR detection across all the samples. The S. mansoni SHERLOCK assay demonstrated higher sensitivity than qPCR and was able to detect infection in mouse serum as early as 3 weeks post-infection. In human samples, S. mansoni SHERLOCK had 100% sensitivity when compared to qPCR of faecal and serum samples. INTERPRETATION: These schistosomiasis diagnostic assays demonstrate the potential of SHERLOCK/CRISPR-based diagnostics to provide highly accurate and field-friendly point-of-care tests that could provide the next generation of diagnostic and surveillance tools for parasitic neglected tropical diseases. FUNDING: Australian Infectious Diseases Research Centre seed grant (2022) and National Health and Medical Research Council (NHMRC) of Australia (APP1194462, APP2008433).

Cost analysis for "The Magic Glasses Philippines" health education package to prevent intestinal worm infections among Filipino schoolchildren.

Background: Soil-transmitted helminth (STH) infections are a significant public health problem affecting over 900 million people globally. Health education has been shown to complement mass drug administration (MDA) for the control of these intestinal worms. We reported recently results of a cluster randomised control trial (RCT) showing the positive impact of the "The Magic Glasses Philippines (MGP)" health education package in reducing STH infections among schoolchildren in intervention schools with ≤15% STH baseline prevalence in Laguna province, the Philippines. To inform decision making on the economic implications of the MGP, we evaluated the in-trial costs and then quantified the costs of scaling up the intervention both regionally and nationally. Methods: Costs were determined for the MGP RCT conducted in 40 schools in Laguna province. We estimated the total cost and the costs incurred per student for the actual RCT and the total costs for regional and national scale-up in all schools regardless of STH endemicity. The costs associated with the implementation of standard health education (SHE) activities and mass drug administration (MDA) were determined with a public sector perspective. Findings: The cost per participating student in the MGP RCT was Php 58.65 (USD 1.15) but if teachers instead of research staff had been involved, the estimated cost would have been considerably lower at Php 39.45 (USD 0.77). Extrapolating the costs for regional scale-up, the costs per student were estimated to be Php 15.24 (USD 0.30). As it is scaled up at the national level to include more schoolchildren, the estimated cost was increased at Php 17.46 (USD 0.34). In scenario 2 and 3, consistently, labour/salary costs associated with the delivery of the MGP contributed most to overall programme expenditure. Furthermore, the estimated average cost per student for SHE and MDA were Php 117.34 (USD 2.30) and Php 58.17 (USD 1.14), respectively. Using national scale up estimates, the cost of combining the MGP with SHE and MDA was Php 192.97 (USD 3.79). Interpretation: These findings suggest that the integration of MGP into the school curriculum would be an affordable and scalable approach to respond to the continuous burden of STH infection among schoolchildren in the Philippines. Funding: National and Medical Research Council, Australia, and the UBS-Optimus Foundation, Switzerland.

Diagnostic performance of a urine-based ELISA assay for the screening of human schistosomiasis japonica: A comparative study.

The current study developed and evaluated the performance of a urine-based enzyme-linked immunosorbent assay (ELISA) for the screening of Schistosoma japonicum infection in a human cohort (n = 412) recruited from endemic areas, Northern Samar, the Philippines. The diagnostic performance of the urine ELISA assay was further compared with the Kato-Katz (KK) technique, serum-based ELISA assays, point-of-care circulating cathodic antigen (POC-CCA) urine cassette test, and droplet digital (dd)PCR assays performed on feces, serum, urine, and saliva samples, which were designated as F_ddPCR, SR_ddPCR, U_ddPCR, and SL_ddPCR, respectively. When urine samples concentrated 16× were assessed, the SjSAP4 + Sj23-LHD-ELISA (U) showed sensitivity/specificity values of 47.2/93.8% for the detection of S. japonicum infection in KK-positive individuals (n = 108). The prevalence of S. japonicum infection in the total cohort determined by the urine ELISA assay was 48.8%, which was lower than that obtained with the F_ddPCR (74.5%, p < 0.001), SR_ddPCR (67.2%, p < 0.001), and SjSAP4 + Sj23-LHD-ELISA (S) (66.0%, p < 0.001), but higher than that determined by the Sj23-LHD-ELISA (S) (24.5%, p < 0.001), POC-CCA assay (12.4%, p < 0.001), and SL_ddPCR (25.5%, p < 0.001). Using the other diagnostic tests as a reference, the urine ELISA assay showed a sensitivity between 47.2 and 56.9%, a specificity between 50.7 and 55.2%, and an accuracy between 49.3 and 53.4%. The concentrated urine SjSAP4 + Sj23-LHD-ELISA developed in the current study was more sensitive than both the KK test and POC-CCA assay, and showed a comparable level of diagnostic accuracy to that of the U_ddPCR. However, its diagnostic performance was less robust than that of the F_ddPCR, SR_ddPCR, and SjSAP4 + Sj23-LHD-ELISA (S) assays. Although they are convenient and involve a highly acceptable non-invasive procedure for clinical sample collection, the insufficient sensitivity of the three urine-based assays (the urine ELISA assay, the U_ddPCR test, and the POC-CCA assay) will limit their value for the routine screening of schistosomiasis japonica in the post mass drug administration (MDA) era, where low-intensity infections are predominant in many endemic areas.

Identification of a linear B-cell epitope on the Schistosoma japonicum saposin protein, SjSAP4: Potential as a component of a multi-epitope diagnostic assay.

BACKGROUND: Schistosoma japonicum is one of three major species of blood flukes causing schistosomiasis, a disease, which continues to be a major public health issue in the Philippines. SjSAP4, a member of a multigene family of saposin-like proteins, is a recognized immunodiagnostic biomarker for schistosomiasis japonica. This study aimed to identify linear B-cell epitopes on SjSAP4 and to validate their potential as components of a multi-epitope assay for the serological diagnosis of schistosomiasis japonica. METHODOLOGY: SjSAP4-derived peptides were expressed as GST-peptide-fused proteins and these were Western blot probed with human serum samples from S. japonicum Kato-Katz (KK)-positive individuals and uninfected controls. A core epitope was further identified by Western blotting through probing a series of truncated peptides with the schistosomiasis patient sera. The diagnostic performance of the core epitope-containing peptides and the full-length SjSAP4 was evaluated by enzyme-linked immunosorbent assay (ELISA) using a panel of sera collected from subjects resident in a schistosomiasis-endemic area of the Philippines. MAIN FINDINGS: As a result of the peptide mapping, one peptide (P15) was found to be highly immunogenic in the KK-positive individuals. We subsequently showed that -S163QCSLVGDIFVDKYLD178- is a core B-cell epitope of P15. Subsequent ELISAs incorporating SjSAP4, SjSAP4-Peptide and SjSP-13V2-Peptide showed a sensitivity of 94.0%, 46.0% and 74.0%, respectively, and a specificity of 97.1%, 100% and 100%, respectively. Notably, complementary recognition of the B-cell epitopes (SjSAP4-Peptide and SjSP-13V2-Peptide) was observed in a subset of the KK-positive individuals. A dual epitope-ELISA (SjSAP4-Peptide + SjSP-13V2-Peptide-ELISA) showed a diagnostic sensitivity of 84.0% and a specificity of 100%. CONCLUSIONS/SIGNIFICANCE: In this study, -S163QCSLVGDIFVDKYLD178- was identified as a dominant linear B-cell epitope on SjSAP4. This peptide and the complementary recognition of other B-cell epitopes using sera from different KK-positive individuals can provide the basis of developing a multi-epitope assay for the serological diagnosis of schistosomiasis.

"The Magic Glasses Philippines": a cluster randomised controlled trial of a health education package for the prevention of intestinal worm infections in schoolchildren.

BACKGROUND: Soil-transmitted helminths (STH) cause substantial disease and disability globally. Health education has proven complementary to school-based drug administration programs for STH control. We determined the generalizability of the impact of "The Magic Glasses" health education package for STH prevention in schoolchildren in Laguna province, the Philippines, having previously shown its positive impact in China. METHODS: We conducted a cluster-randomised controlled trial, in schoolchildren, aged 9-10 years, across 40 schools over one year. Schools were randomly assigned either to the "Magic Glasses Philippines" health education intervention package (consisting of a cartoon video, classroom discussions, drawing and essay competition) complementing the standard health education activities of the Philippines Departments of Health and Education, or to a control group, which involved only the standard health education activities. The primary trial outcomes were the proportion of STH infected schoolchildren and their knowledge, attitude and behaviour of STH assessed in both groups at baseline and through two follow-up surveys undertaken immediately prior to the semi-annual national mass administration of albendazole. The outcomes between the study arms were compared using generalized estimating equation models, accounting for clustering at the school level. The trial is registered with Australian New Zealand Clinical Trials Registry number: ACTRN12616000508471. FINDINGS: At follow-up assessments, the mean knowledge and behaviour scores in the intervention group were, respectively, 5·3 (95% confidence interval [CI]:4·2-6·5; p=<0.001) and 1·1 (95% CI: 0·4-1·7; p=0.002) percentage points higher than the control group. There was no overall effect on helminth infections (any STH; adjusted odds ratio [aOR]:1·0; 95% CI: 0·8-1·3; p=0·856), Ascaris lumbricoides; aOR:1·0; 95% CI: 0·7-1·6; p=0·894, or Trichuris trichiura; aOR:1·7; 95% CI: 0·9-1·6; p=0·315) but sub-group analysis showed a 60% reduction in the odds of any STH infection resulting from the "Magic Glasses" intervention in schools with a baseline prevalence ≤15% (aOR: 0·4; 95% CI: 0·2-0·7; p=0·001). INTERPRETATION: The health-education package demonstrated a modest but statistically significant impact on the students' overall STH knowledge and changes in their behaviour but was only effective in preventing STH infections in intervention schools where the baseline prevalence was ≤15%. FUNDING: National Health and Medical Research Council, Australia, and the UBS-Optimus Foundation, Switzerland.