Performance of circulating cathodic antigen (CCA) urine-dipsticks for rapid detection of intestinal schistosomiasis in schoolchildren from shoreline communities of Lake Victoria

For disease surveillance and mapping within large-scale control programmes, RDTs are becoming popular. For intestinal schistosomiasis, a commercially available urine-dipstick which detects schistosome circulating cathodic antigen (CCA) in host urine is being increasingly applied, however, further validation is needed. In this study, we compared the CCA urine-dipstick test against double thick Kato-Katz faecal smears from 171 schoolchildren examined along the Tanzanian and Kenyan shorelines of Lake Victoria. Diagnostic methods were in broad agreement; the mean prevalence of intestinal schistosomiasis inferred by Kato-Katz examination was 68.6% (95% confidence intervals (CIs) = 60.7-75.7%) and 71.3% (95% CIs = 63.9-78.8%) by CCA urine-dipsticks. There were, however, difficulties in precisely 'calling' the CCA test result, particularly in discrimination of 'trace' reactions as either putative infection positive or putative infection negative, which has important bearing upon estimation of mean infection prevalence; considering 'trace' as infection positive mean prevalence was 94.2% (95% CIs = 89.5-97.2%). A positive association between increasing intensity of the CCA urine-dipstick test band and faecal egg count was observed. Assigning trace reactions as putative infection negative, overall diagnostic sensitivity (SS) of the CCA urine-dipstick was 87.7% (95% CIs = 80.6-93.0%), specificity (SP) was 68.1% (95% CIs = 54.3-80.0%), positive predictive value (PPV) was 86.1% (95% CIs = 78.8-91.7%) and negative predictive value (NPV) was 71.1% (95% CIs = 57.2-82.8%). To assist in objective defining of the CCA urine-dipstick result, we propose the use of a simple colour chart and conclude that the CCA urine-dipstick is a satisfactory alternative, or supplement, to Kato-Katz examination for rapid detection of intestinal schistosomiasis

Paper-based microfluidics for DNA diagnostics of malaria in low resource underserved rural communities

Rapid, low-cost, species-specific diagnosis, based upon DNA testing, is becoming important in the treatment of patients with infectious diseases. Here, we demonstrate an innovation that uses origami to enable multiplexed, sensitive assays that rival polymerase chain reactions (PCR) laboratory assays and provide high-quality, fast precision diagnostics for malaria. The paper-based microfluidic technology proposed here combines vertical flow sample-processing steps, including paper folding for whole-blood sample preparation, with an isothermal amplification and a lateral flow detection, incorporating a simple visualization system. Studies were performed in village schools in Uganda with individual diagnoses being completed in <50 min (faster than the standard laboratory-based PCR). The tests, which enabled the diagnosis of malaria species in patients from a finger prick of whole blood, were both highly sensitive and specific, detecting malaria in 98% of infected individuals in a double-blind first-in-human study. Our method was more sensitive than other field-based, benchmark techniques, including optical microscopy and industry standard rapid immunodiagnostic tests, both performed by experienced local healthcare teams (which detected malaria in 86% and 83% of cases, respectively). All assays were independently validated using a real-time double-blinded reference PCR assay. We not only demonstrate that advanced, low-cost DNA-based sensors can be implemented in underserved communities at the point of need but also highlight the challenges associated with developing and implementing new diagnostic technologies in the field, without access to laboratories or infrastructure

Paper-based microfluidics for DNA diagnostics of malaria in low resource underserved rural communities

Rapid, low-cost, species-specific diagnosis, based upon DNA testing, is becoming important in the treatment of patients with infectious diseases. Here, we demonstrate an innovation that uses origami to enable multiplexed, sensitive assays that rival polymerase chain reactions (PCR) laboratory assays and provide high-quality, fast precision diagnostics for malaria. The paper-based microfluidic technology proposed here combines vertical flow sample-processing steps, including paper folding for whole-blood sample preparation, with an isothermal amplification and a lateral flow detection, incorporating a simple visualization system. Studies were performed in village schools in Uganda with individual diagnoses being completed in &lt;50 min (faster than the standard laboratory-based PCR). The tests, which enabled the diagnosis of malaria species in patients from a finger prick of whole blood, were both highly sensitive and specific, detecting malaria in 98% of infected individuals in a double-blind first-in-human study. Our method was more sensitive than other field-based, benchmark techniques, including optical microscopy and industry standard rapid immunodiagnostic tests, both performed by experienced local healthcare teams (which detected malaria in 86% and 83% of cases, respectively). All assays were independently validated using a real-time double-blinded reference PCR assay. We not only demonstrate that advanced, low-cost DNA-based sensors can be implemented in underserved communities at the point of need but also highlight the challenges associated with developing and implementing new diagnostic technologies in the field, without access to laboratories or infrastructure

Intestinal schistosomiasis in Uganda at high altitude (>1400 m): malacological and epidemiological surveys on Mount Elgon and in Fort Portal crater lakes reveal extra preventive chemotherapy needs

Background Intestinal schistosomiasis is of public health importance in Uganda but communities living above 1400 m are not targeted for control as natural transmission is thought unlikely. To assess altitudinal boundaries and at-risk populations, conjoint malacological and epidemiological surveys were undertaken on Mount Elgon (1139 m–3937 m), in Fort Portal crater lakes and in the Rwenzori Mountains (1123 m–4050 m). Methods Seventy freshwater habitats [Mount Elgon (37), Fort Portal crater lakes (23), Rwenzori Mountains (8) and Lake Albert (2)] were inspected for Biomphalaria species. Water temperature, pH and conductivity were recorded. A parasitological examination of 756 schoolchildren [Mount Elgon (300), Fort Portal crater lakes (456)] by faecal microscopy of duplicate Kato-Katz smears from two consecutive stool samples was bolstered by antigen (urine-CCA dipstick) and antibody (SEA-ELISA) diagnostic assays. Results Biomphalaria spp. was found up to 1951 m on Mount Elgon and 1567 m in the Fort Portal crater lakes. Although no snail from Mount Elgon shed cercariae, molecular analysis judged 7.1% of snails sampled at altitudes above 1400 m as having DNA of Schistosoma mansoni; in Fort Portal crater lakes three snails shed schistosome cercariae. Prevalence of intestinal schistosomiasis as measured in schoolchildren by Kato-Katz (Mount Elgon = 5.3% v. Fort Portal crater lakes = 10.7%), CCA urine-dipsticks (18.3% v. 34.4%) and SEA-ELISA (42.3% v. 63.7%) showed negative associations with increasing altitude with some evidence of infection up to 2000 m. Conclusions Contrary to expectations, these surveys clearly show that natural transmission of intestinal schistosomiasis occurs above 1400 m, possibly extending up to 2000 m. Using spatial epidemiological predictions, this now places some extra six million people at-risk, denoting an expansion of preventive chemotherapy needs in Uganda

Schistosoma mansoni Infections in Young Children: When Are Schistosome Antigens in Urine, Eggs in Stool and Antibodies to Eggs First Detectable?

In sub-Saharan Africa, intestinal schistosomiasis is a debilitating disease caused by a worm infection. To arrest disease progression, de-worming medications are given out, often en masse, to school-aged children. In Uganda, however, much younger children can be infected, and in lakeshore communities both infants and pre-school children can already show signs and symptoms of intestinal schistosomiasis. To change de-worming practices, further information on the occurrence of infections in these younger is needed for evidence-based decision making. Our study applied current methods of disease diagnosis to better define the ‘age of first infection’ and estimate general infection prevalence within a disease-endemic village. Up to 50% of young children were clearly shown to have schistosomiasis and could likely wait up to 3–4 years before obtaining first treatment if present de-worming policies are not changed. In the context of identifying future treatment needs, we propose that antigen detection methods are most suitable

The impact of storage conditions on human stool 16S rRNA microbiome composition and diversity

Background: Multiple factors can influence stool sample integrity upon sample collection. Preservation of faecal samples for microbiome studies is therefore an important step, particularly in tropical regions where resources are limited and high temperatures may significantly influence microbiota profiles. Freezing is the accepted standard to preserve faecal samples however, cold chain methods are often unfeasible in fieldwork scenarios particularly in low and middle-income countries and alternatives are required. This study therefore aimed to address the impact of different preservative methods, time-to-freezing at ambient tropical temperatures, and stool heterogeneity on stool microbiome diversity and composition under real-life physical environments found in resource-limited fieldwork conditions. Methods: Inner and outer stool samples collected from one specimen obtained from three children were stored using different storage preservation methods (raw, ethanol and RNAlater) in a Ugandan field setting. Mixed stool was also stored using these techniques and frozen at different time-to-freezing intervals post-collection from 0–32 h. Metataxonomic profiling was used to profile samples, targeting the V1–V2 regions of 16S rRNA with samples run on a MiSeq platform. Reads were trimmed, combined and aligned to the Greengenes database. Microbial diversity and composition data were generated and analysed using Quantitative Insights Into Microbial Ecology and R software. Results: Child donor was the greatest predictor of microbiome variation between the stool samples, with all samples remaining identifiable to their child of origin despite the stool being stored under a variety of conditions. However, significant differences were observed in composition and diversity between preservation techniques, but intra-preservation technique variation was minimal for all preservation methods, and across the time-to-freezing range (0–32 h) used. Stool heterogeneity yielded no apparent microbiome differences. Conclusions: Stool collected in a fieldwork setting for comparative microbiome analyses should ideally be stored as consistently as possible using the same preservation method throughout

Understanding the relationship between egg- and antigen-based diagnostics of Schistosoma mansoni infection pre- and post-treatment in Uganda.

BACKGROUND: Schistosomiasis is a major socio-economic and public health problem in many sub-Saharan African countries. After large mass drug administration (MDA) campaigns, prevalence of infection rapidly returns to pre-treatment levels. The traditional egg-based diagnostic for schistosome infections, Kato-Katz, is being substituted in many settings by circulating antigen recognition-based diagnostics, usually the point-of-care circulating cathodic antigen test (CCA). The relationship between these diagnostics is poorly understood, particularly after treatment in both drug-efficacy studies and routine monitoring. RESULTS: We created a model of schistosome infections to better understand and quantify the relationship between these two egg- and adult worm antigen-based diagnostics. We focused particularly on the interpretation of "trace" results after CCA testing. Our analyses suggest that CCA is generally a better predictor of prevalence, particularly after treatment, and that trace CCA results are typically associated with truly infected individuals. CONCLUSIONS: Even though prevalence rises to pre-treatment levels only six months after MDAs, our model suggests that the average intensity of infection is much lower, and is probably in part due to a small burden of surviving juveniles from when the treatment occurred. This work helps to better understand CCA diagnostics and the interpretation of post-treatment prevalence estimations

Residence time, water contact, and age-driven Schistosoma mansoni infection in hotspot communities in Uganda

Schistosomiasis is the second most important parasitic infection after malaria in terms of its socioeconomic impact and is endemic in 78 countries. It affects more than 240 million people worldwide, with 90% of cases occurring in sub-Saharan Africa. In Uganda, Schistosomamansoni is the most common species, with more than seven million people infected and 17 million living at risk despite mass drug administration (MDA) of praziquantel initiated more than 16 years ago. There has been a shift in the WHO schistosomiasis goals from controlling morbidity to elimination as a public health problem. Understanding the drivers of infection in persistent transmission hotspots despite ongoing control interventions is paramount. We conducted a cross-sectional epidemiological study of 381 individuals in Bugoto community, Mayuge district, Eastern Uganda, along with a structured survey to ascertain drivers of S. mansoni infection. Bugoto has had community-wide MDA since 2004. We detected a S. mansoni prevalence of 52% across the whole community and a prevalence of 71% in school-age children. This qualifies Bugoto as a highly endemic community according to WHO guidelines. Using a multivariate logistic regression, we found that S. mansoni infection was best explained by age group, longer residence times, and any daily contact with lake water. Schistosomamansoni infection remains a large burden across this community. This study identifies opportunities for interventions that reduce lake water contact, expand treatment eligibility to all at risk, and improve MDA coverage for long-term residents in these settings to control schistosomiasis in persistent transmission hotspots

Fecal occult blood and fecal calprotectin as point-of-care markers of intestinal morbidity in Ugandan children with Schistosoma mansoni infection.

BACKGROUND: Calprotectin is a calcium-binding cytoplasmic protein found in neutrophils and increasingly used as a marker of bowel inflammation. Fecal occult blood (FOB) is also a dependable indicator of bowel morbidity. The objective of our study was to determine the applicability of these tests as surrogate markers of Schistosoma mansoni intestinal morbidity before and after treatment with praziquantel (PZQ). METHODS: 216 children (ages 3-9 years old) from Buliisa District in Lake Albert, Uganda were examined and treated with PZQ at baseline in October 2012 with 211 of them re-examined 24 days later for S. mansoni and other soil transmitted helminths (STH). POC calprotectin and FOB assays were performed at both time points on a subset of children. Associations between the test results and infection were analysed by logistic regression. RESULTS: Fecal calprotectin concentrations of 150-300 µg/g were associated with S. mansoni egg patent infection both at baseline and follow up (OR: 12.5 P = 0.05; OR: 6.8 P = 0.02). FOB had a very strong association with baseline anemia (OR: 9.2 P = 0.03) and medium and high egg intensity schistosomiasis at follow up (OR: 6.6 P = 0.03; OR: 51.3 P = 0.003). Both tests were strongly associated with heavy intensity S. mansoni infections. There was a significant decrease in FOB and calprotectin test positivity after PZQ treatment in those children who had egg patent schistosomiasis at baseline. CONCLUSIONS: Both FOB and calprotectin rapid assays were found to correlate positively and strongly with egg patent S. mansoni infection with a positive ameloriation response after PZQ treatment indicative of short term reversion of morbidity. Both tests were appropriate for use in the field with excellent operational performance and reliability. Due to its lower-cost which makes its scale-up of use affordable, FOB could be immediately adopted as a monitoring tool for PC campaigns for efficacy evaluation before and after treatment

Interpreting ambiguous ‘trace’ results in Schistosoma mansoni CCA Tests: Estimating sensitivity and specificity of ambiguous results with no gold standard

Background The development of new diagnostics is an important tool in the fight against disease. Latent Class Analysis (LCA) is used to estimate the sensitivity and specificity of tests in the absence of a gold standard. The main field diagnostic for Schistosoma mansoni infection, Kato-Katz (KK), is not very sensitive at low infection intensities. A point-of-care circulating cathodic antigen (CCA) test has been shown to be more sensitive than KK. However, CCA can return an ambiguous ‘trace’ result between ‘positive’ and ‘negative’, and much debate has focused on interpretation of traces results. Methodology/Principle findings We show how LCA can be extended to include ambiguous trace results and analyse S. mansoni studies from both Côte d’Ivoire (CdI) and Uganda. We compare the diagnostic performance of KK and CCA and the observed results by each test to the estimated infection prevalence in the population. Prevalence by KK was higher in CdI (13.4%) than in Uganda (6.1%), but prevalence by CCA was similar between countries, both when trace was assumed to be negative (CCAtn: 11.7% in CdI and 9.7% in Uganda) and positive (CCAtp: 20.1% in CdI and 22.5% in Uganda). The estimated sensitivity of CCA was more consistent between countries than the estimated sensitivity of KK, and estimated infection prevalence did not significantly differ between CdI (20.5%) and Uganda (19.1%). The prevalence by CCA with trace as positive did not differ significantly from estimates of infection prevalence in either country, whereas both KK and CCA with trace as negative significantly underestimated infection prevalence in both countries. Conclusions Incorporation of ambiguous results into an LCA enables the effect of different treatment thresholds to be directly assessed and is applicable in many fields. Our results showed that CCA with trace as positive most accurately estimated infection prevalence