Impact of Different Sampling Schemes for Decision Making in Soil-Transmitted Helminthiasis Control Programs.

Starting and stopping preventive chemotherapy (PC) for soil-transmitted helminthiasis is typically based on the prevalence of infection as measured by Kato-Katz (KK) fecal smears. Kato-Katz-based egg counts can vary highly over repeated stool samples and smears. Consequentially, the sensitivity of KK-based surveys depends on the number of stool samples per person and the number of smears per sample. Given finite resources, collecting multiple samples and/or smears means screening fewer individuals, thereby lowering the statistical precision of prevalence estimates. Using population-level data from various epidemiological settings, we assessed the performance of different sampling schemes executed within the confines of the same budget. We recommend the use of single-slide KK for determining prevalence of moderate-to-heavy intensity infection and policy decisions for starting and continuing PC; more sensitive sampling schemes may be required for policy decisions involving stopping PC. Our findings highlight that guidelines should include specific guidance on sampling schemes.

Ascaris suum draft genome.

Parasitic diseases have a devastating, long-term impact on human health, welfare and food production worldwide. More than two billion people are infected with geohelminths, including the roundworms Ascaris (common roundworm), Necator and Ancylostoma (hookworms), and Trichuris (whipworm), mainly in developing or impoverished nations of Asia, Africa and Latin America. In humans, the diseases caused by these parasites result in about 135,000 deaths annually, with a global burden comparable with that of malaria or tuberculosis in disability-adjusted life years. Ascaris alone infects around 1.2 billion people and, in children, causes nutritional deficiency, impaired physical and cognitive development and, in severe cases, death. Ascaris also causes major production losses in pigs owing to reduced growth, failure to thrive and mortality. The Ascaris-swine model makes it possible to study the parasite, its relationship with the host, and ascariasis at the molecular level. To enable such molecular studies, we report the 273 megabase draft genome of Ascaris suum and compare it with other nematode genomes. This genome has low repeat content (4.4%) and encodes about 18,500 protein-coding genes. Notably, the A. suum secretome (about 750 molecules) is rich in peptidases linked to the penetration and degradation of host tissues, and an assemblage of molecules likely to modulate or evade host immune responses. This genome provides a comprehensive resource to the scientific community and underpins the development of new and urgently needed interventions (drugs, vaccines and diagnostic tests) against ascariasis and other nematodiases.

Advances in the diagnosis of Ascaris suum infections in pigs and their possible applications in humans.

Ascariasis is one of the most common parasitic diseases in both humans and pigs. It has been shown to cause growth deficits in both species and to impair cognitive development in children. Notwithstanding its substantial impact on pig economy and public health, diagnosis of ascariasis has mostly relied on the detection of eggs in stool and further development of novel, more sensitive methods has been limited or non-existent. Here, we discuss the currently available techniques for the diagnosis of ascariasis in pigs, their caveats, and the implications of a new serological detection technique for the evaluation of both pig and human ascariasis.

Serological diagnosis of soil-transmitted helminth (Ascaris, Trichuris and hookworm) infections: A scoping review.

BACKGROUND: The World Health Organization emphasizes the importance of integrated monitoring and evaluation in neglected tropical disease (NTD) control programs. Serological assays offer a potential solution for integrated diagnosis of NTDs, particularly for those requiring mass drug administration (MDA) as primary control and elimination strategy. This scoping review aims (i) to provide an overview of assays using serum or plasma to detect infections with soil-transmitted helminths (STHs) in both humans and animals, (ii) to examine the methodologies used in this research field and (iii) to discuss advancements in serological diagnosis of STHs to guide prevention and control programs in veterinary and human medicine. METHODOLOGY: We conducted a systematic search in the Ovid MEDLINE, Embase and Cochrane Library databases, supplemented by a Google search using predefined keywords to identify commercially available serological assays. Additionally, we performed a patent search through Espacenet. PRINCIPAL FINDINGS: We identified 85 relevant literature records spanning over 50 years, with a notable increased interest in serological assay development in recent years. Most of the research efforts concentrated on diagnosing Ascaris infections in both humans and pigs, primarily using ELISA and western blot technologies. Almost all records targeted antibodies as analytes, employing proteins and peptides as analyte detection agents. Approximately 60% of sample sets described pertained to human samples. No commercially available tests for Trichuris or hookworms were identified, while for Ascaris, there are at least seven different ELISAs on the market. CONCLUSIONS: While a substantial number of assays are employed in epidemiological research, the current state of serological diagnosis for guiding STH prevention and control programs is limited. Only two assays designed for pigs are used to inform efficient deworming practices in pig populations. Regarding human diagnosis, none of the existing assays has undergone extensive large-scale validation or integration into routine diagnostics for MDA programs.

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.

Longitudinal assessment of the exposure to Ascaris lumbricoides through copromicroscopy and serology in school children from Jimma Town, Ethiopia.

BACKGROUND: We previously demonstrated that serology holds promise as an alternative diagnostic tool to copromicroscopy to monitor and evaluate deworming programs targeting soil-transmitted helminths (STHs). Here we explored the dynamics of anti-Ascaris antibodies (Ab) and evaluated the Ab-isotype of choice to assess the longitudinal exposure to Ascaris in Ethiopian school children. METHODOLOGY: Between October 2018 and February 2020, stool and blood samples were collected every four months from school children (4 to 6 years of age). Stool samples were analyzed by duplicate Kato-Katz to assess the presence and intensity of any STH infection. Plasma Ab-responses against the total extract of Ascaris suum lung third stage larvae were measured through in-house Ab-ELISA's for seven different Ab-isotypes. PRINCIPAL FINDINGS: At baseline, 42.4% of the 66 children were excreting eggs of any STH, Trichuris (37.9%) being the most prevalent. The cumulative prevalence (proportion of children tested that positive at least once over the entire study period) was 56.1% for Trichuris and 31.8% for Ascaris. For Ascaris, re-infections were frequently observed, whereas for Trichuris, children often remained excreting eggs following drug administration. When measuring anti-Ascaris Ab-levels, the cumulative seroprevalence was generally higher (IgG4: 60.6%; IgG1: 50.0%; IgE: 36.4%). The individual anti-Ascaris IgG4 levels at baseline were positively associated with the fecal egg counts averaged over the study period, the rate of egg-appearance and the number of positive test results. There was no apparent cross-reactivity between the anti-Ascaris IgG4 Ab-ELISA and Trichuris. CONCLUSIONS/SIGNIFICANCE: We demonstrate that the children are exposed to STH before the age of four and that the exposure to Ascaris is underestimated when measured with copromicroscopy. Compared to other Ab-isotypes, IgG4 is the Ab-isotype of choice to measure Ascaris exposure in STH endemic settings. Finally, the results also highlight that measuring anti-Ascaris IgG4 levels holds promise as a tool to identify individuals at higher risk for continued exposure to this STH.

Assessment of environmental contamination with soil-transmitted helminths life stages at school compounds, households and open markets in Jimma Town, Ethiopia.

BACKGROUND: It remains largely unknown where and how infections with soil-transmitted helminths (STHs; Ascaris, Trichuris, Necator and Ancylostoma) occur. We therefore aimed to identify possible sources of infection by assessing the environmental contamination in an STH-endemic area. METHODS: We first performed a series of laboratory experiments designed to optimize a soil straining-flotation method to detect and quantify Ascaris and Trichuris eggs in soil, and to validate the diagnostic performance of the optimized method when followed by microscopy and qPCR. In a second phase, we applied this method to assess the level of STH contamination in 399 environmental samples collected from 10 school compounds, 50 households and 9 open markets in Jimma Town (Ethiopia). Subsequently, we explored associations between the environmental contamination and both the corresponding STH epidemiology at the level of the schools and the household characteristics. Finally, we assessed the knowledge, attitude and practice (KAP) towards STHs in school children. PRINCIPAL FINDINGS: Our soil straining-flotation method has an analytical sensitivity of 50 eggs per 100 grams of soil and egg recovery rate of 36.0% (Ascaris) and 8.0% (Trichuris). The analysis of field samples with both microscopy and qPCR revealed the presence of 8 different helminth species of medical importance, including but not limited to the human STHs. There was a significant association between the environmental contamination and prevalence of any STH infections at the school level only. The KAP indicated a lack of knowledge and awareness of STHs. CONCLUSIONS/SIGNIFICANCE: Our optimized straining-flotation method has a moderate diagnostic performance and revealed that life stages of helminths are ubiquitous in the environment, which might be due to the poor sanitary facilities at both the schools and the households, and a poor level of KAP towards STHs. Further research is required to gain more insights into the contribution of these life stages to transmission.

Comparison of coproprevalence and seroprevalence to guide decision-making in national soil-transmitted helminthiasis control programs: Ethiopia as a case study.

BACKGROUND: WHO recommends periodical assessment of the prevalence of any soil-transmitted helminth (STH) infections to adapt the frequency of mass drug administration targeting STHs. Today, detection of eggs in stool smears (Kato-Katz thick smear) remains the diagnostic standard. However, stool examination (coprology) has important operational drawbacks and impedes integrated surveys of multiple neglected tropical diseases. Therefore, the aim of the present study was to assess the potential of applying serology instead of coprology in STH control program decision-making. METHODOLOGY: An antibody-ELISA based on extract of Ascaris lung stage larvae (AsLungL3-ELISA) was applied in ongoing monitoring activities of the Ethiopian national control program against schistosomiasis and soil-transmitted helminthiasis. Blood and stool samples were collected from over 6,700 students (median age: 11) from 63 schools in 33 woredas (districts) across the country. Stool samples of two consecutive days were analyzed applying duplicate Kato-Katz thick smear. PRINCIPAL FINDINGS: On woreda level, qualitative (seroprevalence) and quantitative (mean optical density ratio) serology results were highly correlated, and hence seroprevalence was chosen as parameter. For 85% of the woredas, prevalence based on serology was higher than those based on coprology. The results suggested cross-reactivity of the AsLungL3-ELISA with Trichuris. When extrapolating the WHO coproprevalence thresholds, there was a moderate agreement (weighted κ = 0.43) in program decision-making. Using the same threshold values would predominantly lead to a higher frequency of drug administration. SIGNIFICANCE: This is the first time that serology for soil-transmitted helminthiasis is applied on such large scale, thereby embedded in a control program context. The results underscore that serology holds promise as a tool to monitor STH control programs. Further research should focus on the optimization of the diagnostic assay and the refinement of serology-specific program decision-making thresholds.

Affordable artificial intelligence-based digital pathology for neglected tropical diseases: A proof-of-concept for the detection of soil-transmitted helminths and Schistosoma mansoni eggs in Kato-Katz stool thick smears.

BACKGROUND: With the World Health Organization's (WHO) publication of the 2021-2030 neglected tropical diseases (NTDs) roadmap, the current gap in global diagnostics became painfully apparent. Improving existing diagnostic standards with state-of-the-art technology and artificial intelligence has the potential to close this gap. METHODOLOGY/PRINCIPAL FINDINGS: We prototyped an artificial intelligence-based digital pathology (AI-DP) device to explore automated scanning and detection of helminth eggs in stool prepared with the Kato-Katz (KK) technique, the current diagnostic standard for diagnosing soil-transmitted helminths (STHs; Ascaris lumbricoides, Trichuris trichiura and hookworms) and Schistosoma mansoni (SCH) infections. First, we embedded a prototype whole slide imaging scanner into field studies in Cambodia, Ethiopia, Kenya and Tanzania. With the scanner, over 300 KK stool thick smears were scanned, resulting in total of 7,780 field-of-view (FOV) images containing 16,990 annotated helminth eggs (Ascaris: 8,600; Trichuris: 4,083; hookworms: 3,623; SCH: 684). Around 90% of the annotated eggs were used to train a deep learning-based object detection model. From an unseen test set of 752 FOV images containing 1,671 manually verified STH and SCH eggs (the remaining 10% of annotated eggs), our trained object detection model extracted and classified helminth eggs from co-infected FOV images in KK stool thick smears, achieving a weighted average precision (± standard deviation) of 94.9% ± 0.8% and a weighted average recall of 96.1% ± 2.1% across all four helminth egg species. CONCLUSIONS/SIGNIFICANCE: We present a proof-of-concept for an AI-DP device for automated scanning and detection of helminth eggs in KK stool thick smears. We identified obstacles that need to be addressed before the diagnostic performance can be evaluated against the target product profiles for both STH and SCH. Given that these obstacles are primarily associated with the required hardware and scanning methodology, opposed to the feasibility of AI-based results, we are hopeful that this research can support the 2030 NTDs road map and eventually other poverty-related diseases for which microscopy is the diagnostic standard.

Identification of antigenic linear peptides in the soil-transmitted helminth and Schistosoma mansoni proteome.

The scientific community identified non stool-based biomarkers as the way forward to support soil-transmitted helminth (STH; Ascaris lumbricoides, Trichuris trichiura and the hookworms Ancylostoma duodenale and Necator americanus) and schistosome (S. mansoni and S. haematobium) deworming programs. This support is needed in making the decision of whether or not to stop preventive chemotherapy intervention efforts and to ultimately transition towards a post-intervention surveillance phase. We applied a two-step micro-array approach to identify antigenic linear epitopes in the STH and S. mansoni proteomes. In a first experiment, we identified antigenic peptides by applying sera from 24 STH and/or S. mansoni infected Ethiopian children on a high-density peptide microarray containing 3.3 million peptides derived from the complete STH and S. mansoni proteomes. A second array experiment with 170,185 peptides that were recognized in the first array was designed to identify non-specific antibody reactivity by applying sera from 24 healthy individuals from Belgium (a non-endemic country). From this array testing cascade, several peptides were identified for STH but none of them appeared to be unique for one species. We therefore concluded that for STH, none of the peptides revealed to be sufficiently sensitive or species specific. For S. mansoni, some promising peptides were identified prompting future investigation. Based on these results, it is unlikely that linear epitopes would be highly useful in detecting species-specific antibody responses to STH in endemic communities. For S. mansoni, one particular peptide of the micro-exon gene 12 (MEG-12) protein deserves further research. In addition, this study emphasizes the need of well-characterized biobanks for biomarker discovery, particularly when the integration of multiple disease programs is envisioned.

A strong effect of individual compliance with mass drug administration for lymphatic filariasis on sustained clearance of soil-transmitted helminth infections.

BACKGROUND: The impact of semiannual mass drug administration (MDA) with albendazole (ALB; 400 mg) alone on lymphatic filariasis (LF) and soil-transmitted helminth (STH) infections was assessed during two trials conducted from 2012 to 2018 in the Republic of Congo and the Democratic Republic of Congo. The collected data were analyzed to evaluate the effect of compliance with ALB treatment on STH infections. METHODS: STH infections were diagnosed with duplicate Kato-Katz thick smears and the results are reported as eggs per gram of stool. All subjects with at least two STH infection assessments were included in the analyses. We used parametric survival models to assess the influence of compliance with ALB treatment on the probability of (i) achieving sustained clearance of an STH infection, and (ii) acquiring an STH infection during the follow-up. RESULTS: Out of 2658 subjects included in the trials, data on 202 participants (701 person-years; PY) with hookworm infection, 211 (651 PY) with Ascaris lumbricoides infection and 270 (1013 PY) with Trichuris trichiura infection were available to calculate the probability of achieving sustained clearance of infection. The effect of ALB was dose related for all three STH. For hookworm, the time required for sustained clearance was longer (4.3 years, P < 0.001) for participants who took zero doses per year and shorter (3.4 years, P = 0.112) for participants who took two doses per year compared to those who took one dose per year (3.7 years). For Ascaris, the time required to obtain sustained clearance followed the same pattern: 6.1 years (P < 0.001) and 3.2 years (P = 0.004) vs 3.6 years for, zero, two and one dose per year, respectively. For Trichuris, less time was required for sustained clearance (4.2 years, P < 0.001) for fully compliant participants, i.e. those who took two doses per year, than for those who only took one dose per year (5.0 years). ALB was more effective in achieving sustained clearance of STH infection in subjects with light baseline infection intensities compared to those with higher egg counts. CONCLUSION: Our results illustrate the importance of MDA compliance at the level of the individual with respect to the STH benefit provided by semiannual ALB MDA, which is used for the elimination of LF in Central Africa.

Characterization of the ß-tubulin gene family in Ascaris lumbricoides and Ascaris suum and its implication for the molecular detection of benzimidazole resistance.

BACKGROUND: The treatment coverage of control programs providing benzimidazole (BZ) drugs to eliminate the morbidity caused by soil-transmitted helminths (STHs) is unprecedently high. This high drug pressure may result in the development of BZ resistance in STHs and so there is an urgent need for surveillance systems detecting molecular markers associated with BZ resistance. A critical prerequisite to develop such systems is an understanding of the gene family encoding ß-tubulin proteins, the principal targets of BZ drugs. METHODOLOGY AND PRINCIPAL FINDINGS: First, the ß-tubulin gene families of Ascaris lumbricoides and Ascaris suum were characterized through the analysis of published genomes. Second, RNA-seq and RT-PCR analyses on cDNA were applied to determine the transcription profiles of the different gene family members. The results revealed that Ascaris species have at least seven different ß-tubulin genes of which two are highly expressed during the entire lifecycle. Third, deep amplicon sequencing was performed on these two genes in more than 200 adult A. lumbricoides (Ethiopia and Tanzania) and A. suum (Belgium) worms, to investigate the intra- and inter-species genetic diversity and the presence of single nucleotide polymorphisms (SNPs) that are associated with BZ resistance in other helminth species; F167Y (TTC>TAC or TTT>TAT), E198A (GAA>GCA or GAG>GCG), E198L (GAA>TTA) and F200Y (TTC>TAC or TTT>TAT). These particular SNPs were absent in the two investigated genes in all three Ascaris populations. SIGNIFICANCE: This study demonstrated the presence of at least seven ß-tubulin genes in Ascaris worms. A new nomenclature was proposed and prioritization of genes for future BZ resistance research was discussed. This is the first comprehensive description of the ß-tubulin gene family in Ascaris and provides a framework to investigate the prevalence and potential role of ß-tubulin sequence polymorphisms in BZ resistance in a more systematic manner than previously possible.

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.

Evaluation of copromicroscopy and serology to measure the exposure to Ascaris infections across age groups and to assess the impact of 3 years of biannual mass drug administration in Jimma Town, Ethiopia.

BACKGROUND: The scientific community has recently summarized the desired characteristics for diagnostic tools across the different phases of a soil-transmitted helminth (STH) mass drug administration (MDA) program. Although serology meets some of the desired criteria, there is a scarcity of data on baseline serological profiles in human populations, both prior to and during MDA programs. METHODS: In this study, we compared the copromicroscopic and the serological infection profiles in 600 school-aged children (SAC) and 600 adults at the advent of the MDA program in Jimma Town, Ethiopia. The serological profiles were examined by two ELISAs that measure IgG4 responses to the Ascaris suum haemoglobin antigen (AsHb) and a somatic extract of lung stage larvae (AsLungL3). Three years into the MDA program, we sampled another group of 600 SAC from the same schools to assess the reduction in prevalence and intensity of Ascaris infections measured by copromicroscopy and serology. PRINCIPAL FINDINGS: Prior to the start of MDA, copromicroscopy revealed an Ascaris prevalence of 31.0% and a mean fecal egg count of 2,919 eggs per gram (EPG) in SAC. Following three years of biannual treatment, the prevalence reduced to 13.2% (57.8% reduction) and the mean fecal egg count to 1,513 EPG (48.1% reduction). This reduction was also reflected in the serological results. The seroprevalence reduced with 40.9% and 27.4% and the mean optical density ratio reduced with 44.2% and 38.2% as measured by the AsHb or AsLungL3 ELISA respectively. We also showed that, despite a decreasing coproprevalence, seroprevalence to Ascaris increased with age. CONCLUSIONS: This study is the first to provide IgG4 response profiles of an endemic population to two different A. suum antigens. The results suggest that exposure to the infectious stages of Ascaris reaches beyond SAC alone. Furthermore, it highlights the possible use of serological assays to monitor changes in STH exposure during MDA programs.

Quantitative lipidomic analysis of Ascaris suum.

Ascaris is a soil-transmitted nematode that causes ascariasis, a neglected tropical disease affecting predominantly children and adolescents in the tropics and subtropics. Approximately 0.8 billion people are affected worldwide, equating to 0.86 million disability-adjusted life-years (DALYs). Exploring the molecular biology of Ascaris is important to gain a better understanding of the host-parasite interactions and disease processes, and supports the development of novel interventions. Although advances have been made in the genomics, transcriptomics and proteomics of Ascaris, its lipidome has received very limited attention. Lipidomics is an important sub-discipline of systems biology, focused on exploring lipids profiles in tissues and cells, and elucidating their biological and metabolic roles. Here, we characterised the lipidomes of key developmental stages and organ systems of Ascaris of porcine origin via high throughput LC-MS/MS. In total, > 500 lipid species belonging to 18 lipid classes within three lipid categories were identified and quantified-in precise molar amounts in relation to the dry weight of worm material-in different developmental stages/sexes and organ systems. The results showed substantial differences in the composition and abundance of lipids with key roles in cellular processes and functions (e.g. energy storage regulation and membrane structure) among distinct stages and among organ systems, likely reflecting differing demands for lipids, depending on stage of growth and development as well as the need to adapt to constantly changing environments within and outside of the host animal. This work provides the first step toward understanding the biology of lipids in Ascaris, with possibilities to work toward designing new interventions against ascariasis.

Detection of Ascaris lumbricoides infection by ABA-1 coproantigen ELISA.

Intestinal worms, or soil-transmitted helminths (STHs), affect hundreds of millions of people in all tropical and subtropical regions of the world. The most prevalent STH is Ascaris lumbricoides. Through large-scale deworming programs, World Health Organization aims to reduce morbidity, caused by moderate-to-heavy intensity infections, below 2%. In order to monitor these control programs, stool samples are examined microscopically for the presence of worm eggs. This procedure requires well-trained personnel and is known to show variability between different operators interpreting the slides. We have investigated whether ABA-1, one of the excretory-secretory products of A. lumbricoides can be used as a coproantigen marker for infection with this parasite. Polyclonal antibodies were generated and a coproantigen ELISA was developed. Using this ELISA, it was found that ABA-1 in stool detected Ascaris infection with a sensitivity of 91.5% and a specificity of 95.3%. Our results also demonstrate that there is a correlation between ABA-1 levels in stool and A. lumbricoides DNA detected in stool. Using a threshold of 18.2 ng/g stool the ABA-1 ELISA correctly assigned 68.4% of infected individuals to the moderate-to-heavy intensity infection group, with a specificity of 97.1%. Furthermore, the levels of ABA-1 in stool were shown to rapidly and strongly decrease upon administration of a standard anthelminthic treatment (single oral dose of 400 mg albendazole). In an Ascaris suum infection model in pigs, it was found that ABA-1 remained undetectable until day 28 and was detected at day 42 or 56, concurrent with the appearance of worm eggs in the stool. This report demonstrates that ABA-1 can be considered an Ascaris -specific coproantigen marker that can be used to monitor infection intensity. It also opens the path for development of point-of-care immunoassay-based tests to determine A. lumbricoides infection in stool at the sample collection site.

Diagnostic sensitivity of direct wet mount microscopy for soil-transmitted helminth infections in Jimma Town, Ethiopia.

INTRODUCTION: Soil-transmitted helminthiasis (STH) remains a major public health problem in school children in Ethiopia. Although direct wet mount microscopy (DWMM) is the means to diagnose parasitic diseases in health care facilities in Ethiopia, it remains unclear what its diagnostic performance is for STH. METHODOLOGY: A cross-sectional study was performed in Jimma Town (Ethiopia) and included 600 children from 10 primary schools. The diagnostic sensitivity of DWMM was compared to a composite reference standard (CRS) consisting of Kato-Katz, McMaster and Mini-FLOTAC. We also explored the impact of intensity of infection (the highest faecal egg counts (FECs; expressed as eggs per gram of stool (EPG)) across the CRS) on the diagnostic sensitivity of DWMM. RESULTS: Based on the CRS, there were 210 Ascaris (35.0%), 312 Trichuris (52.0%) and 102 hookworm cases (17.0%). The median intensity of infections equalled 2,057 EPG for Ascaris, 200 EPG for Trichuris and 110 EPG for hookworms. The sensitivity of DWMM was 73.8% for Ascaris, but was around 17% for both Trichuris and hookworms. The sensitivity significantly increased with intensity of STH. For Ascaris, the odds for detecting an infection intensity of 1,000 EPG was 6.2 times higher than detecting an infection of 100 EPG. For Trichuris and hookworms, these odds ratios were 7.1 and 14. CONCLUSIONS: The diagnostic sensitivity of DWMM is low for STH, but it is able to detect those subjects that are in the highest need of treatment, and hence contributes to the global goal to eliminate STH as a public health problem.

High anti-Ascaris seroprevalence in fattening pigs in Sichuan, China, calls for improved management strategies.

BACKGROUND: Ascariasis, caused by Ascaris suum, is an important soil-transmitted parasitic disease of pigs worldwide. It leads to significant economic losses in the pork industry, as a consequence of low feed conversion efficiency in pigs and liver condemnation at slaughter. Despite ascariasis still being widespread on pig farms in many developing and the industrialised countries, there are surprisingly limited data on porcine ascariasis in China, where nearly half of the world's total pork is produced. METHODS: In the present study, using the recently developed A. suum-haemoglobin (As-Hb) antigen-based serological test, we screened 512 individual serum samples from fattening pigs from 13 farms across seven distinct locations of Sichuan Province in China for anti-Ascaris antibody. RESULTS: The prevalence of anti-Ascaris antibody ranged from 0% to 100% on the distinct farms, with the mean (overall) seroprevalence being > 60%. There was no significant difference in seroprevalence between the intensive and extensive farms. CONCLUSIONS: To our knowledge, this is the first study to measure anti-Ascaris seroprevalence in China. The results of this 'snapshot' investigation indicate that Ascaris infection in commercial pig farms in Sichuan Province is seriously underestimated, encouraging future, large-scale serological studies to assess the distribution and extent of Ascaris exposure and infection in various regions of China and the world.

An in-depth report of quality control on Kato-Katz and data entry in four clinical trials evaluating the efficacy of albendazole against soil-transmitted helminth infections.

BACKGROUND: Efforts to control soil-transmitted helminth (STH) infections have intensified over the past decade. Field-survey data on STH prevalence, infection intensity and drug efficacy is necessary to guide the implementation of control programs and should be of the best possible quality. METHODOLOGY: During four clinical trials designed to evaluate the efficacy of albendazole against STHs in Brazil, Ethiopia, Lao PDR and Tanzania, quality control (QC) was performed on the duplicate Kato-Katz thick smears and the data entry. We analyzed datasets following QC on both fecal egg counts (FECs) and data entry, and compared the prevalence of any STH infection and moderate-to-heavy intensity (MHI) infections and the drug efficacy against STH infections. RESULTS: Across the four study sites, a total of 450 out of 4,830 (9.3%) Kato-Katz thick smears were re-examined. Discrepancies in FECs varied from ~3% (hookworms) to ~6.5% (Ascaris lumbricoides and Trichuris trichiura). The difference in STH prevalence and prevalence of MHI infections using the datasets with and without QC of the FECs did not exceed 0.3%, except for hookworm infections in Tanzania, where we noted a 2.2 percentage point increase in MHI infections (pre-QC: 1.6% vs. post-QC: 3.8%). There was a 100% agreement in the classification of drug efficacy of albendazole against STH between the two datasets. In total, 201 of the 28,980 (0.65%) data entries that were made to digitize the FECs were different between both data-entry clerks. Nevertheless, the overall prevalence of STH, the prevalence of MHI infections and the classification of drug efficacy remained largely unaffected. CONCLUSION/SIGNIFICANCE: In these trials, where staff was informed that QC would take place, minimal changes in study outcomes were reported following QC on FECs or data entry. Nevertheless, imposing QC did reduce the number of errors. Therefore, application of QC together with proper training of the personnel and the availability of clear standard operating procedures is expected to support higher data quality.

The impact of four years of semiannual treatments with albendazole alone on lymphatic filariasis and soil-transmitted helminth infections: A community-based study in the Democratic Republic of the Congo.

BACKGROUND: The World Health Organization now recommends semiannual mass drug administration (MDA) of albendazole with integrated vector management as an option for eliminating lymphatic filariasis (LF) in areas of loiasis-endemic countries where it may not be safe to use diethylcarbamazine or ivermectin in MDA programs. However, the published evidence base to support this policy is thin, and uptake by national programs has been slow. METHODOLOGY/PRINCIPAL FINDINGS: We conducted a community trial to assess the impact of semiannual MDA on lymphatic filariasis and soil-transmitted helminth infections (STH) in two villages in the Bandundu province of the Democratic Republic of the Congo with moderately high prevalences for LF and hookworm infections. MDA with albendazole was provided every six months from June 2014 to December 2017 with treatment coverages of the eligible population (all ≥ 2 year of age) that ranged between 56% and 88%. No adverse effects were reported during the trial. Evaluation at 48 months, (i.e. 6 months after the 8th round of MDA), showed that W. bancrofti microfilaremia (Mf) prevalence in the study communities had decreased between 2014 to 2018 from 12% to 0.9% (p<0.001). The prevalence of W. bancrofti antigenemia was also significantly reduced from 31.6% to 8.5% (p<0.001). MDA with albendazole also reduced hookworm, Ascaris lumbricoides and Trichuris trichiura infection prevalences in the community from 58.6% to 21.2% (p<0.001), from 14.0% to 1.6% and 4.1% to 2.9%, respectively. Hookworm and Ascaris infection intensities were reduced by 93% (p = 0.02) and 57% (p = 0.03), respectively. In contrast, Trichuris infection intensity was not significantly reduced by MDA (p = 0.61) over this time period. CONCLUSION/SIGNIFICANCE: These results provide strong evidence that semiannual MDA with albendazole alone is a safe and effective strategy for LF elimination in Central Africa. Community MDA also had a major impact on STH infections.