Human helminth co-infection: no evidence of common genetic control of hookworm and Schistosoma mansoni infection intensity in a Brazilian community.

Strong statistical associations between soil-transmitted helminths and schistosomes are frequently observed in co-endemic human populations, although the underlying explanations remain poorly understood. This study investigates the contribution of host genetics and domestic environment to hookworm and Schistosoma mansoni infection intensity and evaluates the role of genetic and non-genetic factors in co-variation of infection intensity. Detailed genealogical information allowed assignment of 1303 individuals living in the Brazilian community of Americaninhas, Minas Gerais state, to 25 pedigrees (containing between two and 1159 members) residing in 303 households. The prevalence of co-infection with both hookworms and schistosomes was high (38.5%), with significant correlation between Necator americanus and S. mansoni faecal egg counts. Bivariate variance component analysis demonstrated a modest but significant species-specific heritability for intensity of N. americanus (h(2)=0.196) and S. mansoni infection (h(2)=0.230). However, after accounting for demographic, socio-economic and household risk factors, no evidence for common genetic control of intensity of hookworm and schistosome infection was observed. There was some evidence for residual clustering within households but the majority (63%) of the covariance between N. americanus and S. mansoni infection intensity remained specific to the individual and could not be explained by shared genes, shared environment or other shared demographic, socio-economic or environmental risk factors. Our results emphasize the importance of exposure to hookworm and schistosome infection in driving the association between levels of infection with these species in hosts resident in areas of high transmission and suggest that much of this common exposure occurs outside the home

Assessment of anthelmintic efficacy of mebendazole in school children in six countries where soil-transmitted helminths are endemic

BACKGROUND: Robust reference values for fecal egg count reduction (FECR) rates of the most widely used anthelmintic drugs in preventive chemotherapy (PC) programs for controlling soil-transmitted helminths (STHs; Ascaris lumbricoides, Trichuris trichiura, and hookworm) are still lacking. However, they are urgently needed to ensure detection of reduced efficacies that are predicted to occur due to growing drug pressure. Here, using a standardized methodology, we assessed the FECR rate of a single oral dose of mebendazole (MEB; 500 mg) against STHs in six trials in school children in different locations around the world. Our results are compared with those previously obtained for similarly conducted trials of a single oral dose of albendazole (ALB; 400 mg). METHODOLOGY: The efficacy of MEB, as assessed by FECR, was determined in six trials involving 5,830 school children in Brazil, Cambodia, Cameroon, Ethiopia, United Republic of Tanzania, and Vietnam. The efficacy of MEB was compared to that of ALB as previously assessed in 8,841 school children in India and all the above-mentioned study sites, using identical methodologies. PRINCIPAL FINDINGS: The estimated FECR rate [95% confidence interval] of MEB was highest for A. lumbricoides (97.6% [95.8; 99.5]), followed by hookworm (79.6% [71.0; 88.3]). For T. trichiura, the estimated FECR rate was 63.1% [51.6; 74.6]. Compared to MEB, ALB was significantly more efficacious against hookworm (96.2% [91.1; 100], p CONCLUSIONS/SIGNIFICANCE: A minimum FECR rate of 95% for A. lumbricoides, 70% for hookworm, and 50% for T. trichiura is expected in MEB-dependent PC programs. Lower FECR results may indicate the development of potential drug resistance

Circumventing qPCR inhibition to amplify miRNAs in plasma

Background: Circulating microRNAs (c-miRNAs) have be identified in saliva, urine and blood, which has led to increasing interest in their development as biomarkers for diverse diseases including cancers. One of the key advantages of c-miRNAs over other biomarkers is the ability to be amplified and quantified by quantitative PCR (qPCR). However, at phlebotomy when whole blood is dispensed into heparinized tubes, residual levels of the anti-coagulant lithium heparin may remain in the plasma and hence with RNA isolated from the plasma. This can confound the detection of c-miRNAs by qPCR because it inhibits reverse transcriptase (RT). Here we present a procedure, modified from earlier techniques, to detect c-miRNAs in plasma that improves sensitivity and streamlines performance.Findings: Treatment of total RNA isolated from human blood plasma with Bacteroides heparinase I during reverse transcription at 37°C for one hour improved sensitivity and performance of the qPCR. This is in comparison to no treatment or treatment of the RNA prior to RT, which is the current suggested method and exposes plasma to Flavobacterium heparinum heparinase I for up to 2 hours before RT. This modest alteration improved qPCR performance and resulted in lowered threshold cycles (C) for detection of the target sequence, candidate c-miRNA biomarkers, and controls. It also reduced the expense and number of processing steps, shortening the duration of the assay and minimizing exposure of RNA to elevated temperatures.Conclusion: Incorporating Bacteroides heparinase I treatment into conventional RT protocols targeting c-miRNA in plasma can be expected to expedite the discovery of biomarkers

Advances in the Diagnosis of Human Opisthorchiasis: Development of Opisthorchis viverrini Antigen Detection in Urine

Background Many strategies to control opisthorchiasis have been employed in Thailand, but not in the other neighbouring countries. Specific control methods include mass drug administration (MDA) and health education to reduce raw fish consumption. These control efforts have greatly shifted the epidemiology of Opisthorchis viverrini (OV) infection over the last decade from presenting as densely concentrated heavy infections in single villages to widespread light OV infections distributed over wide geographical areas. Currently, the gold standard detection method for OV infection is formalin ethyl-acetate concentration technique (FECT), which has limited diagnostic sensitivity and diagnostic specificity for light OV infections, with OV eggs often confused with eggs of minute intestinal flukes (MIFs) in feces. In this study, we developed and evaluated the diagnostic performance of a monoclonal antibody-based enzyme-linked immunosorbent assay for the measurement of OV excretory-secretory (ES) antigens in urine (urine OV-ES assay) for the diagnosis of opisthorchiasis compared to the gold standard detection FECT method. Methodology We tested several methods for pre-treating urine samples prior to testing the diagnostic performance of the urine OV-ES assay. Using trichloroacetic acid (TCA) pre-treated urine, we compared detection and quantification of OV infection using the urine OV-ES assay versus FECT in OV-endemic areas in Northeastern Thailand. Receiver operating characteristic (ROC) curves were used to determine the diagnostic sensitivity and specificity of the urine OV-ES assay using TCA pre-treated urine, and to establish diagnostic positivity thresholds. The Positive Predictive Value as well as the likelihood of obtaining a positive test result (LR+) or a negative test result (LR-) were calculated for the established diagnostic positivity threshold. Diagnostic risks (Odds Ratios) were estimated using logistic regression. Results When urine samples were pre-treated with TCA prior to use in the urine OV-ES assay, the analytical sensitivity was significantly improved. Using TCA pre-treatment of urine, the urine OV-ES assay had a limit of detection (LoD) of 39 ng/ml compared to the LoD of 52 ng/mL reported for coprological antigen detection methods. Similarly, the urine OV-ES assay correlated significantly with intensity of OV infection as measured by FECT. The urine OV-ES assay was also able to detect 28 individuals as positive from the 63 (44.4%) individuals previously determined to be negative using FECT. The likelihood of a positive diagnosis of OV infection by urine OV-ES assay increased significantly with the intensity of OV infection as determined by FECT. With reference to FECT, the sensitivity and specificity of the urine OV-ES assay was 81% and 70%, respectively. Conclusion The detection of OV-infection by the urine OV-ES assay showed much greater diagnostic sensitivity and diagnostic specificity than the current gold standard FECT method for the detection and quantification of OV infection. Due to its ease-of-use, and noninvasive sample collection (urine), the urine OV-ES assay offers the potential to revolutionize the diagnosis of liver fluke infection and provide an effective tool for control and elimination of these tumorigenic parasites

BioClojure: A functional library for the manipulation of biological sequences

Motivation: BioClojure is an open-source library for the manipulation of biological sequence data written in the language Clojure. BioClojure aims to provide a functional framework for the processing of biological sequence data that provides simple mechanisms for concurrency and lazy evaluation of large data sets. Results: BioClojure provides parsers and accessors for a range of biological sequence formats, including UniProtXML, Genbank XML, fasta and fastq. In addition it provides wrappers for key analysis programs, including BLAST, SignalP, TMHMM and InterProScan, and parsers for analyzing their output. All interfaces leverage Clojure\u27s functional style and emphasize laziness and composability, so that BioClojure, and user-defined, functions can be chained into simple pipelines that are thread-safe and seamlessly integrate lazy evaluation. Availability: BioClojure is distributed under the Lesser GPL (LGPL) and the source code is freely available from GitHub (https://github.com/s312569/clj-biosequence). Contact: [email protected]

Levels of 8-OxodG Predict Hepatobiliary Pathology in Opisthorchis viverrini Endemic Settings in Thailand.

Opisthorchis viverrini is distinct among helminth infections as it drives a chronic inflammatory response in the intrahepatic bile duct that progresses from advanced periductal fibrosis (APF) to cholangiocarcinoma (CCA). Extensive research shows that oxidative stress (OS) plays a critical role in the transition from chronic O. viverrini infection to CCA. OS also results in the excision of a modified DNA lesion (8-oxodG) into urine, the levels of which can be detected by immunoassay. Herein, we measured concentrations of urine 8-oxodG by immunoassay from the following four groups in the Khon Kaen Cancer Cohort study: (1) O. viverrini negative individuals, (2) O. viverrini positive individuals with no APF as determined by abdominal ultrasound, (3) O. viverrini positive individuals with APF as determined by abdominal ultrasound, and (4) O. viverrini induced cases of CCA. A logistic regression model was used to evaluate the utility of creatinine-adjusted urinary 8-oxodG among these groups, along with demographic, behavioral, and immunological risk factors. Receiver operating characteristic (ROC) curve analysis was used to evaluate the predictive accuracy of urinary 8-oxodG for APF and CCA. Elevated concentrations of 8-oxodG in urine positively associated with APF and CCA in a strongly dose-dependent manner. Urinary 8-oxodG concentrations also accurately predicted whether an individual presented with APF or CCA compared to O. viverrini infected individuals without these pathologies. In conclusion, urinary 8-oxodG is a robust \u27candidate\u27 biomarker of the progression of APF and CCA from chronic opisthorchiasis, which is indicative of the critical role that OS plays in both of these advanced hepatobiliary pathologies. The findings also confirm our previous observations that severe liver pathology occurs early and asymptomatically in residents of O. viverrini endemic regions, where individuals are infected for years (often decades) with this food-borne pathogen. These findings also contribute to an expanding literature on 8-oxodG in an easily accessible bodily fluid (e.g., urine) as a biomarker in the multistage process of inflammation, fibrogenesis, and infection-induced cancer

Impact of gender on the decision to participate in a clinical trial: a cross-sectional study

Background In order for Informed Consent to be ethical and valid each clinical trial participant must be able to make a voluntary decision to participate, free from pressure or coercion. Nonetheless, many factors may influence the decision reached, and such influences may be different for male and female volunteers. Being aware of these differences may help researches develop better processes for obtaining consent that safeguard the right of autonomy for all participants. The goal of this study was to evaluate potential gender-based differences in the factors influencing clinical trial participation. Methods This cross-sectional study was conducted in the Northeast region of Minas Gerais, Brazil, in October 2011. A structured questionnaire was administered to 143 volunteers (48 male, 95 female) screened for participation in a clinical study of an investigational functional food with potential anthelminthic properties. Answers regarding their decision to participate in the study were compared, by gender, using chi-square and Mann Whitney tests. Odds ratios (OR) was used to measure association. Results A majority of subjects (58% of males, 59% of females) listed the desire to collaborate with the development of a product against parasitic worms as their main reason for participation. Females were significantly more likely to report a decision influenced by friends, family, or researchers (OR 3.14, 3.45, and 3.46 respectively, p \u3c 0.005). Females were also significantly more likely to report a decision influenced by general altruistic considerations (OR 8.45, p \u3c 0.005). There was no difference, by gender, in the report of decisions influenced by informational meetings, understanding of the disease, or the availability of medical treatments or exams. There was also no difference in knowledge of the rights of research participants. Conclusion Study results indicate that there is a strong difference between male and female participants regarding social influences on the decision to participate in clinical research. Further research into the impact this may have on autonomy is warranted

Comparative Immunogenicity of Na-GST-1 Human Hookworm Vaccine with Synthetic Glucopyranosyl Lipid Adjuvant (GLA) in BALB/c Mice

More than 740 million people worldwide are infected with Hookworm. Hookworm infection is most prevalent in the poorest of the poor populations of the world, and has serious health effects. Hookworm infection causes blood loss leading to iron deficiency anemia and protein energy malnutrition, which results in a compromised immune response. Consequently, the target human population suffers from an increased susceptibility to infectious diseases including hookworm infection. We have developed recombinant adult hookworm vaccines against hookworm infection to break this vicious cycle. Toll-like receptor (TLR) 4 agonist are known to boost immune response in healthy and immunocompromised individuals. We believe that co-injecting Synthetic Glucopyranosyl Lipid Adjuvant (GLA) a novel TLR-4 agonist with adult hookworm Na-GST-1 + Alhydrogel® vaccine will produce a robust and sustainable immune response in this target human population. Here, we discuss the rationale of using GLA, study designs and the results of the pre-clinical immunogenicity studies of the Human Hookworm Na-GST-1 + Alhydrogel® Vaccine in BALB/c mice with and without GLA. We conclude that, GLA enhanced the immunogenicity of co-administered adult hookworm Na-GST-1 + Alhydrogel® vaccine, producing a strong anti-Na-GST-1 IgG response. These preclinical results lay the foundation of co-administrating GLA with adult hookworm Na-GST-1 + Alhydrogel® vaccine in Phase 1 clinical trial in Brazil

Of monkeys and men: immunomic profiling of sera from humans and non-human primates resistant to schistosomiasis reveals novel potential vaccine candidates.

Schistosoma haematobium affects more than 100 million people throughout Africa and is the causative agent of urogenital schistosomiasis. The parasite is strongly associated with urothelial cancer in infected individuals and as such is designated a group I carcinogen by the International Agency for Research on Cancer. Using a protein microarray containing schistosome proteins, we sought to identify antigens that were the targets of protective IgG1 immune responses in S. haematobium-exposed individuals that acquire drug-induced resistance (DIR) to schistosomiasis after praziquantel treatment. Numerous antigens with known vaccine potential were identified, including calpain (Smp80), tetraspanins, glutathione-S-transferases, and glucose transporters (SGTP1), as well as previously uncharacterized proteins. Reactive IgG1 responses were not elevated in exposed individuals who did not acquire DIR. To complement our human subjects study, we screened for antigen targets of rhesus macaques rendered resistant to S. japonicum by experimental infection followed by self-cure, and discovered a number of new and known vaccine targets, including major targets recognized by our human subjects. This study has further validated the immunomics-based approach to schistosomiasis vaccine antigen discovery and identified numerous novel potential vaccine antigens

Carcinogenic liver fluke secretes extracellular vesicles that promote cholangiocytes to adopt a tumorigenic phenotype

Background. Throughout Asia there is an unprecedented link between cholangiocarcinoma and infection with the liver fluke Opisthorchis viverrini. Multiple processes including chronic inflammation and secretion of parasite proteins into the biliary epithelium drive infection towards cancer. Until now, the mechanism and effects of parasite protein entry into cholangiocytes was unknown. Methods. Various microscopy techniques were used to identify O. viverrini extracellular vesicles (EVs) and their internalization by human cholangiocytes. Using mass spectrometry we characterised the EV proteome and associated changes in cholangiocytes after EV uptake, and detected EV proteins in bile of infected hamsters and humans. Cholangiocyte proliferation and IL-6 secretion was measured to assess the impact of EV internalization. Results. EVs were identified in fluke culture medium and bile of infected hosts. EVs internalized by cholangiocytes drove cell proliferation and IL-6 secretion and induced changes in protein expression associated with endocytosis, wound repair and cancer. Antibodies to an O. viverrinitetraspanin blocked EV uptake and IL-6 secretion by cholangiocytes. Conclusions. This is the first time that EVs from a multicellular pathogen have been identified in host tissues. Our findings imply a role for O. viverrini EVs in pathogenesis and highlight an approach to vaccine development for this infectious cancer