Simple fecal flotation is a superior alternative to guadruple Kato Katz smear examination for the detection of hookworm eggs in human stool

Microscopy-based identification of eggs in stool offers simple, reliable and economical options for assessing the prevalence and intensity of hookworm infections, and for monitoring the success of helminth control programs. This study was conducted to evaluate and compare the diagnostic parameters of the Kato-Katz (KK) and simple sodium nitrate flotation technique (SNF) in terms of detection and quantification of hookworm eggs, with PCR as an additional reference test in stool, collected as part of a baseline cross-sectional study in Cambodia.; Fecal samples collected from 205 people in Dong village, Rovieng district, Preah Vihear province, Cambodia were subjected to KK, SNF and PCR for the detection (and in case of microscopy-based methods, quantification) of hookworm eggs in stool. The prevalence of hookworm detected using a combination of three techniques (gold standard) was 61.0%. PCR displayed a highest sensitivity for hookworm detection (92.0%) followed by SNF (44.0%) and quadruple KK smears (36.0%) compared to the gold standard. The overall eggs per gram feces from SNF tended to be higher than for quadruple KK and the SNF proved superior for detecting low egg burdens.; As a reference, PCR demonstrated the higher sensitivity compared to SNF and the quadruple KK method for detection of hookworm in human stool. For microscopic-based quantification, a single SNF proved superior to the quadruple KK for the detection of hookworm eggs in stool, in particular for low egg burdens. In addition, the SNF is cost-effective and easily accessible in resource poor countries

Genotyping of Giardia duodenalis among children and dogs in a closed socially deprived community from Italy

Molecular characterization of Giardia duodenalis cysts from humans and animals living in well-defined contexts is useful to study the circulation of isolates and represents a tool to evaluate zoonotic infection risk. The presence of giardiasis in children living in a disadvantaged and socially deprived small Rom community, as well in dogs roaming freely in the same context was carried out by microscopic analysis and beta-giardin gene amplification. Five out of 14 children were found positive at microscopic examination for G. duodenalis and six positive at PCR, while eight out of 14 dogs tested both microscopically and molecularly positive for G. duodenalis. Moreover, most of the children and dogs were symptomatic. Molecular characterization of Giardia positive samples from children and dogs showed 99.5% identity with Giardia Assemblage A1. The dog-specific genotypes C and D were not found. The findings of this survey provide the first European evidence to support the possible role of dogs in zoonotic transmission involving children and stray dogs in a closed context with very low standards of hygiene (i.e. Rom community), and these results show the need to monitor the health of marginal populations to safeguard ethnic minority groups

Development and evaluation of a multiplex quantitative real-time Polymerase Chain Reaction for hookworm species in human stool

Hookworm disease caused by; Necator americanus; ,; Ancylostoma duodenale; , and; Ancylostoma ceylanicum; affects half a billion people worldwide. The prevalence and intensity of infection of individual hookworm species are vital for assessing morbidity and generating targeted intervention programs for their control. The present study aims to evaluate a multiplex real-time quantitative PCR (qPCR) assay to determine the prevalence and egg intensity of all three hookworm species and compare this with standard microscopy and published genus-based conventional and real-time multiplex qPCRs. Performance of the diagnostic assays was evaluated using DNA extracted from 192 fecal samples collected as part of a soil-transmitted helminth (STH) survey in northern Cambodia. The prevalence of hookworms as detected by the multiplex hookworm qPCR of 84/192 (43.8%) was significantly higher than that using microscopy of 49/192 (25.5%). The hookworm multiplex qPCR showed very good agreement for the detection of both; N. americanus; (Kappa 0.943) and; Ancylostoma; spp. (Kappa 0.936) with a multiplex STH qPCR. A strong and moderate quantitative correlation between cycle threshold and eggs per gram (EPG) feces was obtained for the hookworm qPCR for seeded DNA egg extracts (; R; 2; ≥ 0.9004) and naturally egg-infected individuals (; R; 2; = 0.6848), respectively. The newly developed hookworm quantitative multiplex qPCR has the potential for application in anthelmintic efficacy trials and for monitoring the success of mass deworming programs targeting individual species of anthroponotic and zoonotic hookworms

Immunoproteomics to identify species-specific antigens in Neospora caninum recognised by infected bovine sera

Bovine neosporosis is a disease of concern due to its global distribution and significant economic impact through massive losses in the dairy and meat industries. To date, there is no effective chemotherapeutic drug or vaccine to prevent neosporosis. Control of this disease is therefore dependent on efficient detection tests that may affect treatment management strategies. This study was conducted to identify the specific immunoreactive proteins of Neospora caninum tachyzoites recognised by sera from cattle infected with N. caninum, Toxoplasma gondii, Cryptosporidium parvum, Babesia bovis and B. bigemina, and by sera from uninfected cattle using two-DE dimensional gel electrophoresis (2-DE) combined with immunoblot and mass spectrometry (LC-MS/MS). Among 70 protein spots that reacted with all infected sera, 20 specific antigenic spots corresponding to 14 different antigenic proteins were recognised by N. caninum-positive sera. Of these immunoreactive antigens, proteins involved in cell proliferation and invasion process were highly immunogenic, including HSP90-like protein, putative microneme 4 (MIC4), actin, elongation factor 1-alpha and armadillo/beta-catenin-like repeat-containing protein. Interestingly, we discovered an unnamed protein product, rhoptry protein (ROP1), possessing strong immunoreactivity against N. caninum but with no data on function available. Moreover, we identified cross-reactive antigens among these apicomplexan parasites, especially N. caninum, T. gondii and C. parvum. Neospora caninum-specific immunodominant proteins were identified for immunodiagnosis and vaccine development. The cross-reactive antigens could be evaluated as potential common vaccine candidates or drug targets to control the diseases caused by these apicomplexan protozoan parasites

One Health – an Ecological and Evolutionary Framework for tackling Neglected Zoonotic Diseases

Understanding the complex population biology and transmission ecology of multihost parasites has been declared as one of the major challenges of biomedical sciences for the 21st century and the Neglected Zoonotic Diseases (NZDs) are perhaps the most neglected of all the Neglected Tropical Diseases (NTDs). Here we consider how multihost parasite transmission and evolutionary dynamics may affect the success of human and animal disease control programmes, particularly neglected diseases of the developing world. We review the different types of zoonotic interactions that occur, both ecological and evolutionary, their potential relevance for current human control activities, and make suggestions for the development of an empirical evidence base and theoretical framework to better understand and predict the outcome of such interactions. In particular, we consider whether preventive chemotherapy, the current mainstay of NTD control, can be successful without a One Health approach. Transmission within and between animal reservoirs and humans can have important ecological and evolutionary consequences, driving the evolution and establishment of drug resistance, as well as providing selective pressures for spill‐over, host switching, hybridizations and introgressions between animal and human parasites. Our aim here is to highlight the importance of both elucidating disease ecology, including identifying key hosts and tailoring control effort accordingly, and understanding parasite evolution, such as precisely how infectious agents may respond and adapt to anthropogenic change. Both elements are essential if we are to alleviate disease risks from NZDs in humans, domestic animals and wildlife

Multilocus Genotyping of Human Giardia Isolates Suggests Limited Zoonotic Transmission and Association between Assemblage B and Flatulence in Children

Giardia intestinalis is a protozoan parasite found world-wide and it is a major cause of diarrhea in humans and other mammals. The genetic variability within G. intestinalis is high with eight distinct genotypes or assemblages (A-H). Here we performed sequence-based multilocus genotyping of around 200 human Giardia isolates. We found evidence of limited zoonotic transmission of certain A subtypes and an association between flatulence and assemblage B infection in children. This shows that it is important to investigate different assemblages and sub-assemblages of G. intestinalis in human infections in order to understand the clinical significance, zoonotic potential, sequence divergence, and transmission pathways of this parasite

The prevalence of Giardia infection in dogs and cats, a systematic review and meta-analysis of prevalence studies from stool samples

Giardia has a wide range of host species and is a common cause of diarrhoeal disease in humans and animals. Companion animals are able to transmit a range of zoonotic diseases to their owners including giardiasis, but the size of this risk is not well known. The aim of this study was to analyse giardiasis prevalence rates in dogs and cats worldwide using a systematic search approach. Meta-analysis enabled to describe associations between Giardia prevalence and various confounding factors. Pooled prevalence rates were 15.2% (95% CI 13.8-16.7%) for dogs and 12% (95% CI 9.2-15.3%) for cats. However, there was very high heterogeneity between studies. Meta-regression showed that the diagnostic method used had a major impact on reported prevalence with studies using ELISA, IFA and PCR reporting prevalence rates between 2.6 and 3.7 times greater than studies using microscopy. Conditional negative binomial regression found that symptomatic animals had higher prevalence rates ratios (PRR) than asymptomatic animals 1.61 (95% CI 1.33-1.94) in dogs and 1.94 (95% CI 1.47-2.56) in cats. Giardia was much more prevalent in young animals. For cats >6 months, PRR=0.47 (0.42-0.53) and in dogs of the same age group PRR=0.36 (0.32-0.41). Additionally, dogs kept as pets were less likely to be positive (PRR=0.56 (0.41-0.77)) but any difference in cats was not significant. Faecal excretion of Giardia is common in dogs and slightly less so in cats. However, the exact rates depend on the diagnostic method used, the age and origin of the animal. What risk such endemic colonisation poses to human health is still unclear as it will depend not only on prevalence rates but also on what assemblages are excreted and how people interact with their pets