A diverse microbial community and common core microbiota associated with the gonad of female Parascaris spp.

The microbiome plays an important role in health, where changes in microbiota composition can have significant downstream effects within the host, and host-microbiota relationships can be exploited to affect health outcomes. Parasitic helminths affect animals globally, but an exploration of their microbiota has been limited, despite the development of anti-Wolbachia drugs to help control infections with some filarial nematodes. The equine ascarids, Parascaris spp., are considered the most pathogenic nematodes affecting juvenile horses and are also the only ascarid parasite to have developed widespread anthelmintic resistance. The aim of this study was to characterize the microbiota of this helminth, focusing on the female gonad, determine a core microbiota for this organ, identify bacterial species, and show bacterial localization to the female gonad via in situ hybridization (ISH). A total of 22 gonads were isolated from female Parascaris spp. collected from three foals, and 9 female parasites were formalin-fixed and paraffin-embedded for ISH. Next-generation sequencing was performed using V3-V4 primers as well as the Swift Amplicon™ 16S+ ITS Panel. Overall, ten genera were identified as members of the Parascaris spp. female gonad and twelve bacterial species were identified. The most prevalent genus was Mycoplasma, followed by Reyranella, and there were no differences in alpha diversity between parasites from different horses. Specific eubacteria staining was identified in both the intestine and within the gonad using ISH. Overall, this study provided in-depth information regarding the female Parascaris spp. microbiota and was the first to identify the core microbiota within a specific parasite organ.

The equine ascarids: resuscitating historic model organisms for modern purposes.

The equine ascarids, Parascaris spp., are important nematode parasites of juvenile horses and were historically model organisms in the field of cell biology, leading to many important discoveries, and are used for the study of chromatin diminution. In veterinary parasitology, Parascaris spp. are important not only because they can cause clinical disease in young horses but also because they are the only ascarid parasites to have developed widespread anthelmintic resistance. Despite this, much of the general biology and mechanisms of anthelmintic resistance are poorly understood. This review condenses known basic biological information and knowledge on the mechanisms of anthelmintic resistance in Parascaris spp., highlighting the importance of foundational research programs. Although two variants of this parasite were recognized based on the number of chromosomes in the 1870s and suggested to be two species in 1890, one of these, P. univalens, appears to have been largely forgotten in the veterinary scientific literature over the past 100 years. We describe how this omission has had a century-long effect on nomenclature and data analysis in the field, highlighting the importance of proper specimen identification in public repositories. A summary of important basic biology, including life cycle, in vitro maintenance, and immunology, is given, and areas of future research for the improvement of knowledge and development of new systems are given. Finally, the limited knowledge regarding anthelmintic resistance in Parascaris spp. is summarized, along with caution regarding assumptions that resistance mechanisms can be applied across clades.

Ascarids exposed: a method for in vitro drug exposure and gene expression analysis of anthelmintic naïve Parascaris spp.

Ascarid parasites infect a variety of hosts and regular anthelmintic treatment is recommended for all species. Parascaris spp. is the only ascarid species with widespread anthelmintic resistance, which allows for the study of resistance mechanisms. The purpose of this study was to establish an in vitro drug exposure protocol for adult anthelmintic-naïve Parascaris spp. and report a preliminary transcriptomic analysis in response to drug exposure. Live worms were harvested from foal necropsies and maintained in RPMI-1640 at 37 °C. Serial dilutions of oxibendazole (OBZ) and ivermectin (IVM) were prepared for in vitro drug exposure, and worm viability was monitored over time. In a second drug trial, worms were used for transcriptomic analysis. The final drug concentrations employed were OBZ at 40.1 µm (10 µg mL-1) and IVM at 1.1 µm (1 µg mL-1) for 24 and 3 h, respectively. The RNA-seq analysis revealed numerous differentially expressed genes, with some being potentially related to drug detoxification and regulatory mechanisms. This report provides a method for in vitro drug exposure and the phenotypic responses for Parascaris spp., which could be extrapolated to other ascarid parasites. Finally, it also provides preliminary transcriptomic data following drug exposure as a reference point for future studies of Parascaris spp.

DETECTION OF AN ARENAVIRUS IN A GROUP OF CAPTIVE WAGLER'S PIT VIPERS (TROPIDOLAEMUS WAGLERI).

A group of eight Wagler's pit vipers (Tropidolaemus wagleri) from a private collection died with respiratory signs within 6 mo of one another. The group consisted of an adult breeding pair that was wild caught and six offspring from this pair. Four of the dead snakes were submitted for gross and histopathology. Signs of bacterial pneumonia were detected in all four examined snakes. No inclusion bodies suggestive of viral infection were found in any of the examined tissues. Polymerase chain reactions for the detection of ferla-, adeno-, reo-, and nidoviruses were all negative, but reptarenaviruses closely related to viruses previously described in boa constrictors (Boa constrictor) with inclusion body disease were detected in two of the four snakes. This is the first description of reptarenaviruses in viperid snakes. The pathogenic role of the virus in illness is unknown.

Long live the worms: methods for maintaining and assessing the viability of intestinal stages of Parascaris spp. in vitro.

In vitro maintenance of helminth parasites enables a variety of molecular, pharmaceutical and immunological analyses. Currently, the nutritional and environmental in vitro requirements of the equine ascarid parasite, Parascaris spp., have not been determined. Additionally, an objective method for assessing viability of Parascaris spp. intestinal stages does not exist. The purpose of this study was to ascertain the in vitro requirements of intestinal stages of Parascaris spp., and to develop a viability assessment method. A total of 1045 worms were maintained in a total of 212 cultures. Worms obtained from naturally infected foals at necropsy were immediately placed in culture flasks containing 200 mL of culture media. A variety of media types, nutrient supplementation and environmental conditions were examined. A motility-based scoring system was used to assess worm viability. Worms maintained in Roswell Park Memorial Institute-1640 had significantly better viability than any other media (P < 0.0001) and all media types supplemented with any of the nutrients examined (P < 0.0001). The use of a platform rocker also significantly improved viability (P = 0.0305). This is the first study to examine the requirements for maintaining Parascaris spp. intestinal stages in vitro and to evaluate their viability based on movement using an objective scoring system.

Baylisascaris procyonis Roundworm Seroprevalence among Wildlife Rehabilitators, United States and Canada, 2012-2015.

Baylisascaris procyonis roundworms can cause potentially fatal neural larva migrans in many species, including humans. However, the clinical spectrum of baylisascariasis is not completely understood. We tested 347 asymptomatic adult wildlife rehabilitators for B. procyonis antibodies; 24 were positive, suggesting that subclinical baylisascariasis is occurring among this population.

Optimal ELISA antigen for the diagnosis of Ascaris suum infection in humans.

Ascarid nematodes, Ascaris suum, Toxocara canis and Toxocara cati, are the most important causative species of larva migrans syndrome (LMS) in humans. Although the diagnosis of ascarid LMS is generally based on serological tests, specific serological tests for A. suum infection have not been fully developed. In the present study, the sensitivity and specificity of three A. suum antigen preparations, i.e., the somatic adult worm antigen (As-SWAP), larval excretory-secretory (ES) antigens derived from infective L3 (AsiL3-ES) and larval ES from tissue migratory L3 (AsmL3-ES), were evaluated for the serodiagnosis of A. suum infection in enzyme-linked immunosorbent assay (ELISA). We found that all A. suum antigen preparations showed positive reaction to all sera from A. suum-infected mice, while only AsmL3-ES obtained 100 % detection of anti-A. suum antibodies in human visceral ascarosis patients. Comparing the reactivity of each A. suum antigen, sera from both A. suum-infected mice and human patients bound to AsiL3-ES significantly weaker than As-SWAP and AsmL3-ES. Moreover, the OD450 values of ELISA with the A. suum antigen preparations and T. canis larval ES antigen (TciL3-ES) were compared in order to discriminate between ascarosis and toxocarosis. Linear discriminant analysis showed that diagnosis based on TciL3-ES and AsmL3-ES ELISA gave the most reliable result for the discrimination of infecting species. In conclusion, the application of AsmL3-ES antigen in ELISA can be recommended for the serodiagnosis of A. suum infection in humans.

Prevalence of Baylisascaris procyonis in wild rodents in central Georgia, USA.

Raccoon roundworm, Baylisascaris procyonis, is a zoonotic parasite of raccoons (Procyon lotor) that needs a One Health approach to better inform risks to human and animal health. The few studies on B. procyonis in wild rodents have primarily focused on white-footed mice (Peromyscus leucopus). This study aimed to determine the prevalence and rodent host range of B. procyonis in Georgia (USA) and investigate differences in prevalence at urban/fragmented sites and rural/agriculture sites. We sampled 99 rodents of five species. Larvae were recovered from seven of 78 (9.0%) white-footed mice with a mean of 4.4 larvae (range 1-12). One mouse had a single larva in the brain. Prevalence was not different between urban and rural sites. This report extends the geographic range of this parasite and confirms that rodents serve as paratenic hosts in the southern range. Therefore, baylisascariasis should be considered a differential for neurologic domestic animals, wildlife, or people in this region.

BEVA primary care clinical guidelines: Equine parasite control.

BACKGROUND: There is a lack of consensus on how best to balance our need to minimise the risk of parasite-associated disease in the individual horse, with the need to limit the use of anthelmintics in the population to preserve their efficacy through delaying further development of resistance. OBJECTIVES: To develop evidence-based guidelines utilising a modified GRADE framework. METHODS: A panel of veterinary scientists with relevant expertise and experience was convened. Relevant research questions were identified and developed with associated search terms being defined. Evidence in the veterinary literature was evaluated using the GRADE evidence-to-decision framework. Literature searches were performed utilising CAB abstracts and PubMed. Where there was insufficient evidence to answer the research question the panel developed practical guidance based on their collective knowledge and experience. RESULTS: Search results are presented, and recommendation or practical guidance were made in response to 37 clinically relevant questions relating to the use of anthelmintics in horses. MAIN LIMITATIONS: There was insufficient evidence to answer many of the questions with any degree of certainty and practical guidance frequently had to be based upon extrapolation of relevant information and the panel members' collective experience and opinions. CONCLUSIONS: Equine parasite control practices and current recommendations have a weak evidence base. These guidelines highlight changes in equine parasite control that should be considered to reduce the threat of parasite-associated disease and delay the development of further anthelmintic resistance.

Enteric parasites in free-living Mediterranean pond turtle (Mauremys leprosa leprosa) in contrasted areas (natural vs polluted) from central-western of Morocco.

The objective of this study is to assess the occurrence of intestinal parasites in Mediterranean pond turtle Mauremys leprosa leprosa collected from three contrasting environments in Morocco. Stool samples from 92 turtles were examined for parasite detection and enumeration. The identified intestinal parasites belong to helminths (oxyurids and ascarid) and protozoa (Entamoebidae). A total of 25 turtles (27.17%) were found to be infected by helminths and/or protozoan parasites. No adult form of these parasites was detected. Eggs of oxyurid and ascarid were detected in individuals of populations studied from Oued Ksob (23.07% and 30.76% of n = 13 turtles) and Oued Zat (34.14% and 24.39% of n = 41 turtles), respectively. For protozoa, Entamoeba cysts were present in turtles in Oued Ksob (15.38% of n = 13 turtles), Oued Zat (12.19% of n = 41 turtles), and Oued Tensift (5.26% of n = 38 turtles) localities. Oxyurid eggs showed the highest intensity at Oued Zat reaching 29.30 ± 59.59 eggs per gram (EPG), versus 12 ± 0.38 EPG for ascaris eggs in Oued Ksob. Entamoeba cysts were detected in lower levels with a maximum of 1.66 ± 1.50 cysts per gram (CPG), in Oued Zat. The prevalence of turtles eliminating eggs was statistically significant between localities for different parasite groups. This study reports for the first time a parasitological characterization of gastrointestinal parasites in wild populations of M. leprosa leprosa from contrasting environments, suggesting a relationship between turtles' infestation and the quality of their habitat.

The turkey ascarid, Ascaridia dissimilis, as a model genetic system.

Parasitic nematodes cause significant effects on humans each year, with the most prevalent being Ascaris lumbricoides. Benzimidazoles (BZ) are the most widely used anthelmintic drug in humans, and although the biology of resistance to this drug class is understood in some species, resistance is poorly characterized in ascarids. Models such as Caenorhabditis elegans were essential in developing our current understanding of BZ resistance, but more closely related model nematodes are needed to understand resistance in ascarids. Here, we propose a new ascarid model species that infects turkeys, Ascaridia dissimilis, to develop a better understanding of BZ resistance.

Parascaris spp. eggs shedding patterns in juvenile horses.

Parascaris spp. infect foals worldwide and foals typically shed eggs in the feces from about three to six months of age, upon which natural immunity is incurred. High levels of anthelmintic resistance of Parascaris spp. are a global concern, and further understanding egg shedding patterns and fecal egg counting (FEC) data variability is of high importance. The aims of this study were to monitor Parascaris spp. egg shedding in untreated foals during 12-23 weeks of age, estimate sources of data variability, and assess precision of two ascarid FEC techniques. Fecal samples were collected weekly from 11 foals born in 2022, from May through November (29 weeks). Six subsamples were extracted from each weekly sample to determine 30 FECs between two techniques: a McMaster technique and an Automated Egg Counting System (AECS). Mixed linear modeling was carried out with age, sex, birth month, seasonality, spring- or summer-born foals, and egg counting technique as explanatory variables. Ascarid FECs were associated with age (p < 0.001), seasonality (p < 0.001), and technique (p < 0.001). The McMaster technique was more precise with a mean coefficient of variation (CV) of 34.57% and a 95% confidence interval (CI) of 30.80%- 38.30% compared to the CV for the AECS, which was 42.22% (CI: 37.70%-46.70%). Seasonality accounted for the highest proportion of variance (PV) of all covariates, but differences in PVs for covariates existed between techniques with foal age and subsample contributing more variance to the McMaster, and individual foal and seasonality contributing more to the AECS. Subsamples and replicate counts accounted for less than 1% of the total data variance. The results highlighted substantial differences in PVs between the two techniques at the subsample (AECS: 57.14%; McMaster: 77.51%) and replicate count levels (AECS: 42.86%; McMaster: 22.49%). While differences in precision were observed between the two FEC techniques, they were negligible in the data set, as the overwhelming majority of the data variability in ascarid FECs was attributed to individual foal, seasonality, and foal age.

Anthelmintic resistance in equine nematodes: Current status and emerging trends.

Anthelmintic resistance is reported in equine nematodes with increasing frequency in recent years, and no new anthelmintic classes have been introduced during the past 40 years. This manuscript reviews published literature describing anthelmintic resistance in cyathostomins, Parascaris spp., and Oxyuris equi with special emphasis on larvicidal efficacy against encysted cyathostomin larvae and strongylid egg reappearance periods (ERP). Resistance to benzimidazoles and pyrimidines is highly prevalent in cyathostomin populations around the world, and macrocyclic lactone resistance has been documented in cyathostomins in recent years as well. Two recent studies have documented resistance to the larvicidal regimen of fenbendazole, whereas the larvicidal efficacy of moxidectin is variable, but with no evidence of a reduction from historic levels. In the 1990s, ERP estimates were 8-10 and 12-16 weeks for ivermectin and moxidectin, respectively, while several studies published after year 2000 found ERPs to be 5 weeks for both compounds. This is a clear change in anthelmintic performance, but it remains unclear if this is due to development of anthelmintic resistance or selection for other biological traits leading to a quicker resumption of strongylid egg shedding following anthelmintic treatment. Macrocyclic lactone resistance is common in Parascaris spp. around the world, but recent reports suggests that resistance to the two other classes should be monitored as well. Finally, O. equi has been reported resistant to ivermectin and moxidectin in countries representing four continents. In conclusion, multi-drug resistance is becoming the norm in managed cyathostomin populations around the world, and a similar pattern may be emerging in Parascaris spp. More work is required to understand the mechanisms behind the shortened ERPs, and researchers and veterinarians around the world are encouraged to routinely monitor anthelmintic efficacy against equine nematodes.

The microbial community associated with Parascaris spp. infecting juvenile horses.

BACKGROUND: Parasitic nematodes, including large roundworms colloquially known as ascarids, affect the health and well-being of livestock animals worldwide. The equine ascarids, Parascaris spp., are important parasites of juvenile horses and the first ascarids to develop widespread anthelmintic resistance. The microbiota has been shown to be an important factor in the fitness of many organisms, including parasitic nematodes, where endosymbiotic Wolbachia have been exploited for treatment of filariasis in humans. METHODS: This study used short-read 16S rRNA sequences and Illumina sequencing to characterize and compare microbiota of whole worm small intestinal stages and microbiota of male and female intestines and gonads. Diversity metrics including alpha and beta diversity, and the differential abundance analyses DESeq2, ANCOM-BC, corncob, and metagenomeSeq were used for comparisons. RESULTS: Alpha and beta diversity of whole worm microbiota did not differ significantly between groups, but Simpson alpha diversity was significantly different between female intestine (FI) and male gonad (MG) (P= 0.0018), and Shannon alpha diversity was significantly different between female and male gonads (P = 0.0130), FI and horse jejunum (HJ) (P = 0.0383), and FI and MG (P= 0.0001). Beta diversity (Fig. 2B) was significantly different between female and male gonads (P = 0.0006), male intestine (MI) and FG (P = 0.0093), and MG and FI (P = 0.0041). When comparing organs, Veillonella was differentially abundant for DESeq2 and ANCOM-BC (p < 0.0001), corncob (P = 0.0008), and metagenomeSeq (P = 0.0118), and Sarcina was differentially abundant across four methods (P < 0.0001). Finally, the microbiota of all individual Parascaris spp. specimens were compared to establish shared microbiota between groups. CONCLUSIONS: Overall, this study provided important information regarding the Parascaris spp. microbiota and provides a first step towards determining whether the microbiota may be a viable target for future parasite control options.

Detection of Giardia and helminths in Western Europe at local K9 (canine) sites (DOGWALKS Study).

BACKGROUND: Intestinal parasite contamination from infected dogs can place other dogs and humans at risk. A study was initiated to estimate the prevalence of canine intestinal parasitism by collecting fecal samples in cities across Western Europe. METHODS: Fresh fecal samples were collected from 2469 dogs visiting 164 parks in 33 cities across 12 countries. Each owner responded to a questionnaire focusing on their dog's signalment and recent anthelmintic treatment history. The collected samples were examined for hookworms, whipworms, ascarids and Giardia using a coproantigen diagnostic immunoassay and microscopy following centrifugal flotation. RESULTS: Nematodes or Giardia were detected in at least one sample from 100% of cities and in 93.3% of parks. Nematodes were detected in 57% of parks. Overall, 22.8% of dogs tested positive for an intestinal parasite, with Giardia being the most commonly identified parasites (17.3% of dogs, 83.5% of parks). For nematode infection, 7.6% of all dogs tested positive, with 9.9% of dogs aged < 1 year infected, 7.7% of those aged 1-3 years, 7.3% of those aged 4-6 years and 6.6% of those aged ≥ 7 years. Among the nematodes detected, ascarids were the most prevalent (3.6% of dogs, parks, 28.7% of parks), being most common in dogs aged < 1 year but also present in older dogs, including those aged ≥ 7 years. Hookworms and whipworms were detected in 3.2% and 2.3% of dogs of all ages, respectively, and in 37.2% and 17.7% of parks, respectively. A larger proportion of fecal samples tested positive with the coproantigen immunoassay than with centrifugal flotation. Positive test results for Giardia were sevenfold higher when both diagnostic tests were used than when centrifugal flotation alone was used, and there were 60% more positive test results for nematodes when both tests were used than when flotation alone was used. Overall, 77.2% of owners reported previous anthelmintic treatment, among whom at least 62.7% failed to follow recommended treatment frequency. Dogs receiving anthelmintic within the previous month had a lower percentage of nematode infection than those in which > 1 month had passed since the previous dose. CONCLUSIONS: The prevalence estimates of intestinal parasite infections in dogs reported here highlight the need for owner education concerning guidelines for regular testing and treatment, even in older dogs. Failure to adhere to guidelines can result in ongoing transmission of these infections, including those with zoonotic potential. Combining coproantigen immunoassay with centrifugal flotation for diagnostic testing and regular anthelmintic treatment are important measures for ensuring optimal intestinal parasite control.

Canine nematode and Giardia spp. infections in dogs in Edmonton, Alberta, the "CANIDA" study.

BACKGROUND: Canine intestinal parasite prevalence may be influenced by geographical region, age, and health status of the dog. Behaviors such as predation, scavenging, or roaming as well as routine administration of anthelmintics also play a role. The purpose of this study was to evaluate fecal test results using zinc sulfate flotation by centrifugation combined with coproantigen testing directed at protein antigens excreted or secreted by hookworms (Ancylostoma spp. Uncinaria stenocephala), ascarids (Toxocara canis, Toxascaris spp. Baylisascaris spp.), whipworms (Trichuris vulpis), and Giardia spp. during active infection in owned dogs visiting dog parks in Western Canada. METHODS: A total of 774 participants were recruited from Edmonton, Alberta, Canada. Canine fecal samples were collected from seven dedicated off-leash dog parks. Participating dog owners responded to a questionnaire regarding their dogs' signalment, previous veterinary history, and use of parasite-preventive products. Fecal samples were tested using zinc sulfate centrifugation combined with coproantigen testing. RESULTS: The overall prevalence of canine intestinal parasites in client-owned dogs was similar to previous studies conducted in the US. Mean age of dogs tested was 4 years, with puppies and older dogs having higher rates of infection than the mean. Fecal flotation centrifugation found 3.2% hookworm, ascarid, whipworm, and Giardia spp.-positive infections. Coproantigen testing identified 5.8% positive infections, including all of the above that were detected using fecal flotation centrifugation. CONCLUSIONS: Coproantigen testing detected more hookworm, ascarid, whipworm, and Giardia spp.-positive samples in addition to detecting all positive results found using fecal flotation centrifugation. Fecal flotation centrifugation combined with coproantigen testing improves sensitivity over flotation alone and may detect pre-patent or sub-clinical infections in dogs visiting public dog parks.

Monitoring equine ascarid and cyathostomin parasites: Evaluating health parameters under different treatment regimens.

BACKGROUND: Strongylid and ascarid parasites are omnipresent in equine stud farms, and ever-increasing levels of anthelmintic resistance are challenging the industry with finding more sustainable and yet effective parasite control programs. OBJECTIVES: To evaluate egg count levels, bodyweight and equine health under defined parasite control protocols in foals and mares at two Standardbred and two Thoroughbred stud farms. STUDY DESIGN: Longitudinal randomised field trial. METHODS: A total of 93 foals were enrolled and split into two treatment groups, and 99 mares were enrolled and assigned to three treatment groups. All horses underwent a health examination, and episodes of colic or diarrhoea were recorded at each faecal collection date. Bodyweights were assessed using a weight tape, and mares were body condition scored. Group A foals (FA) were dewormed at 2 and 5 months of age with a fenbendazole/ivermectin/praziquantel product, while group B foals (FB) were dewormed on a monthly basis, alternating between the above-mentioned product and an oxfendazole/pyrantel embonate product. Group A mares (MA) were dewormed twice with fenbendazole/ivermectin/praziquantel, group B mares (MB) were dewormed with the same product, when egg counts exceeded 300 strongylid eggs per gram, and group C mares (MC) were dewormed every 2 months, alternating between the two products. Health data were collected monthly for 6 months (foals) and bimonthly for 13 months (mares). Data were analysed with mixed linear models and interpreted at the α = 0.05 significance level. RESULTS: There were no significant bodyweight differences between foal groups, but MA mares were significantly lighter than the other two groups. Very few health incidents were recorded. Foals in group FA had significantly higher ascarid and strongylid egg counts, whereas no significant differences were observed between mare groups. MAIN LIMITATIONS: Study duration limited to one season. CONCLUSIONS: Anthelmintic treatment intensity was lowered from the traditional intensive regimes without measurable negative health consequences for mares and foals.

Comparative studies on faecal egg counting techniques used for the detection of gastrointestinal parasites of equines: A systematic review.

Faecal egg counting techniques (FECT) form the cornerstone for the detection of gastrointestinal parasites in equines. For this purpose, several flotation, centrifugation, image- and artificial intelligence-based techniques are used, with varying levels of performance. This review aimed to critically appraise the literature on the assessment and comparison of various coprological techniques and/or modifications of these techniques used for equines and to identify the knowledge gaps and future research directions. We searched three databases for published scientific studies on the assessment and comparison of FECT in equines and included 27 studies in the final synthesis. Overall, the performance parameters of McMaster (81.5%), Mini-FLOTAC® (33.3%) and simple flotation (25.5%) techniques were assessed in most of the studies, with 77.8% of them comparing the performance of at least two or three methods. The detection of strongyle, Parascaris spp. and cestode eggs was assessed for various FECT in 70.4%, 18.5% and 18.5% studies, respectively. A sugar-based flotation solution with a specific gravity of ≥1.2 was found to be the optimal flotation solution for parasitic eggs in the majority of FECT. No uniform or standardised protocol was followed for the comparison of various FECT, and the tested sample size (i.e. equine population and faecal samples) also varied substantially across all studies. To the best of our knowledge, this is the first systematic review to evaluate studies on the comparison of FECT in equines and it highlights important knowledge gaps in the evaluation and comparison of such techniques.

Very low intraspecific sequence variation in selected nuclear and mitochondrial Parascaris univalens genes.

Equines were over decades considered to be infected by two morphologically virtually indistinguishable ascarid species, Parascaris univalens and Parascaris equorum. Reliable species discrimination is only possible using enzyme isoelectric focussing and karyotyping with P. univalens having one and P. equorum two chromosome pairs. However, presumably the complexity of both methods prevented their routine use in nearly all previous studies about prevalence and drug resistance of Parascaris spp. These have barely been performed on the species level although most studies stated presence of one or the other species. Recently, only P. univalens has been identified by karyotyping and the last published study identifying P. equorum dates back to 1989. In order to improve species-specific detection, molecular markers are required. Here, partial 12S rRNA, cytochrome oxidase I (COI) and complete internal transcribed spacer (ITS)-1 and - 2 sequences were obtained from 24 karyotyped Parascaris specimens from Poland and 6 German specimens (not karyotyped) and used in phylogenetic analyses with orthologous sequences from GenBank. All karyotyped specimens were identified as P. univalens. In the phylogenetic analysis, they formed very homogenous clusters for all target genes and in a multi-locus analysis. Within this cluster, almost all sequences from GenBank were also included, no matter if they had been assigned to P. univalens or P. equorum. However, a small number of P. univalens ITS and COI sequences originating from donkeys from a single farm in China formed a highly supported sister cluster suggesting that they might represent another Parascaris genotype or species. Our data also strongly suggest that nearly all ITS and COI sequences previously deposited in GenBank and assigned to P. equorum actually represent P. univalens. The fact that significantly different sequences can be found in Parascaris spp. suggests that PCR-based species diagnosis will be possible once molecular markers have been identified for P. equorum from karyotyped specimens.

Establishment of a serodiagnosis system for the detection of Toxocara spp. and Ascaris suum infection in chickens.

Chickens are considered to act as paratenic hosts for agents, Toxocara canis, T. cati and Ascaris suum; which cause ascarid larva migrans syndrome (ascarid LMS) in humans. In addition, they are the definitive host for Ascaridia galli, considered not to be infective for humans. All ascarid parasites can have a high homology of antigenicity, leading to cross-reactivity in serodiagnostic assays. This study was conducted to establish a procedure for the serological detection of those roundworm infections in chickens. Twenty-five male Julia chickens were divided into five groups (n = 5); T. canis-, T. cati-, Ascaris suum- and Ascaridia galli-infected, and an uninfected control group. In Ascaris suum-soluble worm antigen preparation (As-SWAP) ELISA, all infected groups showed an elevation of anti-ascarid antibodies, indicating the usefulness of As-SWAP as a screening antigen for the detection of ascarid infections. For infecting species identification, T. canis-excretory/secretory (Tc-ES) and Ascaris suum-ES (As-ES) antigen ELISA were conducted by serial dilution sera. Toxocara spp.-infected sera showed stronger binding to Tc-ES than As-ES, while Ascaris suum and Ascaridia galli-infected sera bound to As-ES more strongly than Tc-ES. To discriminate between Ascaris suum and Ascaridia galli infection, sera were pre-incubated with Ascaridia galli-SWAP antigen and applied to Tc-ES and As-ES ELISAs. In this pre-adsorbed ES antigen ELISAs, only the Ascaris suum infected group showed positive binding to As-ES, resulting from the adsorption of cross-reactive antibodies in Ascaridia galli-infected sera. Finally, anti-Toxocara specific antibodies were confirmed by Tc-ES western blot (WB). Toxocara spp.-infected sera showed toxocariasis-specific band pattern in Tc-ES WB, while no specific band appeared on any strip incubated with Ascaris suum, Ascaridia galli-infected and uninfected sera. In conclusion, the serodiagnostic assays evaluated in this study are useful for the detection of ascarid infections in chickens.