A retrospective study of the prevalence in equine postmortems of cranial mesenteric arteritis caused by Strongylus vulgaris in Alberta (2010 to 2022).

Background: Strongylus vulgaris is one of the most pathogenic nematodes affecting equids. Larval migration through the cranial mesenteric artery (CMA) with attendant arteritis and thromboembolism can result in fatal non-strangulating intestinal infarction. Once considered a historical disease, recent studies have described the reemergence of this pathogen in several European countries; however, little is known of the current prevalence of S. vulgaris in the Canadian horse population. Objective: To determine the prevalence of active S. vulgaris cranial mesenteric arteritis in horses submitted for postmortem examination to the Diagnostic Services Unit (DSU) at the University of Calgary Faculty of Veterinary Medicine. Animals and procedure: We conducted a retrospective review of all equine postmortem cases submitted to the DSU between July 1, 2010 and June 30, 2022. Over 12 y, 510 horses > 2 mo of age from Alberta were submitted to the DSU for necropsy. Active cases were defined as those with endarteritis and thrombosis in the CMA or its branches. Those cases with only intimal scarring of the CMA were classified as historical. Results: The prevalence of all CMA lesions (both historical and active) over the study period was 17.3% (88/510). Active S. vulgaris cranial mesenteric arteritis was documented in 6.1% (31/510) of equine postmortems and the sequelae of verminous arteritis were the cause of euthanasia or death in 1.5% (8/510) of the cases submitted. Conclusion and clinical relevance: Even after historically intense efforts to eradicate this parasite, the continued effects of S. vulgaris are demonstrated by the results of this study. Strongylus vulgaris should not be regarded as a parasite of the past and verminous arteritis remains an important differential diagnosis for horses in western Canada presenting with mild colic or dull demeanor and anorexia of duration > 24 h. Furthermore, S. vulgaris should be taken into careful consideration when implementing antiparasitic control strategies. Practitioners should remain current on prevention, diagnosis, and treatment of this potentially reemerging and fatal equine disease.

Étude rétrospective de la prévalence lors d'autopsies équines de l'artérite mésentérique crâniale causée par Strongylus vulgaris en Alberta (2010 à 2022). Contexte: Strongylus vulgaris est l'un des nématodes les plus pathogènes affectant les équidés. La migration des larves à travers l'artère mésentérique crâniale (CMA), accompagnée d'artérite et de thromboembolie, peut entraîner un infarctus intestinal non étranglant mortel. Autrefois considérée comme une maladie historique, des études récentes ont décrit la réémergence de cet agent pathogène dans plusieurs pays européens; cependant, on sait peu de choses sur la prévalence actuelle de S. vulgaris dans la population équine canadienne. Objectif: Déterminer la prévalence de l'artérite mésentérique crâniale active à S. vulgaris chez les chevaux soumis pour examen post mortem au Diagnostic Service Unit (DSU), College of Veterinary Medicine, University of Calgary. Animaux et procédure: Nous avons effectué un examen rétrospectif de tous les cas post-mortem d'équidés soumis au DSU entre le 1er juillet 2010 et le 30 juin 2022. Sur 12 ans, 510 chevaux âgés de plus de 2 mois de l'Alberta ont été soumis au DSU pour autopsie. Les cas actifs ont été définis comme ceux présentant une endartérite et une thrombose dans la CMA ou ses branches. Les cas présentant uniquement des cicatrices à l'intima de la CMA ont été classés comme anciens. Résultats: La prévalence de toutes les lésions de CMA (anciennes et actives) au cours de la période d'étude était de 17,3 % (88/510). Une artérite mésentérique crâniale active à S. vulgaris a été documentée dans 6,1 % (31/510) des autopsies équines et les séquelles de l'artérite vermineuse ont été la cause de l'euthanasie ou du décès dans 1,5 % (8/510) des cas soumis. Conclusion et pertinence clinique: Malgré des efforts historiquement intenses pour éradiquer ce parasite, les effets continus de S. vulgaris sont démontrés par les résultats de cette étude. Strongylus vulgaris ne doit pas être considéré comme un parasite du passé et l'artérite vermineuse demeure un diagnostic différentiel important pour les chevaux de l'ouest du Canada présentant des coliques légères ou un comportement abattu et une anorexie de durée > 24 h. De plus, S. vulgaris doit être attentivement pris en compte lors de la mise en œuvre de stratégies de contrôle antiparasitaire. Les praticiens doivent rester informés de la prévention, du diagnostic et du traitement de cette maladie équine potentiellement ré-émergente et mortelle.(Traduit par Dr Serge Messier).

Identification of third stage larvae of strongyles and molecular diagnosis of Strongylus vulgaris in the feces of Thoroughbred horses kept in training centers in Rio de Janeiro, Brazil.

The aims of the present study were to identify strongyles in the feces of Thoroughbred horses based on larval morphology; to detect Strongylus vulgaris using molecular diagnosis and compare results to those of feces culture; and to determine the association between the presence of S. vulgaris with corresponding animal information (age range, gender, and anthelmintic use). Feces of horses kept in six Training Centers in Rio de Janeiro State, that showed the presence of ≥500 eggs per gram of feces (EPG) were subjected to strongyle identification. Of the 520 fecal samples collected, 35 had an EPG ≥ 500. After fecal culture for L3 larvae identification, DNA was extracted, subjected to PCR to amplify the ITS2 region DNA fragment of S. vulgaris, and sequenced. A total of 3500 larvae were analyzed. Most were classified as small strong (99.7%), with an emphasis on the type A subfamily of Cyathostominae. Forms of S. vulgaris only corresponded to 0.2%. In all, 25 samples showed amplified S. vulgaris DNA products and 11 showed nucleotide sequences with high sequence identity. Fecal culture and PCR results showed poor agreement (kappa = 0.105) for S. vulgaris diagnosis. Age, gender, anthelmintic use, and anthelmintic administration interval were not statistically significant. The present study showed the presence of S. vulgaris in the feces of horses kept in Rio de Janeiro Training Centers, mainly seen via PCR, which has emerged as the most effective tool for diagnosis. This study made it possible to identify strongyles that infect horses in the region, emphasizing upon the necessity for constant monitoring of the animals.

Gastrointestinal parasite community structure in horses after the introduction of selective anthelmintic treatment strategies.

A relatively new method to study the species richness and diversity of nematode parasites in grazing animals is to perform deep sequencing on composite samples containing a mixture of parasites. In this work, we compared species composition of strongyles in two groups of horses as a function of egg count and age, based on a DNA barcoding approach. Faecal egg counts and larval cultures were obtained from nearly 300 horses, i.e., domestic horses (n = 167) and trotters (n = 130) sampled nationwide. The second internal transcribed spacer region (ITS2) of strongyle nematodes in the larval cultures was first amplified using barcoded universal primers and then sequenced on the PacBio platform. Subsequently, bioinformatic sequence analysis was performed using SCATA to assign operational taxonomic units (OTU). Finally, species occurrence and composition were assessed using R. ITS2 sequences were found in the majority (89%) of larval samples. Sequencing yielded an average of 140 (26 to 503) reads per sample. The OTUs were assigned to 28 different taxa, of which all but three could be identified as species. The average relative abundance of the seven most abundant species (all Cyathostominae) accounted for 87% of the combined data set. The three species with the highest prevalence in both horse groups were Cyathostomum catinatum, Cylicocyclus nassatus and Cylicostephanus calicatus, and they were frequently found in different combinations with other species regardless of horse group. Interestingly, this result is largely consistent with a previous Swedish study based on morphological analysis of adult worms. In addition, two migratory strongylids (Strongylus vulgaris and S. edentatus) occurred in few domestic horses and trotters. Except for C. minutus and C. nassatus, which decreased with age, and C. catinatum and S. vulgaris, which increased, no specific trends were observed with respect to horse age. Taken together, these results are broadly consistent with data obtained before the introduction of selective targeted treatment in Sweden in 2007. All in all, our results suggest that this treatment strategy has not led to a significant change in strongyle nematode community structure in Swedish horses. The study also confirms that nemabiome analysis in combination with diversity index analysis is an objective method to study strongyle communities in horses.

First microscopic and molecular parasitological survey of Strongylus vulgaris in Brazilian ponies.

The frequency of gastrointestinal parasites with an emphasis on Strongylus vulgaris was investigated among the Brazilian Pony breed kept on farms in the municipality of Teresópolis, state of Rio de Janeiro. Fecal samples were collected in three stud farms: A (n= 22 animals), B (n= 3), and C (n= 2). Fecal samples were subjected to the quantitative Mini-FLOTAC technique, using three different solutions, and to qualitative techniques. The parasite prevalence was found to be 81.4%. Eggs from strongylids were identified in 74% of the ponies. Eggs of Parascaris spp. were detected in 22.7% of the animals, which were all females of farm A. At this locality, mares were kept with their foals in fenced paddocks all the time. The NaCl solution of d = 1.200 g/ml was generally the one that presented the highest frequency of diagnosis of nematode eggs and the highest mean of fecal eggs per gram. The fecal samples were also subjected to the polymerase chain reaction for amplification of DNA from the ITS2 region for Strongylus vulgaris. Twelve samples presented nucleotide sequences for S. vulgaris. In the end, this study revealed the high frequency (96.3%) of S. vulgaris among ponies on farms in Teresópolis, Rio de Janeiro, Brazil.

The complete ITS2 barcoding region for Strongylus vulgaris and Strongylus edentatus.

Gastrointestinal nematode parasites are of major concern for horses, where Strongylus vulgaris is considered the most pathogenic among the Strongylus species. Diagnosis of S. vulgaris infections can be determined with next generation sequencing techniques, which are inherently dependent on reference sequences. The best marker for parasitic nematodes is internal transcribed spacer 2 (ITS2) and we provide the first complete ITS2 sequences from five morphologically identified S. vulgaris and additional sequences from two S. edentatus. These sequences have high similarity to already published partial sequences and amplicon sequence variants (ASV) based on next generation sequencing (NGS). The ITS2 sequences from S. vulgaris matched available partial ITS2 sequences and the full ASVs, whereas the S. edentatus sequence matched another complete sequence. We also compare Sanger sequencing and NGS methods and conclude that the ITS2 variation is better represented with NGS methods. Based on this, we recommend that further sequencing of morphologically identified specimens of various species should be performed with NGS cover the intraspecific variation in the ITS2.

Molecular diagnostics for gastrointestinal helminths in equids: Past, present and future.

This review is aimed to (i) appraise the literature on the use of molecular techniques for the detection, quantification and differentiation of gastrointestinal helminths (GIH) of equids, (ii) identify the knowledge gaps and, (iii) discuss diagnostic prospects in equine parasitology. Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines for systematic reviews, we retrieved 54 studies (horses: 50/54; donkeys and zebras: 4/54) from four databases. Polymerase chain reaction (PCR) was employed in all of the studies whereas PCR amplicons were sequenced in only 18 of them. Other techniques used (including modifications of PCR) were reverse line blot, quantitative (q)PCR, restriction fragment length polymorphism, nested-PCR, PCR-directed next-generation sequencing, Southern blotting, single strand conformation polymorphism, PCR-enzyme linked immunosorbent assay, matrix-assisted laser desorption/ionisation-time of flight and random amplification of polymorphic DNA. Most of the studies (53/54) used nuclear ribosomal RNA (including the internal transcribed spacers, intergenic spacer, 5.8 S, 18 S, 28 S and 12 S) as target loci while cytochrome c oxidase subunit 1 and random genomic regions were targeted in only three and one studies, respectively. Overall, to date, the majority of molecular studies have focused on the diagnosis and identification of GIHs of equids (i.e. species of Anoplocephala, Craterostomum, cyathostomins, Oesophagodontus, Parascaris, Strongylus, Strongyloides and Triodontophorus), with a recent shift towards investigations on anthelmintic resistance and the use of high-throughput nemabiome metabarcoding. With the increasing reports of anthelmintic resistance in equid GIHs, it is crucial to develop and apply techniques such as advanced metabarcoding for surveillance of parasite populations in order to gain detailed insights into their diversity and sustainable control. To the best of our knowledge, this is the first systematic review that evaluates molecular investigations published on the diagnosis and quantification of equid GIHs and provides useful insights into important knowledge gaps and future research directions in equid molecular parasitology.

Differentiating Strongylus vulgaris-associated, non-strangulating intestinal infarctions from idiopathic peritonitis and acute colitis in horses.

BACKGROUND: Horses with non-strangulating intestinal infarction (NSII) are often misdiagnosed with idiopathic peritonitis or acute colitis. Early diagnosis is essential to ensure early surgical intervention and improve survival. METHODS: Clinical and laboratory data from horses admitted to the University of Copenhagen Large Animal Teaching Hospital with NSII, idiopathic peritonitis or acute colitis between 2009 and 2018 were used for univariate comparisons and a multivariable logistic regression model for prediction of NSII. RESULTS: Two hundred and thirty-one horses were included. A multivariable model for the prediction of NSII included gastric reflux (more than 5 L) (odds ratio [OR] 8.7; 95% confidence interval [CI] 2.1-36.2), abnormal findings palpated per rectum (intestinal dilatations/impactions [OR 4.43; 95% CI 1.43-13.38], colon displacements [OR 23.16; 95% CI 5.26-101.97] or intestinal mass [OR 179.7; 95% CI 23.5-1375.5]), white blood cell count (OR 1.2; 95% CI 1.1-1.4), packed cell volume (OR 0.9; 95% CI 0.8-0.9), age (OR 0.9; 95% CI 0.8-1.0) and heart rate (OR 1.1; 95% CI 1.0-1.1). The model had a low false positive rate (5%), but a high false negative rate (50%). LIMITATIONS: Due to the retrospective nature of the study, sample collection was inconsistent, resulting in missing values. CONCLUSION: The model had some capability in predicting NSII. However, the high risk of false negatives means that exploratory laparotomy should be considered in horses with peritonitis of unknown aetiology in areas where Strongylus vulgaris is prevalent and occurrence of idiopathic peritonitis is low.

Identifying 3rd larval stages of common strongylid and non-strongylid nematodes (class: Nematoda) infecting Egyptian equines based on morphometric analysis.

Strongylid and non-strongylid nematodes are one of the most important parasites infecting equines. The traditional method to identify these nematodes is through coproscopy and fecal culture. Because of the scarcity of data published in Egypt discussing the morphometric features of infective 3rd larvae of these nematodes, this study aims to provide a morphometric key for L3 of common strongylid and non-strongylid nematodes infecting Egyptian equines. For this reason, we cultured fecal samples containing GINs eggs and 3rd larval stages were identified based on their morphology (i.e., shape and number of intestinal cells (IC), shape of the esophagus, and shape of the tail sheath) in addition to computing their dimensions (i.e., length of larvae with sheath, length of the esophagus, length of intestinal cells, and body breadth). We identified 3rd larval stages of four strongylid nematodes (Cyathostomum sensu lato, Strongylus vulgaris, Strongylus equinus, and Strongylus edentatus) as well as two non-strongylid nematodes (Strongyloides westeri, and Trichostrongylus axei). Statistically, our results revealed significant differences in terms of total length, body width, esophagus length, and gut length among 3rd larvae identified in the current study. The combination of both morphological and metric keys will allow the better identification of common strongylid and non-strongylid nematodes infecting equines.

Clinical features and treatment response to differentiate idiopathic peritonitis from non-strangulating intestinal infarction of the pelvic flexure associated with Strongylus vulgaris infection in the horse.

BACKGROUND: Peritonitis in horses secondary to non-strangulating infarction (NSII) has a guarded prognosis, even after intestinal resection. In contrast, horses with idiopathic peritonitis respond well to medical treatment. Affected horses in both cases often show signs of both colic and systemic inflammation, but early diagnosis is crucial for optimal treatment and an accurate prognosis. One cause of NSII is thrombus formation secondary to Strongylus vulgaris larval migration. There has been a documented increase in S. vulgaris prevalence in Sweden since the implementation of selective anthelmintic treatment in 2007, which subsequently could result in a rise in NSII cases. In a retrospective clinical study, medical records from cases diagnosed with NSII of the pelvic flexure or idiopathic peritonitis from three equine referral hospitals in Sweden during 2017-2020 were reviewed. Information including demographic data, relevant medical history, and clinical- and laboratory parameters were obtained from patient records. To facilitate the differentiation between cases of idiopathic peritonitis and cases with confirmed NSII of the pelvic flexure, the aim of the study was to compare clinical and laboratory parameters, clinical progression and initial response to antimicrobial treatment. A secondary aim was to compare survival-rates. RESULTS: Horses with NSII (n = 20) were significantly more likely to present during the winter months with a poorer response to medical treatment within 48 h. Cases of idiopathic peritonitis (n = 107) had a 100% survival rate with medical treatment, although one case required surgical correction of a colon displacement. In comparison, all confirmed NSII cases were non-responsive to antimicrobial treatment, with a survival rate to discharge of 50% after colon resection. Specific rectal findings and peripheral blood neutropenia were strongly associated with NSII. CONCLUSIONS: In Sweden, idiopathic peritonitis cases still predominate over S. vulgaris associated NSII cases and have an excellent survival rate with antimicrobial treatment. However, horses presenting with septic peritonitis during the winter months with a palpable rectal mass and displaying fever and colic signs beyond 48 h of medical treatment are likely to suffer from NSII of the pelvic flexure and should be considered for abdominal surgery.

Parasite dynamics in untreated horses through one calendar year.

BACKGROUND: Horses are host to a plethora of parasites. Knowledge of the seasonality of parasite egg shedding and transmission is important for constructing parasite control programs. However, studies describing these patterns are sparse, and have largely been conducted only in the United Kingdom. This study evaluated strongylid egg shedding patterns and transmission dynamics of Strongylus vulgaris in naturally infected and untreated mares and foals through one calendar year in Kentucky, USA. The study also investigated the existence of a peri-parturient rise (PPR) in strongylid egg counts in foaling mares and collected information about Strongyloides westeri and Parascaris spp. in the foals. METHODS: This study was conducted from January to December 2018. A herd of 18 mares, one stallion, and 14 foals born in 2018 were followed throughout the year. Sera and feces were collected biweekly from all horses, and worm burdens enumerated in 13 foals at necropsy. An S. vulgaris ELISA antibody test was run on all serum samples. Fecal egg counts were determined for all horses, and coproculture and qPCR assay were employed to test for the presence of S. vulgaris in the mature horses. Data were analyzed using the proc glimmix procedure in the SAS 9.4 software program. RESULTS: We found a general lack of seasonality in strongylid egg shedding throughout the year among the mature horses, and no PPR was demonstrated. Shedding of S. vulgaris eggs displayed a higher abundance during the spring, but findings were variable and not statistically significant. Anti-S. vulgaris antibody concentrations did not display significant fluctuations in the mature horses, but evidence of passive transfer of antibodies to the foals was demonstrated, and foals assumed their own production of antibodies starting at approximately 20 weeks of age. Overall, colts shed higher numbers of strongylid, ascarid, and S. westeri eggs than fillies. CONCLUSIONS: This study demonstrated a lack of seasonality in strongylid egg shedding for the study population, which is in stark contrast to previous studies conducted elsewhere. This strongly suggests that more studies should be done investigating these patterns under different climatic conditions.

Reviving a tradition: The Development of Strongylus vulgaris in larval culture.

All horses are susceptible to the equine gastrointestinal parasite, Strongylus vulgaris, which is known to cause significant disease and death. The parasite undergoes development from the egg through the first (L1), second (L2) and third (L3) larval stages outside the horse. The L3 is the infective stage. The universally available technique for detection of S. vulgaris larvae is the larval culture method. This requires a 10-14 day culture period to induce development from egg to L3, followed by Baermannization and identification of the L3s to genus and/or species. It is unknown if the culture duration is necessary or ideal for S. vulgaris identification. The purpose of this study was to perform daily examinations of known S. vulgaris positive fecal samples in coproculture. Fresh feces were collected from a horse known to be shedding S. vulgaris eggs. A total of 140 cultures were set up using 10 g of feces. Cultures remained at room temperature and moistened every other day. Every day, 10 samples were examined, and all larvae were identified to stage, genus/species, and enumerated. Throughout the study, L1, L2, and L3 stages were observed, and S. vulgaris, Strongylus edentatus, Triodontophorus spp., and cyathostomin L3s were identified. Third stage larvae were observed on Day 5, and the mean number of L3s significantly increased on Day 10 (P < .001), and declined thereafter. Strongylus vulgaris was first observed on Day 6 with a mean count of 4.1 (95 % CI: 1.1, 7.1) S. vulgaris larvae, accounting for 4.1 % (95 % CI:1.8, 7) of the total L3s observed. The number of S. vulgaris larvae was significantly higher on Day 10 with a mean of 156.8 (95 % CI: 120.7, 192.9) S. vulgaris larvae (P < .001), and the proportion was also significantly higher with S. vulgaris comprising 50 % (95 % CI: 45.9, 54.8) (P = .006) of the total larvae. However, after 10 days, the mean number of S. vulgaris larvae declined, as did the proportion of S. vulgaris larvae compared to the total number of larvae. Using the described methods, it is possible to identify S. vulgaris as early as 6 days, and the optimal period is 10 days to detect the maximum number of S. vulgaris.

Oxidative stress in Strongylus spp. infected donkeys treated with piperazine citrate versus doramectin.

Background: Parasitic infection is one of the main problems in equidae, particularly donkeys. Aim: This study evaluated the oxidative stress in donkeys infected with Strongylus spp by determining the correlation between antioxidants levels; malondialdehyde (MDA), total antioxidant capacity (TAC), and the severity of parasitic infection. It also compared the therapeutic efficacy of piperazine citrate as an oral anthelmintic drug and Doramectin as an injectable one. Methods: The study was conducted on 40 donkeys naturally infected with Strongylus spp. These donkeys were divided into two groups (20 donkeys each) according to treatment; One group was treated with piperazine citrate (PipTG) and the other with doramectin (DoraTG). Thorough clinical examination, hematological, biochemical, and parasitological assays were performed before (Day 0) and after treatment (Days 7, 14, 21, and 28). All data were statistically analyzed by independent-sample t-test or paired t-test. Results: In both groups, mean values of MDA were significantly reduced, while those of TAC were significantly elevated after treatment on days 7, 14, 21, and 28. These significant changes were reported after treatment between PipTG and DoraTG in favor of DoraTG. Serum concentrations of MDA were significantly reduced, while those of TAC were significantly elevated for DoraTG treatment group when their values were compared with those of PipTG either on days 7, 14, 21, or 28. Significant correlations were reported in PipTG and DoraTG. Negative significant correlations were reported between fecal egg count (FEC) and each of whole blood picture indices (RBCS, Hb, and PCV), serum TAC and faecal egg count reduction percentage FECR%. A positive correlation was seen between FEC and MDA. MDA exhibited a negative correlation with both blood picture and TAC; hence, TAC was positively correlated with these blood picture indices in both PipTG and DoraTG. In PipTG, anthelmintic resistance (R) was present on days 7 and 14, while it was suspected (S) at day 21 then it was absent (N) at day 28. In DoraTG, anthelmintic resistance was suspected (S) on day 7, then it became absent (N) on days 14, 21, and 28 post therapy. Conclusion: The immunological status of the infected donkeys had greatly improved after treatment. The therapeutic efficacy of injectable doramectin was more efficient than that of oral piperazine citrate in Strongylus spp. infected donkeys.

Diagnosing Strongylus vulgaris in pooled fecal samples.

Strongylus vulgaris is the most pathogenic intestinal helminth parasite infecting horses. The migrating larvae in the mesenteric blood vessels can cause non-strangulating intestinal infarctions, which have a guarded prognosis for survival. Infections are typically diagnosed by coproculture, but a PCR test is available in some countries. While it is ideal to test horses individually, many veterinarians and clients wish to pool samples to reduce workload and cost of the diagnostic method. The purpose of this study was to determine if pooling of fecal samples would negatively impact diagnostic performance of the coproculture and the PCR for determination of S. vulgaris infection. Ten horses with strongylid eggs per gram (EPG) >500 and confirmed as either S. vulgaris positive or negative were selected as fecal donors. Eight pools with feces from five horses were created with 0%, 10 %, 20 %, 30 %, 40 %, 50 %, 80 %, and 100 % S. vulgaris positive feces. From each pool, 20 subsamples of 10 g each were collected and analyzed. Half of these samples were set up for coproculture and the other half for PCR. All pools containing 50 % or greater S. vulgaris positive feces were detected positive by both PCR and coproculture. In the pools with less than 50 % S. vulgaris positive feces, the PCR detected 33 positive samples compared to 24 with the coproculture. Three samples from the 0% pool were detected as low-level PCR positives, but this could be due to contamination. These results indicate that diagnosing S. vulgaris on pooled samples is reliable, when at least 50 % of the feces in a pool are from S. vulgaris positive animals. Since S. vulgaris remains relatively rare in managed horses, however, some diagnostic sensitivity is expected to be lost with a pooled sample screening approach. Nonetheless, pooled sample screening on farms could still be considered useful under some circumstances, and the PCR generally performed better at the lower proportions of S. vulgaris positive feces.

Helminth infections in Italian donkeys: Strongylus vulgaris more common than Dictyocaulus arnfieldi.

Donkeys have been used as working animals for transport and farm activities worldwide. Recently, in European countries, there has been an increasing interest in donkeys due to their use as pets, onotherapy or milk production. During 2014-2016, a countrywide survey was conducted to determine prevalence and risk factors of principal helminth infections in 1775 donkeys in 77 Italian farms. A questionnaire on management and parasite control practices was filled out for each farm. Faecal samples were examined using a modified McMaster technique, a centrifugation/flotation method and a sedimentation technique. Pooled coprocultures were performed for differentiation of strongylid eggs. Strongyles were the most common parasites detected (84.9%), followed by Dictyocaulus arnfieldi (6.9%), Oxyuris equi (5.8%), Parascaris spp. (3.6%), Anoplocephala spp. (1.0%), Strongyloides westeri (0.3%). Coprocultures revealed an omnipresence of cyathostomins (100%), followed by Strongylus vulgaris (31.0%), Poteriostomum spp. (25.0%), Triodontophorus spp. (9.0%), Strongylus edentatus (7.0%), Strongylus equinus (5.0%). Logistic regression analysis identified breed, co-pasture with horses, living area, herd size and number of treatments as significantly associated with strongyles. Sex, age, living area and herd size were significantly associated with Parascaris spp. Dictyocaulus arnfieldi was significantly associated with sex, grass, co-pasture with horses, living area and herd size. Strongylus vulgaris was significantly associated with living area and herd size. The mean number of anthelmintic treatments/year was 1.4; most of the donkeys (71.8%) were dewormed using an ivermectin drug. It is important to design parasite programs to specifically address both D. arnfieldi and S. vulgaris in donkeys, and this is especially important if donkeys co-graze with horses.

Cytokine responses to various larval stages of equine strongyles and modulatory effects of the adjuvant G3 in vitro.

AIMS: To generate different larval stages of Strongylus vulgaris and to study cytokine responses in cultures of eqPBMC exposed to defined larval stages of S. vulgaris and cyathostomins with the aim to understand the early immune reaction to these parasites. METHODS AND RESULTS: EqPBMC were exposed to S. vulgaris larvae (L3, exsheated L3 and L4) and cyathostomin L3 and analysed for cytokine gene expression. Procedures for decontamination, culturing and attenuation of larvae were established. Transcription of IL-4, IL-5 and IL-13 was induced by both S. vulgaris and cyathostomin L3. Moulting of S. vulgaris from L3 to L4 stage was accompanied by a shift to high expression of IL-5 and IL-9 (exsheated L3 and L4) and IFN-γ (L4 only). In parallel, the adjuvant G3 modified the cytokine profile induced by both parasites by reducing the expression of IL-4, IL-5 and IL-10 while concomitantly enhancing the expression of IFN-γ. CONCLUSION: The L4 stage of S. vulgaris generated a cytokine profile different from that induced by the earlier L3 stage of S. vulgaris and cyathostomins. This diversity depending on the life cycle stage will have implications for the choice of antigen and adjuvant in future vaccine design.

Strongylus vulgaris: Infection rate and molecular characterization from naturally infected donkeys at Sadat City, Egypt.

Strongylus vulgaris has high pathogenicity to equines. It causes aneurysm and thrombosis in the arteries particularly an anterior mesenteric artery, that is fatal to equines. In this study, we aimed to diagnose microscopically the natural infection of donkeys with Strongylus vulgaris from Sadat City, Minoufiya Governorate, Egypt. Fecal egg culture was used after the diagnosis of strongyle eggs to identify the species. Hematological and biochemical parameters were assessed. Adult worms were collected after post mortem examination of the infected animal. The sequence of ITS-2 was used to confirm the species of the parasite. The infection rate was 15.85% using the microscopical examination. The larval culture confirmed the infection with strongyle eggs as Strongylus vulgaris larvae. The sequence of ITS-2 was highly identical (about 95%) to sequences from Germany, China, and Turkey and occurred in the same genetic clade with the sequence from Germany. In conclusion, the study presented the diagnosis, the changes in the hematological and biochemical parameters in the infected animals, and the genetic characterization of Strongylus vulgaris from Sadat City, Minoufiya Governorate Egypt for the first time.

Eficacy of ivermectin, moxidectin and febendazole in equine in Brazil.

The objectives of the present study were to evaluate the efficacy of three anthelmintic drugs, i.e. fenbendazole, ivermectin and moxidectin; to determine the genera and species of the most abundant strongyles; and to investigate parasite control measures used in herds of horses in the western region of the state of São Paulo, Brazil. This study was onducted between February and December 2013 on 10 farms in this region. Coprological evaluations were conducted for counting the numbers of eggs per gram of feces (EPG) and coprocultures were made in order to identify third-stage larvae (L3) of strongyles. Parasite control measures were investigated by surveying each farm's management and characteristics. A fecal egg count reduction test (FECRT) was used to determine the anthelmintic efficacy based on EPG count results before and after treatment. The FECRT showed that anthelmintic resistance to fenbendazole was present and that ivermectin and moxidectin had high effectiveness in the western region of the state of São Paulo. Identification of L3 revealed that there was high prevalence of cyathostomins among strongyle larvae. The highest prevalence of Strongylus vulgaris occurred on farms where the intervals between deworming were long. The questionnaire applied showed that ivermectin was the drug most used among these farmers.

Compilation of 29 years of postmortem examinations identifies major shifts in equine parasite prevalence from 2000 onwards.

Horses are infected by a wide range of parasite species that form complex communities. Parasite control imposes significant constraints on parasite communities whose monitoring remains, however, difficult to track through time. Postmortem examination is a reliable method to quantify parasite communities. Here, we compiled 1,673 necropsy reports accumulated over 29 years, in the reference necropsy centre from Normandy (France). The burden of non-strongylid species was quantified and the presence of strongylid species was noted. Details of horse deworming history and the cause of death were registered. Building on these data, we investigated the temporal trend in non-strongylid epidemiology and we determined the contribution of parasites to the deaths of horses throughout the study period. Data analyses revealed the seasonal variations of non-strongylid parasite abundance and reduced worm burden in race horses. Beyond these observations, we found a shift in the species responsible for fatal parasitic infection from the year 2000 onward, whereby fatal cyathostominosis and Parascaris spp. infection have replaced cases of death caused by Strongylus vulgaris and tapeworms. A concomitant break in the temporal trend of parasite species prevalence was also found within a 10 year window (1998-2007) that has seen the rise of Parascaris spp. and the decline of both Gasterophilus spp. and tapeworms. A few cases of parasite persistence following deworming were identified, which all occurred after 2000. Altogether, these findings provide insights into major shifts in non-strongylid parasite prevalence and abundance over the last 29 years. They also underscore the critical importance of Parascaris spp. in young equids.

Anthelmintic activity of crude powder and crude aqueous extract of Trachyspermum ammi on gastrointestinal nematodes in donkey (Equus asinus): An in vivo study.

ETHNOPHARMACOLOGICAL RELEVANCE: Trachyspermum ammi has been used traditionally as a popular ethnobotanical plant in human and animal parasitic infestations. Few scientific studies have been conducted on in vitro anthelmintic activity of T. ammi against various helminths and there is no study on its in vivo/in vitro anthelmintic properties against equine helminths. AIM OF THE STUDY: The present study aimed to evaluate in vivo anthelmintic activity of crude powder (CP) and crude aqueous extract (CAE) of T. ammi seeds against gastrointestinal nematodes in the donkey. MATERIALS AND METHODS: Thirty donkeys, naturally and severely infected with Strongyle-type nematodes, were randomly divided into six groups and administered orally with graded doses of CP and CAE at single dose as follow: Two groups were treated with 1 and 3 gkg-1 body weight of CP, two groups were treated with CAE at the equivalent dose rate 1 and 3 gkg-1 bw of CP and one group was treated with 200 µgkg-1 bw of Ivermectin 0.2% (Positive Control). One group received no medication (Negative Control). Efficacy was determined by faecal egg count reduction (FECR) test. The faecal samples were taken from the donkeys on days 7, 14 and 28 post-treatments (PT) and the egg per gram (EPG) of faeces was determined for each animal. FECR for donkeys treated with CP, CAE, and Ivermectin 0.2% was calculated on days 7, 14, and 28 PT. In the present study, CAE was analyzed using gas chromatography-mass spectrometry (GC-MS) to determine CAE chemical components. High-performance liquid chromatography (HPLC) using a marker (thymol) was applied to confirm the existence of thymol as an active principle. RESULTS: Based on the FECR findings, the maximum FECR (71.2%) was observed in the group treated with CAE3 on day 14 PT, while it was 66.2% for CP3 on the same day. The effects of different treatments (P < 0.001), time (P < 0.001) and their interactions (P < 0.05) on EPG showed a significant difference. A dose-dependent response in the anthelmintic activity of CP and CAE was observed. GC-MS analysis indicated that thymol was the major component of CAE (Area = 45.01%) with the respective peak at retention times (Rt) 10.41 min. On HPLC analysis of CAE, thymol was found to be the major constituent of CAE (Area = 58.76%) with the same Rt at 13.233 min. CONCLUSION: The present study indicated that both CP and CAE of T. ammi had the dose-dependent anthelmintic effects. Due to the development of drug resistance against synthetic anti-helminthics, T. ammi seems to be a promising alternative for the treatment and prevention of helminths in livestock.

Fatal Peritoneal Migration of Strongylus edentatus in a Foal.

A 7-month-old female mixed breed foal with a 2-day history of recumbency and inability to open its mouth convulsed acutely and died and was submitted for necropsy examination. The foal was thin and large patches of haemorrhage were present throughout the peritoneal wall, the diaphragmatic surfaces and the retroperitoneum. Numerous nematode larvae were visible on the serosal surfaces and penetrated and embedded into the subserosa associated with the haemorrhages. The dorsal portion of the abdominal diaphragm had a partial tear and large numbers of nematodes were within the muscle fibres. Histologically, the larvae had a smooth cuticle, polymyarian/coelomyarian musculature and multinucleated intestinal cells, and were typically surrounded by haemorrhage, neutrophils, dense fibrovascular connective tissue and rare multinucleated giant cells. Parasitological examination identified the larvae as Strongylus edentatus based on the morphology of the buccal capsule. Additionally, there was severe muscle necrosis of the tongue and liver tissue analysis detected selenium deficiency. S. edentatus infections are uncommon in California, USA, and are typically non-lethal. In this case, the selenium deficiency may have led to immunosuppression, resulting in the hyperinfection with S. edentatus, and to the muscle damage and tear of the diaphragm. Although ivermectin treatment was indicated in the history, inadequate deworming or anthelmintic resistance may have played a role in the severity of infection.