Reduction of egg reappearance period of cyathostomins in naturally infected horses after increasing doses of ivermectin in Brazil: a grim picture for sustainable parasite control.

Cyathostomins are the largest group of parasites in horses that can be controlled by ivermectin (IVM). This study aimed to run a four-dose titration trial of IVM in 28 naturally infected Thoroughbred yearlings. The local Strongyle population had been recorded to be resistant to IVM (200 µg/kg). The parasite fecal egg count (FEC) was performed to investigate the egg reappearance period (ERP) of two and five weeks (w2pt and w5pt) after IVM treatment. FEC was > 1000 on day zero for all groups. Although 100% FEC reduction was reported at w2pt for all concentrations, the FEC at w5pt revealed < 83% efficacy. This study reports the reduction of ERP using the label dose as well as 300, and 400 µg/kg (double dose) of IVM. The protocol allowed IVM to significantly suppress FEC w2pt although not eliminating adult worms, failing to guarantee an extension of its protection period over 8 weeks. Moreover, the FEC at w5pt possibly means the infection was not cleared, and worms reestablished egg laying. We raised the possibility of withdrawing IVM of control programs when the drug has less than 80% FEC reduction at w5pt.

Molecular identification and genetic variability of equine and bovine ocular setariasis in india: molecular profiling by mitochondrial genes.

BACKGROUND: Ocular setariasis is an ectopic infection caused by a parasite under the genus Setaria. Adult worms belong to the Setariidae family and typically reside in the peritoneal cavity of ungulates. However, immature forms of these species may aberrantly migrate to the eyes of cattle, buffalo, goats, horses and several other hosts, leading to corneal opacity and blindness. Here, we have distinguished the Setaria digitata collected from both equine and buffalo hosts based on the morphology, molecular profiling of mitochondrial cytochrome c oxidase subunit 1 (Cox1), cytochrome c oxidase subunit 3 (Cox3) and, Nicotinamide Adenine Dinucleotide dehydrogenase subunit 1 (NAD1) genes. METHODS AND RESULTS: A single filarial worm was collected from the eye of one equine and one bovine host. These worms were then processed for morphological examination and DNA isolation. Cox1, Cox3 and NAD1 genes were amplified using specific primers and subjected to custom sequencing. The sequences were then used for multiple sequence alignment, assessment of entropy, similarity and haplotype diversity analysis. Key morphological features confirmed the worms collected were male and female Setaria digitata from equine and buffalo hosts, respectively. Cox1, Cox3 and NAD1 gene sequence analysis showed a close association of S.digitata Indian isolates with its counterparts from Sri Lanka and China isolates. CONCLUSION: The phylogram of bovine S. digitata sequences shows a close relationship to other equine S. digitata sequences, indicating the need for further in-depth studies on the prevalence of infection across various host species and intermediate hosts. Although the sequence results suggest that S. digitata is likely the causative agent of ocular setariasis in India, additional samples are needed to confirm this conclusion. Comprehensive analysis of the transcriptome and proteome of S. digitata from both bovine and equine hosts is necessary to explore variations in host-parasite interactions. These findings will aid in future parasite identification, investigations into vector prevalence in India, and the development of control measures against ocular setariasis.

First report on seroprevalence of Toxoplasma gondii infection in zoo animals in Algeria.

Toxoplasma gondii is one of the world's most widespread polyxenic protozoan parasites that affect all warm-blooded animals, including humans. This survey aims to study, for the first time in Algeria, the seroprevalence of Toxoplasma infection in zoo animals. The study included eight animal species of which 54 serum samples were collected from 30 Australian goats (Capra hircus), four bulls (Bos taurus), one dromedary (Camelus dromedarius), three cuffed sheep (Ammotragus lervia), seven donkeys (Equus asinus), one pony (Equus ferus), four bearded horses (Equus ferus caballus) and four rabbits (Oryctolagus cuniculus). The presence of antibodies to T. gondii was determined using the ID Screen® Toxoplasmosis Indirect Multispecies ELISA kit (IDVet, Grabels, France). A total of 8/54 (14.8%) samples were seropositive, including 5/28 (17.9%) males and 3/26 (11.5%) females. The seroprevalence was 6.7%, 50%, 25% and 75% in Capra hircus, Bos Taurus, Equus ferus caballus, and Oryctolagus cuniculus, respectively. No cases were observed in Camelus dromedarius, Ammotragus lervia, Equus asinus, and Equus ferus. This study indicates, for the first time in Algeria, the seroprevalence of T. gondii in zoo animals.

Tabanidae (Diptera) collected on horses in a Cerrado biome in the state of Tocantins, Brazil.

Tabanidae (Diptera), popularly known as horse flies, is an important vector group. This is the first study to ascertain the abundance and diversity of horse flies in horses at the cerrado biome of the state of Tocantins, Brazil. Collecting took place in typical Cerrado, and sampling occurred in the dry and rainy seasons. The horseflies were collected from horses using an entomological net. A total of 249 individuals were collected and spread over 25 species. The prevalent species were Stypommisa aripuana (25.8%) and Catachlorops rufescens (6.4%), in the dry period, and Fidena lissorhina (22.5%), Tabanus occidentalis var. dorsovittatus (10%) and Poeciloderas quadripunctatus (6.4%), in the rainy season. The results suggest that tabanids attack horses throughout the dry and rainy seasons, posing a constant threat to their health in the Cerrado of Tocantins.

Seroprevalence of Toxoplasma gondii and Neospora spp. in horse population of Tehran, Iran.

Neospora spp. and Toxoplasma gondii are two closely related protozoan parasites that are widely distributed throughout the world. Horses can act as intermediate hosts for both parasites and can acquire disease. Blood samples were taken from 487 clinically healthy horses from 17 different mechanized stables in Tehran, the capital of Iran, during September and November of 2022. IFAT and ELISA were employed to detect antibodies directed against Neospora spp. and T. gondii. The anti-N. caninum antibodies were detected in 52 of the horses (10.67%) based on IFAT and in 86 of the 487 horses (17.65%) based on the ELISA test. Also, antibodies against T. gondii were detected in 41 horses (8.42%) based on IFAT and in 63 of 487 horses (12.94%) based on the ELISA test. Also, in 6 of the horses (1.23%) based on IFAT and in 13 of the 487 horses (2.67%) based on the ELISA test, double positivity suggested co-infection with both parasites. Gender, age groups, and the presence of dogs for neosporosis, and age groups and the presence of cats for toxoplasmosis, could be considered factors having an influence on the seroprevalences (P < 0.05). The results proved the importance of the urgent implementation of stringent regulatory measures to prevent and control the spread of these parasites.

The first study of the prevalence and genetic diversity of Theileria equi and Babesia caballi in horses in Russia.

Equine piroplasmosis (EP) is a global worldwide infection, which can lead to the death of animals. Despite the causative agents of EP being well studied, there are no data on the distribution and genetic characteristics of EP agents in any region of Russia. In this study, blood samples from 750 horses from Novosibirsk province, Irkutsk province, and Altai region of Russian Siberia were examined for the presence of EP agents. Theileria equi and Babesia caballi were detected in all examined regions, with mean prevalence rates of 60.4% and 7.2%, respectively. The identified pathogens were genetically characterized by the 18S rRNA gene. The determined T. equi sequences were highly conserved and belonged to genotypes A and E, with genotype E being found in 88.6% of genotyped samples. In contrast to T. equi, B. caballi sequences were genetically diverse. Seven sequence variants of B. caballi were identified, and only two of them matched known sequences from the GenBank database. The determined B. caballi sequences belonged to four distinct branches within genotype A. Mixed infections with several variants of B. caballi or with T. equi and B. caballi were common. The conducted phylogenetic analysis based on all available B. caballi sequences of the 18S rRNA gene (> 900 bp) from GenBank and from this study first demonstrated the presence of five monophyletic clusters within genotype A and three clusters within genotype B. Thus, the genetic study of B. caballi from Siberia has significantly expanded the data on the genetic diversity of this pathogen.

Leishmania spp. in equids and their potential vectors in endemic areas of canine leishmaniasis.

Equids may be infected by zoonotic Leishmania spp., including Leishmania infantum, in regions where canine leishmaniasis (CanL) is endemic, and Leishmania martiniquensis, which has been reported in horses from Central Europe. This study was designed to evaluate the occurrence of both Leishmania spp. among equids living in CanL endemic areas of Italy, as well as to identify dipteran vectors from the same habitats. From March to October 2023, blood, serum and tissue samples from skin lesions were collected from equids (n = 98; n = 56 donkeys and n = 42 horses) living in Italy, as well as sand flies and biting midges. Blood samples (n = 98) and skin lesions (n = 56) were tested for Leishmania spp. by conventional and real time PCRs and sera were tested by immunofluorescence antibody tests (IFAT) for both L. infantum and L. martiniquensis. Insects were morphologically identified, and female specimens (n = 268 sand flies, n = 7 biting midges) analyzed for Leishmania DNA, as well as engorged sand flies (n = 16) for blood-meal detection. Two animals with skin lesions (i.e., one donkey and one horse) scored positive for Leishmania spp. DNA, and 19 animals (i.e., 19.4%; n = 13 donkeys and n = 6 horses) were seropositive for L. infantum, with five of them also for L. martiniquensis. Most seropositive animals had no dermatological lesions (i.e., 68.4%) while both animals molecularly positive for Leishmania spp. scored seronegative. Of the 356 sand flies collected, 12 females (i.e., n = 8 Sergentomyia minuta; n = 3 Phlebotomus perniciosus, n = 1 Phlebotomus perfiliewi) were positive for Leishmania spp. DNA, and one out of seven biting midges collected was DNA-positive for L. infantum. Moreover, engorged sand flies scored positive for human and equine DNA. Data suggest that equids living in CanL endemic areas are exposed to Leishmania spp., but their role in the circulation of the parasite needs further investigations.

Detection of tick-borne pathogens in Rhipicephalus bursa ticks collected from the autochthonous Garrano breed of horses in Portugal.

The Garrano is a semi-feral horse breed native to several mountains in the northern Iberian Peninsula. Despite being endangered, this unique breed of pony has managed to survive in the wild and continues to be selectively bred, highlighting their remarkable resilience and adaptability to harsh environments. Wildlife plays a critical role in the survival of tick vectors in their natural habitats and the transfer of tick-borne pathogens, as they can serve as reservoir hosts for many agents and amplifiers for these vectors. The semi-feral lifestyle of the Garrano horses makes them particularly vulnerable to exposure to numerous tick species throughout the year. Therefore, the aim of this study was to investigate the occurrence of Anaplasma, Ehrlichia, Babesia, Theileria, and spotted fever rickettsiae in the Garrano horse ticks to obtain a knowledge of circulating agents in this host population. The collected ticks (n = 455) were identified as Rhipicephalus bursa. DNA specimens were organized in pools of 5 ticks, for molecular screening. Pools PCR results confirmed the presence of Candidatus Rickettsia barbariae (n = 12 for the ompB gene, n = 11 for the ompA gene and n = 6 for the gltA gene), Babesia bigemina (n = 1), Babesia caballi (n = 3), Theileria equi (n = 15) and Theileria haneyi (n = 1).These results confirm the circulation of an emerging rickettsial spotted fever group member, Candidatus R. barbariae, in R. bursa ticks. Our findings demonstrated that Candidatus R. barbariae co-circulates with B. bigemina and T. equi, which are vectored by R. bursa. We are reporting for the first time, the detection of T. haneyi among R. bursa ticks feeding in the Garrano horses in Portugal. Surveillance studies for tick-borne infections are essential to provide information that can facilitate the implementation of preventive and control strategies.

Strongylids of Domestic Horses in Eastern Slovakia: Species Diversity and Evaluation of Particular Factors Affecting Strongylid Communities.

PURPOSE: This study aimed to examine the species diversity and structure of the strongylid community in domestic horses in Eastern Slovakia. Also, an analysis of the impact of age, sex, and collection location factors on the strongyid communities was performed. METHODS: Twenty-seven horses 1.5-21 years old from two farms in eastern Slovakia with different horse-management conditions were studied. Strongylids were collected after horse treatments with Noromectin (0.2 mg ivermectin); 66,170 specimens were collected and identified. Faecal egg count reduction test (FECRT) was performed following fenbendazole (FBZ) and ivermectin (IVM) treatments. RESULTS: Twenty-four strongylid species were found; horses were infected with 6 to 16 (average = 11.7) species. Six cyathostomin species (Cylicocyclus nassatus, Cyathostomum catinatum, C. pateratum, Cylicostephanus longibursatus, C. goldi, C. calicatus) were the most prevalent; C. catinatum was the dominant species in both farms (Berger-Parker index 0.34 and 0.42). The structure of the strongylid community was multimodal with dominant, subdominant, background, and rare species. The Mantel test showed that horse age and sex did not significantly affect the nematode infracommunity composition (p > 0.05), while the impact of the collection location (farm) was significant (p = 0.03). Additionally, C. longibursatus was identified as the species contributing significantly to the observed farm differences. Strong resistance to FBZ was documented on both farms (FECRT was 36.4% and 22.7%); IVM resistance was not observed (FECRT = 100%). CONCLUSION: This study presents the first report on the strongylids parasitizing domestic horses in Eastern Slovakia and gives basic information for further studies of horse parasites and their control in the region.

Effect of cattle and horse feces storage methods on Nematode egg viability and sensitivity for egg hatch test.

The aim of the present study was to validate methods of stool sample conservation for the egg hatch test (EHT). This study involved the use of a bovine naturally infected predominantly by Cooperia spp. and one equine naturally infected predominantly by cyathostomins characterized as susceptible to benzimidazoles in the EHT. Fecal samples were submitted to three treatments: aerobic methods (anaerobic storage in plastic bottles, anaerobic storage in vacuum-sealed bags or aerobic storage in plastic bags), under two temperature conditions (room temperature and refrigeration) analyzed at four different assessment times (48, 72, 96 and 120 h). As the standard test, an assay was also performed within 3 h. The tests were performed in triplicate for each drug concentration and with three experimental repetitions at one-week intervals. Two criteria were used for the storage methods: hatchability in the negative control group and sensitivity of the eggs to thiabendazole, comparing the EC50 and 95% confidence interval for each treatment to those of the standard test and the other repetitions. Bovine samples can be stored for up to 96 h and refrigerated vacuum storage can be used, ensuring hatchability of the negative control and sensitivity of the eggs to thiabendazole. For equine samples, no forms of storage were indicated due to the variation among the repetitions and the reduction in the sensitivity of the eggs to thiabendazole, which could result in a false positive detection of resistance.

Welfare assessment of Thoroughbred horses naturally infected with gastrointestinal parasites in Southern Brazil: Quantifying the host-parasite relationship.

Horse welfare assessment (HWA) does not account for individual or herd parasite infection. This study investigated the connection between HWA and individual parasite fecal egg count (FEC) in 90 Thoroughbred horses. All horses were naturally infected with gastrointestinal parasites and were evaluated for individual welfare indicators and FEC monthly, for 12 months. Horses were divided into three groups of 30 mares, 30 foals aged between 13 and 16 months (G2013), and 30 foals aged between two months and one year (G2014). A horse welfare protocol was developed and 1024 assessments were carried out by five trained assessors. FEC ranged from 0 to 5,760 with 98.8 % showing small strongyle eggs. Body condition scores were ideal in 94.4 % of the evaluations (n = 967), and 95.8 % of all horses had good clinical and behavioral indicators. Despite the variation in FEC, the data found no significant association between FEC and the behavioral indicators. The study suggests that FEC alone should not be used as a determinant of welfare when animals are managed with good nutritional and health management practices.

Development of a microsphere-based immunoassay for the serological diagnosis of equine trypanosomosis.

Trypanozoon infections in equids are caused by three parasite species in the Trypanozoon subgenus: Trypanosoma equiperdum, T. brucei and T. evansi. They are respectively responsible for infectious diseases dourine, nagana and surra. Due to the threat that Trypanozoon infection represents for international horse trading, accurate diagnostic tests are crucial. Current tests suffer from poor sensitivity and specificity, due in the first case to the transient presence of parasites in the blood and in the second, to antigenic cross-reactivity among Trypanozoon subspecies. This study was designed to develop a microsphere-based immunoassay for diagnosing equine trypanosomosis. We tested beads coated with eight Trypanosoma spp. recombinant antigens: enolase, GM6, PFR1, PFR2, ISG65, VSGat, RoTat1.2 and JN2118HU. Of these, GM6 was identified as the best candidate for the serological diagnosis of Trypanozoon infections in equids. Using a receiver operating characteristic (ROC) analysis on 349 equine sera, anti-GM6 antibodies were detected with an AUC value of 0.994 offering a sensitivity of 97.9% and a specificity of 96.0%. Our findings show that the GM6 antigen is a good target for diagnosing equine trypanosomosis using a microsphere-based immunoassay. This promising assay could be a useful alternative to the official diagnostic tool for equine trypanosomosis.

A cross-sectional study of Leishmania spp. in draft horses from the Distrito Federal, Brazil: Seroprevalence, spatial distribution, and associated factors.

Equine leishmaniasis is a neglected tropical disease caused by the protozoan of the Leishmania genus, and it has been reported in several countries around the world, especially Brazil. Therefore, the present investigation aims to conduct a cross-sectional study to determine the prevalence, spatial distribution, and associated factors with seropositivity for Leishmania spp. in draft horses from the Distrito Federal, Brazil. The serological survey was conducted on 411 animals, employing the Indirect Immunofluorescence Test (IFA) and Enzyme-Linked Immunosorbent Assay (ELISA). The Kappa (κ) and gross agreement indexes evaluated the Leishmania spp. seropositivity by IFA and ELISA test. The statistical analysis was performed using the Chi-square test and logistic regression. The spatial analysis showed the areas with the highest number of seropositive and the Moran autocorrelation analyses between the spatial distribution and the epidemiological model's explanatory variables. A 27.01 % co-positivity was observed with a κ index of 52.64 %. The final model considered the variables: access to water bodies (p-value = 0.008, Odds Ratio (OR) = 2.26, Confidence Interval (CI) = 1.24-4.13), the absence of the use of ectoparasiticide (p-value = 0.008, OR = 1.93 CI = 1.18-3.15) and traveling animal (p-value = 0.059, OR = 1.54, CI = 0.98-2.41). The Kernel map showed hot areas with a high concentration of nine positive animals per area and some lighter areas ranging from five to seven positive animals per area where control measures should be performed. The Moran autocorrelation analysis was significant for the variables: traveling animal (Moran's I = 0.540 and pseudo-p-value = 0.001) and the absence of use ectoparasiticide (Moran's I = 0.259 and pseudo-p-value = 0.005). The current study exposes a high seroprevalence of Leishmania spp. in horses in the Distrito Federal, Brazil. Moreover, it proposes that traveling animal, the access to water bodies and the absence of the use of ectoparasiticide are significantly associated with seropositivity for Leishmania spp. in draft horses, which may contribute to the implementation of prophylactic and controls measures where leishmaniasis is already stalled.

Equine attachment site preferences and seasonality of common North American ticks: Amblyomma americanum, Dermacentor albipictus, and Ixodes scapularis.

BACKGROUND: Ticks are common on horses, but recent publications characterizing equine tick infestations in North America are lacking. METHODS: To further understand attachment site preferences of common ticks of horses, and to document the seasonality of equine tick infestation in northeastern Oklahoma, horses from eight farms were evaluated twice a month over a 1-year period. Each horse was systematically inspected beginning at the head and moving caudally to the tail. Attachment sites of ticks were recorded and all ticks collected were identified to species and stage. RESULTS: Horses (26 males and 62 females) enrolled in the study ranged in age from 1 to 23 years (mean = 12, 95% CI 11-13). A total of 2731 ticks were collected; 84.1% (74/88) of the horses were infested (median = 3 ticks) at one or more examinations. Five tick species were identified, including Amblyomma americanum (78.2%; 2136/2731), Ixodes scapularis (18.2%; 497/2731), Dermacentor albipictus brown variant (2.6%; 71/2731), Dermacentor variabilis (0.7%; 20/2731), and Amblyomma maculatum (0.3%; 7/231). Most ticks were adults (83.6%; 2282/2731), but immature A. americanum (436/2136; 20.4%), D. albipictus (12/71; 16.9%), and A. maculatum (n = 1) were occasionally recovered. Amblyomma americanum were most often attached to the inguinal area, and I. scapularis and D. albipictus were most commonly found on the chest and axillary region (P < 0.0001). Ticks were found on horses in every month of the year. The largest number of ticks (638/2731; 23.4%) were collected in May (P < 0.0001). Amblyomma americanum, primarily immature, was the only tick recovered in September, I. scapularis and D. albipictus predominated October through February, and both A. americanum and I. scapularis were common in March. In the warmer months, April through August, A. americanum was the most common tick, followed by D. variabilis and A. maculatum. CONCLUSIONS: This research confirms that ticks common on horses in North America have attachment site preferences and that ticks infest horses in Oklahoma throughout the year, including during the winter. Additional research is warranted to fully understand the risk these infestations pose to equine health.

Risk factors associated with tick infestations on equids in Khyber Pakhtunkhwa, Pakistan, with notes on Rickettsia massiliae detection.

BACKGROUND: Studies on ticks infesting equids are lacking in various parts of the world, including Khyber Pakhtunkhwa (KP), Pakistan. The aim of this study was to investigate the diversity of ticks infesting equids, associated risk factors and rickettsial detection in ticks from equids in KP. METHODS: Inspection of 404 equid hosts from November 2018 to October 2019 resulted in the collection of 550 ticks. Data on tick-associated risk factors were collected from equid owners by means of a questionnaire. After morphological identification, partial DNA sequences of the tick mitochondrial 16S rRNA gene were used for taxonomic confirmation of species. Partial sequences of the gltA and ompA genes were used for Rickettsia detection in ticks. RESULTS: A total of 550 tick specimens were collected on 324 (80.2%) of the equids inspected, of which 161 were horses (50%), 145 (45%) were donkeys and 18 were mules (5%). The ticks were identified as belonging to the following five species: Rhipicephalus microplus (341 specimens, 62% of the total ticks), Rh. haemaphysaloides (126, 23%), Rh. turanicus (39, 7%), Rh. sanguineus (s.l.) (33, 6%) and Hyalomma anatolicum (11, 2%). The most prevalent tick life stage was adult females (279, 51%) followed by adult males (186, 34%) and nymphs (85, 15%). Higher tick infestations were observed on male equids (relative risk [RR] 0.7432, P < 0.0005) and adult equids (RR 1.268, P < 0.0020). Ticks were frequently attached to the axial region of horses (55, 21%), sternum of donkeys (44, 21%) and belly of mules (19, 23%) (P < 0.04). Temporal patterns of tick infestation in association with temperature and humidity were highly significant (P < 0.05). Risk factors, such as animal housing (P < 0.0003), living management (P < 0.006), grazing type (P < 0.01) and location in hilly areas (P < 0.02), significantly enhanced the chances for tick infestation. Tick species analyzed in this study were phylogenetically related to species from Afghanistan, China, South Africa and Taiwan. Partial sequences of the gltA and ompA genes obtained from Rh. microplus and Rh. haemaphysaloides were 100% identical to the spotted fever group pathogen Rickettsia massiliae. CONCLUSIONS: Equids exposed to significant risk factors were infected by one or more of at least five tick species in KP, Pakistan, and some of the ticks harbored the human pathogen R. massiliae.

Effects of Gasterophilus pecorum infestation on the intestinal microbiota of the rewilded Przewalski's horses in China.

Horse botflies have been a threat to the Przewalski's horses in the Kalamaili Nature Reserve in Xinjiang of China since their reintroduction to the original range. As larvae of these parasites could infest the intestine of a horse for months, they could interact with and alter the structure and composition of its intestinal microbiota, affecting adversely its health. Nonetheless, there are no such studies on the rewilded Przewalski's horses yet. For the first time, this study characterizes the composition of the intestinal microbiota of 7 rewilded Przewalski's horses infected severely by Gasterophilus pecorum following and prior to their anthelmintic treatment. Bioinformatics analyses of the sequence data obtained by amplicon high throughput sequencing of bacterial 16S rRNA genes showed that G. pecorum infestation significantly increased the richness of the intestinal microbial community but not its diversity. Firmicutes and Bacteroidetes were found the dominant phyla as in other animals, and the parasitic infestation decreased the F/B ratio largely by over 50%. Large reduction in relative abundances of the two genera Streptococcus and Lactobacillus observed with G. pecorum infestation suggested possible changes in colic and digestion related conditions of the infected horses. Variations on the relative abundance of the genus groups known to be pathogenic or symbiotic showed that adverse impact of the G. pecorum infestation could be associated with reduction of the symbiotic genera Lactobacillus and Bifidobacterium that are probiotics and able to promote immunity against parasitic infection.

Imported Hyalomma ticks in the Netherlands 2018-2020.

BACKGROUND: Ticks of the genus Hyalomma, which are vectors for several tick-borne diseases, are occasionally found in areas outside their endemic range including northern parts of Europe. The objective of this study was to analyse adult Hyalomma ticks that were recently found in the Netherlands. METHODS: Hyalomma ticks were morphologically identified. Cluster analysis, based upon sequence data (cox1 barcoding) for molecular identification, and pathogen detection were performed. Additionally, a cross-sectional survey of horses was conducted to actively search for Hyalomma ticks in summer 2019. Analysis of temperature was done to assess the possibility of (i) introduced engorged nymphs moulting to adults and (ii) establishment of populations in the Netherlands. RESULTS: Seventeen adult Hyalomma ticks (one in 2018, eleven in 2019, five in 2020) were found by citizens and reported. Fifteen ticks were detected on horses and two on humans. Twelve were identified as H. marginatum, one as H. rufipes and four, of which only photographic images were available, as Hyalomma sp. No Crimean-Congo haemorrhagic fever virus or Babesia/Theileria parasites were detected. One adult tick tested positive for Rickettsia aeschlimannii. In the cross-sectional horse survey, no Hyalomma ticks were found. Analysis of temperatures showed that engorged nymphs arriving on migratory birds in spring were able to moult to adults in 2019 and 2020, and that cumulative daily temperatures in the Netherlands were lower than in areas with established H. marginatum populations. CONCLUSIONS: Our results show that Hyalomma ticks are regularly introduced in the Netherlands as nymphs. Under the Dutch weather conditions, these nymphs are able to develop to the adult stage, which can be sighted by vigilant citizens. Only one human pathogen, Rickettsia aeschlimannii, was found in one of the ticks. The risk of introduction of tick-borne diseases via Hyalomma ticks on migratory birds is considered to be low. Establishment of permanent Hyalomma populations is considered unlikely under the current Dutch climatic conditions.

Anthelmintic resistance in Creole horses in the South of Rio Grande do Sul, Brazil / [Resistência anti-helmíntica em equinos da raça Crioula no sul do Rio Grande do Sul, Brasil]

The objective of the study was to evaluate the antiparasitic resistance against horse nematodes in the South of Rio Grande do Sul, Brazil. The results concerning the tests of anthelmintic efficacy on horses, stored in the database of the Parasitic Diseases Study Group (GEEP) - Veterinary Faculty, at the Federal University of Pelotas (UFPel), were carried out in the laboratory from 2018 to 2019. Stool samples were received from farms with breeding of adult female and male Criollo horses naturally infected, located in municipalities in the country's southern region. The antiparasitic agents tested were Triclorfon + Fenbendazole, Closantel + Albendazole, Ivermectin + Praziquantel, Fenbendazole, Ivermectin, Doramectin, Mebendazole and Moxidectin. Techniques such as Gordon and Whitlock, Coproculture and Fecal Egg Count Reduction Test were performed. Of all the antiparasitic drugs tested, it was observed that only treatments with Ivermectin 2% showed desired values. The observed results indicate that resistance to macrocyclic lactones is usual in equine parasites in this Brazilian region, despite the results with isolated Ivermectin.(AU)

O objetivo deste estudo é avaliar a resistência antiparasitária contra nematodeos de equinos no sul do Rio Grande do Sul, Brasil. Os resultados referentes aos testes de eficácia anti-helmíntica em cavalos, armazenados no banco de dados do Grupo de Estudos de Doenças Parasitárias (GEEP) - Faculdade de Veterinária, da Universidade Federal de Pelotas (UFPel), foram realizados em laboratório, no período de 2018 a 2019. Amostras de fezes foram recebidas de fazendas com criação de cavalos Crioulos adultos fêmeas e machos naturalmente infectados, localizadas em municípios da região Sul do país. Os agentes antiparasitários testados foram triclorfon + fenbendazol, closantel + ivermectina + praziquantel, fenbendazol, ivermectina, doramectina, mebendazol e moxidectina. Técnicas como Gordon e Whitlock, coprocultura e teste de redução da contagem de ovos fecais foram realizadas. De todos os antiparasitários testados, observou-se que apenas os tratamentos com ivermectina 2% apresentaram os valores desejados. Os resultados indicam que a resistência às lactonas macrocíclicas é comum em parasitas equinos nessa região brasileira, apesar dos resultados com ivermectina isolada.(AU)

Theileria equi claudin like apicomplexan microneme protein contains neutralization-sensitive epitopes and interacts with components of the equine erythrocyte membrane skeleton.

Theileria equi is a widely distributed apicomplexan parasite that causes severe hemolytic anemia in equid species. There is currently no effective vaccine for control of the parasite and understanding the mechanism that T. equi utilizes to invade host cells may be crucial for vaccine development. Unlike most apicomplexan species studied to date, the role of micronemes in T. equi invasion of host cells is unknown. We therefore assessed the role of the T. equi claudin-like apicomplexan microneme protein (CLAMP) in the invasion of equine erythrocytes as a first step towards understanding the role of this organelle in the parasite. Our findings show that CLAMP is expressed in the merozoite and intra-erythrocytic developmental stages of T. equi and in vitro neutralization experiments suggest that the protein is involved in erythrocyte invasion. Proteomic analyses indicate that CLAMP interacts with the equine erythrocyte α-and ß- spectrin chains in the initial stages of T. equi invasion and maintains these interactions while also associating with the anion-exchange protein, tropomyosin 3, band 4.1 and cytoplasmic actin 1 after invasion. Additionally, serological analyses show that T. equi-infected horses mount robust antibody responses against CLAMP indicating that the protein is immunogenic and therefore represents a potential vaccine candidate.

Genetic variability, cryptic species and phylogenetic relationship of six cyathostomin species based on mitochondrial and nuclear sequences.

Cyathostomins are important intestinal nematode parasites of equines and include 50 accepted species. Their taxonomy has been frequently revised and the presence of cryptic species suggested. Furthermore, usually molecular- and morphology-based phylogenetic analyses give divergent results. In this study, the nucleotide sequences of the nuclear second internal transcribed spacer (ITS-2) and the mitochondrial partial cytochrome c oxidase subunit I (COI) were determined for adults of six cyathostomin species (Coronocyclus coronatus, Coronocyclus labiatus, Cylicocyclus nassatus, Cylicostephanus calicatus, Cylicostephanus longibursatus, Cylicostephanus minutus) collected from different equine species within two geographic regions. Maximum likelihood trees were calculated for ITS-2, COI, and concatenated data. No obvious differentiation was observed between geographic regions or equine host species. As previously reported, Coronocyclus coronatus and Cylicostephanus calicatus revealed a close relationship. Cryptic species were detected in Cylicostephanus minutus and Cylicostephanus calicatus. Cylicocyclus nassatus and Coronocyclus labiatus showed diverse mitochondrial and nuclear haplotypes occurring in different combinations, while Cylicostephanus longibursatus was comparatively homogenous. In conclusion, a combined analysis of nuclear and mitochondrial haplotypes improved resolution of the phylogeny and should be applied to the remaining cyathostomin species and across additional equine host species and geographic regions.