Analyzing the safety of the parasiticide fungus Mucor circinelloides: first insights on its virulence profile and interactions with the avian gut microbial community.

Parasiticide fungi are considered an accurate, sustainable, and safe solution for the biocontrol of animal gastrointestinal (GI) parasites. This research provides an initial characterization of the virulence of the native parasiticide fungus Mucor circinelloides (FMV-FR1) and an assessment of its impact on birds' gut microbes. The genome of this fungus was sequenced to identify the genes coding for virulence factors. Also, this fungus was checked for the phenotypic expression of proteinase, lecithinase, DNase, gelatinase, hemolysin, and biofilm production. Finally, an in vivo trial was developed based on feeding M. circinelloides spores to laying hens and peacocks three times a week. Bird feces were collected for 3 months, with total genomic DNA being extracted and subjected to long-read 16S and 25S-28S sequencing. Genes coding for an iron permease (FTR1), iron receptors (FOB1 and FOB2), ADP-ribosylation factors (ARFs) (ARF2 and ARF6), and a GTPase (CDC42) were identified in this M. circinelloides genome. Also, this fungus was positive only for lecithinase activity. The field trial revealed a fecal microbiome dominated by Firmicutes and Proteobacteria in laying hens, and Firmicutes and Bacteroidetes in peacocks, whereas the fecal mycobiome of both bird species was mainly composed of Ascomycetes and Basidiomycetes fungi. Bacterial and fungal alpha-diversities did not differ between sampling time points after M. circinelloides administrations (P = 0.62 and P = 0.15, respectively). Although findings from this research suggest the lack of virulence of this M. circinelloides parasiticide isolate, more complementary in vitro and in vivo research is needed to conclude about the safety of its administration to birds, aiming at controlling their GI parasites.IMPORTANCEA previous study revealed that the native Mucor circinelloides isolate (FMV-FR1) can develop parasiticide activity toward coccidia oocysts, one of the most pathogenic GI parasites in birds. However, ensuring its safety for birds is of utmost importance, namely by studying its virulence profile and potential effect on commensal gut microbes. This initial study revealed that although this M. circinelloides isolate had genes coding for four types of virulence factors-iron permease, iron receptors, ADP-ribosylation factors, and GTPase-and only expressed phenotypically the enzyme lecithinase, the administration of its spores to laying hens and peacocks did not interfere with the abundances and diversities of their gut commensal bacteria and fungi. Although overall results suggest the lack of virulence of this M. circinelloides isolate, more complementary research is needed to conclude about the safety of its administration to birds in the scope of parasite biocontrol programs.

Severely Asthmatic Horses Residing in a Mediterranean Climate Shed a Significantly Lower Number of Parasite Eggs Compared to Healthy Farm Mates.

The relationship between helminth infection and allergic diseases has long intrigued the scientific community. This interaction was previously studied in a horse family with high incidence of severe equine asthma and in non-related severely asthmatic horses from equine hospital referrals in Switzerland. Our aim was to determine if this interaction would also be observed in a group of non-related client-owned severely asthmatic horses living in a Mediterranean climate and recruited through a first-opinion veterinarian group. Fecal samples from severe equine asthma-affected and healthy horses living in the same farms and subjected to identical environmental and deworming management were evaluated qualitatively and quantitatively. Strongyle-type eggs and Cyathostomum sensu latum larvae were the most abundant parasites in the studied population of horses; no significant differences between the groups were observed regarding the types of egg and infective larvae. However, we observed significant differences in the number of eggs and infective larvae per gram of feces shed, as this number was significantly lower in the SEA group than in the healthy horses. This may indicate that severely asthmatic horses have an intrinsic resistance to gastrointestinal helminths. Further studies in a larger population of horses are required to ascertain the immunological mechanisms responsible for these findings.

Challenges of copro-parasitological surveys in wild Iberian ibex (Capra pyrenaica) populations addressed through a combination of molecular and statistical tools.

Copro-parasitological surveys in wildlife face challenges due to the secretive nature of many species and the unknown performance of the diagnostic tests employed. To overcome these issues, we used a combination of hierarchical models (site-occupancy and N-mixture models) applied to copro-parasitological data obtained from fecal samples assigned to the host species by molecular methods in the Iberian ibex in north-western Iberian Peninsula. The aims were to compare the performance of four diagnostic tests (Mini-FLOTAC, McMaster, Willis flotation, and natural sedimentation) and to use this methodological approach (molecular analysis with hierarchical models) to better estimate positivity proportion and shedding intensity in a wild ibex population. Pooled fecal samples were collected, and those confirmed by molecular analyses to be the host species in question were included in the study. Hierarchical models confirmed different performances of each diagnostic test, with Mini-FLOTAC showing higher sensitivity for eimeriid coccidia, Willis flotation (for proportion positive) and McMaster (for shedding intensity) in gastrointestinal Strongylida, and equal performance of MiniFlotac/Willis flotation (for proportion positive) and MiniFlotac/McMaster (for shedding intensity) in Moniezia spp. This study employed a combination of molecular and statistical methods that improved the estimates of prevalence and shedding intensity and allowed us to compare the performance of four diagnostic tests while assessing the effect of covariates. Such improvements are critical to enhancing inference in non-invasive wildlife copro-parasitological studies.

Isolation of saprophytic filamentous fungi from avian fecal samples and assessment of its predatory activity on coccidian oocysts.

Fungal strains used in the biocontrol of animal gastrointestinal parasites have been mainly isolated from pasture soil, decaying organic matter, and feces from herbivores and carnivores. However, their isolation from birds and assessment of predatory activity against avian GI parasites has been scarce thus far. This research aimed to isolate filamentous fungi from avian fecal samples and evaluate their predatory activity against coccidia. A pool of 58 fecal samples from chickens, laying hens, and peacocks, previously collected between July 2020-April 2021, were used for isolation of filamentous fungi and assessment of their in vitro predatory activity against coccidian oocysts, using Water-Agar medium and coprocultures. The Willis-flotation technique was also performed to obtain concentrated suspensions of oocysts. A total of seven Mucor isolates was obtained, being the only fungal taxa identified, and all presented lytic activity against coccidia. Isolates FR3, QP2 and SJ1 had significant coccidiostatic efficacies (inhibition of sporulation) higher than 70%, while isolates FR1, QP2 and QP1 had coccidicidal efficacies (destruction of the oocysts) of 22%, 14% and 8%, respectively, after 14 days of incubation, being a gradual and time-dependent process. To our knowledge, this is the first report regarding the isolation of native predatory fungi from avian feces and demonstration of their lytic activity against coccidia.

Morphological and Molecular Identification of Physaloptera alata (Nematoda: Spirurida) in a Booted Eagle (Aquila pennata) from Portugal.

Physaloptera spp. are parasitic nematodes that infect the gastrointestinal tracts of many carnivores and omnivores. Although they are distributed worldwide, Physaloptera spp. have not been studied in raptors in Portugal. In this study, we report Physaloptera alata in a booted eagle (Aquila pennata) in Portugal. Adult nematodes were discovered in the gizzard of a young booted eagle, and morphological features were consistent with those of the genus Physaloptera. DNA was extracted and a PCR assay performed to amplify a region of the 18S small subunit of the ribosomal RNA gene and the cytochrome c oxidase subunit I gene. The resulting PCR products were Sanger-sequenced, and comparison with the available sequences in the GenBank database confirmed the initial morphological classification as Physaloptera sp. Phylogenetic analysis clustered the sequence within the Physaloptera group. The presence of this parasite in raptors from Portugal is of particular importance to wildlife rehabilitation centers, disease ecologists, and wildlife professionals. Furthermore, we produced a new genetic sequence and have added it to the GenBank database of parasites in birds of prey.

Anthelmintic Efficacy in Sheep and Goats under Different Management and Deworming Systems in the Region of Lisbon and Tagus Valley, Portugal.

Infections caused by gastrointestinal parasites have been described worldwide as one of the most important issues impacting small ruminant production. The systematic administration of anthelmintic (AH) drugs without following good practice principles has led to an increase in anthelmintic resistance (AR). There is scarce information regarding AH efficacy in small ruminants in Portugal. This study aimed to characterize by in vivo methods the presence and level of AR in four farms in the region of Lisbon and Tagus Valley. All four farms kept small herds in extensive management systems and used different deworming protocols. The active substances used were fenbendazole and a combination of mebendazole plus closantel in a dosage according to the manufacturers' instructions. On day 0 (T0), fecal samples were collected from all study animals, and animals in the treatment group (n = 40) were dewormed with the AH previously chosen by the assistant veterinarian. Animals in the control group (n = 30) did not receive any AH drug. The fecal sample collection was repeated on day 15 (T15), and the control group was treated. Egg counts were performed using the McMaster method for the eggs per gram (EPG), and AR was evaluated by the fecal egg count reduction test (FECRT) with a 95% confidence level (CL). The results from this experiment indicated that the four farms presented AR with two farms to fenbendazole (FECRT of 48% and 85%) and two farms to mebendazole plus closantel (FECRT of 66% and 79%). These results indicate that the gastrointestinal parasites of the four studied Portuguese farms are resistant to benzimidazoles, which suggests an increase in AR regarding nematodes in small ruminant production systems in Portugal.

The global prevalence of Spirometra parasites in snakes, frogs, dogs, and cats: A systematic review and meta-analysis.

BACKGROUND: Spirometra infection is aneglected food- and waterborne disease with worldwide distribution. OBJECTIVES: The present study aims to estimate the global prevalence of Spirometra species in snakes, frogs, dogs and cats. METHODS: Multiple databases (PubMed, Scopus, ProQuest, Web of Science and Google Scholar) were searched for relevant literatures published up to March 2022. RESULTS: Among 131 data sets (including 113 articles) that met the inclusion, 15 investigations reported Spirometra infection in snakes, 23 in frogs, 41 in dogs and 52 in cats. The pooled prevalence (95% confidence interval) in intermediate hosts and definitive hosts was found to be 0.313% and 0.089%, respectively. Based on continent, the infection was most prevalent in Asia for studies on snakes (0.696%) and frogs (0.181%), while Africa (0.224%) and Oceania (0.203%) were the regions with the highest pooled prevalence rates of the infection in dogs and cats, respectively. Among different diagnostic methods, the highest pooled prevalence was related to morphological method for studies on snakes, frog and cats with rate of 0.665%, 0.189% and 0.104%, respectively. Regarding studies on dogs, the highest pooled prevalence was observed for molecular technique (0.101%). CONCLUSIONS: The results presented here revealed the importance of establishing a prevention and control measure focused on protection of aquaculture systems from being contaminated with faeces of dogs and cats, and raising awareness of parasitic zoonotic diseases to decrease the transmission risk.

Preliminary findings on the gastrointestinal parasites of the brown bear (Ursus arctos) in the Cantabrian mountains, Spain.

No study is currently available on the parasitofauna of the population of brown bears (Ursus arctos) inhabiting the Cantabrian Mountains in Spain. The aim of the present study was to obtain novel information on diversity and prevalence of gastrointestinal parasites in these individuals. During August 2016 and from May to July 2017, 14 fecal samples were collected from the western Cantabrian bear subpopulation, in the Somiedo Natural Park, in the Spanish province of Asturias. The prevalence of parasites detected was 71% and two genera were identified: Dicrocoelium sp. and Trichuris sp. Since the impact that pathogens such as endoparasites can have on the health of bears, together with other stressors, is still poorly understood, research efforts that include disease surveillance are critical to the successful protection of this emblematic species. Our preliminary findings require further investigations, with a wider sampling effort, and bring awareness for the need of carrying further studies on this area as a part of a proactive species management plan.

Control of Strongyles in First-Season Grazing Ewe Lambs by Integrating Deworming and Thrice-Weekly Administration of Parasiticidal Fungal Spores.

Parasiticidal fungi have been used in several in vivo experiments in livestock farms worldwide, constituting an effective tool for the biocontrol of gastrointestinal parasites in grazing animals. In the first year of study, two groups of eight first-season pasturing ewe lambs infected by strongyles were dewormed with albendazole, and then, the test group received an oral dose of 106 chlamydospores of Mucor circinelloides and 106 Duddingtonia flagrans individually and thrice a week from mid-September to May (FS1), while the control group remained without fungi (CT1). In the second year, two new groups of first-season grazing ewe lambs were treated with ivermectin and subjected to the same experimental design (FS2 and CT2, respectively). The anthelmintic efficacy was 96.6% (CT1), 95.6% (FS1), 96.1% (CT2), and 95.1% (FS2). The counts of strongyle egg output increased in the control groups (CT1 and CT2) throughout the study and reached numbers higher than 600 eggs per gram of feces (EPG), while in FS1 and FS2, they were <250 EPG. The values of red blood cell parameters registered for CT1 and CT2 were lower than those of the reference standards, while a significant increment was recorded in FS1 and FS2, and values within the physiological range were attained. It is concluded that integrating efficient anthelminthic deworming with rotational pasturing and the regular intake of chlamydospores of M. circinelloides and D. flagrans provides a helpful strategy for maintaining low levels of strongyle egg output in first-season grazing ewe lambs and improves their health status.

Implementation of Mini-FLOTAC in Routine Diagnosis of Coccidia and Helminth Infections in Domestic and Exotic Birds.

Mini-FLOTAC (MF) has recently been proposed for the fecal quantification of gastrointestinal (GI) parasites in birds due to its higher sensitivity and precision in comparison with the McMaster method. The current research aimed to test the use of MF in routine diagnosis of coccidia and helminth infections in several domestic and exotic bird collections in Portugal. Between July 2020 and April 2021, a total of 142 fecal samples from organic layers, peacocks and ratites were collected in four Portuguese bird collections and processed using MF and fecal cultures to identify and calculate GI parasite shedding and prevalence. The McMaster method was also used to compare the shedding levels obtained for both quantitative techniques. MF's relative sensitivity and specificity were also assessed, using McMaster as the reference technique. The implementation of MF resulted in an average Eimeria spp. shedding higher in peacocks from bird collection 2 (502 OPG), followed by peacocks from collection 1 (107 OPG) and organic layers (24 OPG) and peacocks from collection 3 (9 OPG). Peacocks were also positive for Capillaria spp., Trichostrongylus tenuis and Strongyloides pavonis, whereas ostriches and emus were infected by L. douglassii. The MF protocol for exotic animals and the McMaster method did not differ significantly for each parasitic agent and bird species, and MF achieved relative sensitivities and specificities higher than 70% for Galliform Eimeria spp., peacock helminths and ratites' L. douglassii infections. Higher L. douglassii EPG values were identified using the MF protocol for exotic species (2 g of feces/38 mL of sucrose solution), followed by McMaster 2/28, MF 5/45 and MF 2/18. The use of MF allowed for obtaining different intestinal parasitic populations in several bird species and locations, and MF 2/38 is globally proposed as the most suitable protocol for bird fecal samples as an alternative to the McMaster method in the diagnosis of avian intestinal parasitic infections.

Gastrointestinal Parasitism in Przewalski Horses (Equus ferus przewalskii).

PURPOSE: Przewalski's horse (Equus ferus przewalskii) is a subspecies of Equus ferus classified as endangered by the International Union for Conservation of Nature (IUCN), whose gastrointestinal (GI) parasitism has been studied in different populations, in situ and ex situ, during the last decades. We aimed to summarize the information available on the subject. METHODS: In this review, we collected information from distinct published literature, interpreted it, compared it and related the Przewalski's horse biology and ecology with different findings. RESULTS: Many similarities can be found between the Przewalski's horse populations and the domestic horses regarding their gastrointestinal parasite community. However, not all the GI parasites described in the domestic horse have been reported in Przewalski's horse's surveys. The most prevalent parasites found in Przewalski's horses belong to the Strongylidae family, namely the Cyathostominae. CONCLUSIONS: More continuous monitoring studies should be carried on in different in situ and ex situ locations to consistently support the one-off surveys. It might be necessary to better correlate the parasitological findings of this subspecies with other animals that share the same habitat and ecosystem, for instance, intermediate and reservoir hosts, from different animal taxonomic groups.

Non-Invasive Molecular Survey of Sarcoptic Mange in Wildlife: Diagnostic Performance in Wolf Faecal Samples Evaluated by Multi-Event Capture-Recapture Models.

Sarcoptic mange is globally enzootic, and non-invasive methods with high diagnostic specificity for its surveillance in wildlife are lacking. We describe the molecular detection of Sarcoptes scabiei in non-invasively collected faecal samples, targeting the 16S rDNA gene. We applied this method to 843 Iberian wolf Canis lupus signatus faecal samples collected in north-western Portugal (2006-2018). We further integrated this with serological data (61 samples from wolf and 20 from red fox Vulpes vulpes, 1997-2019) in multi-event capture-recapture models. The mean predicted prevalence by the molecular analysis of wolf faecal samples from 2006-2018 was 7.2% (CI95 5.0-9.4%; range: 2.6-11.7%), highest in 2009. The mean predicted seroprevalence in wolves was 24.5% (CI95 18.5-30.6%; range: 13.0-55.0%), peaking in 2006-2009. Multi-event capture-recapture models estimated 100% diagnostic specificity and moderate diagnostic sensitivity (30.0%, CI95 14.0-53.0%) for the molecular method. Mange-infected individually identified wolves showed a tendency for higher mortality versus uninfected wolves (ΔMortality 0.150, CI95 -0.165-0.458). Long-term serology data highlights the endemicity of sarcoptic mange in wild canids but uncovers multi-year epidemics. This study developed and evaluated a novel method for surveying sarcoptic mange in wildlife populations by the molecular detection of S. scabiei in faecal samples, which stands out for its high specificity and non-invasive character.

Gastrointestinal Parasitism in Miranda Donkeys: Epidemiology and Selective Control of Strongyles Infection in the Northeast of Portugal.

In Portugal, equine parasitism in pasture animals is characterized by high parasitic burden and high helminthic biodiversity; both factors are potentially pathogenic for their hosts. The decrease in the number of donkeys over the last years in Portugal, their importance in rural lowland and mountain ecosystems and pastures and the scarce information regarding their parasitism led to this research, which aimed to evaluate the parasitological status of a Miranda donkey breed population, a native breed mainly located in the northeast of Portugal. This study provides better knowledge of their gastrointestinal parasitism, particularly strongyles, and the assessment of a targeted selective treatment (TST) as an alternative control approach of their parasitism. A longitudinal observational study was developed during a period of five years in a population of 62 Miranda donkeys. At first, strategic deworming of these animals was performed every semester, but this was progressively replaced by a TST approach according to the levels of Eggs per Gram (EPG). This new deworming strategy was conducted in association with a regular parasitological monitoring of the animals every three months, being dewormed with ivermectin when egg shedding was higher than 500 EPG. Over the study period, a decrease of the annual prevalence rate of infection by gastrointestinal strongyles was observed, from 35.5% to 19.4%, as well as a negative binomial distribution of parasitic strongyles in donkeys submitted to selective anthelminthic control. The prevalence rate of infection was higher in females (39.5%), in individuals under four years (46.7%) and in those presenting a lower body condition (40.8%). The egg output was higher in animals younger than four years (589.3 EPG) than in those older than 10 years (533.6 EPG) (p < 0.05). However, no differences were observed according to sex during the study period. Results from this study allowed to note the influence of swampy pastures and of the weather changes in the epidemiology of strongylosis in Miranda donkeys. Moreover, it was possible to establish the annual epidemiological curve of strongyle egg shedding, with June being the month with the highest EPG, December having the lowest EPG and March and September showing intermediate numbers. Overall, a lower biodiversity of gastrointestinal parasites was observed. Cyathostomum sensu lato was the most prevalent genus and Strongylus vulgaris was the most observed large strongyle of the Strongylidae family. Trichostrongylus axei and Parascaris sp. were other nematodes with a minor frequency. The higher prevalence of strongyles at the beginning of the study showed a progressive decrease throughout the research period, and also for parasite biodiversity. Therefore, a targeted selective treatment seems to be a rational anthelminthic control approach in Miranda donkey strongyle infection and in other gastrointestinal parasites, since it reduces the antiparasitic treatments, the parasite' prevalence and the EPG level. However, a loss of parasite biodiversity was noted at the end of the study period, as Cyathostominae were the only isolated strongyles. This can be a challenging situation in the long run, taking the ability of these nematodes to adapt easily to any deworming program, meaning that fecal EPG monitoring should be kept as a rule to a rational parasite control program.

Survey of Zoonotic and Non-zoonotic Vector-Borne Pathogens in Military Horses in Lisbon, Portugal.

Vector-borne diseases of zoonotic and/or veterinary relevance have been increasingly reported in horses globally, although data regarding working and military horses is lacking. Portuguese military horses may constitute a risk group for these pathogens, as they frequently work outdoors in various regions of the country. This study included 101 apparently healthy horses belonging to the Portuguese National Republican Guard. Blood samples were analyzed to determine the presence and prevalence of piroplasms, Anaplasmataceae, Rickettsia spp., and filarioid helminths. Overall 32.7% of the horses gave positive results for Theileria equi. Two genotypes of T. equi were verified. No positive results were recorded for Anaplasma spp., Rickettsia spp., filarioid helminthes, and Babesia caballi. As equine piroplasmosis is a severe infectious tick-borne disease responsible for significant losses in equine production and with numerous impacts in the international movement of horses, adequate treatment, and preventive measures are needed to reduce exposure to vectors and future infections.

Parasite infections in a social carnivore: Evidence of their fitness consequences and factors modulating infection load.

There are substantial individual differences in parasite composition and infection load in wildlife populations. Few studies have investigated the factors shaping this heterogeneity in large wild mammals or the impact of parasite infections on Darwinian fitness, particularly in juveniles. A host's parasite composition and infection load can be shaped by factors that determine contact with infective parasite stages and those that determine the host's resistance to infection, such as abiotic and social environmental factors, and age. Host-parasite interactions and synergies between coinfecting parasites may also be important. We test predictions derived from these different processes to investigate factors shaping infection loads (fecal egg/oocyte load) of two energetically costly gastrointestinal parasites: the hookworm Ancylostoma and the intracellular Cystoisospora, in juvenile spotted hyenas (Crocuta crocuta) in the Serengeti National Park, in Tanzania. We also assess whether parasite infections curtail survival to adulthood and longevity. Ancylostoma and Cystoisospora infection loads declined as the number of adult clan members increased, a result consistent with an encounter-reduction effect whereby adults reduced encounters between juveniles and infective larvae, but were not affected by the number of juveniles in a clan. Infection loads decreased with age, possibly because active immune responses to infection improved with age. Differences in parasite load between clans possibly indicate variation in abiotic environmental factors between clan den sites. The survival of juveniles (<365 days old) to adulthood decreased with Ancylostoma load, increased with age, and was modulated by maternal social status. High-ranking individuals with low Ancylostoma loads had a higher survivorship during the first 4 years of life than high-ranking individuals with high Ancylostoma loads. These findings suggest that high infection loads with energetically costly parasites such as hookworms during early life can have negative fitness consequences.

Nuclear and mitochondrial marker sequences reveal close relationship between Coronocyclus coronatus and a potential Cylicostephanus calicatus cryptic species complex.

The Cyathostominae (Nematoda, Strongyloidea) parasitising equines represent a diverse group currently including 50 species. However, their taxonomy has been repeatedly revised and occasionally the presence of cryptic genospecies was suggested. Moreover, molecular- and morphology-based phylogenetic analyses give divergent results. For instance, molecular data have suggested close relationship between Coronocyclus coronatus and Cylicostephanus calicatus, although morphology-based taxonomy places them in different genera. Here, nuclear (internal transcribed spacer 2, ITS-2) and mitochondrial (cytochrome oxidase I, COI) sequences were obtained from the same individual, morphologically identified worms. In both morphospecies, two ITS-2 sequences types were observed: In Cor. coronatus, a small PCR product of 278 bp (nuclear haplotype group nHGBco) was always present but often in combination with a larger 369-370 bp fragment (nHGAco). In Cys. calicatus, either a large 370 bp product (nHGAca) or a short 281 bp amplicon (nHGBca) were found, but never both. Sequence identity between morphospecies was up to 100%. The smaller differed from the larger fragments by deletion of the region 110-198 in Cor. coronatus and 112-203 in Cys. calicatus. In COI, three and five mitochondrial haplotype groups (HGs), mtHG1co-mtHG3co and mtHG1ca-mtHG5ca were identified for Cor. coronatus and Cys. calicatus, respectively. In Cor. coronatus, there was no particular association of mtHG with nuclear genotypes (only nHGBco vs. both nHGBco plus nHGAco). In Cys. calicatus the nHGAca was always associated with the mtHG1ca, mtHG2ca or mtHG5ca whereas nHGBca was exclusively associated with mtHG3ca or mtHG4ca. Despite up to 100% identity in the nHGs, no mixing of mtHGs was observed between both species. Clear separation of certain nHGs with particular mtHGs in Cys. calicatus, despite the fact that the same host individuals were infected with both groups simultaneously, suggests presence of two non-interbreeding genospecies within Cys. calicatus, which needs further confirmation using additional samples from diverse geographical origins.

Identification of third stage larval types of cyathostomins of equids: An improved perspective.

Cyathostomins comprise around 50 parasite species of equids, offering a great challenge regarding their individual identification. The objective of our work was to improve identification of infective third stage larvae (L3) with a morphological key supplemented with detailed scientific illustrations based on our research and available literature. The highlighted features were; the number, arrangement, and shape of intestinal cells (IC), general features and the total body length of the eight different Cyathostomin sensu latum types (Type A, B, C, D, E, F, G, H), Gyalocephalus capitatus, and Posteriostomum spp. Due to variability, we were unable to define final body length measurements to differentiate L3 of cyathostomins. However, IC characteristics displayed a higher difference between L3 types than total body length. Through the number and arrangement of IC, 14 species were classified within three larval types. The classification of L3 into distinct larval types sensu latum gives us the advantage of reducing the number of probable species presented in equine faecal samples using a low-cost technique when monitoring the parasite fauna present in individual horses or on the farm level. The present improved identification key shall increase the diagnostic capabilities of classical equine parasitology techniques, using general L3 morphology thereby pragmatically improving regional and transnational epidemiological and biodiversity studies. The present key may also assist in defining the cyathostomin community in cyathostominosis clinical cases and within drug resistant populations across different management systems and geographical locations.

Teaching Veterinary Parasitology in Portugal in the 21st century - Changes, challenges and opportunities after the Bologna process.

Veterinary education in Portugal began in 1830, but individualised teaching of parasitology started in 1911 at the Faculty of Veterinary Medicine, University of Lisbon (FMV-ULisboa). Parasitology and Parasitic Diseases were taught in the 3rd and 4th years of the veterinary course, respectively. In 2007/2008, a new curriculum was implemented as a consequence of the Bologna Process (BP), with Parasitology converted into "Parasitology I" (General Parasitology, Arthropods and Protozoology) and "Parasitology II" (Helminthology), being taught in the 1st and 2nd semesters of the 2nd year of the course, respectively. "Parasitic Diseases" became a one-semester subject in the 4th year. BP was intended to harmonise university courses, foster mobility and cooperation among European Universities, but instead there have been major challenges in reducing student contact and an erosion of Parasitology I and II as basic subjects, which were moved to 2nd year. In spite of these changes, the way subjects have been taught since then shows that veterinary students are satisfied with Parasitology; academic success ranges from 70% to 90%; the number of Master and PhD theses in Parasitology has increased; and students share their research results in classes and at conferences in our faculty and abroad. This shows how research and teaching of Veterinary Parasitology are tightly inter-linked and critical, presenting an opportunity to motivate future students to study Parasitology and Parasitic Diseases in such a challenging tertiary education environment in Portugal.

Awareness of zoonotic diseases and parasite control practices: a survey of dog and cat owners in Qatar.

BACKGROUND: Qatar is one of the wealthiest and fastest growing economies in the world, experiencing a rapid increase in human and pet populations. Given the paucity of data on prophylactic measures against endo- and ectoparasites of pets in Qatar, as well as on the owners' awareness of zoonotic diseases, a questionnaire was conducted. METHODS: From July to November 2017, 150 multiple-choice questionnaires were administered to dog and/or cat owners who attended two veterinary clinics in Doha. RESULTS: Only 54% (81/150) owners were aware of transmittable diseases between animals and humans. "Zoonosis/zoonotic disease(s)" was unknown for 88% (132/150) of the respondents and almost a quarter had no idea of transmission pathways associated with parasitic diseases. Thirteen owners (8.7%) reported to have suffered from zoonotic diseases (10 had dermatophytosis, 2 cat-scratch disease and 1 an unknown tick-borne disease) and 24.7% had dewormed themselves. Approximately 83% had their pets yearly vaccinated and 51% identified endo- and ectoparasites on their pets. Only 10% had their animal faeces tested for intestinal parasites as requested by a veterinarian. As for internal parasite control, only 19.3% dewormed their pets with the recommended treatment regimen (minimum quarterly); 52.7% (79/150) dewormed every 4 months to 1 year; 10% (15/150) without periodicity and 8% (12/150) had never done it. For external parasite control, only 16% (24/150) treated their pets with ectoparasiticides on a monthly basis; 44.7% (67/150) every 2 months to 1 year; 6.7% (10/150) without periodicity and 24.7% (37/150) had never done it. Approximately two thirds (63.3%) of pets were allowed to sleep in the owner's bed and 60% to lick their owner's face. Almost all pets were fed with dry/canned food, but 4.7% were fed with raw meat. Approximately 79.5% of dog owners collect their pet's faeces from public areas. CONCLUSIONS: These results highlight the need to raise pet owners' awareness towards prophylactic measures to minimize the potential impact of zoonotic diseases on the health of both animals and humans in Qatar.

Environmental contamination with Toxocara spp. eggs in public parks and playground sandpits of Greater Lisbon, Portugal.

Toxocarosis is a zoonotic parasitic disease transmitted from companion animals to humans. Environmental contamination with Toxocara eggs is considered to be the main source of human infections. In Portugal, knowledge regarding the current situation, including density, distribution and environmental contamination by Toxocara spp., is largely unknown. The present study investigated environmental contamination with Toxocara spp. eggs, in soil and faecal samples collected from public parks and playground sandpits in Greater Lisbon, Portugal. A total of 151 soil samples and 135 canine faecal samples were collected from 7 public sandpits and 12 public parks, over a 4 month-period. Soil samples were tested by a modified centrifugation and sedimentation/flotation technique and faecal samples were tested by an adaptation of the Cornell-Wisconsin method. Molecular analysis and sequencing were performed to discriminate Toxocara species in the soil. Overall, 85.7% of the sandpits (6/7) and 50.0% of the parks (6/12) were contaminated with Toxocara spp. eggs. The molecular analysis of soil samples showed that, 85.5% of the sandpits and 34.4% of the parks were contaminated with Toxocara cati eggs. Faecal analysis showed that 12.5% of the sandpits and 3.9% of the parks contained Toxocara canis eggs. In total, 53.0% of soil and 5.9% of faecal samples were positive for Toxocara spp. Additionally, 56.0% of the eggs recovered from the samples were embryonated after 60 days of incubation, therefore considered viable and infective. The average density was 4.2 eggs per hundred grams of soil. Public parks and playground sandpits in the Lisbon area were found to be heavily contaminated with T. cati eggs, representing a serious menace to public health as the studied areas represent common places where people of all ages, particularly children, recreate. This study sounds an alarm bell regarding the necessity to undertake effective measures such as reduction of stray animals, active faecal collection by pet owners, awareness campaigns and control strategies to decrease the high risk to both animal and human health.