Control of gastrointestinal helminths in small ruminants to prevent anthelmintic resistance: the Italian experience.

Helminth infections are ubiquitous in grazing ruminants and cause significant costs due to production losses. Moreover, anthelmintic resistance (AR) in parasites is now widespread throughout Europe and poses a major threat to the sustainability of modern ruminant livestock farming. Epidemiological data on the prevalence and distribution of gastrointestinal nematodes, cestodes and liver- and rumen-flukes in Italian small ruminants are outdated and fragmentary. However, anthelmintics are commonly used to control these infections and often without prior diagnosis. Compared to other European countries, few reports of AR in small ruminants against levamisole, ivermectin and benzimidazoles have been published in Italy, but recent studies suggest that this phenomenon is spreading. Increased and integrated research efforts in developing innovative approaches to control helminth infections and AR are needed and must be tailored to the peculiarities of each context in order to be effectively implemented. This manuscript provides an overview on helminth prevalence and distribution, sustainable treatment strategies and integrated control approaches in small ruminants in Italy. The implementation of targeted treatments and targeted selective treatments is discussed based on different parameters, such as fecal egg count, degree of anaemia (FAMACHA© method), milk production and body condition score. In addition, several Italian studies have also investigated the implementation of alternative strategies such as the use of different natural bioactive compounds or genetic selection for resistance and resilience to helminth infections. These concrete solutions for helminth management in small ruminant farms in the country are reported and discussed, representing a valid example for other Mediterranean countries.

Cardio-pulmonary nematodes of the red fox (Vulpes vulpes) of Sardinia, Italy.

Cardio-pulmonary parasites, such as Angiostrongylus vasorum, Crenosoma vulpis, and Eucoleus aerophilus, pose a significant concern on account of pulmonary and cardiac problems they induce in dogs. While the red fox is known to be a key reservoir host for A. vasorum and may also play a role in transmitting C. vulpis and E. aerophilus, there has been no recent research on these parasites in foxes from Sardinia, with the most current studies dating back to 1986. A survey was conducted on red foxes in Sardinia, where a total of 51 foxes were collected, necropsied, and examined for adult worms in their hearts and lungs. The worms were identified using morphometric analysis and molecular methods. The results showed a 54.9% overall prevalence at dissection: 45.1% of the foxes were positive for E. aerophilus, 17.6% for C. vulpis, and 13.7% for A. vasorum. The molecular analyses validated the morphological characterization. In comparison to previous research, which found 13 out of 85 foxes to be positive for A. vasorum with a prevalence rate of 15.3% and 1 for E. aerophilus with a prevalence of 1.2%, this study showed an increased prevalence of E. aerophilus and C. vulpis, and a decrease in the prevalence of A. vasorum. These results indicate that the red foxes in Sardinia represent a reservoir host for cardio-pulmonary nematodes and it should be considered in the differential diagnosis of respiratory distress syndrome in dogs.

Global epidemiology and molecular biology of Taenia multiceps: a comparative meta-analysis and in silico analysis study.

In the present study, all published data on the epidemiology and molecular characters of Taenia multiceps were systematically collected from relevant databases (e.g. PubMed, Scopus, National Center for Biotechnology Information), and combined in various statistical and genetic analyses as a contribution to a better understanding of the epidemiology of this ubiquitous taeniid worldwide. While 5.8% of the key hosts (dogs) from various countries had T. multiceps, grey wolves displayed the highest prevalence (21.6%) among the definitive hosts. Small ruminants are the main intermediate hosts and carry the coenuri in various locations, but most commonly in the central nervous system (CNS). Cerebral coenuri were confirmed in 53% of sheep exhibiting neurological symptoms, and infected animals often had only a single coenurus in the brain. Sheep had a higher prevalence (8.8%) of CNS coenuri than goats (5.8%); however, extra-CNS coenuri were detected more frequently in goats than in sheep. In either case, the difference between sheep and goats was statistically insignificant. Analysis of 233 partial cytochrome oxidase subunit I nucleotide sections for T. multiceps revealed high haplotype and low nucleotide diversities. Fifty-one haplotypes were detected circulating in 6 geographic populations. China, Iran and Turkey had 2 major haplotypes, whereas Italy and Egypt shared 3. Haplotypes from Greece circulate worldwide, and displayed similar gene flow values when compared with the other populations. There were no distinct patterns for haplotype distribution in relation to the infected hosts or coenuri locations. The existence of genetic variants in T. multiceps was highlighted, but needs further studies.

Association analysis and functional annotation of imputed sequence data within genomic regions influencing resistance to gastro-intestinal parasites detected by an LDLA approach in a nucleus flock of Sarda dairy sheep.

BACKGROUND: Gastroinestinal nematodes (GIN) are one of the major health problem in grazing sheep. Although genetic variability of the resistance to GIN has been documented, traditional selection is hampered by the difficulty of recording phenotypes, usually fecal egg count (FEC). To identify causative mutations or markers in linkage disequilibrium (LD) to be used for selection, the detection of quantitative trait loci (QTL) for FEC based on linkage disequilibrium-linkage analysis (LDLA) was performed on 4097 ewes (from 181 sires) all genotyped with the OvineSNP50 Beadchip. Identified QTL regions (QTLR) were imputed from whole-genome sequences of 56 target animals of the population. An association analysis and a functional annotation of imputed polymorphisms in the identified QTLR were performed to pinpoint functional variants with potential impact on candidate genes identified from ontological classification or differentially expressed in previous studies. RESULTS: After clustering close significant locations, ten QTLR were defined on nine Ovis aries chromosomes (OAR) by LDLA. The ratio between the ANOVA estimators of the QTL variance and the total phenotypic variance ranged from 0.0087 to 0.0176. QTL on OAR4, 12, 19, and 20 were the most significant. The combination of association analysis and functional annotation of sequence data did not highlight any putative causative mutations. None of the most significant SNPs showed a functional effect on genes' transcript. However, in the most significant QTLR, we identified genes that contained polymorphisms with a high or moderate impact, were differentially expressed in previous studies, contributed to enrich the most represented GO process (regulation of immune system process, defense response). Among these, the most likely candidate genes were: TNFRSF1B and SELE on OAR12, IL5RA on OAR19, IL17A, IL17F, TRIM26, TRIM38, TNFRSF21, LOC101118999, VEGFA, and TNF on OAR20. CONCLUSIONS: This study performed on a large experimental population provides a list of candidate genes and polymorphisms which could be used in further validation studies. The expected advancements in the quality of the annotation of the ovine genome and the use of experimental designs based on sequence data and phenotypes from multiple breeds that show different LD extents and gametic phases may help to identify causative mutations.

A survey on Apicomplexa protozoa in sheep slaughtered for human consumption.

Infections with the Apicomplexa Toxoplasma gondii, Neospora caninum, and Sarcocystis spp. are common causes of reproductive disorders in sheep. However, few epidemiological studies regarding co-infections with these three protozoa are reported in sheep in Italy. For this reason, this study aims to evaluate possible co-infections with T. gondii, N. caninum, and Sarcocystis spp. in sheep slaughtered for human consumption. From April to July 2019, individual blood, brain, heart, and diaphragm samples were collected from 138 sheep after slaughtering. The presence of IgG anti-Toxoplasma in serum samples was evaluated through ELISA. DNA of the three protozoa was investigated using specific PCRs. Co-infection with T. gondii, N. caninum and Sarcocystis spp. was found in 66.7% of the examined sheep. Antibodies against T. gondii were found in the 36.2% of serum samples. The presence of T. gondii DNA was detected in the 67.4%, 77.5%, and 21.7% of the brain, heart, and diaphragm samples, respectively. Neospora caninum DNA was found in 72.5% of the examined brain samples. Sarcocystis spp. DNA was detected in 92% and 52.2% of the heart and diaphragm samples, respectively. Sequence analysis of the Sarcocystis spp. revealed the sole presence of Sarcocystis tenella. The present study demonstrates that sheep have a high risk of infection with the three Apicomplexa investigated, suggesting the need to adopt adequate measures to prevent the spread of these parasitic infections considering their clinical and economic impact on ovine production. Furthermore, the possible role sheep play in the zoonotic transmission of toxoplasmosis to humans was highlighted.

First molecular description of Macracanthorhynchus hirudinaceus in wild boars from Italy with pathomorphological and epidemiological insights.

Macracanthorhynchus hirudinaceus is a zoonotic parasite affecting suids worldwide which are the definitive hosts for this helminth species. Macracanthorhynchus hirudinaceus is of significant economic and management concern due to its pathogenicity, causing intestinal obstruction and perforation in the definitive hosts. Current study is the preliminary investigation from Sardinia, Italy, reporting the pathomorphological findings and molecular characterization of M. hirudinaceus in the wild boars (Sus scrofa meridionalis). A total of 59 wild boars were examined showing acanthocephalan infection in 8 (13.6%) animals. In total, 49 parasites were collected with a mean intensity of 6.1. Comparatively higher infection levels were observed for males (16.7%) and young boars (14.3%); however, these epidemiological differences were statistically non-significant. Histopathological examination revealed the presence of a variable number of nodules (∼5 mm) in the intestine of M. hirudinaceus infested animals surrounded by a hyperemic-hemorrhagic halo. Several parasites were recovered from the intestinal lumen attached by the means of characteristic hooks showing necrosis in muscle layers. A moderate number of plump reactive fibroblasts and lesser numbers of fibrocytes were embedded with and at the borders of the inflammatory nodules in a moderate amount of homogeneous intensely eosinophilic fibrillary material rupturing the cell membrane. For molecular characterization, six isolated worms were amplified for the partial mitochondrial cox1 gene showing distinct interindividual variations. This first pathological and molecular description from southern Europe provided new knowledge about the diffusion of M. hirudinaceus in wild boars, furthering the research into the origin and transmission status of M. hirudinaceus in endemic localities.

First description of Eucoleus garfiai (Gallego and Mas-Coma, 1975) in wild boar (Sus scrofa) in Italy.

Eucoleus garfiai (syn. Capillaria garfiai) is a nematode infecting lingual tissue of domestic and wild swine. Prevalence data for this parasite are scant and often related to accidental findings, occurring only in Japan and a few European countries. In this study, an epidemiological survey was performed in order to identify E. garfiai in wild boar from the Campania region, southern Italy. A total of 153 wild boar carcasses were inspected over the course of two hunting seasons (2019-2020). Histological examinations were performed on tongue samples fixed and stained with haematoxylin and eosin. The scraping of dorsal tongue tissue was carried out to collect adult worms for parasitological examination. Out of 153 wild boars, 40 (26.1%, 95% CI: 19.8-33.6%) tested positive for helminths and/or eggs in tongue tissues. Parasites were identified morphologically and identification was confirmed by molecular analysis of the 18S rRNA gene, showing a 99% nucleotide match with E. garfiai sequences available in literature. No statistically significant differences were found according to age, sex nor hunting province. Our findings agree with previous histopathological data confirming the low pathogenic impact of this nematode. The present study represents the first report of E. garfiai in wild boar from Italy.

Cystic echinococcosis in sheep and goats of Lebanon.

Cystic echinococcosis (CE), a zoonotic disease caused by the species complex of Echinococcus granulosus sensu lato (s.l.), is endemic in Middle East and the Mediterranean basin, where pastoral activity is widespread. Despite the chronic endemicity of the disease in Lebanon and neighbouring countries, recent data are scant. The objectives of this survey were to evaluate the current epidemiology of CE in Lebanon, investigate the prevalence and determine the infecting genotypes in locally raised sheep and goats. A multidimensional approach combining post-mortem inspection of slaughtered animals and molecular diagnosis of the parasite was conducted to this end. From 2018 to 2020, 62.9% of the sheep and 20.9% of the goats were found positive for CE. The presence of hydatids varied between organs, showing higher prevalence in the liver of sheep vs the lungs of goats, however, a higher fertility rate of hydatid cyst was observed in lungs for both animals. Molecular diagnosis based on partial mitochondrial cox1 gene (795 bp) showed that the majority of isolates were identified as E. granulosus sensu stricto (98.7%) and only one isolate from goat was identified as Echinococcus canadensis (genotype G7; 1.3%). Echinococcus granulosus s.s. population among the sheep and goats was represented by 22 haplotypes having very little genetic differentiation and relatively moderate haplotype diversity. Population demographics explored through neutrality indices suggested expanding population within the intermediate hosts. These results document the high prevalence of CE in the livestock of Lebanon and reveal for the first time the presence of three different genotypes G1, G3 and G7.

Genetic diversity and transmission patterns of Echinococcus granulosus sensu stricto among domestic ungulates of Sardinia, Italy.

Cystic echinococcosis (CE), a parasitic zoonosis of public health and economic concern, is highly endemic in Sardinia, Italy. The study involved examining the intraspecific variability and demographic structure of Echinococcus granulosus sensu stricto (s.s.) in common hosts of this parasite. Molecular surveillance included the fragment amplification of a partial mitochondrial gene, cox1 (750 bp), for a total of 69 isolates derived from sheep (n = 52), cattle (n = 11), pigs (n = 4), and goats (n = 2). It was ascertained that E. granulosus s.s. was the primary agent of infection among these ungulates and G1 genotype was highly prevalent (79.71%). Considerable intraspecific variation was found, revealing the existence of 22 haplotypes with relatively high haplotype (0.8555 ± 0.033) and low nucleotide diversities (0.00281 ± 0.00030). Population demographics indicated an expanding parasitic population signifying negative deviation from neutrality indices. Little genetic differentiation was found between the subpopulations of E. granulosus s.s. in the island. Moreover, the geographic dispersal of genotypes G1 and G3 also indicated similarity between Sardinian and mainland Echinococcus granulosus s.s. populations reaffirming the sympatric occurrence and efficient transmission of G1 and G3 genotypes. Molecular survey of CE has the potential to yield baseline information on the infective genotypes among the intermediate hosts and helps in devising suitable control strategies for curtailing the disease.

Clinical forms of peritoneal larval cestodiasis by Mesocestoides spp. in dogs: diagnosis, treatment and long term follow-up.

Canine peritoneal larval cestodiasis (CPLC) is a little-known parasitological infestation of the peritoneal cavity of wild and domestic carnivores with Mesocestoides spp. larvae. While adult Mesocestoides tapeworms reside within the small intestine, the larvae occasionally penetrate the host's intestinal wall, causing a potentially life-threatening peritonitis. Severity of infection as well as the host response influences the prognosis significantly, and early diagnosis and treatment are essential. However, due to the lack of specific symptoms, this condition is underdiagnosed and, furthermore, no clear effective treatment has yet been described. The aim of this study is therefore to report two clinical cases of CPLC in dogs and to illustrate their clinical presentation and follow-up to serve as a reference for clinicians and researchers alike. Both animals were presented with abdominal distention as their main complaint. They underwent clinical examination, abdominal ultrasonography, abdominocentesis, and laparotomy followed by biochemical, cytological, parasitological, and molecular examination of the collected samples. After surgical lavage, the dogs received anthelmintic treatment with either fenbendazole (FBZ) or praziquantel (PZQ). Overall, timely and prolonged administration of high doses of FBZ seems to be the most effective treatment method. Irrespective, to date, no treatment capable of complete eradication of the infection and prevention of recurrence of disease has been found. In conclusion, further investigation into appropriate treatment plans as well as diagnostic development is needed.

Seroprevalence and risk factors associated with Leishmania infantum in dogs in Sardinia (Italy), an endemic island for leishmaniasis.

Leishmaniasis is a widespread, vector-borne parasitosis causing clinical manifestations in animals and in humans. In dogs, Canine Leishmaniasis has been reported in as much as 50 countries and the Mediterranean basin is known to be one of the most affected zones. Within these areas, the Island of Sardinia (Italy) has long been considered endemic for leishmaniasis and the presence of two arthropod vectors has recently been reported there. Nevertheless, to date, no epidemiological surveys regarding CanL have been carried out on the island. Hence, for the first time, the seroprevalence and the risk factors were investigated. Blood samples, as well as clinical and general information from 1.147 dogs, were collected and analyzed. Dogs consisted of two distinct populations, namely "owned dogs" and "kennel dogs." Anti-Leishmania IgG antibodies were detected using IFAT and samples were scored as positive at a cut-off dilution of 1:80. Data was analyzed using a Chi-squared test and bivariate and multivariate analyses were performed. Overall, 15.4% of dogs were found to be infected with CanL while only 44.1% of these animals exhibited clinical signs. Owned dogs (27.2%) were found to be infected more often than kennel dogs (10.6%); male dogs were found to be more frequently infected than female dogs and the number of infected animals increases with age. The present survey confirmed the endemic nature of leishmaniasis in Sardinia with a similar seroprevalence as mainland Italy. The results obtained serve as validation for the hypothesis that, in endemic areas, clinical CanL representations constitute only a fraction of the leishmaniasis cases.

Cystic echinococcosis in the endemic island of Sardinia (Italy): has something changed?

Cystic echinococcosis (CE) is a disease of livestock and humans caused by the larval stage of the Echinococcus granulosus sensu lato (s.l.) species complex and occurs especially in areas where sheep breeding is widespread. This survey evaluates, after more than a decade since the last epidemiological survey, the current situation and trend of CE in sheep of the highly endemic island of Sardinia (Italy). From 2012 to 2018, out of a total of 1416 sheep, 65.3% were found positive for CE. Cyst fertility has increased slightly from 10.3% in 2006 (Scala et al. 2006) to 11.7% in the present survey (χ2 = 1.07; P = 0.30018) with a peak of 14.0% in the province of Sassari. Overall, 6815 hydatids were found and classified as sterile (26.2%), fertile (14.4%), caseous (5.3%), or calcified (54.1%) based on their morphology. All isolates were identified as E. granulosus sensu stricto (former G1/G3, strains). Results shown as the implementation of new European Council Regulations and in particular of 21/2004 that established a system for the individual identification and tracking of sheep could be a step forward towards a better control of animal movement, preventing illegal slaughtering on the island as well as in other countries experiencing similar problems. Even if at first sight the general situation of Sardinia regarding CE seems a little better compared with the past, our findings indicate that in some Provinces the epidemiological situation is not improved since 2006.

Ultrasonography for early diagnosis of Toxocara canis infection in puppies.

Toxocara canis is one of the most common intestinal parasites in dogs and represents a highly infectious zoonotic parasite worldwide. Adult worms live in the bowel of dogs, and infections in puppies are commonly acquired transplacentally. The biology of the parasite and the commonly used diagnostic method, based on faecal examination, often prevent an early diagnosis of toxocariasis in puppies. Ultrasonographic diagnosis of intestinal ascariasis could be a feasible alternative method to diagnose T. canis infection in puppies during the prepatent period. The present study evaluated the diagnostic accuracy of ultrasonography for the diagnosis of intestinal toxocariasis in new-born puppies during the prepatent period. Fifty-four new-born puppies were prospectively recruited in the study. Each dog underwent intestinal ultrasonography and copro-microscopic examination every 5 days, starting from the 10th day post-birth. Intestinal ultrasonography visualized adult T. canis nematodes in the puppies' small intestine from the 10th day post-birth. On the 15th day after birth, ultrasonography showed 100% specificity and 85.4% sensitivity in diagnosing T. canis infection, despite negative results deriving from the copro-microscopic examination performed at the same time point. Our results showed that ultrasonography can be used as test for early diagnosis of T. canis infection in new-born puppies during the prepatent period. Early ultrasonographic diagnosis of T. canis infection in puppies could help control the disease in dogs and reduce the zoonotic risk for the human population.

Atypical outbreak of acute coenurosis by Taenia multiceps in a sheep flock.

Herein, we examined the brain of adult ewes and lambs less than 30 days old which were found affected by neurological signs in a flock located in Sardinia (Italy). Gross anatomo-pathological examination of all brains of the animals revealed multiple linear reddish-yellow foci of necrotic purulent inflammation due to oncosphere migration. Histologically, we confirmed a multifocal pyo-granulomatous meningo-encephalitis both in ewes and in lambs, confirming acute coenurosis. Morphological examination and DNA sequencing identified the Taenia multiceps we isolated as Tm1 strain. This report describes for the first time a natural acute coenurosis infection in suckling lambs under 30 days of age.

Seroepidemiological and biomolecular survey on Toxoplasma gondii in Sardinian wild boar (Sus scrofa).

Toxoplasma gondii is a zoonotic parasite able of infecting all warm-blooded animals. Toxoplasmosis is one of the major foodborne diseases globally. The consumption of wild boar (Sus scrofa) meat from recreational hunting has been linked to outbreaks of human toxoplasmosis. The island of Sardinia (Italy) contains a large wild boar population, thus providing an opportunity to assess the distribution of Toxoplasma in this species and the associated risks of transmission to humans. A total of 562 wild boars were screened: heart and meat juice samples were tested for T. gondii DNA via nested-PCR and IgG anti-Toxoplasma by commercial ELISA. Anti-Toxoplasma IgG were detected in 24.6% (138/562) of animals, while 37.2% (209/562) of the heart samples were PCR positive. The prevalence of T. gondii antibodies and DNA highlights the potential role of wild boar as an important reservoir for this parasite. The study suggests that wild boar could play a significant role in spreading the parasite to humans. As wild boar numbers are increasing throughout their range, their potential role in transmitting toxoplasmosis should be communicated to stakeholders, and the impact of different population control methods on disease transmission should be thoroughly assessed to mitigate potential threats effectively.

Hepatobiliary Ascariasis in a Piglet.

PURPOSE: Ascariasis caused by the helminth Ascaris suum is the most common parasitosis of swine worldwide and it may involve all age categories of pigs. The present study reports an unusual localization of A. suum worms in the biliary system of a piglet slaughtered for human consumption. METHODS: The liver was subjected to ultrasound scan and pathological examination. The isolated worms were morphologically examined and the DNA was extracted for the molecular identification of the species involved. RESULTS: A total of 43 preadult nematodes were found within the gallbladder and the bile ducts. Parasites were morphologically identified as belonging to the genus Ascaris and molecularly as A. suum. At gross examination, the liver was moderately enlarged, with the bile ducts severely dilated. A chronic inflammatory infiltrate was noted, often centered around ectatic bile ducts (up to 5 mm in diameter), lined by hyperplastic epithelium and filled with sections of nematodes. The worm sections showed smooth cuticle, coelomyarian musculature, and an intestinal tract lined by columnar, uninucleated cells within a pseudocoelom. The ex vivo ultrasonographic examination of the liver allowed the visualization of several nematodes in the bile duct lumen and could be suggested for in vivo diagnosis. Unfortunately, the absence of the intestine did not allow to define the pathogenesis of the infection. CONCLUSION: Although, given the unusual nature of this finding, it is difficult to identify predisposing factors for this A. suum localization, it suggests that ascariasis should be considered in the differential diagnosis of pigs with hepatobiliary disease.

The groundbreaking impact of digitalization and artificial intelligence in sheep farming.

The integration of digitalization and Artificial Intelligence (AI) has marked the onset of a new era of efficient sheep farming in multiple aspects ranging from the general well-being of sheep to advanced web-based management applications. The resultant improvement in sheep health and consequently better farming yield has already started to benefit both farmers and veterinarians. The predictive analytical models embedded with machine learning (giving sense to machines) has helped better decision-making and has enabled farmers to derive most out of their farms. This is evident in the ability of farmers to remotely monitor livestock health by wearable devices that keep track of animal vital signs and behaviour. Additionally, veterinarians now employ advanced AI-based diagnostics for efficient parasite detection and control. Overall, digitalization and AI have completely transformed traditional farming practices in livestock animals. However, there is a pressing need to optimize digital sheep farming, allowing sheep farmers to appreciate and adopt these innovative systems. To fill this gap, this review aims to provide available digital and AI-based systems designed to aid precision farming of sheep, offering an up-to-date understanding on the subject. Various contemporary techniques, such as sky shepherding, virtual fencing, advanced parasite detection, automated counting and behaviour tracking, anomaly detection, precision nutrition, breeding support, and several mobile-based management applications are currently being utilized in sheep farms and appear to be promising. Although artificial intelligence and machine learning may represent key features in the sustainable development of sheep farming, they present numerous challenges in application.

Laboratory associated zoonotic parasitic infections: a review of main agents and biosecurity measures.

Laboratory workers are exposed to the risk of acquiring infections due to the manipulation of infectious materials. The biological hazard for researchers is seven times higher when compared with hospital and public health laboratory workers. Despite the implementation of standardized practices to control infections, multiple cases of Laboratory Associated Infections (LAIs) usually go unreported. There has been a lack of comprehensive epidemiological data regarding the situation of LAIs for parasitic zoonosis and besides, the available sources are not completely updated. Since most accounts of laboratory infections are organism-specific, this study has focused on common pathogenic/zoonotic species handled at parasitological laboratories and summarising the standard biosecurity protocols for the infectious agents. The main characteristics of Cryptosporidium spp., Entamoeba spp, Giardia duodenalis, Toxoplasma gondii, Leishmania spp., Echinococcus spp., Schistosoma spp., Toxocara canis, Ancylostoma caninum, Strongyloides stercoralis are considered in this review in order to assess the potential risk of developing occupational infections in the workplace along with stating prevention and prophylactic measures for each species. It was concluded that the LAIs from these agents can be prevented by using personal protective measures and good laboratory practices. However, further studies are necessary to better understand the environmental resistance of cysts, oocysts and eggs, with a view to select the most suitable disinfection methods. Furthermore, it is fundamental to constantly update epidemiological data of infection acquired by laboratory workers, to develop accurate risk indicators.

Establishment and application of unbiased in vitro drug screening assays for the identification of compounds against Echinococcus granulosus sensu stricto.

Echinococcus multilocularis and E. granulosus s.l. are the causative agents of alveolar and cystic echinococcosis, respectively. Drug treatment options for these severe and neglected diseases are limited to benzimidazoles, which are not always efficacious, and adverse side effects are reported. Thus, novel and improved treatments are needed. In this study, the previously established platform for E. multilocularis in vitro drug assessment was adapted to E. granulosus s.s. In a first step, in vitro culture protocols for E. granulosus s.s. were established. This resulted in the generation of large amounts of E. granulosus s.s. metacestode vesicles as well as germinal layer (GL) cells. In vitro culture of these cells formed metacestode vesicles displaying structural characteristics of metacestode cysts generated in vivo. Next, drug susceptibilities of E. multilocularis and E. granulosus s.s. protoscoleces, metacestode vesicles and GL cells were comparatively assessed employing established assays including (i) metacestode vesicle damage marker release assay, (ii) metacestode vesicle viability assay, (iii) GL cell viability assay, and (iv) protoscolex motility assay. The standard drugs albendazole, buparvaquone, mefloquine, MMV665807, monepantel, niclosamide and nitazoxanide were included. MMV665807, niclosamide and nitazoxanide were active against the parasite in all four assays against both species. MMV665807 and monepantel were significantly more active against E. multilocularis metacestode vesicles, while albendazole and nitazoxanide were significantly more active against E. multilocularis GL cells. Albendazole displayed activity against E. multilocularis GL cells, but no effects were seen in albendazole-treated E. granulosus s.s. GL cells within five days. Treatment of protoscoleces with albendazole and monepantel had no impact on motility. Similar results were observed for both species with praziquantel and its enantiomers against protoscoleces. In conclusion, in vitro culture techniques and drug screening methods previously established for E. multilocularis were successfully implemented for E. granulosus s.s., allowing comparisons of drug efficacy between the two species. This study provides in vitro culture techniques for the reliable generation of E. granulosus s.s. metacestode vesicles and GL cell cultures and describes the validation of standardized in vitro drug screening methods for E. granulosus s.s.