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Inflammatory bowel disease and allergic asthma, as typical immune-mediated inflammatory diseases (IMIDs), are associated with immune imbalance caused by complex interactions among environmental, genetic and bacterial factors. The changing immune imbalance of IMIDs not only causes serious pathological damages but also increases the difficulty of treatment. Helminths or helminth-derived molecules have been increasingly employed to treat IMIDs due to their immunoregulatory ability. Since helminth infection is not an appropriate treatment direction due to the complex immunoregulation and safety concerns, one of the new therapies is to harness the immunoregulation induced by the identified helminth-derived molecules using immune indexes as a guide. This review discusses the pathogenesis of inflammatory bowel disease and allergic asthma, and summarizes the therapeutic effect of helminths and the immunoregulatory mechanisms induced by helminth-derived molecules proposing therapeutic regimens.

Background Major prevention trials for Alzheimer’s disease (AD) are now focusing on multidomain lifestyle interventions. However, the exact combination of behavioral factors related to AD pathology remains unclear. In 2 cohorts of cognitively unimpaired individuals at risk of AD, we examined which combinations of personality traits, neuropsychiatric symptoms, and cognitive lifestyle (years of education or lifetime cognitive activity) related to the pathological hallmarks of AD, amyloid-β, and tau deposits. Methods A total of 115 older adults with a parental or multiple-sibling family history of sporadic AD (PREVENT-AD [PRe-symptomatic EValuation of Experimental or Novel Treatments for AD] cohort) underwent amyloid and tau positron emission tomography and answered several questionnaires related to behavioral attributes. Separately, we studied 117 mutation carriers from the DIAN (Dominant Inherited Alzheimer Network) study group cohort with amyloid positron emission tomography and behavioral data. Using partial least squares analysis, we identified latent variables relating amyloid or tau pathology with combinations of personality traits, neuropsychiatric symptoms, and cognitive lifestyle. Results In PREVENT-AD, lower neuroticism, neuropsychiatric burden, and higher education were associated with less amyloid deposition (p = .014). Lower neuroticism and neuropsychiatric features, along with higher measures of openness and extraversion, were related to less tau deposition (p = .006). In DIAN, lower neuropsychiatric burden and higher education were also associated with less amyloid (p = .005). The combination of these factors accounted for up to 14% of AD pathology. Conclusions In the preclinical phase of both sporadic and autosomal dominant AD, multiple behavioral features were associated with AD pathology. These results may suggest potential pathways by which multidomain interventions might help delay AD onset or progression.

Trichinella spiralis is recognized for its ability to regulate host immune responses via excretory/secretory (ES) products. Serine protease inhibitors (serpins) play an important role in ES product-mediated immunoregulatory effects during T. spiralis infection. In this study, the immunoregulatory properties of a serpin derived from T. spiralis (Ts-serpin) were explored in BALB/c mice. The results showed that naturally occurring Ts-serpin was detected in the stichosomes of muscle larvae and adult worms. Moreover, enhancing (by injection of a soluble-expressed recombinant Ts-serpin [rTs-serpin]) or blocking (by passive immunization with anti–rTs-serpin serum) the effects of Ts-serpin changed the levels of cytokines related to inflammation induced by T. spiralis infection in the serum, mesenteric lymph nodes, and peritoneal cavity, which then led to a change in the adult worm burden in early T. spiralis infection. Moreover, the phenotypic changes in peritoneal macrophages were found to be related to Ts-serpin–mediated immunoregulation. Furthermore, a STAT6 activation mechanism independent of IL-4Rα has been found to regulate protein-mediated alternative activation of bone marrow–derived macrophages and mimic the immunoregulatory role of Ts-serpin in T. spiralis infection. Finally, the anti-inflammatory properties of rTs-serpin and bone marrow–derived macrophage alternative activation by rTs-serpin were demonstrated using a trinitrobenzene sulfonic acid–induced inflammatory bowel disease model. In summary, a protein-triggered anti-inflammatory mechanism was found to favor the survival of T. spiralis in the early stage of infection and help to elucidate the immunoregulatory effects of T. spiralis on the host immune response.

In September 2017, a severe trichinellosis outbreak occurred in Cambodia after persons consumed raw wild pig meat; 33 persons were infected and 8 died. We collected and analyzed the medical records for 25 patients. Clinical signs and symptoms included myalgia, facial or peripheral edema, asthenia, and fever. We observed increased levels of creatine phosphokinase and aspartate aminotransferase-, as well as eosinophilia. Histopathologic examination of muscle biopsy specimens showed nonencapsulated Trichinella larvae. A Trichinella excretory/secretory antigen ELISA identified Trichinella IgM and IgG. Biopsy samples were digested and larvae were isolated and counted. PCR for the 5S rDNA intergenic spacer region and a multiplex PCR, followed by sequencing identified the parasite as Trichinella papuae. This species was identified in Papua New Guinea during 1999 and in several outbreaks in humans in Thailand. Thus, we identified T. papuae nematodes in humans in Cambodia.

Parasites have developed many strategies to ensure their development, multiplication, and dissemination, including the use of reservoir hosts that are often nondomesticated species. Despite drastic reductions in their populations, wild birds remain widespread worldwide and could constitute some of these reservoirs. We focused on the identification of wild bird species harboring parasite stages in their muscles. Breast muscles of 327 birds of 27 different species were collected at three different sites in France. After artificial digestion, isolated nematode larvae were identified by PCR sequencing or restriction fragment length polymorphism (PCR-RFLP). Toxocara cati was identified mainly in birds of prey. The presence of anti-Toxoplasma antibodies was investigated by modified agglutination test on muscle fluids. Anti-Toxoplasma antibodies were detected in 65 out of 166 samples from various bird species. Avifauna, particularly birds of prey, could help on the surveillance of parasite circulation and play a role as sentinel species.

We describe a small family outbreak of trichinellosis caused by the consumption of raw ham from a wild boar (Sus scrofa) hunted in the northern Alps of France in February 2022. Out of the six people, aged 3–69 years, who consumed the meat, three were confirmed cases, and three were suspected cases. Eosinophilia detected in four people was the hallmark that drove the diagnosis. Three patients presented with myalgia, two with intense and prolonged chest pain, and one with elevated troponin. One patient presented with dermographism during treatment. Anti-Trichinella IgG were detected in three symptomatic individuals after about ten weeks. One patient had negative serology and no symptoms, but was on long-term corticosteroid therapy. Trichinella britovi larvae (8.3 larvae/g) were detected in the wild boar meat remnants. Trichinellosis is rare in France, but this family outbreak is reminiscent of the circulation of this pathogen in wild animals, highlighting the need to inform hunters about the risk of infection linked to the consumption of raw meat of game animals, and about the need for veterinary inspection of game meat. The consumption of raw meat outside controlled circuits is a practice not devoid of risks, which justifies raising the awareness of hunters, doctors, and medical biologists.

Ethnopharmacological Relevance: An ethnobotanical survey was conducted in three regions of the country (two rural and one urban), using a questionnaire focussing on anti-diarrheal therapeutic habits. Methodology: Information about the plants has been recorded (local name, organs or part(s) of the plant used, therapeutic indications, harvesting methods, methods of administration, side effects, etc.). Collection of the plants was done in collaboration with traditional practitioners and identification of the specimens was conducted at the National Floristic Center (Ivory coast). During the ethnobotanical survey, twelve traditional healers and nine breeders were interviewed which all declared to treat diarrhoea with plants. During this survey, twenty-seven species belonging to eighteen different families of plants were reported for anti-diarrhoeal use in humans or animals. They were all harvested on the field and a herbarium of each species was prepared in duplicate, one stored at the National Floristic Centre of the Felix Houphouët-Boigny University and the other at the Pasteur Institute of Ivory coast. Results: Decoction was the most common mode of preparation used by practionioners and the oral route remained also the main way of administration of plants by healers. These plants are also used in other countries for the same or other purpose. Conclusion: This study shows that traditional medicinal plants play an important role in the treatment of diarrhoea in Ivory coast. It provides basis for future studies to assess, biological and chemical potential of these plants.

Background: More than a hundred species of mammals, birds, and reptiles are infected by nematodes of the Trichinella genus worldwide. Although, Trichinella spp. are widely distributed in neighboring countries including Georgia, Azerbaijan, Turkey and Iran, no study was conducted in Armenia since 1980’s. Methods: In 2017-2018, five muscle samples belonging to Armenian lynx, otter, wild boar, fox and wolf were tested for Trichinella spp. and recovered larvae were identified by multiplex PCR technique. Results: Twenty-six larvae/gram and one larva/gram were found in lynx and fox samples respectively. They were identified as T. britovi. Conclusion: So far only two species were identified in Armenia, T. spiralis and T. pseudospiralis, and this is the first time that T. britovi is reported in Armenia.

Summary The wild boar (Sus scrofa) has a wide geographical distribution and can be an important source of Trichinella spp. infection in humans in Romania. The objective of this study was to identify the presence of Trichinella spp. in the wild boar population in Bihor County, Romania. Eighty four plasma and diaphragm samples, collected from wild boars, were included in this study. Artificial digestion, ELISA and Western blot were performed on these specimens. All diaphragm samples were negative for Trichinella larvae in artificial digestion, while in ELISA, 54 (64.2 %) plasma samples were positive and 6 (7.1 %) plasma samples were doubtful. Western blot was performed on 26 plasma samples from which only 6 (23.0 %) gave a positive result. Serological evidences indicate the presence of Trichinella spp. in wild boars from western Romania. Therefore, human consumers might be at risk to ingest Trichinella larvae, even in low numbers.

Trichinellosis is a rare parasitic zoonosis in the European Union. Meat from backyard pigs was the common source for a trichinellosis outbreak caused by Trichinella spiralis, which occurred in France and Serbia in the beginning of 2017. An epidemiological study was conducted in France and Serbia to determine the extent of the outbreak, to identify its source and to implement control measures. Three cases were exposed in Serbia and brought back to France pork delicatessen which they shared with relatives and friends. Around 47 individuals were exposed to the parasitised meat in France and Serbia and 20 cases of trichinellosis were reported (nine in France and 11 in Serbia). Nine of them were female. The diagnosis was delayed, in part because the parasitosis was not known by most physicians, which led to complications in the French cases such as facial paralysis and pulmonary embolism. Health alerts and survey networks are indispensable at a European level to control the disease.

Background The zoonotic nematode Trichinella britovi was discovered in two neighboring Mediterranean islands of Corsica and Sardinia, almost simultaneously at the beginning of the 21st century. An epidemiological link between the two parasite populations was generally assumed. In 2015, an outbreak of trichinellosis in Nice, the South of France, was reportedly caused by the consumption of raw pork delicatessen imported from Corsica. The aims of the present study were to investigate, by multilocus genotype (MLG) analyses, the hypothesis of the common origin of the Corsican and Sardinian T. britovi foci and to trace “from fork to farm” the origin of the pork product, which caused a trichinellosis outbreak in mainland France in 2015. Methods Sixty-three T. britovi isolates were collected from animals and pork products of Sardinia and Corsica islands and from mainland of Italy, France and Spain. We analyzed genetic variability at four polymorphic microsatellite loci by two independent algorithms, the Bayesian and multivariate analyses, to evaluate the genetic relationships of 1367 single larvae. Results Trichinella britovi isolates of the two islands showed different genetic structures and the Bayesian analysis revealed a different membership of the two insular populations. Furthermore, two geographically separate genetic groups were identified among Corsican isolates. Lastly, the origin of the pork delicatessen marketed in Nice was linked to a breeder-butcher in Corsica. Conclusions The low level of genetic admixture of the insular T. britovi isolates suggests that this pathogen colonized the two islands by separate events. On the other hand in Corsica, although the isolates share the same genetic structure, geographically separate isolates showed different membership. We suggest the MLG analysis as a suitable method in supporting epidemiological investigations to trace “from fork to farm” insular populations of T. britovi. Electronic supplementary material The online version of this article (10.1186/s13071-018-2939-9) contains supplementary material, which is available to authorized users.

Trichinella spiralis typically infects domestic swine, wild boar and occasionally horses, has a cosmopolitan distribution, and consequently is most frequently associated with food-borne outbreaks of trichinellosis in humans. Trichinella murrelli is typically found in wild carnivores in temperate areas of North America, where it has been responsible for outbreaks of human trichinellosis due to consumption of infected wild game. There has previously been only indirect evidence of natural infection with T. murrelli in a horse originating from Connecticut and implicated in an outbreak of trichinellosis in France in 1985. We describe a T. murrelli infection detected during routine testing of a horse from the USA imported to Canada for slaughter and export to the European Union (EU). Approximately 5 or more larvae per gram were recovered from digested tongue and diaphragm samples and identified as T. murrelli by PCR. This case provides the first direct evidence of naturally acquired T. murrelli infection in a horse, and further supports the potential food safety risk posed by this parasite species. It is the first instance in Canada of the detection of a Trichinella-infected horse via routine post-mortem testing. Trichinella spiralis-infected horses have been similarly detected by regulatory testing in France, and further details of two such previously reported cases are also provided here. The cases described herein underscore the importance of continued vigilance in quality assured food safety testing of horse meat to mitigate the risk of human trichinellosis.

15 páginas, 3 figuras y 2 tablas., Environmental and anthropogenic changes are expected to promote emergence and spread of pathogens worldwide. Since 2013, human urogenital schistosomiasis is established in Corsica island (France). Schistosomiasis is a parasitic disease affecting both humans and animals. The parasite involved in the Corsican outbreak is a hybrid form between Schistosoma haematobium, a human parasite, and Schistosoma bovis, a livestock parasite. S. bovis has been detected in Corsican livestock few decades ago raising the questions whether hybridization occurred in Corsica and if animals could behave as a reservoir for the recently established parasite lineage. The latter hypothesis has huge epidemiological outcomes since the emergence of a zoonotic lineage of schistosomes would be considerably harder to control and eradicate the disease locally and definitively needs to be verified. In this study we combined a sero-epidemiological survey on ruminants and a rodent trapping campaign to check whether schistosomes could shift on vertebrate hosts other than humans. A total of 3,519 domesticated animals (1,147 cattle; 671 goats and 1,701 sheep) from 160 farms established in 14 municipalities were sampled. From these 3,519 screened animals, 17 were found to be serologically positive but were ultimately considered as false positive after complementary analyses. Additionally, our 7-day extensive rodent trapping (i.e. 1,949 traps placed) resulted in the capture of a total of 34 rats (Rattus rattus) and 4 mice (Mus musculus). Despite the low number of rodents captured, molecular diagnostic tests showed that two of them have been found to be infected by schistosomes. Given the low abundance of rodents and the low parasitic prevalence and intensity among rodents, it is unlikely that neither rats nor ruminants play a significant role in the maintenance of schistosomiasis outbreak in Corsica. Finally, the most likely hypothesis is that local people initially infected in 2013 re-contaminated the river during subsequent summers, however we cannot definitively rule out the possibility of an animal species acting as reservoir host., JB received a grant from the ANSES (Agence nationale de sécurité sanitaire de l’alimentation, de l’environnement et du travail) (https://www.anses.fr/fr). This study is set within the framework of the "Laboratoire d’Excellence (LABEX)" TULIP (ANR-10-LABX-41).

Trichinellosis is a serious foodborne zoonotic disease. It is an important threat to public health in both developing and developed countries. Human infections are strongly associated with consuming undercooked meat containing infective Trichinella larvae. The development of serological tools has enabled seroepidemiological studies and contributed to our knowledge on the importance of this parasite. Serological tests can also help the diagnosis of parasite infections in humans and the surveillance of animals. Generally speaking, serological techniques include detection methods for specific antibodies and for circulating parasite antigens in the serum or tissue fluids. Here, we present a comprehensive review of various methods used in the detection of antibodies against Trichinella and circulating parasite antigens in animals and humans., Highlights • Trichinellosis is considered as a public health problem in several countries around the world. • The development of immunodiagnostic techniques has promoted the conduct of seroepidemiological studies. • This review provides insight into the evolution of these techniques.

Trichinella spiralis typically infects domestic swine, wild boar and occasionally horses, has a cosmopolitan distribution, and consequently is most frequently associated with food-borne outbreaks of trichinellosis in humans. Trichinella murrelli is typically found in wild carnivores in temperate areas of North America, where it has been responsible for outbreaks of human trichinellosis due to consumption of infected wild game. There has previously been only indirect evidence of natural infection with T. murrelli in a horse originating from Connecticut and implicated in an outbreak of trichinellosis in France in 1985. We describe a T. murrelli infection detected during routine testing of a horse from the USA imported to Canada for slaughter and export to the European Union (EU). Approximately 5 or more larvae per gram were recovered from digested tongue and diaphragm samples and identified as T. murrelli by PCR. This case provides the first direct evidence of naturally acquired T. murrelli infection in a horse, and further supports the potential food safety risk posed by this parasite species. It is the first instance in Canada of the detection of a Trichinella-infected horse via routine post-mortem testing. Trichinella spiralis-infected horses have been similarly detected by regulatory testing in France, and further details of two such previously reported cases are also provided here. The cases described herein underscore the importance of continued vigilance in quality assured food safety testing of horse meat to mitigate the risk of human trichinellosis., Highlights • First direct evidence of natural infection with Trichinella murrelli in a horse • First detection in Canada of a Trichinella-infected horse via regulated testing • Trichinella murrelli should be considered a potential food safety risk in horsemeat. • Horse tongue and diaphragm are effective for detecting T. murrelli and T. spiralis. • Screening of horse carcasses by digestion assay mitigates risk of trichinellosis.

Serine proteases form one of the most important families of enzymes and perform significant functions in a broad range of biological processes, such as intra- and extracellular protein metabolism, digestion, blood coagulation, regulation of development, and fertilization. A number of serine proteases have been identified in parasitic helminths that have putative roles in parasite development and nutrition, host tissues and cell invasion, anticoagulation, and immune evasion. In this review, we described the serine proteases that have been identified in parasitic helminths, including nematodes (Trichinella spiralis, T. pseudospiralis, Trichuris muris, Anisakis simplex, Ascaris suum, Onchocerca volvulus, O. lienalis, Brugia malayi, Ancylostoma caninum, and Steinernema carpocapsae), cestodes (Spirometra mansoni, Echinococcus granulosus, and Schistocephalus solidus), and trematodes (Fasciola hepatica, F. gigantica, and Schistosoma mansoni). Moreover, the possible biological functions of these serine proteases in the endogenous biological phenomena of these parasites and in the host-parasite interaction were also discussed.

Giardia duodenalis (syn. G. lamblia, G. intestinalis) is the protozoan parasite responsible for giardiasis, the most common and widely spread intestinal parasitic disease worldwide, affecting both humans and animals. After cysts ingestion (through either contaminated food or water), Giardia excysts in the upper intestinal tract to release replicating trophozoites that are responsible for the production of symptoms. In the gut, Giardia cohabits with the host's microbiota, and several studies have revealed the importance of this gut ecosystem and/or some probiotic bacteria in providing protection against G. duodenalis infection through mechanisms that remain incompletely understood. Recent findings suggest that Bile-Salt-Hydrolase (BSH)-like activities from the probiotic strain of Lactobacillus johnsonii La1 may contribute to the anti-giardial activity displayed by this strain. Here, we cloned and expressed each of the three bsh genes present in the L. johnsonii La1 genome to study their enzymatic and biological properties. While BSH47 and BSH56 were expressed as recombinant active enzymes, no significant enzymatic activity was detected with BSH12. In vitro assays allowed determining the substrate specificities of both BSH47 and BSH56, which were different. Modeling of these BSHs indicated a strong conservation of their 3-D structures despite low conservation of their primary structures. Both recombinant enzymes were able to mediate anti-giardial biological activity against Giardia trophozoites in vitro. Moreover, BSH47 exerted significant anti-giardial effects when tested in a murine model of giardiasis. These results shed new light on the mechanism, whereby active BSH derived from the probiotic strain Lactobacillus johnsonii La1 may yield anti-giardial effects in vitro and in vivo. These findings pave the way toward novel approaches for the treatment of this widely spread but neglected infectious disease, both in human and in veterinary medicine.

Lysine acetylation is a dynamic and highly conserved post-translational modification that plays a critical role in regulating diverse cellular processes. Trichinella spiralis is a foodborne parasite with a considerable socio-economic impact. However, to date, little is known regarding the role of lysine acetylation in this parasitic nematode. In this study, we utilized a proteomic approach involving anti-acetyl lysine-based enrichment and highly sensitive mass spectrometry to identify the global acetylated proteome and investigate lysine acetylation in T. spiralis. In total, 3872 lysine modification sites were identified in 1592 proteins that are involved in a wide variety of biological processes. Consistent with the results of previous studies, a large number of the acetylated proteins appear to be involved in metabolic and biosynthetic processes. Interestingly, according to the functional enrichment analysis, 29 acetylated proteins were associated with phagocytosis, suggesting an important role of lysine acetylation in this process. Among the identified proteins, 15 putative acetylation motifs were detected. The presence of serine downstream of the lysine acetylation site was commonly observed in the regions surrounding the sites. Moreover, protein interaction network analysis revealed that various interactions are regulated by protein acetylation. These data represent the first report of the acetylome of T. spiralis and provide an important resource for further explorations of the role of lysine acetylation in this foodborne pathogen.

A clone, designated L20h-Ts3, was selected by immunoscreening of cDNA libraries of Trichinella spiralis worms collected 14h, 20h and 48h post-infection (p.i.) from mice intestines. L20h-Ts3 encodes the full-length of a conserved hypothetical protein of 13.1kDa involving putative interaction with the immune system. PCR analysis showed that L20h-Ts3 mRNA is constitutively expressed throughout T. spiralis life cycle and not restricted to intestinal stages. The L20h-Ts3 fusion protein was obtained in an Escherichia coli expression system and purified by Ni-affinity chromatography before inoculation into mice in order to produce polyclonal antibodies. Then, immunohistochemical study and Western blot analysis revealed its presence within the stichosome of T. spiralis and in excretory/secretory products strengthening a putative fundamental role for the parasite's survival such as host tissue invasion or modification of the host muscular cell phenotype. L20h-Ts3 fusion protein was recognized in Western blot as soon as 15–20 days p.i. by sera from pigs experimentally infected with 20,000 muscle larvae (ML) of T. spiralis. Thus, an indirect L20h-Ts3 ELISA was designed and evaluated using sera from experimentally infected pigs by comparison with the only ELISA currently available for trichinellosis purposes. A gain of precocity from 7 up to 14 days and detection up to 25 weeks p.i. was possible with the L20h-Ts3 ELISA offering a large window for trichinellosis detection. The L20h-Ts3 ELISA was less effective in the case of low infections in pigs. Nevertheless, these results show that the L20h-Ts3 ELISA has a real interest due to its precocity and stability of detection in time. The association of the L20h-Ts3 fusion protein with other antigenic proteins identified previously could appreciably improve the serological test and facilitate its standardization.

International audience; To date, no information is available about the presence of Cryptosporidium spp. in French sheep, nor their potential role as zoonotic reservoirs. A total of 23 fecal samples were collected from diarrheic lambs (< 11 days old) from seven randomly selected farms. Cryptosporidium-oocysts were detected microscopically with Direct Immunofluorescence Assays (DFA) in 23/23 (100%) of fecal samples. PCR-RFLP of the 18S rRNA gene was used to determine species in all samples, and only Cryptosporidium parvum was identified. Isolates were subtyped by sequencing the 60 kDa glycoprotein (gp60) gene. Two zoonotic subtypes within the IIa subtype family were identified, including IIaA15G2R1 (22/23) and IIaA16G3R1 (1/23). This study reports for the first time the identification and genotyping of zoonotic C. parvum subtypes from lambs in France. Sheep could thus play an important role as potential reservoirs for this zoonotic protist.

Cryptosporidium is an obligate intracellular protist parasite infecting a wide range of vertebrate hosts and causes significant intestinal disease in both animals and humans, as some species are zoonotic. Cattle and especially calves have been identified as one of the most common reservoirs of this protist. However, little is known about the genetics of Cryptosporidium in calves in some regions of France. The aim of this study was to detect and isolate Cryptosporidium spp. in faecal samples from naturally infected pre-weaned calves (≤45 days-old) in France. A total of 35 diarrhoeic pre-weaned calf faecal samples were collected from 26 dairy cattle farms in six departments (French administrative provinces). Cryptosporidium presence was established by microscopically screening samples for oocystes with an immunofluorescent (DFA) staining method. DFA-positive samples were then analysed by PCR-RFLP and 18S rRNA gene sequencing to determine species. Cryptosporidium parvum-positive samples were subtyped via nested PCR analysis of a partial fragment of the 60 kDa glycoprotein (gp60) gene product. Data were then integrated into phylogenetic tree analysis. DFA revealed the presence of Cryptosporidium oocysts in 31 out of 35 (88%) samples. Combined with 18S rRNA gene analysis results, C. parvum was detected in 30 samples. Subtyping analysis in 27/30 samples (90%) of the C. parvum isolates revealed two zoonotic subtype families, IIa (24/27) and IId (3/27). Four subtypes were recognised within the subtype family IIa, including the hypertransmissible IIaA15G2R1 subtype that is the most frequently reported worldwide (21/27), IIaA17G3R1 (1/27), IIaA17G1R1 (1/27), and IIaA19G1R1 (1/27). Two subtypes were recognised within the IId subtype family including IIdA22G1 (2/27) and IIdA27G1 (1/27). These findings illustrate the high occurrence of Cryptosporidium in calves in dairy herds and increase the diversity of molecularly characterised C. parvum isolates with the first description of IIaA17G3R1, IIaA19G1R1, and IId subtypes in France. The presence of zoonotic C. parvum subtype families (IIa, IId) in this study suggests that pre-weaned calves are likely to be a significant reservoir of zoonotic C. parvum, and highlights the importance of animal to human cryptosporidiosis transmission risk. Further molecular studies in calves and small ruminants from other French regions are required to better understand the epidemiology of cryptosporidiosis in France., Highlights • Faecal samples from pre-weaned diarrheal calves were analysed • Cryptosporidium spp. was detected in 30 samples out of 35. • C. parvum was the only species identified • Two zoonotic subtype families were identified: IIa and IId • The hyper-transmissible IIaA15G2R1 was the dominant C. parvum subtype

International audience; Toxoplasma gondii (Nicolle et Manceaux, 1908) is an obligate intracellular, parasitic protozoan within the phylum Apicomplexa that causes toxoplasmosis in mammalian hosts (including humans) and birds. Since meat of wild boar, Sus scrofa (Linnaeus), has been demonstrated to be a potential source of human infection, a careful evaluation of the prevalence of infection with T. gondii in hunted animals is needed to protect public health. In the Var area in southeastern France, we performed a spatio-temporal survey in order to investigate the prevalence of IgG antibodies in wild boars shot by hunters in the Canjuers military camp during two subsequent hunting seasons. Of 841 wild boars screened, antibodies (IgG) to T. gondii (modified agglutination test, cut-off 1 : 6) were found in 141 (16.8%) muscle extract samples. A significant association (p < 0.001) was found between positivity and age, but not gender, and hunting districts. The results obtained indicate that consumption of raw or undercooked meat from wild boars carries an important risk of infection with T. gondii. Wild boars may be considered as a bioindicator of parasite circulation in this ecosystem.

SUMMARYThe aim of this study was to assess the seroprevalence of the Toxoplasma gondii parasite in pork produced in France, and to determine infection risk factors. An innovative survey was designed based on annual numbers of slaughtered pigs from intensive and outdoor farms in France. A total of 1549 samples of cardiac fluids were collected from pig hearts to determine seroprevalence using a Modified Agglutination Test. Of those, 160 hearts were bio-assayed in mice to isolate live parasites. The overall seroprevalence among fattening pigs was 2·9%. The adjusted seroprevalence in pigs from intensive farms was 3·0%; the highest in sows (13·4%); 2·9% in fattening pigs and 2·6% in piglets. Adjusted seroprevalence in fattening animals from outdoor farms was 6·3%. Strains were isolated from 41 animals and all were genotyped by Restriction Fragment Length Polymorphism as type II. Risk-factor analysis showed that the risk of infection was more than three times higher for outdoor pigs, and that sows’ risk was almost five times higher than that of fattening animals. This study provides further evidence of extensive pork infection with T. gondii regardless of breeding systems, indicating that farm conditions are still insufficient to guarantee ‘Toxoplasma-free pork’.

The trematode Alaria alata is a cosmopolite parasite found in red foxes (Vulpes vulpes), the main definitive host in Europe. In contrast only few data are reported in wild boars (Sus scrofa), a paratenic host. The aim of this paper is to describe the importance and distribution of Alaria alata mesocercariae in wild boars, information is given by findings of these larvae during Trichinella mandatory meat inspection on wild boars’ carcasses aimed for human consumption. More than a hundred cases of mesocercariae positive animals are found every year in the East of France. First investigations on the parasite’s resistance to deep-freezing in meat are presented in this work.

Each year, more than 167 million pigs in the European Union (EU) are tested for Trichinella spp. under the current meat hygiene regulations. This imposes large economic costs on countries, yet the vast majority of these pigs test negative and the public health risk in many countries is therefore considered very low. This work reviewed the current Trichinella status across the EU as well as the national level of monitoring and reporting. It also reviewed which animal species were affected by Trichinella and in which species it should be surveyed. This information was used to design a cost-effective surveillance programme that enables a standardised monitoring approach within the EU. The proposed surveillance programme relies on identifying sub-populations of animals with a distinct risk. Low-risk pigs are finisher pigs that originate from so-called controlled housing. All other pigs are considered high-risk pigs. Controlled housing is identified by the application of a specific list of management and husbandry practices. We suggest that member states (MS) be categorised into three classes based on the confidence that Trichinella can be considered absent, in the specified sub-population of pigs above a specified design prevalence which we set to 1 per million pigs. A simple and transparent method is proposed to estimate this confidence, based on the sensitivity of the surveillance system, taking into account the sensitivity of testing and the design prevalence. The probability of detecting a positive case, if present, must be high (>95 or >99%) to ensure that there is a low or negligible risk of transmission to humans through the food chain. In MS where the probability of a positive pig is demonstrated to be negligible, testing of fattening pigs from a sub-population consisting of pigs from controlled housing can be considered unnecessary. Furthermore, reduced testing of finishers from the sub-population consisting of pigs from non-controlled housing might even be considered, if conducted in conjunction with a proportionate sampling scheme and a risk-based wildlife surveillance programme where applicable. The proposed surveillance programme specifies the required number of samples to be taken and found negative, in a MS. A MS with no data or positive findings will initially be allocated to class 1, in which all pigs should be tested. When a MS is able to demonstrate a 95% or 99% confidence that Trichinella is absent, the MS will be allocated to class 2 or 3, in which the testing requirement is lower than in class 1.

International audience; Pepsin powder constitutes a health risk, potentially causing severe allergic reactions to those handling the chemical. A fluid pepsin formulation was produced and tested, first in a preliminary study and then in a ring trial encompassing four European National Reference Laboratories (NRLs). The purpose of each trial was to ascertain and compare the action of pepsin powder with that of the pepsin fluid for digesting meat and liberating encapsulated Trichinella spiralis larvae for subsequent counting. The quality of digestion was furthermore evaluated by assessing the visibility through the digestion fluid and the amount of debris remaining after digestion. For the ring trial, at each laboratory 20 blinded replicate 100-g samples of pork meat containing a known number of encapsulated T. spiralis larvae (0 to 30) were digested by the magnetic stirrer method using either the standard pepsin powder (10 samples) or the pepsin fluid (10 samples). With an average recovery rate of 70 to 80%, all NRLs found the pepsin fluid and pepsin powder to be equally effective. The NRLs also found no difference between the two pepsin formulations with regard to debris remnants or visibility through the digestion fluid. The use of pepsin fluid may therefore constitute an improvement of the digestion procedure for the analysts involved.

African swine fever (ASF) is an asymptomatic infection of warthogs and bushpigs, which has become an emergent disease of domestic pigs, characterized by hemorrhage, lymphopenia, and disseminated intravascular coagulation. It is caused by a large icosohedral double-stranded DNA virus, African swine fever virus (ASFV), with infection of macrophages well characterized in vitro and in vivo. This study shows that virulent isolates of ASFV also infect primary cultures of porcine aortic endothelial cells and bushpig endothelial cells (BPECs) in vitro. Kinetics of early and late gene expression, viral factory formation, replication, and secretion were similar in endothelial cells and macrophages. However, ASFV-infected endothelial cells died by apoptosis, detected morphologically by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling and nuclear condensation and biochemically by poly(ADP-ribose) polymerase (PARP) cleavage at 4 h postinfection (hpi). Immediate-early proinflammatory responses were inhibited, characterized by a lack of E-selectin surface expression and interleukin 6 (IL-6) and IL-8 mRNA synthesis. Moreover, ASFV actively downregulated interferon-induced major histocompatibility complex class I surface expression, a strategy by which viruses evade the immune system. Significantly, Western blot analysis showed that the 65-kDa subunit of the transcription factor NF-κB, a central regulator of the early response to viral infection, decreased by 8 hpi and disappeared by 18 hpi. Both disappearance of NF-κB p65 and cleavage of PARP were reversed by the caspase inhibitor z-VAD-fmk. Interestingly, surface expression and mRNA transcription of tissue factor, an important initiator of the coagulation cascade, increased 4 h after ASFV infection. These data suggest a central role for vascular endothelial cells in the hemorrhagic pathogenesis of the disease. Since BPECs infected with ASFV also undergo apoptosis, resistance of the natural host must involve complex pathological factors other than viral tropism.

In order to identify Trichinella at the species level, the commonly used test is a multiplex PCR, allowing the discrimination of nine out of the twelve taxa described so far. This test is based on five primer pairs amplifying fragments of the large subunit rDNA. Each taxon produces one or two bands of different sizes, resulting in a specific band pattern. By multiplex PCR, Trichinella murrelli shows two bands of 127 bp and 316 bp. However, a third band of 256 bp can occur. This band can lead to misidentification, since it is similar to the 253 bp band displayed by Trichinella britovi. BLAST analysis confirmed that the 256 bp band is from T. murrelli. The aim of this short note is to inform analysts that T. murrelli larvae may display either two-or three-band patterns., Afin d’identifier les Trichinella au niveau de l’espèce, le test couramment utilisé est une PCR multiplex, permettant la discrimination de neuf des douze taxons décrits jusqu’à présent. Ce test est basé sur cinq paires d’amorces amplifiant des fragments de la grande sous-unité l’ADN ribosomal. Chaque taxon produit une ou deux bandes de tailles différentes, résultant en un patron de bandes spécifique. Par PCR multiplex, Trichinella murrelli présente deux bandes de 127 pb et 316 pb. Cependant, une troisième bande de 256 pb peut s’observer. Cette bande peut être la cause d’une erreur d’identification, car elle est similaire à la bande de 253 pb affichée par Trichinella britovi. L’analyse BLAST a confirmé que la bande à 256 pb provient de T. murrelli. Le but de cette note est d’informer les analystes que les larves de T. murrelli peuvent présenter des patrons à deux ou trois bandes.

From 2006 to 2009 we screened 108 red foxes (Vulpes vulpes) and 894 wild boars (Sus scrofa) in Haut-Var, France for Trichinella britovi infection. Prevalences were 2.7 and 0% respectively. The fox may be considered a predictive sentinel for Trichinella in the Haut-Var ecosystem.

Trichosomoides nasalis (Trichinelloidea) is a parasite of Arvicanthis niloticus (Muridae) in Senegal. Female worms that harbour dwarf males in their uteri, occur in the epithelium of the nasal mucosa. Young laboratory-bred A. niloticus were either fed females containing larvated eggs or intraperitoneally injected with motile first-stage larvae recovered from female uteri. Both resulted in successful infection. Organs examined during rodent necropsy were blood and lymphatic circulatory systems (heart, large vessels, lymphnodes), lungs, liver, kidneys, thoracic and abdominal cavities, thoracic and abdominal muscular walls, diaphragm, tongue, and nasal mucosa. Development to adult nasal stages took three weeks. Recovery of newly hatched larvae from the peritoneal fluid at four-eight hours after oral infection suggests a direct passage from the stomach or intestinal wall to the musculature. However, dissemination through the blood, as observed with Trichinella spiralis, cannot be excluded even though newly hatched larvae of T nasalis are twice as thick (15 mu m). Developing larvae were found in histological sections of the striated muscle of the abdominal and thoracic walls, and larvae in fourth moult were dissected from these sites. Adult females were found in the deep nasal mucosa where mating occurred prior to worms settling in the nasal epithelium. The present study shows a remarkable similarity between T nasalis and Trichinella species regarding muscle tropism, but the development of T. nasalis is not arrested at the late first-larval stage and does not induce transformation of infected fibres into nurse cells. T. nasalis seems a potential model to study molecular relations between trichinelloid larvae and infected muscle fibres., Trichosomoides nasalis (Trichinelloidea) est parasite d’Arvicanthis niloticus (Muridae) au Sénégal. Les vers femelles, qui contiennent des mâles nains intra-utérins, résident dans l’épithélium de la muqueuse nasale. De jeunes A. niloticus d’élevage ont ingéré des femelles avec des oeufs larvés, ou ils ont été inoculés par voie intrapéritonéale avec des larves écloses de stade 1, mobiles, extraites des utérus des femelles. Les rongeurs se sont parasités avec les deux protocoles. Les organes examinés à l’autopsie des rongeurs ont été les systèmes sanguin et lymphatique (coeur, gros vaisseaux, ganglions lymphatiques), les poumons, le foie, les reins, les cavités thoracique et abdominale, les parois musculaires thoracique et abdominale, le diaphragme, la langue et la muqueuse nasale. Le développement jusqu’au stade adulte s’effectue en trois semaines. La récolte de larves nouveau-nées dans le liquide péritonéal quatre-huit heures après l’infection orale suggère un passage direct de la paroi digestive vers la musculature. Cependant, une dissémination par le sang, telle qu’elle est observée avec Trichinella spiralis, ne peut être exclue bien que les larves à l’éclosion de T. nasalis soient deux fois plus épaisses (15 μm). Les larves en développement ont été trouvées dans les parois musculaires abdominale et thoracique et elles ont été identifiées sur coupes histologiques dans les fibres musculaires. Les larves en mue 4 ont été récoltées dans l’abdomen et le thorax, ainsi que, pour les femelles seulement, dans la partie profonde de la muqueuse nasale où l’accouplement a lieu, avant l’installation des femelles dans l’épithélium nasal. Cette étude met en évidence une remarquable similitude entre T. nasalis et les espèces de Trichinella – le tropisme musculaire –, cependant, le développement de T. nasalis ne s’arrête pas au stade 1 âgé et il n’induit pas la transformation des fibres infectées en cellules-nourricières. T. nasalis pourrait être un modèle intéressant pour étudier les mécanismes moléculaires des relations entre les larves de trichinelloïdes et les fibres musculaires infectées.

Sir, Trichinellosis is one of the most widespread parasitic zoonoses in the world. Over the past decade, Eastern Europe has experienced a re-emergence of trichinellosis, connected with the social, political and economic changes of the 1990s. A high prevalence of Trichinella infection in domestic animals and in humans has been reported in Bulgaria, Serbia and Montenegro, Romania and Croatia (Cuperlovic et al., 2005; Blaga et al., 2007). Between 2008 and 2010, the presence of trichinellosis was reported on animals in Serbia and Montenegro, in Bulgaria. Human cases of trichinellosis were also recently reported in Serbia and Montenegro, Bulgaria, Albania (OIE, WAHID database). Update data concerning Trichinella infection in animals and humans in Kosovo are not available. Kosovo is located in the heart of South East Europe, in the middle of the Balkan Peninsula. Kosovo is situated between the 42° and 44° parallels of Northern hemisphere and between the 20° and 22° meridians. Kosovo has common borders with the aforementioned countries of Eastern Europe. To improve the knowledge of the epidemiology of trichinellosis in this area, the detection of Trichinella spp. was performed on muscles collected on carcasses of stray dogs from the outskirts of the city of Mitrovica, north Kosovo. In October 2008, muscle samples were collected from 25 stray dogs from the suburbs of Mitrovica, a city located in the Northern part of Kosovo (19° 36’ 49’’ E; 44° 58’ 56’’ N). They were shot in a landfill situated between the cities of Mitrovica and Zvecan (20° 50’ 23’’ E; 42° 54’ 24’’ N). The dogs included in the study were shot as part of a rabies prevention program, by hunters employed by the civilian authorities of the city. Muscles samples were taken from masseter muscles (about 20% of whole samples) dorsal muscles (about 40% of the samples) and diaphragm muscles and pillars (about 40% of the samples). Muscle samples were kept frozen at -20 °C until further processing in the French national reference laboratory for Food borne parasite, Maisons-Alfort, France. Trichinella larvae were identified using a multiplex PCR as described by Pozio and la Rosa (Pozio & la Rosa, 2003). All dogs included in the study were crossbreed (hound, sheepdog), medium – weighed and mature (Table I). Sex (male or female) was not recorded at the time of sampling. Table I. Results of the search and identification of Trichinella larvae in muscles samples collected on 25 dogs from Mitrovica, north Kosovo. Overall, 16% (4/25) of the dogs were infected by Trichinella. Trichinella britovi was identified in all the positive samples. This result proves the circulation of the parasite in this area. In Kosovo, stray dogs live in the outskirts of the cities and villages. Because of this periurban location, they can create a link between wild and domestic animals, as described by Pozio (Pozio, 2007). The area located on north Mitrovica is occupied by forests. Here, stray dogs come into contact with wildlife. Various species of wild animals are reservoir hosts for T. britovi, transmitted within a sylvatic cycle (Gottstein et al., 2009). Peroral infection of stray dogs may occur after ingestion of muscle tissue from infectious prey animals (rodents, birds...) or by consumption of infectious tissues from a carrion of homologous (cannibalism) or heterologous species (boars, foxes, wolves, birds, rodents...). One of the most important biological factors promoting transmission of the parasite is the physiological ability of muscle-stage larvae to survive in decaying carcasses/carrions. Encapsulated larvae of T. britovi in mouse carcasses packed in plastic vials have been found to be infective for laboratory animals up to 45 days at room temperature even if the muscle is completely liquefied (Pozio, unpublished data). Moreover, muscle larvae of Trichinella spp. can survive even at low freezing temperatures, maintaining infectivity for future hosts. T. britovi can survive in frozen carrion for up to one year (Dick & Pozio, 2001). The survival of muscle larvae after freezing occurs mainly when these larvae parasitize striated muscles of carnivores (bears, foxes, dogs...) (Gottstein et al., 2009). These well-established environmental adaptations go towards increasing the survival of the parasite in nature and perpetuating the transmission of the parasite to wildlife and stray dogs. Stray dogs also go near and even enter in the city of Mitrovica to find out food, especially during cold season (from October to March-April). They eat mainly garbage: rubbish cans from Mitrovica and from the villages around Mitrovica; landfill located near Mitrovica, where all the garbage from the outskirts of Mitrovica are deposited. Garbage are not buried and are stored open-cast in the landfill. They are therefore easily accessible to feral animals. Because Mitrovica is located near a Serb area, where swine herds are bred and pork meats are consumed, garbage may contain pork scraps and even pork carcasses. In several countries of Central Eastern Europe, the transient breakdown of governmental veterinary services and state farms, accompanied by economic problems and war, have resulted in sharp increase in the incidence of Trichinella infection among domestic pig herds, with prevalence rates reaching 50% in some villages in 1990s (Murrell & Pozio, 2000). In the neighbouring Serbia, the consequences have been a 300% increase in pig infection and a concomitant large increase of human cases of trichinellosis (Murrell & Pozio, 2000). Human trichinellosis may be acquired through the consumption of raw or undercooked meat (e.g., pork, horse and game meat) containing infective larvae of Trichinella spp. (Gottstein et al., 2009). Immediate cause of recent human outbreaks in Eastern Europe was the consumption of traditionally prepared sausages (Dupouy-Camet, 2006). Pig contamination mainly occurs by eating scraps of infectious meats mixed with food, by eating rodents present in the pig farm or by caudophagy. Horse can also play a part in infection of stray dogs in Kosovo. Kosovo is mostly rural and horses are used as draught animals. Stray dogs can become infected by consumption of carcasses of infected horses. In the past 30 years, horse meat has been identified as the main source of human trichinellosis in Europe, with more than 3,350 cases reported in 14 outbreaks (Pozio et al., 2005; Boireau et al., 2000). Horses fattening with pork scraps before slaughtering or consumption of rodents crushed and mixed with the fodder by accident are sources of contamination of horses (Murrell et al., 2004). Considering the high parasitic loads revealed on a number of dogs included in the study, stray dogs from the outskirts of Mitrovica probably become infected from several sources of contamination. As they are carnivorous, they concentrate the parasite. The cycle of transmission of Trichinella spp. in animals in Kosovo is not precisely known as knowledge is only partial. Further studies would be necessary, including sampling on wild and out-of-town animals (rodents, stray cats...). In this study, local stray dogs represent bio-indicators for a biological risk. In areas where epidemiological data are unknown or incomplete, local animals can be useful sentinels for humans, for sanitary risks of environmental origin. Because there are a lot of potential reservoir hosts of the parasite, especially among wildlife, Trichinella infection cannot be eradicated. Nevertheless the disease can be efficiently prevented in humans by means of individual and public health measures. Improvement of meat control and education of the consumers (to cook potentially infected meat thoroughly) are key preventive measures. Travellers should also be informed of the risks of illegal importations of traditional meat products from Central and Eastern European countries where trichinellosis is endemic.

International audience; Three cases of human trichinellosis due to Trichinella britovi were reported in 2015 in the Southeast of France resulting from consumption of raw pork sausages (figatelli) prepared in Corsica. Fourteen other people ate figatelli from the same batch but were not infected due to the figatelli being well cooked. This is the first reported human trichinellosis outbreak due to consumption of Corsican sausages prepared from uncontrolled pork. Consumption of raw figatelli is a common tradition in Corsica. As a result, the health recommendation to cook the product well is not always applied. In the present case, the figatelli product label was not sufficiently visible to advise consumers of the risks associated with uncooked pork.

International audience; Routine diagnosis of animal trichinellosis for food safety and trade relies on a method of artificial digestion to free Trichinella muscle larvae from meat for subsequent identification by microscopy. As part of a quality control system, the French National Reference Laboratory (NRL) initiated ring trials to determine the sensitivity of the test performed in the 72 routine diagnostic laboratories in France. A method was devised to obtain calibrated meat samples containing known numbers of capsules with Trichinella spiralis muscle larvae. This method was based on an incomplete artificial digestion of Trichinella-infected mice carcasses to allow the collection of intact Trichinella capsules. Capsules were placed into a meatball of 100 +/- 2 g of pork and horsemeat to produce proficiency samples. Three categories of samples were prepared: small (3 to 5 capsules), medium (7 to 10), and large (12 to 15). The sensitivity was expressed as the percentage of muscle larvae recovered from each proficiency sample. Reproducibility was tested with ring trials organized between two NRLs (France and Canada), and a reference sensitivity of 84.9% was established. National ring trials were then organized in France, with the 72 routine diagnostic laboratories each receiving four proficiency samples per session. After five sessions, an improvement in the digest test sensitivity was observed. Results at the fifth session indicated sensitivities of 78.60% +/- 23.70%, 81.19% +/- 19.59%, and 80.52% +/- 14.71% muscle larvae for small, medium, and large samples, respectively. This study supports the use of proficiency samples to accurately evaluate the performance of routine diagnostic laboratories that conduct digestion tests for animal trichinellosis diagnosis.

Since the beginning of 2006, the French Trichinella National Reference Centre has been notified of three incidents of trichinellosis due to the consumption of wild boar meat