Molecular epidemiology of <Emphasis Type="Italic">Trichinella</Emphasis> spp. in three Baltic countries: Lithuania,Latvia, and Estonia

Meat of domestic pigs and wild boars has been the significant source of emerged human trichinellosis in Lithuania, Latvia, and Estonia over the past two decades. However, there is very little known on the occurrence of Trichinella spp. in main wildlife reservoirs and its transmission in domestic and sylvatic cycles in these countries. The present study demonstrated considerably higher endemicity of Trichinella spp. in main sylvatic reservoirs (28.9–42% in foxes (Vulpes vulpes) and raccoon dogs (Nyctereutes procyonoides) in all three countries than previously reported. Molecular identification of Trichinella larvae from more than 500 sylvatic and domestic animals revealed four Trichinella species (Trichinella spiralis, Trichinella britovi, Trichinella nativa, and Trichinella pseudospiralis) sympatric in a relatively small area and several as the first records for the respective countries. The nonencapsulating T. pseudospiralis is found for the first time in the Eastern Europe. Sylvatic T. britovi was found in domestic pigs in Lithuania and Latvia (16 and 57.1%, respectively) and only in these countries, domestic T. spiralis was detected in sylvatic animals in areas where domestic trichinellosis was registered. The study suggests that transmission of Trichinella between domestic and sylvatic cycles in Lithuania and Latvia is favored by improper human behavior, e.g., pig and slaughter waste management.     

Trichinella infection in wild animals from endemic regions of Argentina

Natural infection with Trichinella has been described in more than 150 mammalian species. However, few reports of Trichinella infection in wild animals have come from Argentina. In this study, muscle tissue was obtained from wild animals in Argentina with the aim of evaluating the presence of Trichinella. A total of 169 muscle samples were collected to determine the presence of Trichinella larvae by artificial digestion. The 169 muscle samples originated from 12 species including 36 opossums (Didelphis albiventris), 19 armadillos (Chaetophractus villosus), 9 capybaras (Hydrocaeris hydrocaeris), 1 puma (Puma concolor), 3 grey fox (Lycalopex gymnocercus), 6 coypus (Myocastor coypus), 6 skunks (Conepatus chinga), 2 ferrets (Galictis cuja), 66 rats (Rattus norvegicus), 6 mice (Mus musculus), 12 wild boars (Sus scrofa), and 3 wild cats (Felis geoffroyi). Trichinella infection was detected in 1 puma [2 larvae per gram (LPG)], 3 wild boars (8–420 LPG), 3 armadillos (0.04–0.08 LPG), and 9 rats (0.1 to 150 LPG). Only 3 Trichinella isolates, of 1 rat and 2 wild boars from Neuquén, were identified as Trichinella spiralis by nested PCR. The presence of Trichinella infection among wild animal populations suggests a sylvatic cycle of transmission in Argentina, which can serve as a reservoir for humans and domestic animals. Further, evidence of high prevalence in rats emphasizes the need to improve pig management, mainly in small individual farms without adequate technology, to enhance the quality of feeds, and to improve veterinary services to avoid exposure of pigs to Trichinella.     

A preliminary investigation on the infectivity of<Emphasis Type="Italic"> Trichinella</Emphasis> larvae in traditional preparations of walrus meat

This study evaluated the infectivity of Trichinella nativa in freshly frozen walrus meat and traditionally aged walrus meat (igunaq) associated with two human outbreaks of trichinellosis in the Canadian Arctic. Trichinella larvae recovered from walrus meat stored at –20°C for up to 20 months remained infective for guinea pigs inoculated with 135 or 716 larval doses. However, none of the 4–5 and 10-month-old igunaq preparations contained infective T. nativa larvae as measured by bioassays using mice and guinea pigs at inoculation doses ranging from 6 to 500 larvae. This indicates that the degradation process that occurred in the field can be sufficient to either kill Trichinella larvae or render them non-infective for mice and guinea pigs. Further research is needed to evaluate the food safety risk of traditional walrus igunaq aged under different field conditions and storage times.     

First report of <Emphasis Type="Italic">Trichinella britovi</Emphasis> in Serbia

In Europe, Serbia ranks among countries with a high prevalence of Trichinella infection in pigs, which continues to be a serious human health problem. While in some Balkan countries, more than one Trichinella species/genotype has been described in both the sylvatic and domestic cycles, these data are lacking for Serbia. To date, only a few Serbian isolates of Trichinella have been genetically specified, and all were classified as T. spiralis. Although transmission of Trichinella from domestic pigs to wildlife could be assumed, neither the infection status nor the species of Trichinella circulating among wildlife in Serbia has been investigated. This study shows the presence of two Trichinella species, T. spiralis and T. britovi, in wild animals originating from five districts in Serbia, where Trichinella infections in domestic pigs and humans have been recorded. Trichinella spiralis was detected in jackals (n = 3), red foxes (n = 2) and a wild cat (n = 1). We also established that wolves (n = 4) and red foxes (n = 2) serve as sylvatic reservoirs for T. britovi. This is the first report on the presence of T. britovi in Serbia.     

Development and Evaluation of a Western Blot Kit for Diagnosis of Human Trichinellosis

We evaluated industrially prepared Western blot strips designed to avoid the cross-reactions observed with indirect immunofluorescence and enzyme-linked immunosorbent assays used for the serodiagnosis of trichinellosis. The antigen preparations were crude extracts of Trichinella spiralis. The Western blot profile characteristic of trichinellosis was characterized by comparing 60 sera from patients infected by Trichinella to 11 sera from healthy subjects, 51 sera from patients with other proven parasitic diseases (cysticercosis, schistosomiasis, strongyloidosis, fascioliasis, toxocariasis, liver amebiasis, anisakiasis, filariasis, toxoplasmosis, hydatidosis, or malaria), and 23 sera from patients with autoantibodies. Specific 43- to 44-kDa and 64-kDa bands were obtained with all of the sera from 51 patients with acute trichinellosis, in 4 out of 9 patients at the early stages of the disease, and in only 1 control patient, who had suspected anisakiasis and in whom trichinellosis could not be ruled out by muscle biopsy.     

In vitro effects of resveratrol on Trichinella spiralis

Resveratrol, a natural phytoalexin found mainly in grapes, possesses antibacterial, antifungal, and antiviral properties. However, there is no information about its effects on helminths such as Trichinella sp. In the present study, we investigated the effects of resveratrol on the viability of Trichinella spiralis life stages in vitro. Adult forms, newborn larvae (NBL), and muscle larvae (ML) were incubated with resveratrol at concentrations varying from 12.5 to 200 μg/ml. Resveratrol showed significant anthelmintic activity against NBL and adult forms of Trichinella, but not against ML. Our results suggest that resveratrol may be useful as a therapeutic agent to treat trichinellosis in early stages and warrant its further assessment in animal models of disease.     

A novel detection method for Alaria alata mesocercariae in meat

Distomum musculorum suis (DMS), the mesocercarial stage of the trematode Alaria alata, can cause severe damages within their hosts, and since several reports about cases of human larval alariosis have been published, it became apparent that infected game animals and in particular wild boars are a potential source of infection for both humans and animals. A final statement concerning the health risks for consumers could not be given due to the lack of information about both the prevalence of DMS and the suitability of Trichinella inspection methods to detect this parasite in wild boar meat. Our studies concentrate on (1) the verification of suitability of the official digestion methods for Trichinella spp. for DMS detection in wild boars, (2) development, optimization, and validation of methods, and (3) the distribution of the parasites within their paratenic hosts. A total of 868 individual samples/digests from 48 wild boars were analyzed by the reference method for Trichinella detection in meat samples according to regulation (EC) No. 2075/2005. In addition to the official protocol, a method modification with Pankreatin© and bile acid was applied for analysis of adipose tissue samples (n?=?89). On the basis of our results, a new detection method based on a larvae migration technique was developed and used for detection of DMS in 574 single samples. Furthermore, the distribution patterns of DMS in wild boars in a total of 1377 single sample migrations/digestions from 35 positive animals were analyzed by application of all three methods. The official digestion method for Trichinella spp. in wild boars meat is inapplicable for the detection of A. alata mesocercariae as it shows shortcomings in both digestion and sampling. A direct comparison between the newly developed A. alata mesocercariae migration technique and the official digestion method for Trichinella spp. based on 574 single samples from 18 animals clearly shows that the sensitivity to detect A. alata developmental stages in tissues of wild boars of the new method is nearly 60% higher compared with the magnetic stirrer method for pooled sample digestion as laid down in regulation (EC) No. 2075/2005. Among other advantages, this method offers a simple, highly applicable, fast, and cost effective way to detect DMS in wild boars which is already applicable in routine veterinary inspection.     

Specific IgG4 response directed against the 45-kDa glycoprotein in trichinellosis: a re-evaluation of patients 15 years after infection

The aim of this study was to evaluate the humoral immune response in late human trichinellosis with particular attention to the presence of IgG4 antibodies directed against the Trichinella-45-kDa glycoprotein (gp). This study re-evaluates subjects 15 years after they were involved in a trichinellosis outbreak that occurred in Central Italy following the consumption of raw boar meat infected with Trichinella britovi. The results show that ELISA tests using the E/S antigen identified five IgM- and eight IgG-positive patients and no IgA-positive patients. Tests using immunoblot (IB) with E/S antigens identified three IgM-, five IgA-, seven- IgG1- and three IgG4-positive sera. When the purified 45-kDa gp was used as an antigen, the IB revealed that six of the ten sera tested were positive for IgG4. Sera were also evaluated with a commercial kit, revealing that 11 of 12 patients had a highly sensitive reactivity against Trichinella proteins (64 and 44–43 kDa). In conclusion, humoral immune response against Trichinella is still present in these patients 15 years after the initial infection, including an IgG4 response directed to the 45-kDa gp.     

First report of Trichinella pseudospiralis in Poland, in red foxes (Vulpes vulpes)

Nematode worms of the genus Trichinella are one of the most widespread zoonotic pathogens. Natural transmission between hosts can only occur through the ingestion of infected meat. To date, two Trichinella species are known to be etiological agents of disease among domestic animals and wildlife in Poland: T. spiralis and T. britovi. In the last decades, since the administration of an oral vaccination against rabies, the red fox population in Poland has increased exponentially. The study area covers the Nowy Targ region: a mountainous area (585–1138 m above the sea) in southern Poland. Of 24 red foxes examined in the study, four were infected with Trichinella isolates: three were identified as T. britovi and one as T. pseudospiralis. The muscle of red foxes infected with T. britovi harboured 2.75, 3.11, 4.4 LPG and with T. pseudospiralis 0.36 LPG. Trichinella larvae were identified at species level by genomic and mitochondrial multiplex PCR, the products of which were sequenced for comparison with other sequences available in GenBank. The sequences obtained from the Polish T. pseudospiralis isolate, deposited in GenBank under the accession numbers JQ809660.1 and JQ809661.1, matched sequences already published in GenBank. Sequence comparison showed a 100% match with the large subunit ribosomal RNA gene of T. pseudospiralis isolate ISS 013, and a 96–95% match with those of T. pseudospiralis isolates ISS 141 and ISS 470. This is the first report of the identification of T. pseudospiralis larvae from red fox in Poland.     

Epidemiology of trichinosis in Italy: correlation between the wild cycle and man

In Italy trichinellosis is a zoonosis caused by Trichinella nelsoni, with an exclusively sylvatic cycle involving wild and synanthropic animals (fox, wolf, stray dog, wild boar, foraging pig). The recent increase and diffusion of wild boar population in all Italy, have caused two human epidemic foci of trichinellosis involving about 100 persons in the years 1985-1986. The custom of eating fox meat have caused other human infections. The results of serological examinations of wild boar population showed the role played by this animal in the epidemiology of the disease. The examination of rodent population has pointed out the infection in these mammals (rat, porcupine). The epidemiological implications of wild cycle on man are reported.     

Clinical and laboratory profile of trichinellosis from a non-endemic country

Trichinellosis is a zoonotic parasitic infection caused by nematodes of the genus Trichinella. Humans mostly get infected by eating raw/under-cooked pork. In India, it has been reported mostly as sporadic cases especially from North Eastern and Eastern part of country.In this study, we retrospectively analyzed the socio-demographic pattern, clinical presentation, laboratory profile and treatment response in Trichinella-infected patients visiting our tertiary care center which mainly caters to patients from North India. For this retrospective laboratory-based analysis, patients diagnosed on the basis of positive anti-Trichinella IgG antibodies between 1^st June 2008, and 31^st May 2019 were included.A total of 11 positive trichinellosis cases were detected, of whom majority were children who presented with history of fever, gastrointestinal symptoms, myalgia, eosinophilia along with hepatomegaly and pulmonary manifestations. No CNS involvement was seen in any of the patients although it is commonly associated with Trichinella infection. All patients recovered uneventfully after antihelminthic treatment.     

Trichinella sp. in red foxes (Vulpes vulpes) from Catalonia, NE Spain

European legislation allows the official recognition of Trichinella-free pig holdings, provided Trichinella sp. infection is absent from humans and prevalence of Trichinella sp. infection in red foxes (Vulpes vulpes) is below 0.1% in the area, region or country. Tibialis anterior muscle samples from 1,319 red foxes captured in Catalonia (NE Spain) between 1998 and 2007 were analyzed for Trichinella sp. using the digestion method. Four foxes resulted positive (one in 1999, one in 2002 and two in 2006), accounting for a low prevalence (0.3%). However, this prevalence was concentrated in mountain or rural areas with a low sample size, reaching high local prevalences. The two positive samples in 2006 were characterized as Trichinella britovi, and a sylvatic cycle of trichinellosis seems to occur, at least in the rural insufficiently sampled regions of Catalonia. Overall, the results obtained do not currently allow the establishment of Trichinella-free pig holdings in the study area, but further research is needed to better know the prevalence and cycle of Trichinella sp. in Catalonia.     

Imported Human Trichinellosis: Sequential IgG4 Antibody Response to<Emphasis Type="Italic"> Trichinella spiralis</Emphasis>

Two out of five members of one household presented with clinical signs of trichinellosis after their return to the Netherlands. The family had consumed Trichinella-infected pork in Montenegro, formerly Yugoslavia. Serological tests were performed at 1, 2, 6, and 18 months after ingestion of the infected meat. Trichinella-specific IgM, IgG, and IgA antibodies measured in sera from symptomatic and asymptomatic patients remained positive up to 18 months after ingestion. The measured IgG4 antibody response directed to a 45 kDa Trichinella spiralis antigen also persisted 18 months after ingestion for three of the family members.     

Validation of a Western Blot for the detection of anti-<Emphasis Type="Italic">Trichinella</Emphasis> spp. antibodies in domestic pigs

Trichinellosis is a zoonotic disease in humans caused by Trichinella spp. According to international regulations and guidelines, serological surveillance can be used to demonstrate the absence of Trichinella spp. in a defined domestic pig population. Most enzyme-linked immunosorbent assay (ELISA) tests presently available do not yield 100% specificity, and therefore, a complementary test is needed to confirm the diagnosis of any initial ELISA seropositivity. The goal of the present study was to evaluate the sensitivity and specificity of a Western Blot assay based on somatic Trichinella spiralis muscle stage (L1) antigen using Bayesian modeling techniques. A total of 295 meat juice and serum samples from pigs negative for Trichinella larvae by artificial digestion, including 74 potentially cross-reactive sera of pigs with other nematode infections, and 93 meat juice samples from pigs infected with Trichinella larvae were included in the study. The diagnostic sensitivity and specificity of the Western Blot were ranged from 95.8% to 96.0% and from 99.5% to 99.6%, respectively. A sensitivity analysis showed that the model outcomes were hardly influenced by changes in the prior distributions, providing a high confidence in the outcomes of the models. This validation study demonstrated that the Western Blot is a suitable method to confirm samples that reacted positively in an initial ELISA. Frey and Schuppers contributed equally to this work. This work was funded by the Swiss Federal Veterinary Office, grant number 1.06.03.     

An outbreak of trichinellosis with detection of Trichinella larvae in leftover wild boar meat

The clinical diagnosis of trichinellosis can be difficult due to lack of pathognomonic signs or symptoms. In Korea, since the first report of human infection by Trichinella spiralis in 1997 following the consumption of raw badger meat, there have been occasional trichinellosis outbreaks. We describe an outbreak of 12 cases of trichinellosis in Korea and implicate raw wild boar meat as the culprit. A total of 27 larvae of Trichinella (0.54 larvae per gram of meat) were recovered from the leftover raw wild boar meat.     

An outbreak of trichinellosis in farmed wild boar in Finland

Nine farmed wild boar out of 25 slaughtered from a single farm were condemned at meat inspection because of trichinellosis. With RAPD-PCR, Trichinella spiralis was identified in all positive wild boar. Out of the available serum samples (n=7), all wild boar which had failed the meat inspection showed seroconversion in ELISA and Western blotting, as did one additional animal which had passed the inspection. The animals became infected during an invasion of rats from an improperly closed dump near the farm. Unfortunately, by the time trichinellosis was discovered in the wild boar, the invasion had already been brought under control; thus, no samples from rats were available. However, having lived through the rat invasion was shown to be a risk factor for trichinellosis in wild boar (relative risk, RR=6.3). In wildlife samples from surrounding areas, sylvatic trichinellosis was found to be very common (74%; n= 19 red foxes). Intriguingly, the prevalent species in trichinella-positive foxes differed from that in wild boar, Trichinella nativa and T. spiralis being found in 12 foxes and in one fox, respectively.     

Ocular system involvement in the course of human trichinellosis. Pathological and diagnostic aspects

Pathomechanism of lesions in the course of trichinellosis was discussed, representing a sum of immunopathologicaI, pathomorphological and biochemical phenomena. Particular attention was devoted to clinical pathology of the visual organ, which prevails at the acute stage of trichinellosis. In evaluation of clinical signs/symptoms manifested in the visual organ traits of its anatomic, morphological structure, function of the eyeball muscles and eyeball vascular system were taken into account. Ocular lesions in the course of trichinellosis reflect in principle angiomyositis due to immunopathology resulting from migration of Trichinella larvae to many organs and to structures of visual system.     

Biology of Alaria spp. and human exposition risk to Alaria mesocercariae—a review

Recent incidental background findings of Alaria alata mesocercariae [“Distomum muscularis suis,” Duncker, 1896] in meat of wild boars during official Trichinella inspection initiated a re-assessment of the potential human health risk as posed by this parasite. The present review of the literature on Alaria biology shows that the human exposition risk should no longer be accepted to be negligible, as it demonstrates a general lack of knowledge in relevant areas of Alaria biology confounding any risk analysis. Sound risk assessment needs future studies which should concentrate on the most pressing questions of (1) the optimization and/or development of methods for reliable Alaria mesocercariae detection, (2) the distribution of the mesocercariae within their paratenic hosts, i.e., identification of potential predilection sites, particularly in wild boars, and (3) their prevalence in sylvatic populations of animals with respect to their introduction into the human food chain. Further, the degree and possibly also the species specificity of Alaria mesocercariae tenacity within the paratenic hosts and respective meat as pertaining to food technological treatments need to be elucidated. While these questions remain unanswered, it is an incontrovertible fact that Alaria mesocercariae have a potentially high human pathogenicity by both occupational and alimentary exposition.     

Molecular identification of Trichinella britovi in martens (Martes martes) and badgers (Meles meles); new host records in Poland

Trichinella larvae were detected in a marten (Martes martes) and a badger (Meles meles) in Poland. The animals were found dead following car accidents. All examined animals derived from the Mazurian Lake district, north-east Poland, near the village Kosewo Górne where Trichinella infection were earlier confirmed in wildlife; red foxes and wild boars. The muscle samples were examined by artificial pepsin-HCl digestion method. The parasites were identified as Trichinella britovi by multiplex polymerase chain reaction method. Larvae were found in two out of three martens and one out of seven examined badgers. This is the first report of the identification of Trichinella britovi larvae from martens and badgers in Poland.