Sylvatic and domestic Trichinella spp. in wild boars; infectivity, muscle larvae distribution, and antibody response

Thirty-six wild boars were inoculated with Trichinella spiralis, Trichinella nativa, Trichinella britovi, Trichinella pseudospiralis (USSR), T. pseudospiralis (USA), T. pseudospiralis (AUST), Trichinella murrelli, Trichinella T6, and Trichinella nelsoni. The wild boars were killed at 5 and 10 wk postinoculation (PI), and the number of muscle larvae per g (lpg) of tissue was determined for 18 muscles or muscle groups. Five weeks PI, all Trichinella genotypes had established as muscle larvae, but their infectivity varied widely: T. spiralis established in high numbers (mean = 296 lpg), T. britovi, T. nelsoni, and 1 of the T. pseudospiralis genotypes (AUST) in moderate numbers (mean = 53-74 lpg), whereas the remaining genotypes were poorly infective (mean 2-16 lpg). Because of considerable weight gain of the wild boars, an estimated total larval burden (live weight x lpg) was calculated for each animal. The total larval burden did not change significantly over time for T. spiralis, T. murrelli, T. britovi, T. nelsoni, and T. pseudospiralis (USA and USSR), whereas a significant reduction could be demonstrated for T. nativa, Trichinella T6, and T. pseudospiralis (AUST). Diaphragm and tongue were predilection sites in wild boars, independent of Trichinella genotype and infection level. At low infection levels, a greater percentage of larvae were found in diaphragm and tongue at 10 wk than 5 wk PI. Antibody responses increased rapidly between weeks 3 and 5 PI. For T. spiralis and T. nelsoni, the high antibody level persisted throughout the experimental period, but for T. nativa, T. britovi, T. murrelli, or Trichinella T6, the levels declined. For T. pseudospiralis, the antibody response increased more gradually between weeks 3 to 10 PI. Infection with all genotypes of Trichinella were detected using any of 7 excretory-secretory antigens, which points to the potential use of 1 common antigen for epidemiological studies on Trichinella in wild boars. In conclusion, T. spiralis is highly infective to wild boars, T. britovi, T. nelsoni, T. pseudospiralis (USA), and T. pseudospiralis (USSR) are moderately infective, and T. nativa, T. murrelli, T. pseudospiralis (AUST), and Trichinella T6 are poorly adapted to this host species.     

Helminth infections of wild boars (Sus scrofa) in the Bursa province of Turkey

The present study aimed to investigate the status of helminth infections in wild boars in the Bursa province of Turkey. For this purpose, during 2007-2008, 27 wild boars were necropsied and examined for helminths. Individual samples of tongue and diaphragm from 27 necropsied wild boars and an additional 22 tongue and diaphragm samples provided by hunters were examined by trichinoscopy and artificial digestion for Trichinella spp. larvae. Twenty animals (74%) were identified as being infected with at least one helminth species. Twelve species of helminths were detected, with the following prevalence rates: Metastrongylus apri (59%), Metastrongylus salmi (52%), Metastrongylus pudendotectus (52%), Dicrocoelium dendriticum (33%), Globocephalus urosubulatus (22%), Macracanthorhynchus hirudinaceus (19%), Gongylonema pulchrum (11%), Physocephalus sexalatus (7%), Trichuris suis (7%), Ascarops strongylina (4%), Hyostrongylus rubidus (4%) and Taenia hydatigena larvae (4%). Generally, lungworms were the predominant helminths. The highest mean abundance was observed for M. pudendotectus, and the lowest was determined for T. hydatigena larvae. Significant differences in the prevalence and intensity were found for D. dendriticum with respect to host age and sex, respectively. The mean intensity of M. pudendotectus was significantly influenced by the sex and age of the wild boars. This study is the first report describing the presence of M. salmi, M. pudendotectus, D. dendriticum, G. urosubulatus, M. hirudinaceus, P. sexalatus, A. strongylina and H. rubidus in wild boars in Turkey. All analysed muscle samples were negative for Trichinella spp. larvae.     

Trichinella nativa in sylvatic wild boars.

Of 17 Trichinella isolates from domestic pigs and wild boars (Sus scrofa) in regions where Trichinella nativa is widespread among sylvatic animals, two wild boars from Estonia were found to be naturally infected with this Trichinella species. The other 15 animals were infected with Trichinella spiralis. Trichinella nativa is tolerant to freezing when in the muscles of carnivores. The biological characteristics and temperature tolerance of this species in swine need to be further investigated if pork is certified for consumption following freezing.     

Predilection Muscles and Physical Condition of Raccoon Dogs (Nyctereutes procyonoides) Experimentally Infected with Trichinella spiralis and Trichinella nativa

The predilection muscles of Trichinella spiralis and T. nativa were studied in 2 experimental groups of 6 raccoon dogs (Nyctereutes procyonoides), the third group serving as a control for clinical signs. The infection dose for both parasites was 1 larva/g body weight. After 12 weeks, the animals were euthanized and 13 sampling sites were analysed by the digestion method. Larvae were found in all sampled skeleton muscles of the infected animals, but not in the specimens from the heart or intestinal musculature. Both parasite species reproduced equally well in the raccoon dog. The median density of infection in positive tissues was 353 larvae per gram (lpg) with T. spiralis and 343 lpg with T. nativa. All the infected animals had the highest larvae numbers in the carpal flexors (M. flexor carpi ulnaris). Also tongue and eye muscles had high infection levels. There were no significant differences in the predilection sites between these 2 parasite species. Trichinellosis increased the relative amount of fat, but not the body weight in the captive raccoon dogs. Thus, Trichinella as a muscle parasite might have catabolic effect on these animals.     

A Study on the Predilection Sites of Trichinella spiralis Muscle Larvae in Experimentally Infected Foxes (Alopex lagopus,Vulpes vulpes)

Studies were carried out on the predilection sites of Trichinella spiralis muscle larvae in experimentally infected arctic foxes (Alopex lagopus) and silver foxes (Vulpes vulpes) reared in cages. The highest number of larvae per gramme tissue was found in the muscles of the legs, eyes, diaphragm, and tongue. The 2 fox species showed no significant differences with regard to predilection sites.     

Sylvatic trichinellosis in southwestern Spain

The epidemiology of Trichinella spp. in their main sylvatic hosts, wild boar (Sus scrofa ferus and red fox (Vulpes vulpes), in Extremadura (southwestern Spain) was studied. We examined 88 Trichinella spp.-positive wild boar muscle-tissue samples from a total of 29,333 killed animals, referred to the Veterinary Parasitology Department (University of Extremadura, Spain) by the Extremadura Veterinary Service. Additionally, 227 red foxes killed during the hunting season and thus not subject to veterinary controls were examined for trichinellosis. Trichinella spp. larvae were found in six (3%) of the red foxes. All samples were examined using direct diagnostic techniques, including trichinoscopy and artificial digestion. The mean intensity of infection was 74.8 larvae/g (LPG) of muscle tissue in wild boars, compared to 30.6 LPG in foxes. Trichinella spiralis (sensu stricto) predominated over T. britovi in wild boars. Random amplified polymorphic DNA (RAPD) and alloenzyme typing showed that 74% of infected wild boars had only T. spiralis, 21% had only T. britovi, and 5% showed mixed infections. In contrast, 33% of infected foxes were infected only with T. spiralis, while 67% had T. britovi, suggesting a clear predominance of the latter in foxes. We suspect the existence of a paranthropic or sylvatic cycle in large areas of this region; given the ease of transfer between sylvatic and domestic or semi-domestic animals, this implies a high epidemiological risk.     

Experimental Trichinella infection in seals

The susceptibility of seals to infection with Trichinella nativa and the cold tolerant characteristics of muscle larvae in seal meat were evaluated. Two grey seals, Halichoerus grypus, were inoculated with 5000 (100 larvae/kg) T. nativa larvae and two grey seals with 50000 (1000 larvae/kg). One seal from each dose group and two control seals were killed at 5 and 10 weeks post-inoculation (p.i.). At 5 weeks p.i., infection was established in both low and high dose seals with mean larval densities of 68 and 472 larvae per gram (lpg), respectively, using eight different muscles for analyses. At 10 weeks p.i., mean larval densities were 531 and 2649 lpg, respectively, suggesting an extended persistence of intestinal worms. In seals with high larval density infections, the distribution of larvae in various muscles was uniform, but in one seal with a low larval density infection, predilection sites of larvae included muscle groups with a relative high blood flow, i.e. diaphragm, intercostal and rear flipper muscles. Trichinella-specific antibody levels, as measured by ELISA, increased during the 10 week experimental period. Infected seal muscle was stored at 5, -5 and -18 degrees C for 1, 4 and 8 weeks. Muscle larvae released from stored seal muscle by artificial digestion were inoculated into mice to assess viability and infectivity. Larvae from seal muscle 10 weeks p.i. tolerated -18 degrees C for 8 weeks but larvae from seal muscle 5 weeks p.i. tolerated only 1 week at -18 degrees C, supporting the hypothesis that freeze tolerance increases with the age of the host-parasite tissue complex. The expressed susceptibility to infection, extended production of larvae, antibody response and freeze tolerance of T. nativa in seals are new findings from the first experimental Trichinella infection in any marine mammal and suggest that pinnipeds (phocids, otariiids or walrus) may acquire Trichinella infection by scavenging even small amounts of infected tissue left by hunters or predators.     

Trichinella papuae n.sp. (Nematoda), a new non-encapsulated species from domestic and sylvatic swine of Papua New Guinea

Encapsulated and non-encapsulated species of the genus Trichinella are widespread in sylvatic animals in almost all zoogeographical regions. In sylvatic animals from Tasmania (Australian region), only the non-encapsulated species Trichinella pseudospiralis has been reported. Between 1988 and 1998, non-encapsulated larvae of Trichinella were detected in five domestic pigs and six wild boars from a remote area of Papua New Guinea. Morphological, biological, and molecular studies carried out on one strain isolated from a wild boar in 1997 suggest that these parasites belong to a new species, which has been named Trichinella papuae n.sp. This species can be identified by the morphology of muscle larvae, which lack a nurse cell in host muscles, and whose total length is one-third greater than that of the other non-encapsulated species, T. pseudospiralis. Adults of T. papuae do not cross with adults of the other species and genotypes. Muscle larvae of T. papuae are unable to infect birds, whereas those of T. pseudospiralis do. The expansion segment V of the large subunit of the ribosomal DNA differs from that of the other species and genotypes. All of these features allow for the easy identification of T. papuae, even in poorly equipped laboratories. The discovery and identification of a second non-encapsulated species in the Australian region strongly supports the existence of two evolutionary lines in the genus Trichinella, which differ in terms of the capacity of larvae to induce a modification of the muscle cell into a nurse cell.     

Predilection sites of Trichinella spiralis larvae in naturally infected horses.

A total of 120 muscle tissues from three horses naturally infected with Trichinella spiralis were examined. The head was the most infected site. In particular, the muscles harbouring the highest number of larvae were: musculus buccinator (12, 411 and 1183 larvae g-1), the tongue (11, 615 and 1749 larvae g-1), m. levator labii maxillaris (17,582 and 1676 larvae g-1), and the masseter (4.9, 289 and 821 larvae g-1). Compared with the diaphragm, the number of larvae per gram was from 3.5 to 6.8 times higher in the tongue, from 3.5 to 6.5 higher in m. levator labii maxillaris, and from 2.5 to 4.6 higher in m. buccinator. Of the examined muscles, the diaphragm had from the 6th to the 15th highest level of infection (3.1, 166 and 256 larvae g-1). Published data from experimentally infected horses confirm these results, suggesting that efforts to detect predilection sites should focus on the head muscles.     

Trichinella britovi etiological agent of sylvatic trichinellosis in the Republic of Guinea (West Africa) and a re-evaluation of geographical distribution for encapsulated species in Africa

In West Africa, Trichinella infection was documented in humans and animals from Senegal in the 1960s, and the biological characters of one isolate showed a lower infectivity to domestic pigs and rodents when compared with that of a Trichinella spiralis pig isolate from Europe. To identify the Trichinella species present in West Africa, a survey was conducted in a total of 160 wild animals in the Republic of Guinea. Three Viverridae, one true civet (Viverra civetta) and two African palm civets (Nandinia binotata) from the Fouta Djallon Massif, Pilimini Subprefecture, were found positive by artificial digestion of muscle samples. Trichinella larvae from these three viverrids were identified as Trichinella britovi and no difference was detected in three examined sequences from these African isolates and the reference strain of T. britovi from Europe, indicating common ancestry, an historically continuous geographic distribution, and recent isolation for African and European populations. The detection of T. britovi in West Africa modifies our knowledge about the distribution of encapsulated species of Trichinella in Africa. Thus, Trichinella nelsoni is now considered to have a distribution limited to the Eastern part of the Afrotropical region from Kenya to South Africa. This provides a plausible explanation for the presence of Trichinella T8 in Namibia and South Africa, and further suggests that T. britovi could be the Trichinella species circulating among wild animals of Northern Africa.     

Prevalence and spatiotemporal distribution of African swine fever in Lithuania, 2014–2017

Background

The emergence in 2014 and persistence of African Swine Fever (ASF) in Lithuania has been linked to infected wild boar movement and close contact with the carcasses of other infected wild boars. Over time the number of reported cases of ASF in wild boars gradually increased, but no detailed epidemiological data has been available. Therefore, the objective of the present study was to determine ASF virus prevalence in wild boars and domestic pigs during the 2014–2017 period and further explore the current geographical distribution of the virus.

Results

Our study results show that ASF virus prevalence in hunted wild boars using PCR analysis increased from 0.83% (95% CI 0.69–0.98) to 2.27% (95% CI 2.05–2.48) from 2014 to 2016 respectively. However, there was a dramatic jump in the number of ASF positive wild boars cases in 2017 resulting in prevalence of 12.39% (95% CI 11.91–12.86) (p <?0.05).The average prevalence of ASF-specific antibodies in wild boar population during years 2014–2017 was 0.45% (95% CI 0.39–0.51) based on ELISA test results.Prevalence of ASF virus in domestic pigs ranged from 0.24% (95% CI 0.17% - 0.32) in 2015 to 2.74% (95% CI 2.33% - 3.15) in 2017. The average seasonal prevalence of ASF virus in pigs was statistically significant (p?<?0.05) and ranged from 0% in spring to 3.68% (95% CI 3.32–4.05) in summer. Correlation between the pig density and number of recorded pig ASF cases in affected regions was only found in 2017 (R =?0.78, p?<?0.05). No correlation was detected between the wild boar density and number of recorded pig or wild boar ASF - positive cases.

Conclusions

This study provides the first results of ASF virus prevalence changes in Lithuania during the 2014–2017. The overall results confirm the relatively high prevalence of ASF virus in wild boar that was gradually increasing from 2014 to 2017. In the last year of study, the number of ASF positive cases in both domestic pigs and wild boars had unexpectedly increased several times. A better understanding of current status of the disease will enable better control and prevent further spread of ASF virus in Western Europe.

     

BsaH Ⅰ对猪激素敏感脂肪酶基因外显子Ⅰ的PCR-RFLP分析

以华北野猪、东北野猪和山西黑猪、长白猪、大白猪、马身猪共计287头猪作为研究对象,对其HSL基因外显子Ⅰ区域进行了PCR-RFLP多态性研究,发现不同品种猪间存在多态性。瘦肉型大白猪、长白猪全部表现为GG基因型;山西黑猪表现为AA、AG和GG三种基因型;脂肪型地方猪种马身猪为单一的AA基因型;华北杂种野猪、东北纯种野猪及杂种野猪表现为AG和GG两种基因型。等位基因A、G及三种基因型的频率在不同猪种中不同。该研究首次对华北及东北野猪HSL基因进行了多态性研究,丰富了国内外对野猪的分子生物学研究,为野猪遗传资源的合理开发利用提供了依据。     

Trichinella murrelli in scavenging mammals from south-central Wisconsin, USA

Tissues and serum from 59 raccoons (Procyon lotor), 42 coyotes (Canis latrans), and seven Striped Skunks (Mephitis mephitis) collected in Dane and Iowa Counties, Wisconsin, USA, between October 2005 and March 2006 were microscopically and serologically examined for the presence of Trichinella spp. Encapsulated larvae were found on compression slides prepared from tongue tissues from a few animals. Complete tissue digestion of tongues revealed that 19% of the raccoons, 26% of the coyotes, and none of the seven skunks tested were infected with Trichinella spp. Cats were subsequently experimentally infected by feeding them the raccoon tissues containing muscle larvae, and muscle larvae isolated from the collected tongues were experimentally transmitted to mice. Multiplex polymerase chain reaction analysis of the isolated muscle larvae demonstrated two distinct bands migrating at 127 base pairs (bp) and 316 bp in all samples, which together are diagnostic for Trichinella murrelli; the isolates were assigned Istituto Superiore di Sanita (ISS) codes ISS1656 through ISS1667, and ISS1708 through ISS1710 by the International Trichinella Reference Centre. These findings extend the geographic range of T. murrelli into Wisconsin, USA.     

Serosurveillance for selected infectious disease agents in wild boars (Sus scrofa) and outdoor pigs in Switzerland

The large abundance of free-ranging wild boars (Sus scrofa) and a trend towards animal friendly outdoor management of domestic pigs lead to an increasing probability of disease transmission between those animal populations. In 2001, an active monitoring was started for classical swine fever (CSF), Aujeszky’s disease (AD) and porcine brucellosis (PB) in wild boars in Switzerland. The objective of this programme was to document the serological status of wild boars regarding the selected pathogens. To continue this serosurveillance, 1,060 wild boar samples were collected during two regular hunting seasons in 2004–2005. Furthermore, in a pilot study, 61 outdoor pigs from 14 farms located in areas with high wild boar densities were sampled in 2004 and serologically tested for AD and PB. All wild boar samples were negative for CSF. Seroprevalence for AD was 2.83% (95% CI 1.91–4.02%). Seroprevalence for PB was 13.5% (95% CI 10.7–16.7%) for the Rose Bengal test and 11.05% (95% CI 8.82–13.61%) for the indirect ELISA. There was no serological evidence for AD in domestic pigs. All tested animals from 13 piggeries were seronegative for PB, but three pigs from the same farm showed doubtful results. Further investigations on the farm did not indicate the presence of PB in the herd. These findings urge the need for better diagnostic tools to obtain reliable results concerning PB prevalence. Since contact and following transmission of infectious agents between infected wild boars and outdoor pigs might occur in the future, it is advisable to include outdoor pigs in areas at risk in routine surveillance programmes.     

Genetic relationship and distribution of the Japanese wild boar (Sus scrofa leucomystax) and Ryukyu wild boar (Sus scrofa riukiuanus) analysed by mitochondrial DNA

Mitochondrial genetic variations were used to investigate the relationships between two Japanese wild boars, Japanese wild boar (Sus scrofa leucomystax) and Ryukyu wild boar (S.s. riukiuanus). Nucleotide sequences of the control (27 haplotypes) and cytochrome b (cyt-b) regions (19 haplotypes) were determined from 59 Japanese wild boars, 13 Ryukyu wild boars and 22 other boars and pigs. From phylogenetic analyses, the mtDNA of Ryukyu wild boar has a distinct lineage from that of Japanese wild boar, which was classified into the Asian pig lineage. This result suggests that the Ryukyu wild boar has a separate origin from the Japanese wild boar.     

Importance of the wild boar in the epidemiology of wild trichinellosis in Piedmont and Liguria

Muscle samples from 1,508 wild boars (Sus scrofa) harvested in Piedmont and Liguria between 1987 and 1990 were examined for larvae of Trichinella sp. by trichinelloscopy and peptic digestion. No wild boar was infected, though trichinellosis was present in foxes (Vulpes vulpes) from the same areas (14 positives out of 608) with the highest prevalence in the mountain range (25.0%). Infection was also not found in 40 stone martens, 5 badgers and 3 stoats. Five vulpine isolates were identified as belonging to T3 zymodeme. These results show that free-ranging wild boars, although being a source of trichinellosis infection for man in Italy (9.2% of the 584 cases diagnosed since 1961), have no significance in maintaining the sylvatic cycle of this parasite in the study area. The data agree with the low infectivity of Italian strains of fox origin for swine seen in the experimental infections carried out so far.     

Associations between Trichinella species and host species in Finland

Examination of 627 wild animals--raccoon dogs (Nyctereutes procyonoides), red foxes (Vulpes vulpes), European lynxes (Lynx lynx), brown bears (Ursus arctos), wolves (Canis lupus), and badgers (Meles meles)--revealed Trichinella spp. The prevalence varied according to geographical region of Finland (north; southwest, SW; and southeast, SE) and was the highest among lynxes (70%, SW). The risk of trichinellosis was higher in the SE (odds ratio, OR, 19.4) and SW regions (OR 14.3), as compared with the northern region (OR 1), with no difference between the former 2 regions. Foxes (OR 2.1) and lynxes (OR 1.9) had a higher risk than raccoon dogs (OR 1) of being infected. The distribution of different Trichinella species was evaluated in 87 wild and domestic mammals by multiplex polymerase chain reaction. Trichinella spiralis was detected more often in domestic and synanthropic animals than in sylvatic hosts. Trichinella nativa was detected only in wildlife. Trichinella pseudospiralis was found both in sylvatic and synanthropic hosts. Trichinella britovi was detected only in mixed infections with other Trichinella species. The raccoon dog was the sole host for all 4 Trichinella species and also carried the most intense infections.     

Mitochondrial Genetic Variation in Chinese Pigs and Wild Boars

The mitochondrial DNAs (mtDNAs) from 30 pig breeds (29 Chinese native breeds and 1 European breed) and wild boars were investigated for restriction fragment length polymorphisms (RFLPs) to determine the phylogenetic relationships and genetic diversity among pig breeds and wild boars. Of the 24 enzymes used, 8 (AvaI, BclI, BglII, EcoRI, EcoRV, ScaI, StuI, and XbaI) detected polymorphisms. By combining the cleavage patterns for each enzyme, 108 individuals were sorted into eight mtDNA mitotypes. There are two haplotype lineages in domestic pigs, i.e., Chinese and European lineages. The pairwise nucleotide sequence divergence was calculated to be 0.56% between Chinese pigs and European pigs, suggesting that they might have diverged from a common ancestor approximately 280,000 years ago. The wild boars showed more extensive genetic variation, four mitotypes were detected in six wild boars. In addition, one of the Zhejiang wild boars was found to share the same mitotype with Chinese native pigs. A UPGMA tree based on genetic distance among mitotypes indicated that mtDNAs of Chinese pigs and European pigs are clearly divided into two clusters, and Chinese wild boars are more closely related to the Chinese pigs. Our results provide molecular evidence to support the previous hypothesis that pigs may be derived from two maternal origins, Asian and European wild boars. Chinese native pig breeds may have a single origin.     

The epidemiological investigation of Trichinella infection in brown rats (Rattus norvegicus) and domestic pigs in Croatia suggests that rats are not a reservoir at the farm level

Whether the brown rat (Rattus norvegicus) is a reservoir of Trichinella spp. infection or merely an accidental host, which may be vector of Trichinella spp., continues to be debated. We estimated the prevalence of Trichinella sp. infection in brown rat populations and in domestic pigs in 2 villages in Croatia, where Trichinella sp. infection in pigs has been endemic in the past 10 yr. Trichinella spiralis larvae, identified by a multiplex polymerase chain reaction analyses, were the only species detected in both rats and pigs. In 2001 and 2002, 2,287 rats were collected on 60 farms with different levels of sanitation and with, or without, T. spiralis-infected pigs. The prevalence of infection in rats ranged from 0.2 to 10.7%. Infected rats were detected only on farms with T. spiralis-positive pigs and low sanitation or formerly with low sanitation (P = 0.007, Fisher's exact test), yet no infected rat was detected on farms with T. spiralis-negative pigs. The finding that no infected rat was found on farms with T. spiralis-negative pigs suggests that, in the investigated area, the brown rat is not a reservoir but only a victim of improper pig slaughtering.