Bulgarian Journal of Veterinary Medicine, 2017 ONLINE FIRST ISSN 1311-1477; DOI: 10.15547/bjvm.2038 Original article SPECIES COMPOSITION OF TRICHINELLA IN DOMESTIC AND WILD ANIMALS IN BULGARIA N. LALKOVSKI National Diagnostic and Research Veterinary Medical Institute, Sofia, Bulgaria Summary Lalkovski, N., 2017. Species composition of Trichinella in domestic and wild animals in Bulgaria. Bulg. J. Vet. Med. (online first). Four Trichinella species cause trichinellosis in Europe: Trichinella spiralis, Trichinella britovi, Trichinella nativа and Trichinella pseudospiralis. The aim of our study was to determine the prepon- derance of Trichinella species in Bulgaria. The research covered the period 2010–2016. Molecular analysis was performed with 120 Trichinella isolates. Two species were discovered: Trichinella bri- tovi and Trichinella spiralis. T. britovi predominated over T. spiralis – 113 isolates (94.17%) and 7 (5.83%) respectively. Both species were identified in domestic pigs and wild boars, with T. bri- tovi:T.spiralis ratios in 45:1 in wild boars and 1:1 in domestic pigs. T. britovi was the geographically more widespread species. It was found in samples from domestic and wild animals from all over the country, while T. spiralis has only been found in several areas. Key words: bear, badger, domestic pig, fox, jackal, Trichinella britovi, Trichinella spi- ralis, wild boar, wolf INTRODUCTION Trichinellosis is a foodborne disease that trichinellosis (Pozio & Zarlenga, 2005). affects skeletal muscle tissue of wild and According to FAO/WHO/OIE Guidelines domestic carnivores and omnivores. The 11 genotypes are identified in the genus causative agents of this disease are ne- Trichinella (Anonymous, 2007). Two matodes belonging to the genus Trichi- main clades are recognised in the genus: nella, family Trichinellidae. By the mid- the first that encapsulates in host muscle dle of last century it was thought that the tissue and a second that does not. The cause of trichinellosis in all animal species representatives of eight genotypes have was the same – Trichinella spiralis. How- species names. The encapsulated clade ever, the implementation of molecular includes 5 species: T. spiralis – genotype biology methods proved the existence of T1, T. nativa – genotype T2, T. britovi – a significant diversity of species and dif- genotype T3, T. murrelli – genotype T5 ferences in the taxonomy, the geographi- and T. nelsoni – genotype T7 and three cal dissemination and the epidemiology of genotypes (Trichinella T6, T8 and T9), Species composition of Trichinella in domestic and wild animals in Bulgaria which invades mammals. The non- authors identified T. britovi in 4 wild encapsulated (acapsular) clade includes boars and T. spiralis in only one sample three taxonomically distinct species: T. from wild boar. Between 1995 and 2002, pseudospiralis – genotype T4, T. papuae 18 human outbreaks have been recorded – genotype T10 and T. zimbabwensis – (Petkova et al., 2008). In seven of them genotype T11. The first one invades the etiological agent was identified as T. mammals and birds and the other two spe- spiralis and in 11 outbreaks as T. britovi. cies – mammals and reptiles. The exis- In wild boars the authors determined only tence of genotype T12 isolated from a T. britovi whereas in domestic pigs – both cougar in Argentina is supposed (Krivo- species. kapich et al., 2008). The purpose of this research was to From epidemiological point of view conduct a more extensive survey on spe- there are two cycles of trichinellosis: sy- cies composition and distribution of nantropic (domestic) cycle where T. spi- agents of trichinellosis in domestic and in ralis is usually involved and in rare cases wild animals in our country. T. britovi, as well as a sylvatic cycle spe- cific for all Trichinella species and geno- MATERIALS AND METHODS types. The most commonly spread etiological The research covered the period 2010– agents of trichinellosis in European wild 2016. The isolated larvae of Trichinella and domestic animals are T. spiralis and were detected during the routine official T. britovi. Two other species – T. nativа control of samples obtained from meat and T. pseudospiralis are of minor impor- products, slaughtered domestic pigs, wild tance as pathogens in domestic animals pigs and shot wild carnivores. The studies (Pozio, 2007). According to Pozio et al. were carried out following the reference (2009) T. britovi is more widespread in method of digestion using a magnetic stir- the Member States of the European Union rer according to Regulation (EU) 2015/ and it can be found in 62.5% to 100% of 1375. A total number of 120 isolates were the samples. T. spiralis is widespread in typified and sorted as followed: 8 from Finland, Germany, Poland and Spain – domestic pigs, 92 from wild boars, 5 from 56.3% to 92.8% of the samples. In Roma- foxes, 3 from jackals, 3 from wolves, 3 nia, the ratio is 49.2% to 50.8% in favour from badgers, 1 from a bear, 3 from raw of T. britovi. T. britovi is more widespread dried sausage made of wild boar meat and than T. spiralis in wild carnivores – 89% 2 from minced meat (made of mixture of vs. 11% respectively. According to this domestic pig meat, beef meat/horse meat). research T. spiralis is more commonly Trichinella spp. larvae were identified at found in wild boars (62% vs. 38%), do- the species level by multiplex PCR, which mestic pigs (82% vs. 18%) and rodents allowed differentiating the eight species of (75% vs. 25% respectively). Trichinella and one of the genotypes – T6. In Bulgaria, Kurdova et al. (2004) in- Ninety one Trichinella spp. isolates vestigated parasite isolates obtained upon were tested in the European Union Refer- 15 epidemic outbreaks. Obtained results ence Laboratory for Parasites (EURLP) in showed the predominance of T. britovi Italy according to the laboratory protocol (etiological agent in 10 outbreaks) while (http://www.iss.it/binary/crlp/ cont/MI_02 T. spiralis was found in 5 outbreaks. The _WEB_SITE_REV_6.pdf). 2 BJVM, ××, No × N. Lalkovski Table 1. Species composition and distribution of causes of trichinosis in domestic and wild animals in Bulgaria in the period 2010–2016 District Hosts and sources Isolates number Species Blagoevgrad Wild boar 32 Trichinella britovi Burgas Wild boar 2 Trichinella britovi Jackal 1 Trichinella britovi Varna Wild boar 1 Trichinella britovi Minced meat (pig+beef ) 1 Trichinella spiralis Veliko Tarnovo Wild boar 3 Trichinella britovi Vidin Wild boar 2 Trichinella britovi Fox 1 Trichinella britovi Vratsa Wild boar 1 Trichinella britovi Gabrovo Wild boar 8 Trichinella britovi Wild boar 1 Trichinella spiralis Dobrich No isolates Kardzhaly Wild boar 6 Trichinella britovi Kyustendil Wild boar 3 Trichinella britovi Lovech Wild boar 1 Trichinella britovi Wild boar 1 Trichinella spiralis Montana Fox 1 Trichinella britovi Pazardzhik Badger 1 Trichinella britovi Fox 1 Trichinella britovi Pernik Wild boar 1 Trichinella britovi Pleven No isolates Plovdiv Wild boar 3 Trichinella britovi Jackal 1 Trichinella britovi Razgrad No isolates Rousse Wild boar 1 Trichinella britovi Sliven Wild boar 7 Trichinella britovi Smolyan Wild boar 2 Trichinella britovi Wolf 2 Trichinella britovi Fox 1 Trichinella britovi Silistra Wild boar 2 Trichinella britovi Fox 1 Trichinella britovi Sofia-town Wild boar 7 Trichinella britovi Domestic pig 2 Trichinella spiralis Domestic pig 2 Trichinella britovi Raw dried sausage 2 Trichinella britovi Sofia-district Wild boar 3 Trichinella britovi Domestic pig 1 Trichinella britovi Domestic pig 1 Trichinella spiralis Wolf 1 Trichinella britovi Minced meat (pig+ horse) 1 Trichinella spiralis Raw dried sausage 1 Trichinella britovi Stara Zagora Wild boar 2 Trichinella britovi Badger 1 Trichinella britovi Bear 1 Trichinella britovi Targovishte Wild boar 1 Trichinella britovi Haskovo Wild boar 2 Trichinella britovi Shumen Domestic pig (free ranging) 2 Trichinella britovi Jackal 1 Trichinella britovi Badger 1 Trichinella britovi Yambol No isolates BJVM, ××, No × 3 Species composition of Trichinella in domestic and wild animals in Bulgaria Twenty-nine samples were tested in the re widespread Trichinella species (Fig. 1) National Reference Laboratory of Parasitic found in samples from domestic pigs, wild Zoonoses, Sofia (Lalkovski et al., 2011). boars and wild carnivores from 24 dis- tricts of the country, while T. spiralis has been found in only 5 districts. RESULTS The results of the multiplex PCR for Tri- DISCUSSION chinella spp. isolates identification are shown in Table 1. The above mentioned results showed that Molecular analysis was performed trichinellosis in domestic pigs, wild boars with 120 larval isolates from 24 districts and wild carnivores in the country was of the country. Two species were disco- caused by two Trichinella species – T. vered: Trichinella britovi and Trichinella spiralis and T. britovi. According to Kur- spiralis. T. britovi was found in 113 dova et al. (2004) the same two species (94.17%) and T. spiralis in 7 (5.83%) caused human trichinellosis outbreaks. samples. Both types were found in domes- Our results confirm the opinion of Pozio tic pigs and wild boars. Concerning spe- et al. (2009), that in most countries of the cies prevalence, there was a predominance European Union, including Bulgaria, T. of T. britovi (97.83%) in wild boars com- britovi was the more prevalent species. T. pared to T. spiralis (2.17%). In domestic spiralis was detected in meat and meat pigs the ratio T. britovi:T.spiralis was 1:1. products in five backyard domestic pigs. In wild carnivores only T. britovi was Given that four of them originated from found. T. britovi is the geographically mo- two adjacent districts (Sofia-town and Vidin ROMANIA Ro u s se Dobrich Pleven Varna Shumen SERBIA Vratsa Veliko Tarnovo Gabrovo Sofia Sliven Pernik Burgas Stara Zagora Yambol Kyustendil Plovdiv Pazardzhik Blagoevgrad Haskovo TURKEY 50 km GREECE 30 mi Fig. 1. Geographical distribution of Trichnella britovi and Trichnella spiralis in Bulgaria (2010–2016); ● Trichinella britovi; ■ Trichinella spiralis 4 BJVM, ××, No × N.