Trichinella Infection in Culled Wild Boar (Sus scrofa) from El Palmar National Park, Argentina, and Exposure Risk in Humans and Dogs Consuming Wild Boar Meat.

Trichinellosis is a foodborne disease caused by ingestion of raw or undercooked meat containing Trichinella spp. larvae. Consumption of wild boar (Sus scrofa) meat represents an important source of human trichinellosis worldwide. In El Palmar National Park (EPNP), Argentina, invasive alien wild boars are controlled and meat from culled animals is released for public consumption following on-site artificial digestion (AD) testing. Meat trimmings and offal from the control program are often used as food for dogs (Canis familiaris). We evaluated infection and exposure to Trichinella spp. in wild boars from EPNP, as well as exposure to Trichinella spp. and associated risk factors in dogs and human consumers of wild boar meat. Trichinella spp. larvae were detected in muscle samples from 5/49 wild boars by AD (10.2%; 95% confidence interval [CI], 3.8%-23%), with a mean burden of 0.24 larvae per gram (lpg; range, 0.06-0.95 lpg). Anti-Trichinella antibodies were not detected in wild boar serum samples (n=42). In dogs, 12/34 were seropositive to Trichinella spp. (35.29%; 95%, CI, 20.3%-53.5%). Immunoglobulin (Ig) G antibodies were not detected in human serum samples (n=63). Our results reveal the presence, albeit at low prevalence, of Trichinella spp. in wild boars and exposure in dogs fed game offal. These findings suggest that the low prevalence and parasitic load in wild boars, together with the best practices applied by EPNP culling program personnel, contribute to keeping the risk of infection in people low. The dog results highlight that the parasite is circulating in the area, and therefore the risk of infection is not negligible. We recommend the implementation of an animal surveillance strategy in order to monitor the evolution of this zoonosis in the study area.

Search for antibodies against Trichinella in two synanthropic Procyonidae species from southeast Mexico: white-nosed coatis (Nasua narica) and raccoons (Procyon lotor).

Trichinella is a nematode that are spread by the consumption of parasitized meat. Carnivora, a mammalian order, serve as key hosts for this parasite. However, evidence of Trichinella in wildlife from the Neotropics is extremely scarce, with reports documenting its presence only for five carnivore species: two Felidae, one Otariidae and two Mustelidae. Other widely distributed species that are consumed as bushmeat, such as Procyonidae, have not been studied in this context. A long-term study was performed for antibodies against Trichinella in coatis (Nasua narica) and common raccoons (Procyon lotor) in southeastern Mexico. Between the summer of 2009 to the winter 2013, a total of 291 coati samples and 125 raccoon samples were collected from a tropical green area located within an urban zone. An Enzyme-linked immunosorbent assay (ELISA) was used to detect antibodies against the excretory and secretory products of Trichinella spiralis muscle larva. ELISA-positive samples were further confirmed by Western Blot analysis. Results showed no evidence of antibodies during the first two years of study. However, in 2011, a sudden appearance of anti-Trichinella occurred. The seroprevalence reached its highest peak of 43% for coatis during winter 2013 and 53% for raccoons in summer 2013. This is the first study that provides evidence of Trichinella circulation within a neotropical procyonid community.

Absence of Trichinella spp. larvae in carcasses of road-killed wild animals in Paraná state, Brazil.

Trichinella spp. are zoonotic parasites that are widely distributed in warm-blooded carnivores and omnivores, including humans. Until the present moment, Brazil has been considered by World Animal Health Organization free from the domestic cycle of trichinellosis, whereas the parasite's sylvatic cycle has the status of infection in limited zones. However, neighboring countries such as Argentina have reports of parasite larvae in the wild fauna. The present study aimed to determine the occurrence of Trichinella spp. in road-killed wild animals in Paraná, Brazil. Biological samples from 71 wild animals-29 Didelphis albiventris, 11 Nasua nasua, ten Cerdocyon thous, seven Dasypus novemcinctus, six Leopardus guttulus, six Sphiggurus spinosus and two Puma concolor-collected from November 2016 to November 2021 were subjected to artificial digestion, following the methodology described in the REGULATION (EC) No. 2075/2005. No Trichinella spp. larvae were detected in the carcasses of the road-killed wild animals. However, considering the wide spectrum of possible reservoirs that could act as a link between the sylvatic and domestic cycles and considering the current Brazilian status of sylvatic trichinellosis in limited zones, frequent monitoring of wild fauna remains necessary.

The genus Trichinella and its presence in wildlife worldwide: A review.

The genus Trichinella has a worldwide distribution, infecting people, domestic animals, and wildlife. It includes 13 genotypes, which are geographically delimited; Trichinella is transmitted to people through the ingestion of undercooked meat. Historically, it has been associated with pigs, but most Trichinella species affect wildlife, and cases of trichinellosis due to the consumption of game meat have been emerging. Therefore, it is important to monitor the sources of transmission to domestic animals and humans. The objective of this work was to analyse reports of Trichinella spp. in wild/feral animals around the world to identify the needs of future research in the epidemiology of the sylvatic cycle. A search of studies published until 2021 was conducted using Web of Science and SciELO. In the Palearctic, the most commonly studied hosts were wild boars and red foxes, and hosts with the highest prevalence rates were polar bears and martens. In the Nearctic, red foxes and black bears were the most frequently studied hosts, and the highest prevalence was found for wolverines and brown bears. In the Neotropics, positive reports were only identified in two countries, with wild boars being the most commonly studied species, and armadillos featuring the highest prevalence. In the Afrotropics, Trichinella limits its presence to Sub-Saharan Africa, where lions are the most studied hosts, and spotted hyenas have the highest prevalence. In the Indo-Malaya and Australasia ecozones, information on wildlife is scarce; the Norwegian rat is the most frequently studied host, and the Tasmanian devil has the highest prevalence of infection. In the last decade, research on world wildlife has increased which is associated with more frequent trichinellosis outbreaks caused by the consumption of wild meat. The results suggest the need to increase research in developing countries, particularly where more diverse sources of meat are available for human consumption.

Absence of Trichinella spp. larvae in carcasses of road-killed wild animals in Paraná state, Brazil / Ausência de larvas de Trichinella spp. em carcaças de animais silvestres atropelados no estado do Paraná, Brasil

Trichinella spp. are zoonotic parasites that are widely distributed in warm-blooded carnivores and omnivores, including humans. Until the present moment, Brazil has been considered by World Animal Health Organization free from the domestic cycle of trichinellosis, whereas the parasite's sylvatic cycle has the status of infection in limited zones. However, neighboring countries such as Argentina have reports of parasite larvae in the wild fauna. The present study aimed to determine the occurrence of Trichinella spp. in road-killed wild animals in Paraná, Brazil. Biological samples from 71 wild animals—29 Didelphis albiventris, 11 Nasua nasua, ten Cerdocyon thous, seven Dasypus novemcinctus, six Leopardus guttulus, six Sphiggurus spinosus and two Puma concolor—collected from November 2016 to November 2021 were subjected to artificial digestion, following the methodology described in the REGULATION (EC) No. 2075/2005. No Trichinella spp. larvae were detected in the carcasses of the road-killed wild animals. However, considering the wide spectrum of possible reservoirs that could act as a link between the sylvatic and domestic cycles and considering the current Brazilian status of sylvatic trichinellosis in limited zones, frequent monitoring of wild fauna remains necessary.(AU)

Trichinella spp. é um parasito zoonótico de distribuição mundial detectado em carnívoros e onívoros de sangue quente e humanos. Até o momento, o Brasil é considerado pela Organização Mundial de Saúde Animal livre do ciclo doméstico da doença, enquanto o ciclo silvestre do parasito tem status de infecção em áreas limitadas. No entanto, em países vizinhos como a Argentina, larvas do parasito foram encontradas na fauna silvestre. O presente estudo teve como objetivo determinar a ocorrência de Trichinella spp. em animais silvestres atropelados no Paraná, Brasil. Amostras biológicas de 71 animais silvestres - 29 Didelphis albiventris, 11 Nasua-nasua, dez Cerdocyon thous, sete Dasypus novemcinctus, seis Leopardus gattulus, seis Sphiggurus spinosus e dois Puma concolor- coletadas de novembro de 2016 a novembro de 2021 foram submetidas à digestão artificial, seguindo a metodologia descrita do REGULAMENTO (CE) Nº2075/2005. Não foram encontradas larvas de Trichinella spp. nas carcaças dos animais selvagens atropelados. No entanto, considerando o amplo espectro de possíveis reservatórios que podem atuar como elo entre os ciclos silvestre e doméstico e considerando a condição atual da triquinelose silvestre no Brasil em áreas limitadas, o monitoramento frequente da fauna silvestre é necessário.(AU)

Reprint of: A review on Trichinella infection in South America.

Trichinella spp. causes human trichinellosis by means of the consumption of raw or inadequately treated meat from domestic or game animals. In the Americas, as well as in other continents, Trichinella infection is a health issue for humans and has a negative impact on the pork meat market, generated by people's fear of becoming infected with the parasite. The distribution of human cases and the sources of this disease in humans and animals were analysed in this report, which summarizes the information available regarding Trichinella infection in animals and humans in South America. Within South America, human infection with Trichinella was documented in Argentina and Chile during the period 2005-2019. Trichinellosis is endemic in these countries for, with human cases and foci in domestic and wild animals. In Argentina, human cases occur throughout the country, with foci found in pigs and wild animals. In Argentina, during the period 2012-2018, the number of suspected human cases reached 6,662. T. spiralis was identified in one South American sea lion (Otaria flavescens) from Patagonia, Argentina, for the first time in the region in 2018. In Chile, 258 human cases of trichinellosis were confirmed during the period 2005-2015; out of those 258 cases, most samples which tested positive for Trichinella spp. (29.5 %) were detected in the Metropolitan district (Santiago de Chile and outskirts), and 17.4 % in The Lake district. Regarding age brackets, people between 30-49 years of age showed the most cases (40.1 %). In Brazil, the infection is absent in domestic species but it has been found in wild boars (Sus scrofa) but limited to one or more region of the country. Within the animal species destined for food in South America, those that showed higher parasitical loads were pigs and wild boars, while armadillos (Chaetophractus villosus) and peccaries (Tayassu tajacu) showed very low Trichinella spp. larvae loads (0.04-0.1 larvae/g). Antibodies against Trichinella spp. have been detected in pigs from Ecuador and Bolivia. In Bolivia, antibodies were also found in humans. Peru, Colombia and Uruguay have no documented presence of Trichinella spp. in animals and humans. There is insufficient information regarding the presence of Trichinella spp. in domestic and wild animals, as well as in humans, since only a very limited number of surveys have been carried out. No papers with information on Trichinella spp. circulating in animals or humans have been published regarding the situation in Guyana, Surinam, French Guiana, Venezuela and Paraguay. Considering the growth of the guinea pig meat market in the Andean region, and the high prevalence of the disease reported in free range pigs and wild boars, as well as other game animal species, it is important to focus on the role of biosecurity and risk management, while improving meat market regulations, and detection of infection prior to consumption, in order to reduce the risk of transmission of this zoonotic disease to humans.

Temporal and geographic analysis of trichinellosis incidence in Chile with risk assessment.

Trichinellosis is a foodborne disease caused by several Trichinella species around the world. In Chile, the domestic cycle was fairly well-studied in previous decades, but has been neglected in recent years. The aims of this study were to analyze, geographically, the incidence of trichinellosis in Chile to assess the relative risk and to analyze the incidence rate fluctuation in the last decades. Using temporal data spanning 1964-2019, as well as geographical data from 2010 to 2019, the time series of cases was analyzed with ARIMA models to explore trends and periodicity. The Dickey-Fuller test was used to study trends, and the Portmanteau test was used to study white noise in the model residuals. The Besag-York-Mollie (BYM) model was used to create Bayesian maps of the level of risk relative to that expected by the overall population. The association of the relative risk with the number of farmed swine was assessed with Spearman's correlation. The number of annual cases varied between 5 and 220 (mean: 65.13); the annual rate of reported cases varied between 0.03 and 1.9 cases per 105 inhabitants (mean: 0.53). The cases of trichinellosis in Chile showed a downward trend that has become more evident since the 1980s. No periodicities were detected via the autocorrelation function. Communes (the smallest geographical administrative subdivision) with high incidence rates and high relative risk were mostly observed in the Araucanía region. The relative risk of the commune was significantly associated with the number of farmed pigs and boar (Sus scrofa Linnaeus, 1758). The results allowed us to state that trichinellosis is not a (re)emerging disease in Chile, but the severe economic poverty rate of the Mapuche Indigenous peoples and the high number of backyard and free-ranging pigs seem to be associated with the high risk of trichinellosis in the Araucanía region.

Survey of Trichinella in American minks (Neovison vison Schreber, 1777) and wild rodents (Muridae and Cricetidae) in Chile.

Trichinellosis is a zoonosis with worldwide distribution. Several hosts have been recognized around the world; however, there is a lack of knowledge of the role of feral mammals in Chile in its transmission. Herein, we tested muscle samples from 555 individuals among American minks (Neovison vison Schreber, 1777. n = 100) and several myomorph rodent species (Muridae and Cricetidae. n = 455) from southern Chile by artificial digestion to detect Trichinella larvae and identified the larvae at the species level through molecular analyses. Rodents were captured in agricultural and wild protected areas of several administrative regions (hereafter: region), while minks were captured in agricultural areas of the Los Ríos region. Trichinella spiralis larvae were detected in a synanthropic black rat (0.24%) of the Ñuble region and in seven minks (7%) trapped in agricultural areas of the Los Ríos region. The present results suggest that T. spiralis circulation is restricted to synanthropic rodents and minks living in agricultural areas where the parasite circulates among domesticated pigs. This study represents the first record of T. spiralis in a mustelid mammal in South America, increasing the number of feral species that could participate in the reservoir.

Trichinella spiralis in a cougar (Puma concolor) hunted by poachers in Chile.

Trichinellosis is a zoonosis caused by nematodes of the Trichinella genus, of which 10 species have been described. Species identification when larvae is found is only possible using molecular tools. Meat from pigs and game animals not subjected to veterinary inspection are the main sources of human infections. The hunting of native carnivores is prohibited in Chile due to conservation issues and the fact that those animals favor pest control. The illegal hunting of a cougar (Puma concolor) occurred in September 2020. Herein, the molecular identification of Trichinella larvae, by analyzing nuclear (expansion segment V) and mitochondrial (cytochrome C oxidase subunit I) sequences are described. Both the amplification of the expansion segment V region and the phylogenetic analysis of a segment of a fragment of the cytochrome c-oxidase subunit I sequence confirmed that the larvae belonged to T. spiralis. The case described herein represents the first evidence of illegal hunting of a protected mammal infected with Trichinella in Chile, highlighting the 'One Health' perspective to face this disease in the rural-sylvatic interphase.

Study of Trichinella patagoniensis in wild boars.

Trichinellosis is a zoonotic disease, which represents a significant public health concern in some South American countries, such as Argentina and Chile. Its impact is essentially due to absence of adequate control measures on meat from game animals, as well as the presence of illegal slaughterhouses and the trade of meat products without being tested for this parasite. In Argentina, trichinellosis is an endemic disease. At present, Trichinella spiralis, Trichinella patagoniensis, Trichinella pseudospiralis, and Trichinella britovi have been detected in animals from Argentina. Until now, T. patagoniensis had only been found in mountain cougars (Puma concolor) in Argentina but there is limited information available. The present study intends to determine susceptibility, serological response and distribution of muscle larvae in wild boars infected with T. patagoniensis, T. spiralis and T. pseudospiralis. For each of the Trichinella species five wild boars were inoculated with 20,000 muscle larvae. Except for two specimens which died during the experiment, the animals were euthanized 19 weeks post infection (pi). Blood samples were collected throughout the study in order to determine the antibody kinetics. Also, nine muscle samples from each specimen were taken and analysed for determination of larval distribution. Additionally, four muscle samples were used to obtain muscle juices. Wild boars infected with T. patagoniensis showed little to no larvae in the muscle samples analysed while animals infected with T. spiralis and T. pseudospiralis had a significantly high larval load in all the samples analysed. Optical density (OD) values remained above the cut-off value throughout the experiment. This is the first study to characterize the biological aspects of T. patagoniensis in wild boars.

Trichinella spiralis in a cougar (Puma concolor) hunted by poachers in Chile / Trichinella spiralis em um puma (Puma concolor) caçado por caçadores furtivos no Chile

Abstract Trichinellosis is a zoonosis caused by nematodes of the Trichinella genus, of which 10 species have been described. Species identification when larvae is found is only possible using molecular tools. Meat from pigs and game animals not subjected to veterinary inspection are the main sources of human infections. The hunting of native carnivores is prohibited in Chile due to conservation issues and the fact that those animals favor pest control. The illegal hunting of a cougar (Puma concolor) occurred in September 2020. Herein, the molecular identification of Trichinella larvae, by analyzing nuclear (expansion segment V) and mitochondrial (cytochrome C oxidase subunit I) sequences are described. Both the amplification of the expansion segment V region and the phylogenetic analysis of a segment of a fragment of the cytochrome c-oxidase subunit I sequence confirmed that the larvae belonged to T. spiralis. The case described herein represents the first evidence of illegal hunting of a protected mammal infected with Trichinella in Chile, highlighting the 'One Health' perspective to face this disease in the rural-sylvatic interphase.

Resumo A triquinelose é uma doença causada por nematoides do gênero Trichinella. Dez espécies foram descritas. A identificação das espécies a partir das larvas só foi possível com ferramentas moleculares. O abate doméstico de suínos e o consumo de animais de caça, sem inspeção veterinária, são as principais fontes de infecção humana. A caça de carnívoros nativos é proibida no Chile por questões de conservação e pelo fato de que esses animais favorecem o controle de pragas. A caça ilegal de um puma (Puma concolor) aconteceu em setembro de 2020. No Chile, a identificação molecular de larvas de Trichinella, obtida pelo exame de sequências nucleares (segmento de expansão V) e mitocondriais (citocromo C oxidase subunidade I) são descritas. Tanto a amplificação da região do segmento de expansão V quanto a análise filogenética de um segmento de um fragmento da sequência da subunidade I da Citocromo c-oxidase, confirmaram que as larvas pertenciam a T. spiralis. O caso aqui descrito representa a primeira evidência de caça ilegal de um mamífero protegido infectado com Trichinella no Chile, destacando a perspectiva de "Saúde Única" para enfrentar esta doença na interfase rural-silvestre.

Statistical approach to Trichinella infection in horses handled by rural slaughterhouses across five distinctive socioeconomic regions in Mexico

Abstract This work aimed to investigate the prevalence of Trichinella infection in horses (Equus ferus caballus) handled by rural slaughterhouses across five distinctive socioeconomic regions in Mexico. Serum samples were obtained by non-probabilistic convenience sampling in the Eastern, Southern Central and Western regions (100 samples of each). Additionally, muscle tissue samples were collected from the East (n = 45), Southeastern (n = 88), Southern Central (n = 39) and Southwestern (n = 11) regions. Antibodies were determined by Western blot and the muscle tissue was examined by artificial digestion. A global antibody prevalence of 2% was obtained. Regionally, a prevalence of 5% was observed in the East and 1% in the Southern Central region. No antibodies were detected in the West region and no larvae were found in the muscle tissue samples. These findings support the low presence of Trichinella in Mexican horses, which can positively impact the Mexican horse meat trade.

Resumen Se investigó la prevalencia de Trichinella en caballos (Equus ferus caballus) demataderos rurales en cinco regiones socioeconómicas de México. Los sueros se obtuvieron por Trichinella muestreo de conveniencia no probabilístico en las regiones Este, Centrosur y Oeste (100 muestras de cada una). Además se colectaron muestras de tejido muscular de animales de las regionesEste (n = 45), Centro Sur (n = 39), Sudeste (n = 88) y Sudoeste (n = 11). Los anticuerpos se determinaron por Western blot y el tejido muscular se examinó por digestión artificial. Se obtuvo una prevalencia global de anticuerpos del 2%. A nivel regional se observó una prevalencia del 5% enel Este y del 1% en el Centrosur. No se detectaron anticuerpos en el Oeste y no se encontraron larvas en el tejido muscular. Los datos respaldan la baja presencia de Trichinella en caballos mexicanos, lo que puede impactar positivamente en su comercio.

Trichinella spiralis y Trichinella patagoniensis: estudio de la carga globular aplicando el método de Azul Alcian / Trichinella spiralis and Trichinella patagoniensis: study of globular charge applying Alcian Blue method / Trichinella spiralis e Trichinella patagoniensis: estudo da carga globular pelo método de Azul de Alcian

Resumen Los residuos de ácido siálico de las glicoproteínas de superficie son los principales responsables de la carga negativa eritrocitaria. El objetivo de este trabajo fue estudiar alteraciones de carga globular producidas por Trichinella spiralis y Trichinella patagoniensis. Se trabajó con concentrados de larvas musculares de ambas especies y con eritrocitos frescos. Se incubó el sedimento globular con igual volumen de concentrado larval (1 y 2 horas). Los controles fueron incubados con solución salina. Se aplicó el método de Azul Alcian y se determinó el coeficiente experimental de carga aniónica eritrocitaria (CexpCAE). Los resultados mostraron que la carga disminuyó con el aumento del tiempo de tratamiento para ambas especies. Los valores de CexpCAE de las suspensiones incubadas con T. spiralis fueron menores que con T. patagoniensis, indicando que T. spiralis produjo mayor disminución de carga que T. patagoniensis. Se concluye que la desialización producida por ambas especies no es la misma, lo que sugiere que la relación hospedador-parásito que se establecería in vivo sería distinta.

Abstract The sialic acid residues of the surface glycoproteins are mainly responsible for the erythrocyte negative charge. The objective of this work was to study alterations of globular charge produced by Trichinella spiralis and Trichinella patagoniensis. Work was carried out on muscle larvae concentrates of both species and fresh erythrocytes. The treatment was performed by incubating the globular pellet with equal volume of larval concentrate (1 and 2 hours). Controls were incubated with saline solution. The Alcian Blue method was applied and the experimental coefficient of erythrocyte anion charge (expCEAC) was determined. The results showed that the globular charge decreased with increasing treatment time for both species. The expCEAC values of the suspensions incubated with T. spiralis were lower than with T. patagoniensis, indicating that T. spiralis produced a greater decrease in charge than T. patagoniensis. It is concluded that the desialization produced by both species is not the same, suggesting that the host-parasite relationship that would be established in vivo would be different.

Resumo Os resíduos de ácido siálico das glicoproteínas de superfície são os principais responsáveis pela carga negativa dos eritrócitos. O objetivo desse trabalho foi estudar as alterações da carga globular produzidas por Trichinella spiralis e Trichinella patagoniensis. Trabalhamos com concentrados de larvas musculares de ambas as espécies e com eritrócitos frescos. O tratamento foi realizado incubando o sedimento globular com igual volume de concentrado larval (1 e 2 horas), Os controles foram incubados com solução salina. Foi aplicado o método de Azul de Alcian e se determinou o coeficiente experimental de carga aniônica de eritrócitos (CexpCAE). Os resultados mostraram que a carga diminuiu com o aumento do tempo de tratamento para ambas as espécies. Os valores de CexpCAE das suspensões incubadas com T. spiralis foram menores que com T. patagoniensis, indicando que T. spiralis produziu uma diminuição maior na carga que T. patagoniensis. Conclui-se que a dessalinização produzida por ambas as espécies não é a mesma, sugerindo que a relação hospedeiro parasita que seria estabelecida in vivo é diferente.

Statistical approach to Trichinella infection in horses handled by rural slaughterhouses across five distinctive socioeconomic regions in Mexico.

This work aimed to investigate the prevalence of Trichinella infection in horses (Equus ferus caballus) handled by rural slaughterhouses across five distinctive socioeconomic regions in Mexico. Serum samples were obtained by non-probabilistic convenience sampling in the Eastern, Southern Central and Western regions (100 samples of each). Additionally, muscle tissue samples were collected from the East (n=45), Southeastern (n=88), Southern Central (n=39) and Southwestern (n=11) regions. Antibodies were determined by Western blot and the muscle tissue was examined by artificial digestion. A global antibody prevalence of 2% was obtained. Regionally, a prevalence of 5% was observed in the East and 1% in the Southern Central region. No antibodies were detected in the West region and no larvae were found in the muscle tissue samples. These findings support the low presence of Trichinella in Mexican horses, which can positively impact the Mexican horse meat trade.

Trichinella spiralis y Trichinella patagoniensis: cinética de desialización eritrocitaria / Trichinella spiralis and Trichinella patagoniensis: kinetics of erythrocyte desialization / Trichinella spiralis e Trichinella patagoniensis: cinética de dessialização eritrocitária

El objetivo del trabajo fue estudiar la cinética de desialización eritrocitaria producida por larvas infectantes de Trichinella spiralis y Trichinella patagoniensis. Se trabajó con 7 suspensiones eritrocitarias incubadas con 1.000±200 larvas musculares/mL, durante 2 horas, tomando muestra al tiempo inicial y cada 15 minutos. Los respectivos eritrocitos controles se incubaron de la misma manera con solución salina. Se aplicaron el método de titulación por Polibrene calculando el CexpST y un análisis de varianza (ANOVA) con las comparaciones múltiples según Tukey. Los resultados mostraron que el valor promedio de CexpST disminuyó con el aumento del tiempo de incubación, para ambas especies. En el tratamiento con T. spiralis no hubo diferencias significativas entre el valor medio del coeficiente a tiempo 60 y 75 minutos, mientras que con T. patagoniensis, a 45 y 60 minutos. Todas las restantes diferencias fueron significativas. La comparación entre los tratamientos, para cada uno de los tiempos, mostró que al tiempo inicial el coeficiente promedio no difirió entre las especies, pero que a todos los otros tiempos fue significativamente menor en la incubación de los eritrocitos con T. spiralis. Se concluye que la relación hospedador-parásito que se establece en ambos casos es distinta y probablemente también la capacidad de adaptación y de daño al hombre.

The objective of this work was to study the kinetics of erythrocyte desialization produced by infective larvae of Trichinella spiralis and Trichinella patagoniensis. It was performed on 7 erythrocyte suspensions incubated with 1,000±200 muscle larvae/ mL for 120 minutes, taking samples at the initial time and every 15 minutes. The respective control erythrocytes were incubated in the same way with saline solution. The Polybrene Titration method calculating the CexpST and variance analysis (ANOVA) with the multiple comparisons according to Tukey were applied. The results showed that the average value of CexpST decreased with the increase in incubation time, for both species. There were no significant differences between the mean value of the coefficient at 60 and 75 minutes in the treatment with T. spiralis, while neither were there any differences between 45 and 60 minutes in the incubation with T. patagoniensis. All other differences were significant. The comparison between the two treatments, for each of the times, showed that at the initial time the average coefficient did not differ between the species, but at all other times it was significantly lower in the incubation of the erythrocytes with T. spiralis. It is concluded that the parasite host relationship that is established in both cases is different and probably also is the ability to adapt and cause harm to man.

O objetivo do trabalho foi estudar a cinética de dessialização eritrocitária. produzida por larvas infectantes de Trichinella spiralis e Trichinella patagoniensis. O trabalho foi feito com 7 suspensões eritrocitárias incubadas com 1.000±200 larvas musculares/mL por 2 horas, colhendo amostras no tempo inicial e a cada 15 minutos. Os respectivos eritrócitos-controle foram incubados da mesma forma com solução salina. Foi aplicado o método de titulação por Polibreno calculando o CexpST e também uma análise da variância (ANOVA) com as comparações múltiplas de acordo com Tukey. Os resultados mostraram que o valor médio de CexpST diminuiu com o aumento do tempo de incubação para ambas as espécies. No tratamento com T. spiralis não houve diferenças significativas entre o valor médio do coeficiente no tempo 60 e 75 minutos, ao passo que com T. patagoniensis, aos 45 e 60 minutos. Todas as diferenças restantes foram significativas. A comparação entre os tratamentos, para cada um dos tempos, mostrou que no tempo inicial o coeficiente médio não diferiu entre as espécies, mas que em todos os outros tempos foi significativamente menor na incubação dos eritrócitos com T. spiralis. A conclusão é que a relação hospedeiro-parasita, estabelecida em ambos os casos, é diferente e provavelmente também a capacidade de adaptação e dano ao homem.

Synthetic gene as target to assess the sensitivity of PCR to detect Trichinella spp. larvae in meat from a non-endemic region.

Trichinellosis is a zoonotic disease exotic in Brazil but commonly found worldwide including South American countries like Argentina. International trading of swine meat needs an official Trichinella-free diagnosis commonly carried out by pepsin-HCl digestion of diaphragm tissue fragments followed by microscopic examination for the presence or absence of Trichinella larvae. The easiness of this diagnostic method allows it to be performed at slaughtering plants but, in contrast, it lacks sensitivity and does not allow species differentiation, which is fundamental for determining geographical and species distribution of different genotypes. In our study, we aimed to evaluate a highly sensitive diagnostic method based on the polymerase chain reaction (PCR) that would allow us to detect and classify different species of Trichinella. Thus, we designed a synthetic gene and selected five sets of primers targeting specific regions of the Trichinella genome. The synthetic gene was cloned into a plasmid and then used to optimize PCR conditions. Using our PCR, we were able to detect 0.001 pg of the synthetic gene, which corresponded to 0.01 larvae. Then, we collected 175 samples of Suidae (domestic and wild boars) diaphragm fragments that were pooled into groups, digested with pepsin-HCl, and had the DNA extracted for analysis by PCR. The clinical samples evaluated were negative by PCR. Our results indicate that the PCR-based method might be a useful diagnostic method complementary to the pepsin-HCl digestion method currently in use, mostly in non-endemic areas.

Detection of Trichinella britovi in pork sausage suspected to be implicated in a human outbreak in Mendoza, Argentina.

Of the three Trichinella species described in South America, T. spiralis, T. pseudospiralis and T. patagoniensis, only the former has been implicated in human infections from consumption of pork-derived products. During a presumed trichinellosis outbreak in 2012 in Mendoza, Argentina, we evaluated the serological responses of three patients who had eaten the incriminated food and had signs and symptoms compatible with trichinellosis, using ELISA. We also analyzed potentially contaminated pork sausage by artificial digestion technique and recovered Trichinella muscle larvae, which were identified to the species level using a PCR multiplex assay and by sequencing a region of the mitochondrial gene coding cytochrome oxidase subunit I. No antibodies were detected in the sera of the patients, probably because the samples were collected during the immunological window period. According to molecular identification, all larvae from the sausage corresponded to T. britovi. Trichinella britovi is reported here for the first time in the American Continent, and represents the only cold-tolerant member of the genus in the Neotropics. This species was most likely introduced from Europe to South America during Spanish colonization through pigs, wild boars and/or rats.

Efecto de Trichinella spiralis y de Trichinella patagoniensis n.sp. sobre la desialización / Effect of Trichinella spiralis and Trichinella patagoniensis n.sp. on desialization

El ácido siálico tiene importantes funciones biológicas, muchas de las cuales determinan su participación en la respuesta inmune. El objetivo del trabajo fue comparar el efecto de Trichinella spiralis y Trichinella patagoniensis n.sp. sobre la desialización eritrocitaria. Se trabajó con 10 concentrados de larvas musculares de T. spiralis y 10 de T. patagoniensis de la misma concentración larval. Se realizó el tratamiento incubando el sedimento de eritrocitos frescos con igual volumen de concentrado larval (37 ºC), tomando muestra a los 30, 60 y 90 minutos. Los controles fueron incubados de la misma forma con solución salina. Se aplicó el método de Titulación de la Agregación por Polibrene y se determinó el CexpST. Los resultados mostraron que el valor medio del CexpST en los eritrocitos tratados con T. spiralis fue significativamente menor que en los glóbulos tratados con T. patagoniensis, para todos los tiempos estudiados. El aumento del tiempo de tratamiento también disminuyó significativamente el valor medio del CexpST para las dos especies. Éste fue significativamente menor a los 90 minutos de incubación que a los 60 minutos y éstos a su vez menores que a los 30 minutos. Se concluye que T. spiralis provocó mayor desialización eritrocitaria que T. patagoniensis en las condiciones experimentales estudiadas.

Sialic acid has important biological functions, many of which determine its participation in the immune response. The objective of this paper was to compare the effect of Trichinella spiralis and Trichinella patagoniensis n.sp. on erythrocyte desialization. Work was performed on 10 larval concentrates of muscle larvae of T. spiralis and 10 of T. patagoniensis of the same larval concentration. The treatment was carried out incubating the sediment of fresh erythrocytes with an equal volume of larval concentrate (37 °C), taking samples at 30, 60 and 90 minutes. The controls were incubated in the same way treated with saline solution. Titration of aggregation by Polybrene Method was applied and the CexpST was determined. The results showed that the mean value of CexpST in erythrocytes with T. spiralis was significantly lower than in the globules treated with T. patagoniensis, for all the studied times. The increase in treatment time also significantly decreased the mean value of CexpST for the two species, being significantly lower at 90 minutes of incubation than at 60 minutes and these in turn lower than at 30 minutes. It is concluded that T. spiralis caused greater erythrocyte desialization than T. patagoniensis in the experimental conditions studied.

O ácido siálico tem importantes funções biológicas, muitas das quais determinam sua participação na resposta imune. O objetivo foi comparar o efeito de Trichinella spiralis e Trichinella patagoniensis n.sp. sobre a dessialização eritrocitária. Trabalhou-se com 10 concentrados de larvas musculares de T. spiralis e 10 de T. patagoniensis da mesma concentração larval. Realizou-se o tratamento incubando o sedimento de eritrócitos frescos com igual volume de concentrado larval (37 ºC), tomando amostra aos 30, 60 e 90 minutos. Os controles foram incubados da mesma forma com solução salina. Foi aplicado o método de Titulação da Agregação por Polibrene e se determinouo CexpST. Os resultados mostraram que o valor médio do CexpST nos eritrócitos Tratados com T. spiralis foi significativamente menor que nos glóbulos tratados com T. patagoniensis, para todos os tempos estudados. O aumento do tempo de tratamento também diminuiu significativamente o valor médio do CexpST para as duas espécies, sendo significativamente menor aos 90 minutos de incubação que aos 60 minutos e eles por sua vez menores que aos 30 minutos. Conclui-se que T. spiralis provocou maior dessialização eritrocitária que T. patagoniensis nas condições experimentais estudadas.

Prevalencia de Trichinella Spiralis en carnes de cerdo comercializadas en el mercado de la ciudad de Posadas y caracterización de su producción primaria / Prevalence of Trichinella Spiralis in pork meat sold in the market of the city of Posadas and characterization of its primary production

INTRODUCCIÓN En los últimos años, el consumo de carne de cerdo en Argentina denota un incremento sostenido. En Misiones, gran parte de la producción proviene de unidades familiares rurales que abastecen al mercado interno. La triquinosis es una zoonosis cuyo agente es Trichinella spiralis, y está ligada con la producción porcina. Su contagio humano se produce principalmente por ingestión de carne cruda o mal cocida de cerdo o sus derivados. OBJETIVOS Determinar la prevalencia de Trichinella spiralis en carnes de cerdo destinadas a la comercialización en mercados de la ciudad de Posadas y caracterizar su producción primaria. MÉTODOS Se recolectaron muestras de carne de cerdo de cuatro mercados de Posadas, que provenían de granjas productoras de diferentes localidades, y se procesaron en busca del parásito por la técnica de digestión de muestras agrupadas con utilización de un agitador magnético, remitiendo resultados previamente a su comercialización. Se realizaron encuestas estandarizadas en dichas granjas para la caracterización productiva. Los datos se procesaron con planillas de cálculo de uso libre. RESULTADOS Tras el análisis de 1235 muestras, la prevalencia obtenida fue de 0%. Las muestras provenían de 8 establecimientos productores de 5 municipios. La caracterización productiva arrojó los siguientes resultados escaso aislamiento sanitario (25%, n=8), desconocimiento de las enfermedades animales (100%, n=8), presencia de fauna sinantrópica y silvestre en las proximidades con abundante vegetación que pueda servir de refugio (75%, n=8), adecuada asistencia profesional (100%, n=8) y realización de tratamientos para el control de roedores (100%, n=8). DISCUSIÓN Los resultados contribuyen a mejorar el conocimiento epidemiológico regional y la planificación estratégica de acciones preventivas. La replicación de este modelo de control sanitario previo a la comercialización se constituye como una herramienta de vigilancia sanitaria eficaz frente a esta zoonosis.

Multiplex TaqMan qPCR assay for specific identification of encapsulated Trichinella species prevalent in North America.

BACKGROUND Human trichinellosis is a foodborne parasitic zoonotic disease caused by ingestion of raw or undercooked meat infected with nematode larvae of the genus Trichinella. In the USA, sporadic cases and outbreaks caused by the consumption of wild game meat infected with Trichinella have been reported. The current methods for diagnosis such as serology and microscopy are not specific, may result in false negative results, and cannot differentiate encapsulated Trichinella larvae to species level. The molecular protocols currently available for the differentiation of all encapsulate Trichinella species prevalent in North America have some limitations such as the inability to identify and resolve the presence of several Trichinella species in a single test. OBJECTIVES/METHODS In this study we developed and evaluated a multiplex TaqMan quantitative real-time polymerase chain reaction (qPCR) assay, which can simultaneously detect, identify and differentiate all species of encapsulated Trichinella occurring in North America i.e., T. nativa, T. spiralis, T. murrelli and Trichinella T6, even in cases of multiple infection in a single sample. We investigated two human biopsies and 35 wild animal meat samples considered as having a high likelihood of harboring Trichinella larvae obtained from the United States during 2009-2017. FINDINGS Using the multiplex assay describe here, 22 (59%) samples that tested positive contained Trichinella spp., were identified as: T. nativa (n = 7, including a human biopsy), T. spiralis (n = 9, including a human biopsy), T. murrelli (n = 3), Trichinella T6 (n = 1). Results also included two rare mixed infection cases in bears, a T. nativa/T. spiralis from Alaska and a T. spiralis/Trichinella T6 from California. The species identifications were confirmed using a conventional PCR targeting the rRNA ITS1-ITS2 region, followed by DNA sequencing analysis. The estimated limit of detection (LOD) was approximately seven larvae per gram of meat. MAIN CONCLUSIONS Differentiation of Trichinella spp. is needed to improve efforts on identification of case, optimize food safety control and better understand the geographic distribution of Trichinella species. The Trichinella qPCR multiplex proved to be a robust, easy to perform assay and is presented as an improved technique for identification of all known encapsulated species occurring in North America continent.