Análisis proteómico de los productos de excreción-secreción de cuatro aislados de Trichinella spiralis obtenidos de hospederos accidentales / Proteomic analysis of the excretion-secretion products of four Trichinella spiralis isolates obtained from accidental hosts

Resumen: Introducción: Trichinella spiralis es un nemátodo tisular que se aloja en el músculo esquelético de humanos y otros mamíferos y causa una serie de alteraciones fisiológicas. Las proteínas de los productos de excreción-secreción de T. spiralis juegan un papel importante en la aparición y regulación de estas alteraciones. Sin embargo, aún no se conoce el efecto de estos productos en la infección e invasión del parásito al hospedero. Métodos: Mediante un análisis electroforético en una dimensión, Western blot y espectrometría de masas, se evaluaron las diferencias y similitudes entre proteínas antigénicas y de superficie de cuatro aislados de T. spiralis obtenidos de hospederos accidentales (perros) y la cepa de referencia aislada de cerdos (MSUS/MEX/91/CM). Resultados: Utilizando ontología de genes, se encontraron cinco proteínas exclusivas de los hospederos accidentales. Después del análisis, se encontró que estas proteínas forman parte de la matriz extracelular del parásito, cuentan con actividad catalítica y están implicadas en la adhesión a las células del hospedero. La actividad antigénica de las cuatro cepas aisladas de hospederos accidentales es idéntica a la reportada para T. spiralis, visualizándose el triplete antigénico característico de 43, 45 y 47 kDa. Conclusiones: Las proteínas exclusivas de los hospederos accidentales proveen información para entender el mecanismo de acción de este parásito para penetrar el músculo y evadir la respuesta inmune en el hospedero.

Abstract: Background: Trichinella spiralis is an intestinal and tissue nematode specific for mammalian skeletal muscle, causing a series of physiological alterations. T. spiralis excretory-secretion products play an important role in the appearance and regulation of these alterations. However, the effect of these products on the infection and invasion of the parasite to the host is unknown. Methods: Differences and similarities between antigenic proteins and surface proteins of four accidental hosts isolates (dogs) of T. spiralis and the reference strain isolated from pigs (MSUS/MEX/91/CM) were assessed by electrophoresis, western blot and mass spectrometry. Results: Using gene ontology, five proteins exclusive to the accidental hosts were analyzed. The results showed that these proteins are part of the extracellular matrix of the parasite, present catalytic activity, and bind to host cells. The antigenic activity the four strains showed the antigenic triplet characteristic of T. spiralis of 43, 45 and 47 kDa. Conclusions: Five proteins exclusive to dog isolates provided information to understand the mechanism of action of this parasite to penetrate the muscle and evade the immune response in the host.

Avaliação da presença de Trichinella spiralis em equinos abatidos em Araguari, MG / Evaluation of the presence of Trichinella spiralis in horses slaughtered in Araguari, MG

A larva do nematódeo Trichinella spiralis é responsável por infectar animais como porcos, javalis e cavalos e por causar grave doença em seres humanos. Surtos devido ao consumo da carne de cavalo são mais comuns na Europa, principalmente na França e Itália, onde existe a cultura de ingeri-la crua. Apesar de no Brasil não ter relatos sobre a infecção desse parasita em animais e seres humanos, os abatedouros exportadores para alguns mercados, como Europa e Estados Unidos, são obrigados a pesquisar o parasita. No presente estudo, a presença de larvas de Trichinella spiralis foi avaliada em 14.852 cavalos abatidos entre dezembro de 2014 a julho de 2016 em um frigorífico de Araguari sob inspeção federal. Os animais eram procedentes dos estados de Minas Gerais, Goiás e Bahia. A técnica empregada foi a de digestão artificial, em que se utilizou o músculo masseter, seguindo as legislações da União Europeia: Diretiva 91/497/CEE, Diretiva 77/96/CEE, Regulamento (CE) N° 2075/2005 e suas alterações. A larva não foi encontrada em nenhuma amostra analisada.

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.

Cross-sectional study of Trichinella spp in pigs in CDR, Nepal using pepsin digestion and ELISA serology.

This epidemiological cross-sectional study was conducted in five major pig-producing districts of CDR, Nepal from November 2006 to April 2007. A total of 576 slaughtered pigs were randomly selected and diaphragmatic crus muscles (n = 551), corresponding sera (n = 487) and sera only (n = 25) were collected from them. Meat samples were examined by pepsin digestion to evaluate for larvae of Trichinella spp. The sera were investigated using ELISA to evaluate for antibodies against T. spiralis. The doubtful and positive sera from the ELISA test were investigated by end-point ELISA, and the Western blot was used for confirmatory diagnosis. Pepsin digestion did not detect the larvae of Trichinella spp. AB-ELISA showed 2 positive and 14 doubtful results, out of 344 sera analyzed. End-point ELISA and the Western blot had revealed that all 16 samples were truly negative. A questionnaire revealed no rodent control (70%), left over feeding practices (65%), dumping of farm waste (82.5%) and uncooked meat was being used as feed (100%). This study reveals that Trichinella spp has a low prevalence.

Application of Giemsa stain for easy detection of Trichinella spiralis muscle larvae

The application of Giemsa technique to stain compressed diaphragm samples obtained from rodents experimentally infected with Trichinella spiralis is described. Diaphragm samples from rats heavily infected with 20 muscle larvae per gram of body weight (20 ML/gbw) were cut into several pieces and stained with Giemsa; on the other hand, whole diaphragms from slightly infected mice (1 ML/gbw) were also stained with Giemsa. Besides, muscle samples were also stained with Giemsa. Observation at 10 x magnification revealed that both ML and nurse cells (NC) look as bluish structures clearly contrasting with the pinkish color of the non-infected muscle fibers. NC in the diaphragms of mice could be easily observed at naked eye as blue points contrasting with the pink surrounding areas formed by the non-infected muscle fibers. Among NC observed in the diaphragms of rats infected with 20 ML/gbw, 4.4% was multiple infection. These findings were confirmed in sectioned and hematoxylin-eosin stained specimens. This data could be usefulness for a rapid diagnosis of trichinellosis in post-mortem mammals without magnification procedures.

Application of Giemsa stain for easy detection of Trichinella spiralis muscle larvae

The application of Giemsa technique to stain compressed diaphragm samples obtained from rodents experimentally infected with Trichinella spiralis is described. Diaphragm samples from rats heavily infected with 20 muscle larvae per gram of body weight (20 ML/gbw) were cut into several pieces and stained with Giemsa; on the other hand, whole diaphragms from slightly infected mice (1 ML/gbw) were also stained with Giemsa. Besides, muscle samples were also stained with Giemsa. Observation at 10 x magnification revealed that both ML and nurse cells (NC) look as bluish structures clearly contrasting with the pinkish color of the non-infected muscle fibers. NC in the diaphragms of mice could be easily observed at naked eye as blue points contrasting with the pink surrounding areas formed by the non-infected muscle fibers. Among NC observed in the diaphragms of rats infected with 20 ML/gbw, 4.4% was multiple infection. These findings were confirmed in sectioned and hematoxylin-eosin stained specimens. This data could be usefulness for a rapid diagnosis of trichinellosis in post-mortem mammals without magnification procedures.

First record of human trichinosis in Chile associated with consumption of wild boar (Sus scrofa)

The first South American case of human trichinosis, resulting from the consumption of roast wild boar (Sus scrofa) is reported in Chile. The patient presented fever, diarrhea, myalgias, facial edema, sub-conjunctival reddening, photophobia, eosinophilia, and elevated glutamic oxalacetic transaminase. The diagnosis was confirmed by two immunoenzymatic tests (ELISA) using somatic and excretion-secretion antigens.

The effects of Trichinella spiralis infection on renal function in rats.

Trichinella spiralis infection was induced in rats by oral feeding of infective larvae. Four weeks later, renal function, including renal plasma flow (RPF), glomerular filtration rate (GFR), excretion rate of protein, sodium and potassium were determined using clearance technics. There were no significant changes in these parameters. However, plasma urea nitrogen was significantly higher in the infected group, suggesting that either an impaired regulation of renal tubular urea transport or an increased skeletal muscle breakdown is likely.

Búsqueda de anticuerpos contra trichinella spiralis en roedores de libre desplazamiento capturados en un zoológico de la Ciudad de México / Search for antibodies against trichinella spiralis in free roaming rodents caught in a zoological park from Mexico City

A serological survey to search for antibodies against T. spiralis was performed in free roaming rats (n=64) and mice (n=35) caught in zoological park from Mexico City. Serum samples were analyzed by ELISA and immunoelectrotransfer blot assay (EIBT). None serum show positive absorbance values in ELISA nor recognized T. spiralis specific antigenic fractions in EIBT. However, two rat samples recognized three antigens of 31,37 y 55 kDa, while one of them reacted with two additional antigens of 64 and 67 kDa. As it is known that the antigen epitope profiles varied among trichinella species, it could be possible that in rats, there is 3 percent of antibody prevalence agains trichinella sp.; however, due that other organisms could induce the production of cross-reacting antibodies, such conclusion can not be supported at all. These results suggest that T. spiralis was not part of helminthological fauna in these rodents

Brotes epizoóticos de triquinosis en dos criaderos de cerdos de la Región Metropolitana, Chile / Epizootic outbreaks of trichinosis in 2 pig farms from the Metropolitan Region, Chile

In Chile swine trichinosis has presented a progressive decreasing in the last two decades of XX century. T. spiralis pig infection descended from an average of 0,683 per 1000 in 1980-1984 to 0,315 in 1985-1989 and to 0,115 in 1990-1996. In the particular case of Metropolitan Region this decreasing has been more marked: from an average of 0,058 per 1000 in 1990-1994 to 0,003 in 1995-1999. Between the end of june 1999 and middle january 2000 in Metropolitan Region abattoirs T. spiralis was detected in 15 (4,9 percent) out of 306 swine from two pigsties located in El Monte (E.M) and Padre Hurtado (P.H) 45 and 30 km south-west from Santiago. In the same period another four pigs from the same premises were found infected in abattoirs of other regions. During inspection visits it was stated that both pig farms had deficient sanitary conditions. Phototrichinoscopy was positive in three out of five rattus norvegicus collected in E.M. In pigsty PH the examination of diaphragm samples of 25 dogs and 17 cats resulted negative. In the premises originating T. spirali infected swine the Metropolitan Environmetal Health Service Abattoirs Program carries out and epidemiological vigilance consisting in the follow-up of animls destined for slaughtering in order to initiate prophylactic actions oriented to eliminate eventual sources of trichinosis infection for human and rearing pigs

Triquinosis: importancia del caso índice / Trichinosis: importance of the index case

Se reporta el caso de un paciente varón de 28 años, previamente sano, proveniente de Curicó, que tras haber ingerido carne de cerdo faenado en domicilio, inició un cuadro clínico y exámenes de laboratorio altamente sugerentes triquinosis. Se notificó de inmediato a la sección de Epidemiología del ministerio de Salud, con lo que se logró precozmente encontrar al resto de los afectados y decomisar la carne contaminada para evitar nuevos casos, destacando de esta forma la importancia del caso índice. Posteriormente se apoyó el diagnóstico con test de ELISA para trichinella spiralis, y se inició el tratamiento anti parasitario con albendazol y sintomático, con lo cual el paciente evolucionó satisfactoriamente. Se destaca la importancia del diagnóstico precoz y de la oportuna notificación a las autoridades sanitarias cuando se tiene un cuadro clínico compatible, y exámenes de laboratorio generales, como hemograma y los niveles de CPK alterados, aun cuando no se disponga de exámenes serológicos específicos, como ELISA para triquinosis

Evaluation of fast-elisa for the diagnosis of experimental trichinosis

In order to compare between FAST-ELISA and ELISA for the diagnosis of experimental trichinosis and study the kinetics of antibody and eosinophilic responses, 6 New Zealand rabbits were infected orally by Trichinella spiralis larvae. Blood was collected every other day for the first 2 weeks, then weekly for 11 weeks post infection. T. spiralis crude larval antigen was prepared for coating of ELISA plates and FAST-ELISA beads. Blood was examined for eosinophilic count and for serum antibody level by ELISA and FAST-ELISA techniques. The burden of infection was assessed by counting encysted larvae in muscle samples of the infected rabbits. By FAST-ELISA antibodies were detected 7 days post infection [PI], while with ELISA technique antibodies were detected after 10 days. Both tests detected maximum antibody levels on the 4th week. The eosinophilic count reached its peak by the 2nd week. There was a significant inverse correlation between the mean eosinophilic count and the mean larval count. FAST- ELISA proved to be more sensitive than ELISA in early detection of infection, besides being a simple, fast and sensitive assay for antibody detection against T. spiralis

Frecuencia de triquinosis humana en Concepción y Arauco, Chile: estudios en personas autopsiadas en el Servicio Médico Legal de Concepción: junio 1996 a marzo 1997 / Frequency of human trichinosis in Concepción and Arauco, Chile: study in autopsied persons at the Servicio Médico Legal, Concepción: june 1996 to march 1997

The frequency of trichinosis was determined in 300 dead persons, residents in the provinces of Concepción and Arauco, and requiring necropsy at the Servicio Medico Legal of Concepción. Diaphragm muscle samples were taken from june 1996 to march 1997. The diagnosis was made by trichinoscopy and artificial digestion techniques. Five samples (1.67 percent) were positive. There was not differences between sexes. Positive cases were found mainly among 30 to 49 years old people belonging to middle-lower class. Both diagnostic techniques showed diferent sensitivity as a consequence of the parasitic load and on the contrary, the artificial digestion was less sensitiva when the cyts were fully calcified

Tendencia actual de la epidemiología de la triquinosis humana en Chile / Present trends of human trichinosis epidemilogy in Chile

Surveillance of the epidemiology of human triquinosis in Chile has been maintained during the last 30 years by the Department of Parasitology. Incidence and prevalence have been followed-up by analysing Ministry of Health annual reports and periodical phototrichinoscopic examination of diaphragm samples from individuals autopsied at the Santiago Medico Legal Service. A decrease has been observed on: incidence from 1.4 per 100.000 in the 1960s to 0.7 in the 1980s down to 0.3 in 1993 and 1994 and to 0.5 in 1995, and prevalence from 3.4 percent 2.8 percent, 2.0 percent and 0.8 percent in 1972, 1982, 1992 and 1997 respectively. A predominance of the prevalence in the groups of more age, with the observation of calcified Trichinella spiralis larvae should be indicating a decline of new infections in the general population. This decrease of frequence of human trichinosis in Chile is possible due to the interaction of two main factors, efficacy and higher implementation of control measures and significant advances in porcine technology with a resulting provision to consumers of pork from young animals practically trichinosis free

Actualidades sobre la triquinelosis equina / Current facts about horse trichinellosis

La triquinelosis es una zoonosis causada por parásitos del género Trichinella que es trasmitida principalmente por la ingestión de carne de animales como el cerdo, oso y zorro entre otros. Sin embargo, en Europa se han reportado varios brotes debido al comsumo de carne de caballo. La presencia del parásito no ha sido demostrada de manera directa en la carne de estos animales, sin embargo, la identificación de las especies de Trichinella (T. spiralis T. britovi y T. nativa) involucradas en estos brotes ha sido posible a partir de biopsias tomadas de individuos que consumieron carne de caballo. Recientemente, se identificaron por primera vez larvas de T. spiralis en caballos sacrificados en un rastro del Estado de México, presentando así evidencia directa de la infección de estos animales con el parásito. Por otro lado empleando extractos totales o antígenos TSL-1 de T. spiralis se han detectado anticuerpos en contra de Trichinella en caballos sacrificados en rastros de diferentes paises de Europa así como en México. Asimismo, la infección con varias especies de Trichinella se ha logrado reproducir experimentalmente en caballos y los resultados obtenidos son importantes en el desarrollo de métodos de diagnóstico que permitan estimas la prevalencia de esta infección en caballos cuya carne se destina para el consumo tanto animal como humano y eventualmente instrumentar medias para el control de la trasmisión de la triquinelosis por carne de caballos

Detection of circulating trichinella pseudospiralis antigen by enzyme linked immunosorbent assay [ELISA]

A modified double antibody sandwich ELISA was used as a sensitive technique for demonstrating the appearance of circulating antigen of T. pseudospiralis in the sera of infected mice. The sera were tested at 5 days and 2, 4, 6, 7, 8, 10 and 12 weeks PI. The antigen was first detected on the 5th day PI; its level increased to reach the 1st peak at 4 weeks, then declined at 6 weeks. A 2nd peak was observed at 8 weeks, followed by a sharp decline to give negative OD reading at 10 weeks and increased to a moderate level at 12 weeks PI

Effect of synanthic and ivomec on trichinella spiralis with reference to their serological evaluation

Three groups, each of 10 rats, were experimentally infected with Trichinella spiralis larvae [250/rat]. On the 5th week post-infection animals of two groups were treated with a single dose of synanthic [oxfendazole] and ivomec MK-933 at a dose of 5 mg and 0.2 mg/kg b.w., respectively. Efficacy rate of the former was 84.2% and for the later was 72.5% depending on the number of living larvae in the diaphragm. Anti Trichinella spiralis hemagglutinative antibodies were monitored before and after treatment. They were markedly declined two weeks after treatment. This makes passive hemagglutination test a feasible test to evaluate the drug efficacies either in laboratory or field studies

Epidemiología de la triquinosis en Chile: estudio de la prevalencia mediante reacciones inmunodiagnósticas / Epidemiology of trichinosis in Chile: study of prevalence by immunodiagnostic reactions

In the period 1983-1994 a series of seroepidemiological surveys by using immunodiagnostic tests for trichinosis, was carried out in 138 localities and health institutions and health institutions from the 13 regions of the country. Thus, a total of 12,882 randomly selected persons, with different sex and age distribution, were submitted to precipitin test and indirect hemagglutination test for trichinosis. One hundred and ninety one (1.5 percent) persons resulted positive for trichinosis. Higher rates of infection were observed in central and southern Chile. No differences of positivity, according to age sex, but constant increase parallel to age of the individual were detected. The general prevalence rate of infection of 1.5 percent is in reasonable accordance with the prevalence of 2.0 percent found in human corpses in 1992

Brote de triquinosis en la comuna de Purranque X Región, Chile: octubre/noviembre 1992 / Outbreak of trichinosis in Purranque county X Region, Chile: october/november 1992

An outbreak of trichinosis occurred in Purranque county, X Region, Chile, between october and november of 1992, which involved 36 persons. The incubation period, determinated by the clinical picture and laboratory assays, fluctuated between 10 and 12 days, myalgias (88,9 for percent) and palpebral oedema (86,1 por percent) were the most important symptoms, followed by fever (44,4 for percent) and headache (33,3 for percent). Eosinophils count ranged from two to 42 for percent the first week, and this value raised to 55 for percent the second week of the outbreak. Anti-trichinella spiralis antibodies were determinated by some serological tests such as: precipitin test (PT), bentonite flocculation test (BFT) and indirect hemagglutination test (IHAT) in sera IHAT (21,4 for percent) and BFT (3,6 for percent). After 15 days, the three tests were perfomed in sera from 12 patients. At that time, the positivity was elevated in all of them: PT (100,0 for percent), BFT (66,7 for percent) and IHAT (91,7 for percent). It is believed that the outbreak had its origin in infected pork meat that was consumed raw or insufficiently cooked without a previous veterinary inspection