Anti-Trichinella antibodies detected in chronically infected horses by IFA and Western blot, but not by ELISA.

In the Balkan countries, where trichinellosis is a re-emerging zoonosis, it is of great importance to determine Trichinella infection prevalence among the major hosts, including horses. One method for monitoring prevalence is serological surveillance; however, the validity of serological methods in horses is not well understood. The dynamics of anti-Trichinella IgG production and circulating excretory/secretory (ES) antigens were investigated in three horses experimentally-infected with Trichinella spiralis. Horses were slaughtered at 32 week post infection (p.i.). Low worm burdens were found in all three animals. Anti-Trichinella IgG was detected up to 32 weeks p.i. by an indirect immunofluorescence assay (IFA) and by Western blot (Wb), but not by ELISA. The ELISA test detected antibodies for only a short period of time (up to 18 weeks p.i. using ES antigen or up to 20 weeks p.i. using tyvelose-BSA antigen). The presence of circulating muscle larvae ES antigen in sera of infected horses was observed by dot blot from the 4th week p.i. up to the 32nd week p.i.

The need for implementation of International Commission on Trichinellosis recommendations, quality assurance standards, and proficiency sample programs in meat inspection for trichinellosis in Serbia.

Implementation of methods to control inspection for Trichinella in meat recommended by International Commission on Trichinellosis (ICT), particularly the introduction of the quality assurance standards and proficiency panels for certified analysts is extremely important in Serbia and other countries where Trichinellosis is endemic. In spite of existing regulations, including the inspection of 0.5 g samples of diaphragm by the compression method or by artificial digestion of 1g samples, in Serbia 280 people were diagnosed with clinical trichinellosis after consumption of inspected meat during the period 2001--2002. These outbreaks, which occurred in the municipalities of Kumane, Surcin and Bogatic, were a consequence of inadequate application of inspection methods and insufficient education of some veterinary inspectors. The problem of inadequate veterinary inspection in Serbia can be overcome by strict application of the ICT recommendations for the control of Trichinella with specific emphasis on implementing the quality assurance system (QAS) and proficiency sampling (PS/--PP/panel).

Re-emergence of trichinellosis in southeastern Europe due to political and economic changes.

The countries of southeastern Europe including the Balkan region and bordering countries - Albania, Bulgaria, Bosnia and Herzegovina, Croatia, Greece, Hungary, Macedonia, Romania, Serbia and Montenegro, Slovenia, and the European part of Turkey - occupy a very important strategic position and represent a land bridge between Europe and Asia. In the majority of southeastern European countries, cases of trichinellosis among the human and animal populations were described in the late 19th or early 20th centuries. Trichinella infections among wildlife were also described in the aforementioned countries. Today, the prevalence of trichinellosis is different between the Balkans and bordering countries. A high prevalence of trichinellosis in domestic animals and humans has been reported in Bulgaria, Serbia and Montenegro, Romania and Croatia. A moderate prevalence was found in Bosnia and Herzegovina. In Hungary, human trichinellosis has not been present for a long period of time. However, sporadic cases were recorded in swine over the last 2 years. Trichinellosis has not been found among domestic animals and humans in Greece and Macedonia in recent years while in Turkey and Slovenia human trichinellosis is sporadic. The re-emergence of trichinellosis is connected with the changes in the social and political systems in Bulgaria and Romania. In Serbia and Montenegro as well in Croatia, however, a re-emergence of trichinellosis was due not only to political and social changes but also to wars that took place in these countries during the last years of the 20th century. Social, economic and political factors responsible for the re-emergence of trichinellosis in southeast European countries are discussed in this communication.

Epidemiology of Trichinella infection in the horse: the risk from animal product feeding practices.

A discovery in 2002 of a Trichinella spiralis-infected horse in Serbia offered an opportunity to conduct needed epidemiological studies on how horses, considered herbivores, acquire a meat-borne parasite. This enigma has persisted since the first human outbreaks from infected horse meat occurred in then 1970s. The trace back of the infected horse to a farm owner was carried out. Interviews and investigations on the farm led to the conclusion that the owner had fed the horse food waste in order to condition the horse prior to sale. Further investigations were then carried out to determine the frequency of such practices among horse owners. Based on interviews of horse producers at local horse markets, it was revealed that the feeding of animal products to horses was a common practice. Further, it was alleged that many horses, particularly those in poor nutritional condition would readily consume meat. A subsequent series of trials involving the experimental feeding of 219 horses demonstrated that 32% would consume meat patties. To confirm that horses would eat infected meat under normal farm conditions, three horses were offered infected ground pork balls containing 1100 larvae. All three became infected, and at necropsy at 32 weeks later, were still positive by indirect IFA testing, but not by ELISA using an excretory-secretory (ES) antigen. This result indicates that further study is needed on the nature of the antigen(s) used for potential serological monitoring and surveillance of horse trichinellosis, especially the importance of antigenic diversity. The experimentally-infected horses also had very low infection levels (larvae per gram of muscle) at 32 weeks of infection, and although the public health consequences are unknown, the question of whether current recommended inspection procedures based on pepsin digestion of selected muscle samples require sufficient quantities of muscle should be addressed. It is concluded that horses are more willing to consume meat than realized and that the intentional feeding of animal products and kitchen waste is a common occurrence among horse owners in Serbia (and elsewhere?). This is a high risk practice which demands closer scrutiny by veterinary and food safety authorities, including the implementation of rules and procedures to ensure that such feeds are rendered safe for horses, as is now required for feeding to swine.

Social, political, and economic factors responsible for the reemergence of trichinellosis in Serbia: a case study.

Over the past decade, eastern Europe has experienced a resurgence of trichinellosis. A recent outbreak in Serbia, Yugoslavia, from December 2001 to January 2002, involving 309 people, revealed many of the causes for this reemergence. Epidemiological investigations indicate that the immediate cause of the recent outbreak was the consumption of smoked sausages produced by a small slaughterhouse or meat processor. However, failure of in-house meat inspection procedures and quality assurance as well as oversight by official veterinary control were also responsible. Further analysis of this breakdown in the food safety net revealed additional general factors that have yielded a seriously deficient veterinary control system, and these are factors that are relevant to the problems experienced throughout eastern Europe and other regions. The recent civil war that led to the breakup of the former Federation of Yugoslavia resulted in severe economic and demographic changes, including high inflation and external economic sanctions. This led to (1) the loss of large numbers of experienced veterinary control officers and their replacement with inexperienced personnel, (2) a change in the swine industry with reduction in the number of large establishments with in-house inspection and replacement with more than 1,000 small abattoirs, too small to afford full-time in-house inspection, and (3) an increase in smallholder pig farming with reduced government oversight to ensure high standards in pig-rearing practices (infection risk management). The consequences of these events have been a 300% increase in Serbian pig infection and a concomittant large increase in human outbreaks. Before 1990, swine trichinellosis in Serbia was confined to 4 small districts, but today about one third of the Republic is considered endemic for trichinellosis. The reemergence of trichinellosis in Serbia illustrates the ability of this zoonosis to "leak" through a poorly maintained food safety barrier and the vulnerability of effective veterinary control to national and international events.

The lateral flow card test: an alternative method for the detection of Trichinella infection in swine.

A novel lateral flow card (TS-Card pork) test was developed for the serological detection of Trichinella infected pigs. Based on extensive studies performed in Romania during 1999-2000 this test proved to be highly specific sensitive, rapid (3-12 minutes) and easy to use (no need for laboratory facilities). It can be used both for the detection of Trichinella infection in carcasses and for epizooliological studies using a variety of samples including whole or dried blood, serum, or tissue fluids. The TS-Card pork test, used as a screening test, can be the foundation of an on-farm or field based inspection system to significantly improve food safety in countries with a high prevalence of Trichinella in pigs or other food animal species. The results presented are also promising for application of the test in an on-line laboratory based inspection system since the speed of the test allows sufficient time to rail out suspected hog carcasses during the slaughter process.

Present status of trichinellosis in Yugoslavia: Serbia.

Trichinellosis was recognized almost one century ago as a health and animal husbandry problem in Serbia. In the last 10 years, trichinellosis has been expanding from three endemic regions (Srem, Macva and Negotinska Krajina) to neighbouring regions. The infection rate detected by veterinary inspection in 1999 year was 0.17% in slaughtered swines. Simultaneously, the number of infected humans increased three-five times in comparison with the period 1980-1990. For instance, 555 individuals were registered in 1999 as infected after the consummation of non-inspected pork from domestic swine or wild boar. Prevalence of trichinellosis in wild animals was examined more than a 20 years ago. The trichinellosis in horses has not been detected in the country, but infected horses imported from Serbia were detected in France and Italy.

International Commission on Trichinellosis: recommendations on methods for the control of Trichinella in domestic and wild animals intended for human consumption.

This document provides a uniform set of recommendations for the control of Trichinella at all levels (on the farm, at slaughter and in processed meats). These recommendations are based on the best scientific information available and represent the official position of the International Commission on Trichinellosis regarding acceptable control methods. These recommendations are subject to change as new scientific information becomes available.

[Susceptibility and the immune response to Trichinella spiralis infection]. / Prijemsivost i imunski odgovor na infekciju Trichinella spiralis-om.

The aim of this study was to assess a possible association between parameters of immune response and susceptibility/resistance to T. spiralis infection. The immune response to T. spiralis was monitored in two inbred strains of mice (BALB/c and C57Bl/6), as well as in the most important natural host--swine, by analysing the changes in main lymphocyte populations, the appearance of specific antibodies and worm burden recovery. In the murine model, the lower level of muscle larvae worm burden recovery in mice expressed greater resistance (BALB/c) was associated with manifested clinical signs, earlier development of humoral antibody response and a persistently higher level of CD4/CD8 T cell ratio, compared to T. spiralis-susceptible mice (C57Bl/6). In swine, high susceptibility to infection was also associated with lack of clinical symptoms, a delayed development of an antibody response and persistently low CD4/CD8 T cell ratio.

Epidemiology of swine trichinellosis in Yugoslavia.

Intensive epidemiological and seroepidemiological studies of Trichinella spiralis infection in swine have been carried out in the borough of Kladovo, one of the four regions in Yugoslavia where trichinellosis is endemic. A high prevalence (4.7%) of trichinellosis was discovered in three neighboring parishes adjoining the bank of the river Danube. Both geographically and epidemiologically these three parishes constitute a very unique condition that is very vulnerable to T. spiralis infection. Studies of 7,409 swine showed that the ELISA test for the detection of specific antibodies to T. spiralis in swine was highly reliable.

Current status of food-borne parasitic zoonoses--eastern Europe.

For this review, the countries of Eastern Europe are the USSR and Poland (in the eastern part of the European continent), what was formerly East Germany, Czechoslovakia, and Hungary (in the north and center), and Yugoslavia, Romania, Bulgaria, and Albania (to the south). During 1989 and 1990, all of these countries experienced remarkable political and social change, thereby making the term "Eastern Europe" a political anachronism. There are a great many differences among these countries in terms of human behavior, parasites, environment and animal husbandry practices. In spite of recent political changes, however, livestock husbandry practices, which affect the status of food-borne parasites, will not likely change in the near future. Some of the characteristics of the eastern European countries allow the epidemiology of food parasitic zoonoses to be discussed as a common problem.

Peripheral blood mononuclear cell subsets during Trichinella spiralis infection in pigs.

The immune response of 'Yugoslav meat breed' pigs inoculated with low doses of Trichinella spiralis muscle larvae was followed over two to nine weeks of primary infection, by analysing changes in peripheral blood mononuclear cell subsets, the development of a humoral antibody response and muscle larvae burden. During the course of the infection, infected animals showed a persistent elevation of both CD4+ and CD8+ T cell subsets from days 15 to 60 after the parasite exposure. During this time, the number of peripheral blood mononuclear cells expressing major histocompatibility complex class II antigens was also increased, while no significant differences were found in the number of circulating monocytes/macrophages and B cells over time. Humoral antibody responses to muscle larvae excretory-secretory products were evident as early as 41 days after infection, while the muscle larvae were recovered as early as 27 days after infection. The increased levels of CD4+ and CD8+ T cell subsets, as well as cells expressing major histocompatibility complex class II antigens in pigs exposed to T spiralis, may be indicative of some considerable alterations in cell subsets that are involved in the regulation of the swine immune response to this parasite.

Comparative efficacy of antigen and antibody detection tests for human trichinellosis.

Sera collected from patients with suspected or confirmed exposure to Trichinella spiralis were tested for circulating parasite antigens and antiparasite antibodies. Using an immunoradiometric assay, excretory--secretory antigens from muscle-stage larvae of T. spiralis were detected in the sera of 47% of 62 patients with clinical trichinellosis and 13% of 39 patients without clinical signs but suspected of exposure to infected meat. In comparison, antibodies were detected using an indirect immunofluorescent test in the circulation of 100% of the 62 patients with clinical trichinellosis and 46% of the 39 patients with suspected exposure. The presence of antibodies specific to excretory-secretory products of T. spiralis muscle larvae was confirmed in the majority of the samples tested by a monoclonal antibody-based competitive inhibition assay. These results indicate that antibody detection is a more sensitive diagnostic method for human trichinellosis, but that antigen detection might be a useful confirmatory test because it is a direct demonstration of parasite products in the circulation.

Immunoglobulin levels in seminal plasma of oligospermic patients.

The levels of immunoglobulins in the seminal plasma of healthy males were compared with those of oligospermic patients. The results show markedly increased levels of IgG in oligospermic patients. The presence of IgM, which has not been detected in the seminal plasma of healthy males, was estimated in some of the oligospermic patients. The authors recommend this simple screening test to be carried out in every male infertility follow up.