American Trypanosomiasis Infection in Fattening Pigs from the South-East of Mexico

Authors

  • M. Jiménez-Coello,

    1.  Laboratorio de Biologia Celular, CA Biomedicina de Enfermedades Infecciosas y Parasitarias, C.I.R. ‘Dr. Hideyo Noguchi’, Universidad Autonoma de Yucatan, Yucatan, Mexico
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  • K. Y. Acosta-Viana,

    1.  Laboratorio de Biologia Celular, CA Biomedicina de Enfermedades Infecciosas y Parasitarias, C.I.R. ‘Dr. Hideyo Noguchi’, Universidad Autonoma de Yucatan, Yucatan, Mexico
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  • E. Guzman-Marin,

    1.  Laboratorio de Biologia Celular, CA Biomedicina de Enfermedades Infecciosas y Parasitarias, C.I.R. ‘Dr. Hideyo Noguchi’, Universidad Autonoma de Yucatan, Yucatan, Mexico
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  • A. Ortega-Pacheco

    1.  Departamento de Salud Animal y Medicina Preventiva. CA Salud Animal, Facultad de Medicina Veterinaria y Zootecnia, Universidad Autonoma de Yucatan, Yucatan, Mexico
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  • The work was carried out in Universidad Autonoma de Yucatan, Yucatan, Mexico.

A. Ortega-Pacheco. Departamento de Medicina y Cirugia, CA Salud Animal. Facultad de Medicina Veterinaria y Zootecnia, Universidad Autonoma de Yucatan, Km. 15.5 Carr. Merida-Xmatkuil AP 4-116 Merida, Yucatan, Mexico. Tel.: +52 (999) 9423200, ext 16; Fax: +52(999) 423205;
E-mail: opacheco@uady.mx

Summary

American Trypanosomiasis (AT) is an infectious parasitic disease produced by the protozoa Trypanosoma cruzi (T. cruzi). Infection is acquired by vectorial via but can also be transmitted congenitally, by ingestion of an infected host, by transfusion with contaminated blood or transplant of organs from an infected donor. Currently, AT is widely distributed from the South of the United States to South America. In Mexico, the presence of the parasite has been reported throughout the country where several reservoirs such as dogs, opossums, rats and cats have been identified. Yucatan is in the south-east of Mexico where AT is endemic and has been reported since 1940s. There is little information about the role of pigs as reservoirs of T. cruzi. The frequency of specific antibodies against T. cruzi was determined in fattening pigs from Yucatan, Mexico. After sampling in the 3 main areas of pig production in the state, IgG ELISA and Western blot were performed to identify seropositive cases. Association of farm size, farm area and production system with infected pigs was evaluated. From 273 sampled pigs, 5.4% (n = 15) positive cases were found. No association with evaluated factors and infected pigs was found. Pigs are also reservoirs of T. cruzi in the studied area. These findings are considered important to improve vectorial control in the area in order to avoid the parasite infection in animal populations destined for human consumption and avoid further transmission to humans.

Impacts

  •  Pigs are important reservoirs of Trypanosoma cruzi in commercial pigs destined to human consumption.
  •  Infected pigs may be a source of human infection when uncooked meat is consumed or manipulated during acute infections.
  •  Vector control should be implemented around all commercial pig farms in the State to prevent exposition.

Introduction

American trypanosomiasis or Chagas disease is a parasitic disease caused by the flagellate protozoan Trypanosoma cruzi and is found from the southern United States to Argentina. This parasite can infect humans and more than 150 wild and domestic mammals. The transmission is through a haematophagous insect vector well established in all Latin-American countries, but can also be transmitted less frequently by the transplacental route, orally, and by blood transfusions or organ transplants. Until now, there is no effective drug treatment or immune to prevent or control infection, which gives it special importance and seeks to prevent transmission in humans. Domestic and wild animals are important reservoir to maintain the circulation of the protozoa in the environment (i.e. opossums, raccoons, dogs, cats, etc.). Domestic and wild pigs (Sus scrofa) as other mammals are susceptible to be infected with T. cruzi and become reservoirs of the protozoa, but no information about its role has been reported.

The aim of this study was to determine the prevalence of IgG-specific antibodies against T. cruzi in commercial fattening pig farms in an endemic area of Mexico.

Materials and Methods

Study area

The study was conducted in the state of Yucatan, located in south-eastern Mexico (19°30′ and 21°35′ north latitude and 90°24′ west longitude). The climate is tropical sub-humid with heavy summer rains. The maximum monthly temperature varies between 35 and 40°C, with an average temperature of 26.6°C. The relative humidity varies between 65% and 100% taking the mean value over 78%. The annual rainfall is from 415 to 1290 mm (INEGI, 2002).

Size of the sample

A two-stage cross-sectional study was performed during September to December 2009. Two-hundred and seventy-three pigs from 39 farms, between 18 and 20 weeks of age, were randomly selected. A proportional number of farms from the three main pig production regions of the State (West, Centre and South) were included. The sample size to estimate prevalence was determined by the formula: inline image. Considering a confidence level of 95% (z = 1.96), absolute precision of 5% (d), a design effect (D) of 2 and an estimated prevalence of 10% (p). The estimated prevalence was obtained from a pilot study that included 45 animals. The number of animals sampled at each farm (b = 7) was calculated based on the formula: b = √ (ce/cd) (1 − re/re), where ce cost of a sample into two clusters (10), cd sample cost of two units of interest in a same cluster (1), and the correlation intraconglomerates (re) was (0.04) (Segura-Correa and Honhold, 2000). The number of farms (m = n/b) was calculated by dividing the sample size (n = 273) between the number of animals sampled in each establishment (b = 7).

Sample collection

Individual blood samples were collected using vacutainer® tubes without EDTA to obtain serum. Blood samples were centrifuged at 2500 g for 10 min and were stored at −20°C until further evaluation.

Indirect enzyme-linked immunosorbent assay and western blot assays

For the specific detection of IgG antibodies against T. cruzi, a commercial indirect enzyme-linked immunosorbent assay was used (WienerLab ChagaTest). The technique used was adapted to that described by Jiménez-Coello et al. (2010) using anti-IgG pig antibody labelled with horseradish peroxidase (HRP; Santa Cruz Inc., Santa Cruz, CA, USA) on 96-well plate coated with recombinant proteins of T. cruzi. Serum samples were diluted to a ratio of 1 : 100 in phosphate-buffered saline (PBS; pH 7.2), and the secondary goat anti-IgG pig antibody HRP labelled was used at a dilution of 1 : 5000.

Sera from previously evaluated dogs and BALB/c artificially infected mice showing high anti-IgG antibodies titres by ELISA (1 : 1024) and PCR positive against T. cruzi were used as positive controls, and sera pool from 10 healthy pigs previously tested by triplicate with ELISA IgM, IgG and PCR were used as negative controls. On the basis of the ELISA, subjects were diagnosed as either positive or negative for specific IgG antibodies to T. cruzi. The optical density (OD) was measured in a spectrophotometer at 450 nm (Multiskan Multisoft Primary EIA®) and was used to compute the per cent positivity (PP) using the following formula: inline imagePer cent positivity of 15% or above was considered as positive.

For confirmation of the diagnosis ELISA seropositive cases, the western blot (WB) technique was performed (Ramos-Ligonio et al., 2006; Jiménez-Coello et al., 2008), in which Tulahuen strain epimastigotes of T. cruzi were used as antigen. Samples were considered positives to WB based on an established criterion. A serum sample was considered positive when it recognized at least five antigenic bands from a group of 10 with the highest frequency of recognition; the result was considered indeterminate when the sample recognized of 1–4 antigenic bands and was negative when the serum sample showed no reactivity (Teixeira et al., 1994; Jiménez-Coello et al., 2008).

Statistical analysis

Univariate analysis (X2) was used to determine the association of possible risk factors with the presence of specific antibodies to Chagas’ disease. Risk factors considered were farm size (≤400 or >400), production system (full cycle or fattening farms) and farm region (West, Central or Sotuh). Odds ratio (OR) and 95% confidence intervals were also calculated using the EpiInfo program (Dean et al., 1994). The level of significance was set at < 0.05.

Results

From 273 sampled pigs, there were found a 5.4% (n = 15) positive cases. All positive cases were from different farms. There was no association between seropositivity and any of the evaluated risk factors (> 0.05; Table 1).

Table 1.   Risk factors evaluated to determine association with seropositivity against Trypanosoma cruzi in fattening pigs from Yucatan, Mexico
Risk FactorCategoryPositivesPrevalence (%)ORIC (95%) P-value
Farm Size≤40055.050.90.32–2.670.89
>400105.43
ProductionFull cycle94.30.580.21–1.640.32
SystemFattening67.2
Farm regionWest55.41 0.67
Center44.20.780.21–2.8
South66.91.20.40–4.00

Discussion

American trypanosomiasis is a widespread infection in the Americas in a large number of domestic mammals that serve as reservoirs that maintain the infection because they are in direct contact with vectors and human settlements. There are no formal epidemiological studies that demonstrate the role of the domestic pig as a reservoir of T. cruzi. However, natural and experimental infections have been described (Salazar-Schettino et al., 1997; Solaymani-Mohammadi and Petri, 2006). These reports point to the pig as a potential reservoir host for infections of T. cruzi in humans, which have an important role in the maintenance of domestic and peridomestic cycle of infection.

The prevalence of infection with T. cruzi found in this study is considered low. However, this is the first formal epidemiological report showing the presence of this protozoan distribution in a major pig production area from Mexico. Previously, the presence of natural infection in pigs in Mexico has been described (Salazar-Schettino et al., 1997). The presence of pigs naturally infected with T. cruzi has been reported in other Latin American countries like Brazil (Costa et al., 1998) and Paraguay (Fujita et al., 1994).

The infection of pigs with T. cruzi in the study area is probably due to exposure to the vector Triatoma dimidiata, which has been described as the main vector transmitting T. cruzi in south-eastern Mexico where the disease is considered endemic (Guzman-Tapia et al., 2005), unlike the vector transmitter in domestic pigs in Brazil (Panstrongylus geniculatus and Triatoma brazilienzis; Valente et al., 1998).

Apparently, this protozoal infection in pigs does not have considerable economic importance because it does not affect the production parameters (i.e. daily weight gain). The pathogenesis of the disease in pigs has not been demonstrated, but as in other mammals, clinical signs during the acute phase may result in fatal consequences. On the other hand, because of the short lifetime of fattening pigs, the indeterminate or chronic phase of the disease is never reached. However, the parasite strains circulating in the region show heart and muscle tissue tropism of infected hosts (amastigote intracellular stage) (Barrera et al., 2001). Although the presence of infection in pigs is low, the consumption of undercooked pork meat should be avoided, and precautions in handling raw meat are recommended because during the acute stage of infection, the parasite is circulating in the bloodstream (trypomastigote stage in the blood).

It is concluded that the domestic pig in the area of study is an important reservoir of T. cruzi because it is in an endemic area where the vector is widely distributed and recommended vector control measures in pork production farms to reduce the risk of transmission for this zoonotic disease.

Acknowledgements

We gratefully acknowledge to D. Saez-Peniche and M.E. Rosado-Barrera for their technical assistance.

Conflicts of interest

The authors have not declared any potential conflicts.

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