Toxocara canis: larval migration dynamics, detection of antibody reactivity to larval excretory-secretory antigens and clinical findings during experimental infection of gerbils (Meriones unguiculatus).

In this study, Mongolian gerbils were used to analyse features of Toxocara infection that included larval migration, humoral immune responses to Toxocara canis excretory-secretory antigens (TES) and aspects of host physiology. At day 10 post-infection (p.i.) most larvae were in the intestine and the lungs while later the total number of larvae was higher in the carcass tissue; the number of larvae per gram of tissue was lower elsewhere other than in the brain. Infected animals showed several neurological abnormalities, an early increase in leukocyte and neutrophil levels, two peaks of peripheral eosinophilia (5 and 40 d.p.i.) and high antibody levels against TES in the circulation and in the vitreous humor. A sequential recognition of eight T.canis larval antigens with MW from 24 to 200 kDa was detected by Western blot. The results obtained in this study further support the use of gerbils as an experimental model for systemic, ocular and cerebral toxocariasis.

Sequential exposure and assembly of cyst wall filaments on the surface of encysting Giardia duodenalis.

The mode of appearance and assembly of cyst wall filaments on the surface of Giardia duodenalis trophozoites committed to encyst was analysed by scanning and transmission electron microscopy (SEM and TEM) and by fluorescence microscopy (FM). SEM showed a progressive appearance of fibril patches, predominantly on the anterior area of ventral and dorsal surfaces, which then spread and coalesced. By TEM, ruthenium red (RR) displayed staining in encysting cells as rodlike spots of variable diameter (3-25 nm), possibly microfibril tips with polyanionic moieties, that displayed tangential associations and random orientations over the cell membrane. In FM assays, the 1,10-phenanthroline derivative of ruthenium red (RR/oPHE) was a specific ligand for these assembling fibrils and this staining was significantly blocked by N-acetylgalactosamine (GalNac) and galactosamine (GalN). Interestingly, RR staining was lost when the cyst wall was completely assembled and thickened as observed by TEM and FM. Kinetic FM assays, in which a mAb specific for a 26 kDa Giardia cyst wall polypeptide was used concomitantly with RR/oPHE staining, showed a differential pattern for the appearance and reactivity of polypeptide and assembling GalN/GalNac-rich moieties of Giardia cyst wall.

Epidemiology of trichinellosis in Mexico, Central and South America.

Trichinella species are widely distributed throughout the world and are found in a large number of carnivorous animals, humans and incidental hosts. The data presented in this review show that Trichinella infection has been reported in both humans and animals in Mexico, Argentina and Chile since the end of the 19th century, and more recently in Bolivia. This parasitic infection is still a public health problem in countries such as Argentina and Chile. Although efforts have focused on the control and prevention of trichinellosis in these countries, there were still human cases and outbreaks of trichinellosis reported. Diagnosis of infection in animals such as pigs still includes, in many endemic areas, the use of trichinoscopy. In Argentina, however, artificial digestion has been recently introduced in slaughterhouses to detect Trichinella infection in pigs. In some endemic areas in Mexico, the use of serological assays for human trichinellosis and pig infections have resulted in improved detection. Most of the outbreaks of human trichinellosis in Mexico, Argentina and Chile have occurred as a result of the consumption of undercooked pork or pork products from animals raised under poor hygienic conditions and which are clandestinely slaughtered. In several studies, rats, dogs and cats have been found to be infected with Trichinella and may be considered a risk for transmission of the infection to pigs or other animals intended for human consumption. Another potential source of transmission of Trichinella to humans is horsemeat; however, information related to horse trichinellosis in Latin-American countries is scarce. In most cases the etiological agent of human trichinellosis in Central and South America has been reported to be Trichinella spiralis; however, only in a few cases has the parasite species been properly identified. Based on the reports available, it is clear that there is a need to carry out better controlled epidemiological studies to determine the true prevalence of the infection in this region of the world. Also, more sensitive methods of diagnosis and improvements in conditions for pig production as well as better sanitary inspection systems, are needed for controlling and preventing transmission of trichinellosis in these countries.

Histopathological investigation of experimental ocular toxocariasis in gerbils.

Male gerbils were inoculated intragastrically with embryonated Toxocara canis eggs. They were euthanised, and their eyes were excised at different days p.i. to identify the number of larvae as well as lesions resulting in these organs. In most animals, larvae were detected from day 5 to day 60 p.i. (end of the study). From days 10 to 20 p.i., larvae and haemorrhage were observed in the choroid and in the ciliary process. At days 30 and 40 p.i., some eyes had larvae surrounded by an infiltrate of neutrophils, oedema, haemorrhages and tissue damage in the retina or the ciliary process. On day 60 p.i., there were granulomatous lesions in the retina. This study provides a model for the study of ocular toxocariasis.

Detection of Trichinella infection in slaughter horses by ELISA and western blot analysis.

In order to determine the presence of Trichinella infections in horses slaughtered at an abattoir in Mexico, 147 serum samples were examined by two immunoenzymatic methods. Specific antibodies were detected by ELISA in 7% of the serum samples at a dilution 1:400 and in 10% at lower dilutions (1:20, 1:40) using Trichinella spiralis muscle larvae (ML) excretory/secretory (E/S) products. Serum samples from four naturally infected horses (confirmed by direct methods) gave negative O.D. values in an ELISA at a 1:400 dilution and only two of them were positive at a 1:20 and 1:40 dilutions. Serum samples from experimentally infected horses reacted by Western blotting with ML components with molecular weights of 47, 52, 59, 67, 72 and 105 kDa which correspond to the TSL-1 antigens. Serum samples from the four naturally infected horses and from the abattoir horses that were positive in ELISA using E/S antigens recognized several ML components, some of them reacted with all the TSL-1 antigens mentioned above and others recognized preferentially two or three of these molecules. Since the serologic assays may not offer the sensitivity required in the diagnosis of horses trichinellosis and the direct methods had not always been useful in the detection of larvae in horsemeat related to trichinellosis outbreaks in Europe, it is proposed that additional assays are performed to determine Trichinella infection in horses. These can include detection of parasite antigens by ELISA and Dot ELISA or PCR, which in turn may also help to determine the presence of the parasite in early and late infections of horses.

[Updates on equine trichinellosis]. / Actualidades sobre la triquinelosis equina.

Trichinellosis is a zoonosis caused by parasites of the genus Trichinella. Transmission of trichinellosis to humans has been shown to occur mainly by the ingestion of meat from pigs, bears of foxes parasitized with muscle larvae of this parasite. However, in Europe, the major human outbreaks of the disease have occurred due to the ingestion of parasitized horse meat. Although the larvae were not isolated from the horse meat, the identification of larvae as T. nativa, T. britovi and T. spiralis was done in biopsy samples obtained from infected individuals. More recently T. spiralis muscle larvae have been isolated and identified, for the first time, in muscle tissue of horses slaughtered at an abattoir in the State of Mexico. Furthermore, in ELISA assays using total extracts or TSL-1 antigens, circulating antibodies against Trichinella have been detected in horses slaughtered at abattoirs from various countries in Europe and Mexico. On the other hand, the experimental infection of horses with parasites of the genes Trichinella has been achieved by several research groups and data obtained regarding the kinetics of antibody production in these animals are important in the development of sensitive and specific diagnostic assays for horse trichinellosis. This will allow to determine the frequency of this infection in horses which are used for animal and human feeding. These assays will also be very helpful for designing strategies to control transmission on the disease by horse meat.

Detection of Trichinella spiralis muscle larvae in naturally infected horses.

Human trichinellosis outbreaks related to horsemeat consumption have been reported in France and Italy in recent years. In order to determine if Trichinella is present in horses slaughtered at an abattoir in the State of Mexico, diaphragm muscle tissue samples (22-37 g) from 80 horses were examined by artificial digestion. Four of these samples had larvae that were characterized as Trichinella sp. by morphological criteria and as Trichinella spiralis by the polymerase chain reaction.

[Characterization of surface antigens of the nematode parasite Trichinella spiralis: study of its role in protection mechanisms and their usefulness in the diagnosis of trichinosis]. / La caracterización de los antígenos de superficie del nemátodo parásito Trichinella spiralis: estudio de su participación en los mecanismos de protección y su utilidad en el diagnóstico de la triquinosis.

Among the most important aspects in the study of trichinosis are the development of specific and sensitive diagnostic methods for the detection of the infection by the parasite as well as the characterization of antigens from Trichinella spiralis that induce protection in the host. In the context, the characterization of surface stichosome and excretory secretory antigens of T. spiralis muscle larvae has been an important issue due to the high immunogenicity of such components in most hosts so far studied. In this work, we have been able to identify and characterize molecules from both compartments of muscle larvae. These components have been used in assays for specific detection of T. spiralis infections particularly in pigs, as well as in assays to evaluate their role in the induction of protection in mice.

[Current aspects of trichinosis diagnosis]. / Aspectos actuales sobre el diagnóstico de la triquinelosis.

An important approach to the study of trichinellosis is the development of sensitive diagnostic methods that allow the detection of Trichinella spiralis. Recently, ELISA assays that use surface/stichosome and/or secretion/excretion antigens from muscle larvae have been proved to be highly sensitive and specific in the diagnosis of the infection. Furthermore, the high immunodominance of carbohydrate residues on these molecules in a broad host range make them useful diagnostic markers for trichinellosis.

Study of human Entamoeba histolytica infection by zymodeme, indirect hemagglutination and electroimmunotransfer blot assays.

Human amebiasis is a parasitic infection which involves asymptomatic as well as intestinal and extraintestinal symptomatic stages in individuals harbouring Entamoeba histolytica. Several factors have been proposed to explain the variability in the outcome of the disease. Among these are differences in virulence of E. histolytica strains and methods such as zymodeme analysis have been used to differentiate invasive from non invasive strains of this parasite (Sargeaunt and Williams, 1978). In order to establish a possible correlation between zymodeme analysis and serological data in human cases of amebiasis, a cross-sectional epidemiological study was carried out in the community area of Cadereyta, State of Queretaro, Mexico. In this study, fecal and serum samples from individuals were also tested by Indirect Hemagglutination assay (IHA) to determine antibody titre and by a standardized Electroimmunotransfer blot assay (EITB) for recognition of E. histolytica specific antigens. Zymodemes were determined in a total of 81 samples. Of these, 27 had a pathogen zymodeme (PZ) while 54 had a non pathogen zymodeme (NPZ). Values obtained by IHA varied from negative to titres up to 1:256 in serum samples tested. Reactivity to E. histolytica antigens determined by EITB was observed in all sera. Patterns of antigen recognition were complex and showed reactivity to several parasite molecules. Among these, components with M. Wt. of 165, 119, and doublets of 98-100 and 50-52 Kd were more frequently recognized by antibodies present in the sera tested. In general, antibody titres detected by IHA did not showed a direct correlation with zymodeme analysis. Samples with PZ or NPZ had similar variable levels of reactivity to E. histolytica antigens.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of an immunoblot methodology for the detection of relevant Entamoeba histolytica antigens by antibodies induced in human amebiasis.

The complexity of the clinical spectrum in human amebiasis and the high variability in laboratory methods used to detect Entamoeba histolytica infections have impeded the collection and evaluation of reliable epidemiological data. Thus, more sensitive, specific and standardized methods are needed in order to accurately identify infections with this parasite. An important step in the development of serological diagnostic methods is the identification and isolation of specific parasite antigens which are immunogenic in the host. In this work, we have standardized an electroimmunotransfer blot technique to characterize E. histolytica antigens recognized by antibodies present during human amebic infections. An important aspect was an investigation of technical variations in the preparation of cell lysates including the use of different protease inhibitors and solubilizing agents. The highest yield of protein was achieved by homogenization of trophozoites in the presence of 10 mM p-hydroxymercuribenzoate (pHMB) as a protease inhibitor and by lysis using Triton X-100 and a mixture of protease inhibitors. Recovery of degraded vs non-degraded proteins in the cell extracts was evaluated by gradient polyacrylamide gel electrophoresis. Both quantitative and qualitative differences were noted between the different methods of preparing soluble cell extracts. A more complete set of antigenic components was obtained by homogenization and use of pHMB. Thus parasite extracts prepared by this method were selected for protein transfer. In this, the optimal protein concentration was of 120 micrograms of protein per cm of gel width and efficient transfer of proteins to nitrocellulose sheets was achieved at 100 V for 2 hrs and at 4 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)