Action of colchicine on hepatic schistosomal granuloma

Amorphous material and altered collagen fragments within dilated secretory vesicles and cisternae of fibroblast cytoplasm were the main ultrastructural changes seen in hepatic periovular granulomas formed in mice infected with Schistosoma mansoni and treated with colchicine. Despite promoting ultrastructural changes in the fibroblasts found in hepatic periovular granulomas, colchicine administration to infected mice did not significantly change the light microscopic appearance of the hepatic schistosomal lesions, did not diminish the amount of total hepatic collagen, and did not change the collagen isotypes in the granulomas, as observed after a comparative study with non-colchicine treated infected control mice. When administered to mice two weeks after curative treatment of schistosomiasis with praziquantel, colchicine did not seem to increase extracellular collagen degradation or to induce a more rapid resorption of hepatic periovular granulomas, although still promoting ultrastructura alterations in fibroblasts.

Histogical and ultrastructural aspects of the brindley's glands of pantrongylus megistus (Burmeister, 1835) (Hemiptera: Reduviidae)

The Brindley's glands of Panstrongylus megistus were studied under the antomic, histologic and ultrastructural point of view. These glands located in the insect's methatorax are paired and have an opening near the third parir of the feet. Beside this aperture, ther are evaporation areas. Shape, sixe and aspect of the gland vary according to the feeding status. The glands are composed by a tubular part corresponding to the duct and a sack-like portion corrsponding to the secretory part. By electron microscopy we observed that the basal part of the epithelium has many interdigitations associated with mitochondria. On the apical surface where epicuticular foldings are located an electonlucent space is often seen. The glands are composed of the following elements: 1) superficial epithelial cells, located just below the apical surface foldings; 2) secretory cells; which are long and have an intracellular canalicule which changes according to the functional state of the cell; 3) a collecting duct to the secretory cells and covered with an epicuticle, reaching up to the gland's lumen; and 4) cells around the duct.

The reproductive capacity of Panstrongylus megistus (Hemiptera: Reduviidae) males treated with precocenes

Reproductive performance including mating, semen transfer, egg production and hatching rate were investigated in pairs of Panstrongylus megistus in wich the male was treated with either precocene II or ethoxyprecocene II, topically applied shortly after imaginal molt. Eighty percent of the males were not able to mate for a period as long as 40 days after treatment. Those males wich did mate (20%) produced small spermatophores; a few (10%) were unable to transfer spermatozoa to female, while the others (10%) exhibited reproductive performance similar to that of non-treated males. The cumulative effects of the precocene on the different stages of reproduction resulted in a drastic reduction of offspring production. The anti-allatal compound precocene seems to inhibit successful reproduction through a corpus allatum inactivation and subsequent lack of secretory activity of the male accessory glands.

Eokines: synthesis, storage and release from human eosinophils

Eosinophils are prominent inflammatory cells in asthma and other allergic disorders, as well as in helminthic parasite infections. Recently, eosinophils have been reported to synthesize and store a range of regulatory proteins within their secretory granules (eokines). Eokines comprise a group of cytokines, chemokines, and growth factors which are elaborated by eosinophils. These proteins, and the messages which encode them, appear to be identical to those produced by lymphocytes and other tissues. Interestingly, immunoreactivity to many of these eokines has been found to co-localize to the eosinophil´s secretory granules. In this review, we have discussed the repertoire of 18 eokines so far identified in eosinophils, and focused on four of these, namely, interleukin-2 (IL-2), IL-4, granulocyte/macrophage colony-stimulating factor (GM-CSF), and RANTES. These four eokines co-localize to the crystalloid granules in eosinophils, as shown in studies using subcellular fractionation and immunogold labeling in electron microscopy. During stimulation by physiological triggers, for example, with serum-coated particles, eosinophils release these mediators into the surrounding supernatant. In addition, eokines are likely to be synthesized within eosinophils rather than taken up by endocytosis, as show in detection of mRNA for each of these proteins using in situ hybridization, RT-PCR, and in the case of RANTES, in situ RT-PCR. Eokines synthesis and release from eosinophils challenges the commonly held notion that these cells act downstream of key elements in immune system, and indicate that they may instead belong to the afferent arm of immunity.

Human toxocarosis. Its seroprevalence in the City of La Plata

Toxocara canis is very common in dogs throughout the world. It is the primary cause of visceral larva migrans (VLM) in humans. Soil contaminated with T. canis embryonated eggs is the main source of infection of man. Our objective was to describe Toxocara seroprevalence in humans in the city of La Plata associated with some determinants such asage, presence or absence of clinical manifestations and risk factors. Blood samples were collected at random from 156 patients of different sex and age, with and without clinical symptoms compatible with the disease. The diagnostic technique ELISA test was performed with the Bordier Affinity Products Commercial Kit, using excretory-secretory Toxocara antigen with a sensitivity higher than 90%. The values were positive in 39% of the studied population. In the analysis according to age, the younger group presented significant differences with respect to the older one (Chi-square p<0.05). Positive patients presented clinical symptoms compatible with the disease (84%), and 41% presented some risk factor. The level of positivity obtained indicates a certain risk of being infectes mainly in patients younger than 15 years old. The authors agree that an early identification and treatment of VLM may save a life.

Isotype specific immune responses in murine experimental toxocariasis

In this work, a murine experimental model of toxocariasis has been developed in BALB/c, C57BL/10 and C3H murine strains orally inoculated with 4,000 Toxocara canis embryonated eggs, in order to investigate the isotype-specific immune responses against excretory-secretory antigens from larvae. T. canis specific IgG+M, IgM, IgG, IgA, IgG1, IgG2a and IgG3 were tested by ELISA. The dynamics of the specific immunoglobulins (IgG+IgM) production showed a contrasting profile regarding the murine strain. Conversely to the results obtained with the IgM isotype, the IgG antibody class showed similar patterns to those obtained with IgG+IgM antibodies, only in the case of the BALB/c strain, being different and much higher than the obtained with IgG+IgM antibodies, when the C3H murine strain was used. The antibodies IgG+IgM tested in BALB/c and C57BL/10 were both of the IgM and IgG isotypes. Conversely, in the C3H strain only IgG specific antibody levels were detected. The IgG1 subclass responses showed a similar profile in the three murine strains studied, with high values in BALB/c, as in the case of the IgG responses.

Preliminary analysis of Sm14 in distinct fractions of Schistosoma mansoni adult worm extract

In previous studies it was shown that the recombinant molecule, r-Sm14, induces high levels of protection against Schistosoma mansoni infection in two outbred animal models and immune crossprotection against infection by Fasciola hepatica in Swiss outbred mice. r-Sm14 was derived from a living worm extract, called SE, and is being developed as the molecular basis of an anti-helminth bivalent vaccine against the two parasites, for medical and veterinary application. Present data refer to SDS-PAGE and Western Blotting analysis of four different preparations of S. mansoni adult worms focusing Sm14 identification. The extracts correspond to the initial fraction of the SE extraction process, containing products released by living worms (SEi); SE2, reextraction of adult worms in PBS; and SE of separated male and female adult worms. In all extracts it was possible to detect the component of 14 kDa, that was recognized by specific anti-rSm14 antibody raised in rabbits.

Laboratory diagnosis of Schistosomiasis in areas of low transmission: a review of a line of research

After 57 years of successful control of schistosomiasis in Venezuela, the prevalence and intensity of infection have declined. Approximately 80% of the individuals eliminate less than 100 eggs/g of stools, therefore morbidity is mild and the majority are asymptomatic. The sensitivity of Kato-Katz decreases to approximately 60%. Available serological methods for the detection of circulating antigens only reach a 70% of sensitivity. Tests based on the detection of antibodies by immunoenzymatic assays have been improved. The circumoval precipitine test has shown a high sensitivity (97%), specificity (100%), and correlation with oviposition, being considered the best confirmatory diagnostic test. Additionally to the classical immunoenzymatic assays, the development of the alkaline phosphatase immunoassay, allowed to reach a 100% specificity with an 89% sensitivity. Recently, we have developed a modified ELISA in which the soluble egg antigen is treated with sodium metaperiodate (SMP-ELISA) in order to eliminate the glycosilated epitopes responsible for the false positive reactions. The specificity and sensitivity reaches 97% and 99%, respectively. Synthetic peptides from the excretory-secretory enzymes, cathepsin B (Sm31) legumain (Sm32) and cathepsin D (Sm45), have been synthesized. The combination of two peptides derived from the Sm31 have been evaluated, reaching a sensitivity of 96% when analyzed independently and with a 100% specificity. Antibodies raised in rabbits against peptides derived from the Sm31 and Sm32 are currently evaluated in two different antigen-capture-based assays. The development of a simple, cheap and reliable test that correlates with parasite activity is a major goal.

Prevalence of Helicobacter pylori infection in Warao lineage communities of Delta Amacuro State, Venezuela

The purpose of this study was the evaluation of Helicobacter pylori infections in children and adults from two indigenous communities of Delta Amacuro State, Venezuela, that differ in hygienic conditions of the housing. The evaluation was performed in 98 children (mean age 7 ± 3.37 years) and their mothers (33.96 ± 13.77 years) from two communities of Warao lineage. Anti-H. pylori serum IgG and secretory anti-H. pylori IgA antibodies were de-termined, as well as total secretory IgA and H. pylori antigens in feces. Serological prevalence of H. pylori infection was 38% in children and 84% their in mothers. Children from the community that had the most deficient sanitary and hygienic conditions had significantly lower titers of specific IgG antibodies and total secretory IgA (P < 0.0001) and a high percentage of them had H. pylori antigens in their feces (P < 0.0001). The levels of specific IgA were similar in both groups. The results indicate that in these populations there is a high prevalence of H. pylori infection and that poor hygienic conditions can increase the risk of infection and damage to the gastrointestinal tract.

Study of the antibody response against Mycobacterium tuberculosis antigens in Warao Amerindian children in Venezuela

This study was aimed at investigating alternate methods for serodiagnosis of tuberculosis (TB), which are needed because bacteriologic diagnosis of childhood TB is difficult. A selection of 80 serum and saliva samples were tested from Warao indigenous children under 15 years of age; 34 high TB suspects (28 positive and 6 negative for the tuberculin skin test, TST) and 46 healthy contact children (32 positive and 14 negative for the TST). Several enzyme-linked immunosorbent assay (ELISA) serological tests were developed to test for Mycobacterium tuberculosis-specific antibodies, including serum IgA, IgG, IgE, and secretory IgA (sIgA) in saliva against 3 specific antigens (PPD, HSP60, 38 kDa). Of these, 2 antigens, PPD and 38 kDa, showed significantly higher reactivity. The sensitivity and specificity of these tests for diagnosis remained limited, between 26.5% and 38.2%, and 77.4% and 97%, respectively. Of all the samples studied and combinations realized between all isotypes and antigens combined with 3 isotypes (anti-PPD IgG, IgE, and anti-38kDa sIgA) managed to detect the largest number of patients, showing an improved sensitivity level of 64.7%, although specificity levels dropped to 81.8%. These results were compared with the Omega diagnostics commercial kit results. The commercial kits showed significantly lower reactivity (sensitivity of 20% and 13.33% to Myco G and Complex Plus, respectively) and a specificity of 100%. This study shows that in indigenous populations of Venezuela, where invasive procedures cannot be used to select samples but evaluation with a chest X-ray for radiological studies is available, the combination of 3 specific isotypes may be a useful tool to increase diagnostic accuracy with pulmonary TB in this population, when used together with clinical and epidemiological criteria.

Human toxocariasis: a seroepidemiological survey in schoolchildren of Sorocaba, Brazil

A seroepidemiological survey for toxocariasis, among 180 schoolchildren of the public schools of Sorocaba City, state of São Paulo, Brazil, was carried out from August 2000 to July 2001. ELISA test was performed using excretory and secretory antigens for the detection of IgG anti-Toxocara antibodies. Information regarding the children was obtained from the parents or legal guardians. The results showed that the mean age was 5.4 ± 1.4 years, the infection coefficient (IC) was 38.3 and the infection risk was higher among the children living in the city outskirts (IC = 47.4) where the socioeconomic conditions were worse than in the central region of the city (IC = 11.1). There was an association between higher frequency of seroreactivity in the ELISA test and the condition of living in a house with a yard and/or unpaved street. The same was observed in relation to a history of enteroparasitism. There was also an association between a seronegative ELISA test and previous treatment of pet dogs and/or cats with vermifuge. Based on these results, the authors propose that public health programs should include anthelmintic for dogs and cats during the antirabies vaccination campaigns, in order to diminish environmental contamination with Toxocara spp. eggs and consequently human infection.

Shared and non-shared antigens from three different extracts of the metacestode of Echinococcus granulosus

Hydatid cyst fluid (HCF), somatic antigens (S-Ag) and excretory-secretory products (ES-Ag) of Echinococcus granulosus protoscoleces are used as the main antigenic sources for immunodiagnosis of human and dog echinococcosis. In order to determine their non-shared as well as their shared antigenic components, these extracts were studied by ELISA-inhibition and immunoblot-inhibition. Assays were carried out using homologous rabbit polyclonal antisera, human sera from individuals with surgically confirmed hydatidosis, and sera from dogs naturally infected with E. granulosus. High levels of cross-reactivity were observed for all antigenic extracts, but especially for ES-Ag and S-Ag. Canine antibodies evidenced lesser avidity for their specific antigens than antibodies from human origin. The major antigenic components shared by HCF, S-Ag, and ES-Ag have apparent molecular masses of 4-6, 20-24, 52, 80, and 100-104 kDa, including doublets of 41/45, 54/57, and 65/68 kDa. Non-shared polypeptides of each antigenic extract of E. granulosus were identified, having apparent masses of 108 and 78 kDa for HCF, of 124, 94, 83, and 75 kDa for S-Ag, and of 89, 66, 42, 39, 37, and 35 kDa for ES-Ag.

Nitric oxide and the resolution of inflammation: implications for atherosclerosis

The ubiquitous free radical, nitric oxide (NO), plays an important role in many biological processes including the regulation of the inflammatory response. Alterations in NO synthesis by endogenous systems likely influence inflammatory processes occurring in a wide range of diseases including many in the cardiovascular system (e.g. atherosclerosis). Progression of inflammatory conditions depends not only upon the recruitment and activation of inflammatory cells but also upon their subsequent removal from the inflammatory milieu. Apoptosis, or programmed cell death, is a fundamental process regulating inflammatory cell survival and is critically involved in ensuring the successful resolution of an inflammatory response. Apoptosis results in shutdown of secretory pathways and renders effete, but potentially highly histotoxic, cells instantly recognisable for non-inflammatory clearance by phagocytes (e.g., macrophages). However, dysregulation of apoptosis and phagocytic clearance mechanisms can have drastic consequences for development and resolution of inflammatory processes. In this review we highlight the complexities of NO-mediated regulation of inflammatory cell apoptosis and clearance by phagocytes and discuss the molecular mechanisms controlling these NO mediated effects. We believe that manipulation of pathways involving NO may have previously unrecognised therapeutic potential for limiting or resolving inflammatory and cardiovascular disease.

Evaluation of the dot enzyme-linked immunosorbent assay in comparison with standard ELISA for the immunodiagnosis of human toxocariasis

A dot enzyme-linked immunosorbent assay (dot-ELISA) was standardized using excretory-secretory antigens of Toxocara canis for the rapid immunodiagnosis of human toxocariasis. Thirty patients with clinical signs of toxocariasis, 20 cases with other parasitic diseases, and 40 healthy subjects were tested. A total of 0.2 ng of antigen per dot, serum dilution of 1:160 and dilution conjugate of 1:1000 were found optimal. The sensitivity and specificity of the assay were 100 and 95%, respectively. Comparable sensitivity of dot-ELISA and the standard ELISA was obtained, but only 3 cross-reactions occurred in the dot-ELISA, compared with 6 in the standard ELISA. Dot-ELISA is simple to perform, rapid, and low cost. Large-scale screening studies should be done to evaluate its usefulness under field conditions.

Preliminary results in the immunodiagnosis of tuberculosis in children based on T cell responses to ESAT-6 and PPD antigens

The aim of this work was to study the difference in interferon gamma (IFN-gamma) production by T lymphocytes after early secretory antigen target 6 (ESAT-6) or purified protein derivate (PPD) stimulation in whole blood culture supernatants from children with suspected tuberculosis (TB) disease (n = 21), latent TB infection (n = 16) and negative controls (NC) (n = 22) from an endemic area in Brazil. The concentration of IFN-gamma (pg/ml) was measured by enzyme linked immunosorbent assay and the differences in the IFN-gamma levels for each group were compared and evaluated using an unpaired Student's t-test; p values < 0.05 were considered significant. Measurement of IFN-gamma levels after ESAT-6 stimulation raised the possibility of early diagnosis in the latent TB group (p = 0.0030). Nevertheless, the same group showed similar responses to the NC group (p > 0.05) after PPD stimulation. The IFN-gamma assay using ESAT-6 as an antigenic stimulus has the potential to be used as a tool for the immunodiagnosis of early TB in children.