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.

Specific antibody levels and antigenic recognition of Wistar rats inoculated with distinct isolates of Trypanosoma evansi

"Mal de Cadeiras", an enzootic disease caused by Trypanosoma evansi, is one of the most important trypanosomiases in the Brazilian Pantanal region. The disease affects mainly horses, which are widely used in extensive cattle production, an activity of greatest economical significance for the region. The parasite also infects sylvan (coatis and capybaras) and domestic (dogs) animals, respectively considered wild and domestic reservoirs of T. evansi. For a better understanding of the interaction of T. evansi with its rodent host, we evaluated the differences in the specific antibody level patterns and in the parasitic peptides recognition patterns of experimentally infected Wistar rats. The rats experimentally infected with T. evansi isolates obtained from coatis, dogs and horses were submitted to indirect immunofluorescence test (IgM e IgG) and Western blotting. The serological titers for IgM and IgG ranged between 1:40 and 1:160. The most recognized polypeptide profiles were in a range of 17 and 74 kDa. Our data suggest that the humoral immune response in Wistar rats is not sufficient for granting an effective control of T. evansi infections.

Specific situations related to acute schistosomiasis in Pernambuco, Brazil

The present work reports on two epidemiological episodes resulting in acute schistosomiasis involving wealthy persons living in the State of Pernambuco, Brazil. The authors discuss the epidemiological, clinical and serologic characteristics of the acute infections and also the way in which the conditions for transmission occurred.

Influence of specific treatment on the morbidity of schistosomiasis mansoni in an endemic area of Minas Gerais, Brazil

The authors describe the evolution of schistosomiasis mansoni in inhabitants of Capitão Andrade, Minas Gerais, Brazil, from 1973 to 1994. The prevalence of infection was 60.8% in 1973, 36.2% in 1984, and 27.3% in 1994. The evolution of the clinical forms of the disease in this group was as follows: unchanged in 76.7%, clinical progression in 8.4% and clinical regression in 14.9%. The reduction of the prevalence and severity of Schistosoma mansoni infection over the 21 years period, can be attributed to treatment of infected subjects performed in the area and to the installation of piped water in their dwellings.

Comparison between specific and multiplex reverse transcription-polymerase chain reaction for detection of hepatitis A virus, poliovirus and rotavirus in experimentally seeded oysters

Outbreaks of gastroenteritis have occurred among consumers of raw or undercooked shellfish harvested from faecally polluted waters. A multiplex reverse transcription-polymerase chain reaction (RT-PCR) was applied for the simultaneous detection of hepatitis A virus (HAV), poliovirus (PV) and simian rotavirus (RV-SA11) and compared with specific primers for each genome sequence. Three amplified DNA products representing HAV (192 bp), PV (394 bp) and RV (278 bp) were identified when positive controls were used. However, when tested on experimentally contaminated raw oysters, this method was not able to detect the three viruses simultaneously. This is probably due to the low concentration of viral RNAs present in oyster extract which were partially lost during the extracts preparation.

Detection of Cryptosporidium - specific coproantigen in human immunodeficiency virus/acquired immunodeficiency syndrome patients by using a commercially available immunoenzymatic assay

The aim of this study was to verify the occurrence of Cryptosporidium infection in 52 human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) patients (group 1) and 38 clinically healthy individuals (group 2) by using enzyme immunoassay (EIA). All fecal samples collected were submitted to the Baermann, Lutz, and Ritchie methods, the Safranin/Methylene Blue, and Weber's chromotrope modified Trichrome staining techniques, and EIA. In group 1, parasitological staining techniques and EIA were both positive for Cryptosporidium sp. infection in 3/52 (5.8%) samples and both negative in 45/52 (86.5%) samples, while 4/52 (7.7%) samples were positive in EIA and negative in parasitological staining techniques. Concerning group 2, all samples were negative by EIA and microscopy for Cryptosporidium infection. In conclusion, EIA may be an alternative method for detecting Cryptosporidium-specific coproantigen in HIV/AIDS patients.

Stage-specific activity of potential antimalarial compounds measured in vitro by flow cytometry in comparison to optical microscopy and hypoxanthine uptake

The evaluation of new antimalarial agents using older methods of monitoring sensitivity to antimalarial drugs are laborious and poorly suited to discriminate stage-specific activity. We used flow cytometry to study the effect of established antimalarial compounds, cysteine protease inhibitors, and a quinolone against asexual stages of Plasmodium falciparum. Cultured P. falciparum parasites were treated for 48 h with different drug concentrations and the parasitemia was determined by flow cytometry methods after DNA staining with propidium iodide. P. falciparum erythrocytic life cycle stages were readily distinguished by flow cytometry. Activities of established and new antimalarial compounds measured by flow cytometry were equivalent to results obtained with microscopy and metabolite uptake assays. The antimalarial activity of all compounds was higher against P. falciparum trophozoite stages. Advantages of flow cytometry analysis over traditional assays included higher throughput for data collection, insight into the stage-specificity of antimalarial activity avoiding use of radioactive isotopes.

Detection of hepatitis A, B, and C virus-specific antibodies using oral fluid for epidemiological studies

In this report, we examine the adaptability of commercially available serological kits to detect antibodies markers for viral hepatitis in oral fluid samples. We also assessed the prevalence of hepatitis A, B, and C virus-specific antibodies, and related risk factors for these infectious diseases through sensitivity of the tests in saliva samples to evaluate if oral fluid can be an alternative tool to substitute serum in diagnosis of acute viral hepatitis and in epidemiological studies. One hundred and ten paired serum and saliva specimens from suspect patients of having acute hepatitis were collected to detect antibodies to hepatitis A (total and IgM), hepatitis B (anti-HBs, total anti-HBc and IgM anti-HBc), and hepatitis C (anti-HCV) using commercially available enzyme-linked immunossorbent assay (EIA). In relation to serum samples, oral fluid assay sensitivity and specificity were as follows: 87 and 100% for total anti-HAV, 79 and 100% for anti-HAV IgM, 6 and 95% for anti-HBs, 13 and 100% for total anti-HBc, 100 and 100% for anti-HBc IgM, and 75 and 100% for anti-HCV. The consistency observed between antibodies tests in saliva and expected risk factors for hepatitis A and C suggests that the saliva method could replace serum in epidemiological studies for hepatitis A and C.

Remodeling of hepatic vascular changes after specific chemotherapy of schistosomal periportal fibrosis

Hepatosplenic schistosomiasis was the first human disease in which the possibility of extensive long standing hepatic fibrosis being degraded and removed has been demonstrated. When such changes occurred, the main signs of portal hypertension (splenomegaly, esophageal varices) progressively disappeared, implying that a profound vascular remodeling was concomitantly occurring. Hepatic vascular alterations associated with advanced schistosomiasis have already been investigated. Obstruction of the intrahepatic portal vein branches, plus marked angiogenesis and compensatory hyperplasia and hypertrophy of the arterial tree are the main changes present. However, there are no data revealing how these vascular changes behave during the process of fibrosis regression. Here the mouse model of pipestem fibrosis was used in an investigation about these vascular alterations during the course of the infection, and also after treatment and cure of the disease. Animals representing the two polar hepatic forms of the infection were included: (1) "isolated granulomas" characterized by isolated periovular granulomas sparsely distributed throughout the hepatica parenchyma; and (2) 'pipestem fibrosis' with periovular granulomas and fibrosis being concentrated within portal spaces, before and after treatment, were studied by means of histological and vascular injection-corrosion techniques. Instances of widespread portal vein obstruction of several types were commonly found in the livers of the untreated animals. These obstructive lesions were soon repaired, and completely disappeared four months following specific treatment of schistosomiasis. Treatment was accomplished by the simultaneous administration of praziquantel and oxamniquine. The most impressive results were revealed by the technique of injection of colored masses into the portal system, followed by corrosion in strong acid. The vascular lesions of non-treated pipestem fibrosis were represented in the plastic casts by considerable diminution of the fine peripheral portal vein radicles, plus dilatation of periportal collaterals. Four months after treatment, this last picture appeared replaced by tufts of newly interwoven vessels formed along the main portal vein branches, disclosing a strong angiomatoid reparative change. Understanding about the cellular elements at play during fibro-vascular repairing changes of hepatic schistosomiais represents a matter of considerable scientific and conceptual importance. At present time one may only speculate about the participation of some type of natural stem-cell capable of restoring the diseased liver back to normal once the cause of the disorder has been eliminated.

Aspartic protease activities of schistosomes cleave mammalian hemoglobins in a host-specific manner

We examined the efficiency of digestion of hemoglobin from four mammalian species, human, cow, sheep, and horse by acidic extracts of mixed sex adults of Schistosoma japonicum and S. mansoni. Activity ascribable to aspartic protease(s) from S. japonicum and S. mansoni cleaved human hemoglobin. In addition, aspartic protease activities from S. japonicum cleaved hemoglobin from bovine, sheep, and horse blood more efficiently than did the activity from extracts of S. mansoni. These findings support the hypothesis that substrate specificity of hemoglobin-degrading proteases employed by blood feeding helminth parasites influences parasite host species range; differences in amino acid sequences in key sites of the parasite proteases interact less or more efficiently with the hemoglobins of permissive or non-permissive hosts.

Merozoite surface protein 2 allelic variation influences the specific antibody response during acute malaria in individuals from a Brazilian endemic area

The antibody response to Plasmodium falciparum parasites of naturally infected population is critical to elucidate the role of polymorphic alleles in malaria. Thus, we evaluated the impact of antigenic diversity of repetitive and family dimorphic domains of the merozoite surface protein 2 (MSP-2) on immune response of 96 individuals living in Peixoto de Azevedo (MT-Brazil), by ELISA using recombinant MSP-2 proteins. The majority of these individuals were carrying FC27-type infections. IgG antibody responses were predominantly directed to FC27 parasites and were correlated to the extension of polymorphism presented by each MSP-2 region. This finding demonstrated the impact of the genetic polymorphism on antibody response and therefore, its importance on malaria vaccine efficacy.

Trypanosoma cruzi: a stage-specific calpain-like protein is induced after various kinds of stress

Calpains are calcium-dependent cysteine proteinases found in all living organisms and are involved in diverse cellular processes. Calpain-like proteins have been reported after in silico analysis of the Tritryps genome and are believed to play important roles in cell functions of trypanosomatids. We describe the characterization of a member of this family, which is differentially expressed during the life-cycle of Trypanosoma cruzi.

Genus-specific kinetoplast-DNA PCR and parasite culture for the diagnosis of localised cutaneous leishmaniasis: applications for clinical trials under field conditions in Brazil

The positivities of two methods for the diagnosis of localised cutaneous leishmaniasis (CL) were estimated in 280 patients enrolled in a clinical trial. The trial was conducted in an endemic area of Leishmania (Viannia) braziliensis and trial participants were patients with skin ulcers and positive leishmanin skin tests. Patients underwent aspirative skin punctures of the ulcerated lesions and lymph nodes for in vitro cultures, which were processed under field conditions at the local health centre. Skin lesion biopsies were tested at a reference laboratory using kinetoplastid DNA (kDNA)-PCR to detect DNA. The median time required to obtain a positive culture from the skin samples was seven days and the contamination rate of the samples was 1.8%. The positivities of the cultures from skin lesions, kDNA-PCR and the combination of the two methods were 78.2% (95% CI: 73-82.6%), 89.3% (95% CI: 85.1-92.4%) and 97.1% (95% CI: 94.5-98.5%). We conclude that parasite culture is a feasible method for the detection of Leishmania in field conditions and that the combination of culture and PCR has a potential role for the diagnosis of CL in candidates for clinical trials.

Comparison of HPV genotyping by type-specific PCR and sequencing

Human papillomavirus (HPV) infection is the most common sexually transmitted disease worldwide and there is a strong link between certain high-risk viral types and cervical carcinogenesis. Although there are several typing methods, it is still unclear which test is the best. This study compared the effectiveness of type-specific PCR (TS-PCR) and sequencing, with a focus on their clinical application. A total of 260 cervical samples from HPV-positive patients were tested for types 6, 11, 16, 18, 31, 33 and 35 using TS-PCR and sequencing. The genotype was identified in 36% of cases by TS-PCR and in 75% by sequencing. Sequencing was four times more likely to identify the viral type in positive samples than TS-PCR (p = 0.00). Despite being more effective for virus genotyping, sequencing was unable to identify viral types in multiple infections. Combining both techniques resulted in highly sensitive detection (87% of cases), showing that they are complementary methods. HPV genotyping is an important step in HPV management, helping to identify patients with a higher risk of developing cervical cancer and contributing to the development of type-specific vaccines.

Specific schistosomiasis treatment as a strategy for disease control

The great hope for schistosomiasis treatment began with the development of oxamniquine and praziquantel. These drugs can be administered orally in a single dose and have a high curative power with minor side effects. In this study, we carried out a field experiment involving a population of 3,782 people. The population was examined at four localities in Minas Gerais within the valleys of the Doce and Jequitinhonha Rivers. In this cohort, there were 1,790 patients infected with Schistosoma mansoni (47.3%) and we showed that only 1,403 (78.4%) could be treated with oxamniquine in a single dose of 12.5-20 mg/kg orally. The other 387 (21.6%) were not treated during the first stage because of contraindications (pregnancy or impeditive diseases), absences or refusals. It was observed that, on average, 8.8-17% of the infected patients continued to excrete S. mansoni eggs at the end of the 2nd month after treatment and 30-32% of the cohort was infected by the end of the 24th month. In one of the areas that we followed-up for a total of 30 years, the prevalence of the infection with S. mansoni fell from 60.8-19.3% and the hepatosplenic form of the disease dropped from 5.8-1.3%. We conclude that specific treatment of schistosomiasis reduces the prevalence of infection in the short-term and the morbidity due to schistosomiasis in medium to long-term time frames, but does not help to control disease transmission.