T-Cell Immunophenotyping and Cytokine Production Analysis in Patients with Chagas Disease 4 Years after Benznidazole Treatment.

The major problem with Chagas disease is evolution of the chronic indeterminate form to a progressive cardiac disease. Treatment diminishes parasitemia but not clinical progression, and the immunological features involved are unclear. Here, we studied the clinical course and the immune response in patients with chronic-phase Chagas disease at 48 months after benznidazole treatment. Progression to the cardiac form of Chagas disease or its aggravation was associated with higher in vitro antigen-specific production of interferon gamma (IFN-γ) in patients with cardiac Chagas disease than in patients with the indeterminate form. Predominance of IFN-γ production over interleukin-10 (IL-10) production in antigen-specific cultures was associated with cardiac involvement. Significantly higher numbers of antigen-specific T helper 1 cells (T-Bet+ IFN-γ+) and a significantly higher IFN-γ+/IL-10+ ratio were observed in patients with cardiac Chagas disease than in patients with the indeterminate form. Cardiac damage was associated with higher numbers of T helper cells than cytotoxic T lymphocytes producing IFN-γ. Patients with cardiac Chagas disease had predominant CD25- and CD25low T regulatory (Treg) subpopulations, whereas patients with the indeterminate form manifested a higher relative mean percentage of CD25high Treg subpopulations. These findings suggest that at 48 months after benznidazole treatment, the disease can worsen or progress to the cardiac form. The progression may be related to increased IFN-γ production (mostly from CD4+ T cells) relative to IL-10 production and increased Treg percentages. Patients with the indeterminate form of Chagas disease show a more balanced ratio of proinflammatory and anti-inflammatory cytokines.

Evaluation of the multispecies coalescent method to explore intra-Trypanosoma cruzi I relationships and genetic diversity.

Chagas Disease is a zoonosis caused by the parasite Trypanosoma cruzi. Several high-resolution markers have subdivided T. cruzi taxon into at least seven lineages or Discrete Typing Units (DTUs) (TcI-TcVI and TcBat). Trypanosoma cruzi I is the most diverse and geographically widespread DTU. Recently a TcI genotype related to domestic cycles was proposed and named as TcIDOM. Herein, we combined traditional markers and housekeeping genes and applied a Multispecies Coalescent method to explore intra-TcI relationships, lineage boundaries and genetic diversity in a random set of isolates and DNA sequences retrieved from Genbank from different countries in the Americas. We found further evidence supporting TcIDOM as an independent and emerging genotype of TcI at least in Colombia and Venezuela. We also found evidence of high phylogenetic incongruence between parasite's gene trees (including introgression) and embedded species trees, and a lack of genetic structure among geography and hosts, illustrating the complex dynamics and epidemiology of TcI across the Americas. These findings provide novel insights into T. cruzi systematics and epidemiology and support the need to assess parasite diversity and lineage boundaries through hypothesis testing using different approaches to those traditionally employed, including the Bayesian Multispecies coalescent method.

Identification of bat trypanosomes from Minas Gerais state, Brazil, based on 18S rDNA and Cathepsin-L-like targets.

Several bat species can be infected by trypanosomes, but there is not much information about which of these parasites infect bats from Triângulo Mineiro and Alto Paranaíba, Minas Gerais state, Brazil, a formerly endemic region for Trypanosoma cruzi, the causative agent of Chagas disease. The aim of this study was to describe, characterize, and identify the presence of trypanosomes in bats. The captured bats (448) belong to four families and to 19 different species. Of those, 37 bats were found to be positive for trypanosomes by microhematocrit, (infection rate 8.3%) and 27 were positive after hemoculture analysis. Initially, the isolates were identified by PCR (18S rDNA, 24Sα rDNA, spliced leader, COII RFLP-PCR) using primers originally designed for T. cruzi. PCRs (18S rDNA, 24Sα rDNA) showed compatible bands for TcI, whereas COII RFLP-PCR showed a similar pattern associated to TcII. However, there was no DNA amplification using spliced leader as a target, revealing a discrepancy between the results. Phylogenetic analysis of Cathepsin L-like and 18S rDNA sequences proved that 15 of the isolates corresponded to Trypanosoma cruzi marinkellei and one to Trypanosoma dionisii. These results revealed that the diversity of trypanosome species in a region considered endemic for Chagas disease is greater than previous descriptions. All this can confirm the necessity of using DNA sequencing approaches in order to determinate trypanosomes species isolated from bats.

Leishmania infection in bats from a non-endemic region of Leishmaniasis in Brazil.

Leishmaniasis is a complex of zoonotic diseases caused by parasites of the genus Leishmania, which can develop in domestic as well as wild animals and humans throughout the world. Currently, this disease is spreading in rural and urban areas of non-endemic regions in Brazil. Recently, bats have gained epidemiological significance in leishmaniasis due to its close relationship with human settlements. In this study, we investigated the presence of Leishmania spp. DNA in blood samples from 448 bats belonging to four families representing 20 species that were captured in the Triangulo Mineiro and Alto Paranaiba areas of Minas Gerais State (non-endemic areas for leishmaniasis), Brazil. Leishmania spp. DNA was detected in 8·0% of the blood samples, 41·6% of which were Leishmania infantum, 38·9% Leishmania amazonensis and 19·4% Leishmania braziliensis. No positive correlation was found between Leishmania spp. and bat food source. The species with more infection rates were the insectivorous bats Eumops perotis; 22·2% (4/18) of which tested positive for Leishmania DNA. The presence of Leishmania in the bat blood samples, as observed in this study, represents epidemiological importance due to the absence of Leishmaniasis cases in the region.

Effects of cholinergic stimulation with pyridostigmine bromide on chronic chagasic cardiomyopathic mice.

The aim of the present study was to assess the effects of an anticholinesterase agent, pyridostigmine bromide (Pyrido), on experimental chronic Chagas heart disease in mice. To this end, male C57BL/6J mice noninfected (control:Con) or chronically infected (5 months) with Trypanosoma cruzi (chagasic:Chg) were treated or not (NT) with Pyrido for one month. At the end of this period, electrocardiogram (ECG); cardiac autonomic function; heart histopathology; serum cytokines; and the presence of blood and tissue parasites by means of immunohistochemistry and PCR were assessed. In NT-Chg mice, significant changes in the electrocardiographic, autonomic, and cardiac histopathological profiles were observed confirming a chronic inflammatory response. Treatment with Pyrido in Chagasic mice caused a significant reduction of myocardial inflammatory infiltration, fibrosis, and hypertrophy, which was accompanied by a decrease in serum levels of IFNγ with no change in IL-10 levels, suggesting a shift of immune response toward an anti-inflammatory profile. Lower nondifferent numbers of parasite DNA copies were observed in both treated and nontreated chagasic mice. In conclusion, our findings confirm the marked neuroimmunomodulatory role played by the parasympathetic autonomic nervous system in the evolution of the inflammatory-immune response to T. cruzi during experimental chronic Chagas heart disease in mice.

Species-specific markers for the differential diagnosis of Trypanosoma cruzi and Trypanosoma rangeli and polymorphisms detection in Trypanosoma rangeli.

Trypanosoma cruzi and Trypanosoma rangeli are kinetoplastid parasites which are able to infect humans in Central and South America. Misdiagnosis between these trypanosomes can be avoided by targeting barcoding sequences or genes of each organism. This work aims to analyze the feasibility of using species-specific markers for identification of intraspecific polymorphisms and as target for diagnostic methods by PCR. Accordingly, primers which are able to specifically detect T. cruzi or T. rangeli genomic DNA were characterized. The use of intergenic regions, generally divergent in the trypanosomatids, and the serine carboxypeptidase gene were successful. Using T. rangeli genomic sequences for the identification of group-specific polymorphisms and a polymorphic AT(n) dinucleotide repeat permitted the classification of the strains into two groups, which are entirely coincident with T. rangeli main lineages, KP1 (+) and KP1 (-), previously determined by kinetoplast DNA (kDNA) characterization. The sequences analyzed totalize 622 bp (382 bp represent a hypothetical protein sequence, and 240 bp represent an anonymous sequence), and of these, 581 (93.3%) are conserved sites and 41 bp (6.7%) are polymorphic, with 9 transitions (21.9%), 2 transversions (4.9%), and 30 (73.2%) insertion/deletion events. Taken together, the species-specific markers analyzed may be useful for the development of new strategies for the accurate diagnosis of infections. Furthermore, the identification of T. rangeli polymorphisms has a direct impact in the understanding of the population structure of this parasite.

A DTU-dependent blood parasitism and a DTU-independent tissue parasitism during mixed infection of Trypanosoma cruzi in immunosuppressed mice.

Trypanosoma cruzi (Tc) diversity is determined by different biological, genetic, and biochemical markers and has been grouped into six discrete typing units (DTUs) or taxonomic groups (TcI-TcVI). This variability, coupled with natural reinfection or the hosts' immunosuppression, may play an important role in the pathogenesis of Chagas disease. Therefore, we evaluated the blood and tissue parasitism and genetic profile of mice coinfected with the TcII (JG) strain and TcI AQ1-7 (AQ) or MUTUM (MT) strains during the acute and chronic phases of the disease and during immunosuppression. T. cruzi blood populations in mixed infections were clearly associated with the TcII strain during acute and chronic phases or during immunosuppression. However, in tissues, the parasite populations were distributed according to the strain and the stage of infection. TcII populations overlapped TcI strains during the acute phase; in contrast, during chronic phase, both TcI strains were more prevalent than the TcII strain. The immunosuppression induced selective exacerbation of parasite populations, leading to reactivation of the TcII strain when associated with the AQ, but not with MT strain. Thus, a differential distribution of T. cruzi populations in blood and tissues with overlapping according to the stage of infection and strain used was observed. Blood parasitism was associated with the DTU TcII and tissue parasitism with a specific parasite strain and not with DTUs. Finally, to our knowledge, this is the first study to analyze subpatent blood parasitism and to simultaneously identify different T. cruzi populations in tissues and blood.

Homogeneity of Trypanosoma cruzi I, II, and III populations and the overlap of wild and domestic transmission cycles by Triatoma brasiliensis in northeastern Brazil.

The genetic variability of 24 Trypanosoma cruzi isolates from humans (11) and triatomines (13) in northeastern Brazil was analyzed by random amplified polymorphic DNA (RAPD) and compared with taxonomic groups, host, and geographical origin of the parasite. TcI (12.5%), TcII (45.8%), and TcIII (41.6%) showed a similarity coefficient (SC) of 0.74 using the mean of three primers and 0.80, 0.75, and 0.66 for λgt11-F, M13-40F, and L15996 primers, respectively. The samples were clustered according to their phylogenetic origin in two polymorphic and divergent branches: one associated with TcI and the other with two subbranches corresponding to TcII and TcIII. TcI was only identified in humans and correlated with the Id homogenous group (0.80 SC). TcII from humans and Triatoma brasiliensis showed 0.86 SC and was clustered according monoclonal or polyclonal populations with similar RAPD profiles detected among the vector and/or humans in different municipalities. TcIII was isolated exclusively in sylvatic cycles from T. brasiliensis and Panstrongylus lutzi and showed low variability (0.84 SC) and high homology mainly among isolated populations at the same locality. The homology of T. cruzi among different hosts and locations suggests the distribution of principal clones circulating and reveals an overlapping between the sylvatic and domestic cycles in this area, where T. brasiliensis infected with TcII acts as link in both environments. This species is important to maintain TcII and TcIII in wild cycles and deserves particular attention due an emergent risk of these populations being introduced into the domestic cycle; moreover, its clinical and epidemiological implications remain unknown.

Trypanosoma cruzi experimental congenital transmission associated with TcV and TcI subpatent maternal parasitemia.

The congenital transmission of Chagas disease is associated with an increase in parasitemia during pregnancy, maternal and fetal immunity, and populations of Trypanosoma cruzi. In this study, the biological behavior of TcI and TcV (isolated from a human congenital case) strains and their potential for experimental congenital transmission were evaluated in female BALB/C mice. Parasitemia was estimated by fresh blood examination, semiquantitative microhematocrit, and hemoculture, while congenital transmission was evaluated by culture in the liver infusion tryptose medium and by polymerase chain reaction (PCR) of the pups' tissues on postnatal day 7 and of the pups' blood sample at 30 days after birth. Infection was detected in 100 % of the females. Both strains showed subpatent parasitemia, which was higher for TcV infection. The presence of amastigote nest was detected only in an animal infected with TcI. The inflammatory process was more frequent (p = 0.001) in the tissues of the animals infected with TcV (58.6 %) than TcI (31.1 %). The fertility rates of females mated after 35 days postinfection were similar (90 % for TcV, 88.9 % for TcI; p = 0.938). Parasitemia did not change during pregnancy. The average number of pups/female was greater (p = 0.03) in mice with TcV infection (8.30) than in those with TcI infection (4.78). Congenital transmission was detected exclusively by PCR in 50.9 % of the pups, 46.6 % for TcV and 58.1 % for TcI. The PCR positivity for TcI was higher in the blood than in the tissue (p = 0.003). These results demonstrate the T. cruzi experimental congenital infection associated with subpatent maternal parasitemia of TcI and TcV.

Antifungal susceptibility, enzymatic activity, PCR-fingerprinting and ITS sequencing of environmental Cryptococcus laurentii isolates from Uberaba, Minas Gerais, Brazil.

Cryptococcus laurentii has been classically considered a saprophytic species, although several cases of human infection have been already reported. This study aimed to evaluate the phospholipase, proteinase and hemolysins activity, the antifungal susceptibility profile, the genetic variability by M13 and (GACA)(4) fingerprinting and the internal transcribe spacer (ITS) sequencing of 38 C. laurentii isolates recovered from captive bird droppings and surrounding hospital areas. All of them exhibited phospholipase activity, while the hemolytic activity was evidenced in 34 (89.4%) isolates. None of them exhibited proteinase activity. Twenty-seven isolates (71.1%) presented susceptibility dose dependent to fluconazole. Most isolates (94.7%) were susceptible to voriconazole, while one (2.65%) was resistant to this drug. Twenty-one (55.3%) isolates showed reduced susceptibility to itraconazole while nine (23.7%) were resistant. Three (7.9%) and five (13.1%) isolates exhibited resistance to ketoconazole and amphotericin B, respectively. Most C. laurentii fingerprinting obtained with M13 and (GACA)(4) showed high heterogeneity. By using the two primers, seven (18.4%) isolates grouped as A (CL2, CL7, and CL8), B (CL35, CL38) and C (CL29, CL30) with 100% similarity. Different from most variable surrounding hospital isolates, all but one of the pet shops strains clustered with the two primers, although they had been recovered from different neighborhoods. All isolates were identified as C. laurentii phylogenetic group I by ITS sequencing. Thus, the presence of virulence factors, a decreased antifungal susceptibility and a heterogeneous molecular pattern of the C. laurentii isolates here described suggests this species can be a potential pathogen in the context of the immunocompromised population.

RAPD analysis with the primer L15996 of Brazilian clinical and environmental Cryptococcus neoformans isolates.

Different methods have been used to perform the molecular characterization of Cryptococcus neoformans. Among them, RAPD analysis is able to separate isolates of the same species and genotypes. This study aimed to evaluate clinical and environmental C. neoformans isolates from Minas Gerais, Brazil by RAPD and correlate the genetic profiles with the ones obtained by URA5-RFLP, virulence factors and antifungal susceptibility patterns. Forty-five environmental (31 from areas surrounding hospital and 14 from captive bird droppings from pet-shops) and 29 clinical C. neoformans isolates were evaluated. Antifungal susceptibility tests (Clinical and Laboratory Standards Institute), URA5-RFLP analysis and the assessment of virulence factors were performed according to their original descriptions. RAPD profiles were obtained using the L15996 primer (5'-CTCCACCATTAGCACCCAAAGC-3'). RAPD analysis generated two to 20 bands for all studied isolates. The isolates presented similarities ranging from 10.8 to 100.0%. Considering a minimum identity score of 50%, four clusters were formed. Cluster I contained 10 pet-shops bird dropping isolates, cluster II contained 22 clinical isolates most of them recovered from cerebrospinal fluid, cluster III contained 14 isolates from hospital surroundings and cluster IV contained 12 environmental isolates most from hospital surroundings. Fourteen isolates were not grouped. The RAPD profiles were clustered according to their source and URA5-RFLP pattern. No correlation between virulence factors or antifungal susceptibility profile with the obtained RAPD profiles was observed.

The relationship between heart rate variability and serum cytokines in chronic chagasic patients with persistent parasitemia.

BACKGROUND: Persistent parasitemia, immunological, and autonomic nervous system impairments may play an important role in the evolution and clinical outcome of the chronic phase of Chagas' disease by triggering functional cardiovascular changes. METHODS: Three groups were evaluated: 17 chronic chagasic patients with the indeterminate form (IChD), 12 chronic chagasic patients with cardiac forms (ChHD), and 29 individuals as a healthy control group. Parasitemia was assessed by polymerase chain reaction; hemoculture, heart rate variability by linear and nonlinear methods, and interleukin (IL)-1ß, IL-4, IL-6, IL-10, IL-12, IL-13, IL-17, and tumor necrosis factor-α, and interferon (IFN)-γ serum cytokines were assessed by enzyme-linked immune assay. RESULTS: Twenty-nine chronic chagasic patients were positive for parasitemia (17 IChD and 12 ChHD). Heart rate variability parameters in baseline condition and after cold face test were significantly decreased in chagasic patients compared to controls. Tilt tests showed no alteration. However, using nonlinear indices, ChHD patients presented lower values compared to IChD and controls. Differences in the expression of serum cytokines were observed between chagasic patients and controls. However, among the groups, ChHD presented higher median values of IL-10 and lower of IFN-γ compared to IChD. CONCLUSION: Both chagasic groups present an autonomic impairment using linear methods. The nonlinear methods revealed that the ChHD group had a higher cardiovascular risk. Serum cytokine concentrations between chagasic patients were similar. However, ChHD showed higher concentrations of IL-10 and lower of IFN-γ, suggesting some established process of immune regulation.

Polymorphisms in the 18S rDNA gene of Cystoisospora belli and clinical features of cystoisosporosis in HIV-infected patients.

Intraspecific variability among Cystoisospora belli isolates and its clinical implications in human cystoisosporosis have not been established. In this study, the restriction fragment length polymorphisms in a 1.8-kb amplicon of the small subunit ribosomal DNA (SSU rDNA) of the parasite was investigated in 20 C. belli-positive stool samples obtained from 15 HIV-infected patients. Diarrheic syndrome was observed in all patients with cystoisosporosis and the number of diarrheic episodes per patient during hospitalization ranged from 1 to 26 (mean of 9.64 ± 9.30), with a mean duration of 2 to 12 days (mean of 5.90 ± 3 days). Three restriction profiles (RF) were generated with MboII digestion, which were named RFI, RFII, and RFIII. Two isolates obtained from a patient with extraintestinal cystoisosporosis showed distinct restriction profiles with MboII. This study demonstrates that patients can be infected with different C. belli genotypes, and this information may be useful for identifying new C. belli genotypes infecting humans.

Sequencing and analysis of chromosomal extremities of Trypanosoma rangeli in comparison with Trypanosoma cruzi lineages.

The aim of this study was to investigate the genetic variability of sequences present in the chromosome ends of Trypanosoma rangeli strains defined by the presence (+) or absence (-) of KP1 minicircles, and to compare the mean terminal restriction fragment (TRF) lengths to those of Trypanosoma cruzi populations representative of groups TcI, TcII, TcIV, and TcVI. Southern blots containing RsaI-digested genomic DNA of T. rangeli KP1(+) strains, T. rangeli KP1(-) strains, and T. cruzi strains were probed with the previously described subtelomeric sequences (170 bp) of T. rangeli and with telomeric hexamer repeats. Mean TRF length analysis showed that the chromosome ends of T. rangeli are distinctly organized, with TRFs ranging from 1.3 to 9 kb for KP1(+) strains and from 0.3 to 5.0 kb for KP1(-) strains. In T. cruzi, TRF length ranged from 0.2 to 9 kb and no association with the genotype of the parasite could be established. Sequence analysis of the 170-bp amplicons revealed the occurrence of sequence polymorphisms in the subtelomeric region between and within KP1(+) and KP1(-) strains. The GTT triplet was detected in all KP1(+) strains, except for strain Cas4, but not in any of the KP1(-) strains. The dendrogram constructed by alignment of all T. rangeli strains showed the division into two main groups, mainly related to the presence or absence of the KP1 minicircle. In conclusion, the present results extend the genotype differences demonstrated by kDNA and karyotype analysis in T. rangeli to the chromosome ends of the parasite.

IgG Subclass Analysis in Patients with Chagas Disease 4 Years After Benznidazole Treatment.

BACKGROUND: In humans, Trypanosoma cruzi infection is controlled by a complex immune response. Immunoglobulin G (IgG) is important for opsonizing blood trypomastigotes, activating the classic complement pathway, and reducing parasitemia. The trypanocidal activity of benznidazole is recognized, but its effects on the prevention and progression of Chagas disease is not well understood OBJECTIVE: We aimed to evaluate the levels of total IgG and cross-specific IgG subclasses in patients with chronic Chagas disease of different clinical forms before and after 4 years of benznidazole treatment. METHODS: Eight individuals with the indeterminate form and nine with the cardiac form who completed the treatment protocol were evaluated. The levels of total IgG and IgG1, IgG2, IgG3, and IgG4 isotypes were quantified in the serum of each individual using the fluorescent immunosorbent assay. The results are expressed as relative fluorescence unit. RESULTS: Patients with chronic Chagas disease presented decreased levels of total IgG at 48 months after benznidazole treatment. Increased IgG1 and decreased IgG3 levels were observed in patients with the cardiac form and those with exacerbated clinical forms. In addition, a decrease in the IgG3/IgG1 ratio was observed in individuals with the cardiac form of Chagas disease. CONCLUSIONS: Benznidazole administration in the chronic phase differentially changes IgG subclasses in patients with cardiac and indeterminate forms, and monitoring the IgG3 level may indicate the possible prognosis to the cardiac form or worsening of the already established clinical form.

Socioepidemiological screening of serologically ineligible blood donors due to Chagas disease for the definition of inconclusive cases.

Epidemiological screening combined with serological tests has become an important tool at blood banks for the characterization of donors with or without Trypanosoma cruzi infection. Thus, the objective of the present study was to describe the sociodemographic and epidemiological characteristics of blood donors with non-negative serology for T. cruzito determine possible risk factors associated with serological ineligibility. Sociodemographic and epidemiological data were collected by analysis of patient histories and interviews. The data were analyzed descriptively using absolute and relative frequencies and odds ratio (OR) evaluation. The frequency of serological ineligibility was 0.28%, with a predominance of inconclusive reactions (52%) and seropositivity among first-time donors (OR = 607), donors older than 30 years (OR = 3.7), females (OR = 1.9), donors from risk areas (OR = 4) and subjects living in rural areas (OR = 1.7). The risk of seropositivity was higher among donors who had contact with the triatomine vector (OR = 11.7) and those with a family history of Chagas disease (OR = 4.8). The results demonstrate the value of detailed clinical-epidemiological screening as an auxiliary tool for serological definition that, together with more specific and more sensitive laboratory methods, will guarantee a higher efficacy in the selection of donors at blood centres.

Trypanosoma rangeli 28Sß Ribosomal Gene Allows Intra and Interspecific Molecular Differentiation.

Trypanosoma rangeli is an avirulent flagellate protozoan that could mislead correct diagnosis of Trypanosoma cruzi infection, the causative agent of Chagas' disease, given their high similarity. Besides, T. rangeli presents two genetic groups, whose differentiation is achieved mainly by molecular approaches. In this context, ribosomal DNA (rDNA) is a useful target for intra and interspecific molecular differentiation. Analyzing the rDNA of T. rangeli and comparison with other trypanosomatid species, two highly divergent regions (Trß1 and Trß2) within the 28Sß gene were found. Those regions were amplified and sequenced in KP1(+) and KP1(-) strains of T. rangeli, revealing group-specific polymorphisms useful for intraspecific distinction through restriction fragment length polymorphism technique. Also, amplification of Trß1 allowed differentiation between T. rangeli and T. cruzi. Trß2 predicted restriction length profile, allowed differentiation between T. rangeli, T. cruzi, Trypanosoma brucei, and Leishmania braziliensis, increasing the use of Trß1 and Trß2 beyond a molecular approach for T. rangeli genotyping, but also as a useful target for trypanosomatid classification.

Interstitial cells of Cajal in chagasic megaesophagus.

Chagasic visceromegalies are the most important digestive manifestations of Chagas disease and are characterized by motor disorders and dilation of organs such as esophagus and colon. One of the theories raised to explain the physiopathogenesis of chagasic megas is the plexus theory. Recent studies have shown a reduction of interstitial cells of Cajal (ICCs) in the colon of chagasic patients. These cells are present throughout the gastrointestinal tract and are considered to be pacemaker cells, that is, they are responsible for coordinating peristalsis and for mediating nerve impulses. In view of the lack of studies on these cells in megaesophagus and the previous observation of a reduction of ICCs in chagasic megacolons, we compared the distribution of ICCs in the esophagus of chagasic and nonchagasic patients to contribute to a better understanding of the physiopathogenesis of this esophageal disease. Esophageal biopsy samples from 10 chagasic and 5 nonchagasic patients were used. Cells were identified with the anti-CD117 antibody. The number of ICCs was quantified in longitudinal and circular muscle layers and myenteric plexus. The results were analyzed statistically by comparison of means. An intense reduction in the number of ICCs was observed in muscle layers and in the myenteric plexus of patients with megaesophagus. We conclude that there is an intense reduction of ICCs in the esophagus of chagasic patients when compared to nonchagasic patients, a finding supporting the important role of these cells in gastrointestinal tract motility. A deficiency in these cells might be implied in the genesis of megaesophagus.

Absence of experimental cross-protection induced by a Trypanosoma cruzi-like strain isolated from bats.

This study evaluated the possibility of inoculation and reinoculation with a trypanosomatid isolated from bats that is morphologically, biologically and molecularly similar to Trypanosoma cruzi, to protect against infection by virulent strains. Non-isogenic mice were divided into 24 groups that received from zero to three inoculations of Trypanosoma cruzi-like strain RM1, in the presence or absence of Freunds adjuvant, and were challenged with the VIC or JG strains of Trypanosoma cruzi. Parasitemia and survival were monitored and animals were sacrificed for histopathological analysis. Animals immunized with Trypanosoma cruzi-like strain RM1 presented decreased parasitemia, independently of the number of inoculations or the presence of adjuvant. In spite of this reduction, these animals did not present any protection against histopathological lesions. Severe eosinophilic infiltrate was observed and was correlated with the number of inoculations of Trypanosoma cruzi-like strain RM1. These findings suggest that prior inoculation with this strain did not protect against infection but, rather, aggravated the tissue inflammatory process.

Seasonal profile and level of CD4+ lymphocytes in the occurrence of cryptosporidiosis and cystoisosporidiosis in HIV/AIDS patients in the Triângulo Mineiro region, Brazil.

Patients with AIDS are particularly susceptible to infection with intestinal coccidia. In this study the prevalence of infections with Cryptosporidium sp and Cystoisospora belli were evaluated among HIV/AIDS patients in the Triângulo Mineiro region, Brazil. Between July 1993 and June 2003 faecal samples from 359 patients were collected and stained by a modified Ziehl-Neelsen method, resulting in 19.7% of positivity for coccidian (8.6% with Cryptosporidium sp, 10.3% with Cystoisospora belli and 0.8% with both coccidian). Patients with diarrhoea and T CD4+ lymphocyte levels < or =200 cells/mm3 presented higher frequency of these protozoans, demonstrating the opportunistic profile of these infections and its relationship with the immunological status of the individual. It was not possible to determine the influence of HAART, since only 8.5% of the patients positive for coccidian received this therapy regularly. Parasitism by Cryptosporidium sp was more frequent between December and February and thus was characterised by a seasonal pattern of infection, which was not observed with Cystoisospora belli.