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O Fernandes

Publications and source records attributed to O Fernandes.

At least 19 recordsLinked to original sources
PubMed2006

Stable infection of primates with Trypanosoma cruzi I and II.

In order to better comprehend the putative association between genotype Trypanosoma cruzi II and primates, an evaluation of the infection in free ranging primates and specimens born in captivity from different geographical areas, the Amazon and the Atlantic forest, was carried out. Seroprevalences of the T. cruzi infection among the primates was similar in both biomes (45.5% and 46%). The parasites were isolated from 8 and 4 different species of primates, respectively from the Amazon and Atlantic forest. Multi-locus enzyme electrophoresis (MLEE) typed the isolates from Amazon as zymodeme 1. Mini-exon gene analysis characterized all these isolates as T. cruzi I, the main genotype circulating in the region. In the Atlantic forest, primates infected with TCI and TCII, as well as a mixed infection (TCI and TCII), were detected. These findings prove that primates may maintain stable infections by both genotypes. Moreover, data show that T. cruzi can occur in a wide range of primate genera, independent of their social behaviour, niches or habitats. Considering the high seroprevalence and stability of T. cruzi infection among the primates, these animals play an important role in the maintenance of the parasite in nature.

Animals↗
PubMed2005

A zymographic study of metalloprotease activities in extracts and extracellular secretions of Leishmania (Viannia) braziliensis strains.

Proteolytic activities of 5 strains of Leishmania (Viannia) braziliensis isolated from Brazilian and Colombian patients, presenting distinct clinical manifestations, were characterized and compared using whole-promastigote extracts and extracellular secretions. Zymographic assays concerning whole-cell extracts and supernatants resulted in the detection of high molecular weight bands, ranging from 50 to 125 kDa. Proteolytic activities from both whole-cell extracts and supernatants were optimal in a pH range 5.5 to 9.0 for all analysed strains. Such protease activities were inhibited when 10 mM 1,10-phenanthroline was assayed, strongly suggesting that the enzymes responsible for hydrolysis of the substrate belong to the metalloproteases class. Distinct profiles of metalloproteases were observed among the studied L. (V.) braziliensis strains. Differences among the microorganisms might be related to the geographical origin of the strains and/or to the clinical presentation.

Animals↗
PubMed2005

Trypanosoma cruzi infection in wild mammals of the National Park 'Serra da Capivara' and its surroundings (Piaui, Brazil), an area endemic for Chagas disease.

We studied the prevalence of Trypanosoma cruzi infection among eight species of wild small mammals (n=289) in an area where human cases of infection/disease have occurred. Dogs (n=52) and goats (n=56) were also surveyed. The study was carried out inside a biological reserve, the National Park 'Serra da Capivara' and its surroundings in Piaui State, Brazil. The marsupial Didelphis albiventris and the caviomorph rodent Trichomys apereoides were found to be the most important reservoirs in the study area. Trichomys apereoides was the most abundant species (80%) and D. albiventris the most frequently infected (61%). Both T. cruzi I and T. cruzi II genotypes were isolated from these species. One specimen of Tr. apereoides displayed a mixed T. cruzi I/zymodeme 3 infection. Serum prevalence among dogs suggests that they may be involved in the maintenance of the parasite in the peridomestic environment, in contrast to goats, which are not apparently of any epidemiological importance. The distinct distribution and patterns of infection observed in the study areas suggest that even in the same biome, epidemiological studies or determination of control measures must take into account ecological peculiarities.

Animals↗
PubMed2004

[Epidemiological, social, and sanitary aspects in an area of the Rio Negro, State of Amazonas, with special reference to intestinal parasites and Chagas' disease].

A cross-sectional study was carried out on the residents of one in every four dwellings in the town of Barcelos (in the northern part of the State of Amazonas, on the right bank of the Rio Negro, 490 kilometers from Manaus by river), in order to evaluate social and sanitary conditions and specific indicators for intestinal parasites and Chagas' infection. During the survey, two questionnaires were applied, a household one to evaluate social and sanitary aspects, and an individual one, for social and epidemiological evaluation of the population conditions. A conglomerate family sample of 171 dwellings was studied. From each of the 658 habitants, a sample was requested for stool examination by Lutz sedimentation and Baermann-Moraes-Coutinho techniques modified by Willcox & Coura (1989), and blood was collected in filter paper for immunofluorescence test by Camargo (1966) and Souza & Camargo (1966) methods modified by Petana & Willcox (1975). The stool examination showed 69.4% of samples with one or more parasites. Ascaris lumbricoides was predominant with 51% of positivity and Entamoeba histolytica, although surveyed by a non-specific method, was present in 19.7%. Surprisingly, 20.1% of the 658 sera samples were reactive for T. cruzi antibodies at a dilution of 1:20 and 13.7% at 1:40. There was a strong correlation between this result and the level of human contact with wild triatomines, known locally as "piasava lice", and we succeeded in isolating by xenodiagnosis one strain of T. cruzi from one patient, a sixty-one-year old man (n. 209 -1), a native of the area, with positive serology for Chagas' disease and who worked in agriculture and transporting piasava and was very familiar with "piasava lice".

English Abstract↗
PubMed2004

Comparison of four serological tests for the diagnosis of Chagas disease in a Colombian endemic area.

The performance of 4 serological tests for the diagnosis of Chagas disease was evaluated in Santander, Colombia, a region still presenting active transmission. Serum samples from 638 individuals were submitted to an enzyme immunoassay test (EIA), using total lysate of a local Trypanosoma cruzi strain and 52.5% were positive (335/638). A subset of this group (94 positive individuals and 90 seronegatives) was randomly selected for further serological confirmation. Three additional tests were used--indirect immunofluorescence (IIF) and 2 distinct enzyme-linked immunosorbent assays using total lysate of the Y strain (EIA BM) and a mixture of 2 recombinant antigens (EIA RA). Seventy-nine patients were seropositive in all tests (84.0%-79/94). The number of positive sera with the IIF, EIA RA and EIA BM was 84/94 (89.4%), 80/94 (85.1%) and 79/94 (84.0%), respectively. In 15 out of the 94 EIA seropositive patients (16.0%), 10 individuals were negative in all 3 tests (10.6%-10/94). One was negative in the EIA BM and positive in the other two tests (1.1%-1/94) and 4 patients were positive, solely, in the IIF assay (4.3%-4/94). Relative to the 90 EIA negative individuals, 89 were confirmed in all other tests (98.9%-89/90). One individual, although seronegative in the IIF, was positive in both confirmatory EIA tests (1.1%-1/90). In addition, 120 blood specimens were submitted to PCR amplification. This group consisted of 79 confirmed seropositive cases, 16 individuals with discordant serological results and 25 validated seronegative individuals. The PCR was able to detect the presence of parasite DNA in 67 out of the 79 seropositive patients (84.8%), in 8 individuals with discordant serology (50.0%) and in only one seronegative individual (4.0%). The results pointed to the necessity for performing more than one serological test, preferentially with antigens from autochthonous strains, to achieve a reliable diagnosis of Chagas disease in Colombia.

Animals↗
PubMed2004

Distinct patterns of Trypanosoma cruzi infection in Leontopithecus rosalia in distinct Atlantic coastal rainforest fragments in Rio de Janeiro--Brazil.

Previous studies on infection of Trypanosoma cruzi in the Poço das Antas Biological Reserve population of wild free-ranging Leontopithecus rosalia have shown the presence of genotype T. cruzi II, associated in Brazil with human disease. Herein, this study has been extended, the infection being evaluated in L. rosalia of 3 different tamarin populations, inhabiting distinct forest areas located in the same Atlantic Coastal Rainforest. Edentata, Marsupialia, Rodentia and Chiroptera were examined exclusively in the Poço das Antas Biological Reserve. Excluding Chiroptera, T. cruzi infection was found in all orders. Biochemical and molecular characterization demonstrated that golden lion tamarins maintained stable infections by T. cruzi II. The isolates from the other mammals corresponded to T. cruzi I, suggesting independent transmission cycles occurring among the sylvatic mammals inside Poço das Antas Biological Reserve. Significant differences in the infection patterns presented by the 3 populations of wild and captive-born golden lion tamarins were noticed. In Poço das Antas a considerably higher number of positive haemocultures from tamarins with positive serological titres was observed in comparison to those obtained from other areas. The implications for conservation and public health of an active sylvatic cycle in the Atlantic Coastal Rainforest of Rio de Janeiro are discussed.

Animals↗
PubMed2002

Two main clusters within Trypanosoma cruzi zymodeme 3 are defined by distinct regions of the ribosomal RNA cistron.

Trypanosoma cruzi is currently classified into 2 major phylogenetic lineages, T. cruzi I and II, that correlate with the formerly described zymodeme 1 and 2, respectively. Another isoenzymic group (zymodeme 3-Z3) was also described. In this study, we analysed the genetic diversity among Z3 isolates of the Brazilian Amazon by restriction fragment length polymorphism of the intergenic transcribed spacers (ITSs) of the ribosomal RNA cistron and the size of the divergent domain D7 of the 24Salpha rRNA gene. DNAs from 12 T. cruzi Z3 isolates obtained from humans (2), Panstrongylus geniculatus (1), and Rhodnius brethesi (9) were submitted to PCR amplification of the ITSs plus the 5.8S rDNA. The PCR products were digested with 4 distinct endonucleases and the profiles analysed by a numerical methodology. The phenetic dendrogram revealed a clear dichotomy in the Z3 group, defining 2 groups that were named Z3-A and Z3-B. Dimorphism was also found in the band sizes of the amplified D7 divergent domain of the 24Salpha rDNA, which showed a perfect correlation with the ITSs clustering. The organization of the ribosomal cistron was investigated by Southern blotting and shown to be conserved in the genome of the 2 Z3 groups. This study shows that the rDNA cistron allows the definition of 2 distinct subclusters in Z3 isolates.

Animals↗
PubMed2002

The genetic diversity of Brazilian Trypanosoma cruzi isolates and the phylogenetic positioning of zymodeme 3, based on the internal transcribed spacer of the ribosomal gene.

There is a high level of genetic diversity among the clinical isolates of the taxon Trypanosoma cruzi. The results of recent studies on well defined gene loci have revealed that T. cruzi can be divided into two major phylogenetic lineages, designated T. cruzi I and T. cruzi II. Further intra-lineage polymorphisms were explored, in clinical isolates from five representative regions of Brazil, using the internal transcribed spacers that flank the 5.8S ribosomal RNA gene. These regions were amplified in a PCR and the products were digested with six restriction enzymes before being subjected to RFLP analysis in polyacrylamide gels. The results were used to construct a complex evolutionary tree, in which the isolates were clustered into two major phylogenetic lineages with a low level of similarity. To investigate the evolutionary relationships between the isolates, the PCR products were cloned and sequenced. The results of the subsequent phylogenetic analysis confirmed the presence of the two major groups of T. cruzi and showed that zymodeme 3, a third iso-enzymatic sub-group, is closer to T. cruzi I than to T. cruzi II.

Animals↗
PubMed2001

Sensitivity of the polymerase chain reaction for the diagnosis of cutaneous leishmaniasis due to Leishmania (Viannia) guyanensis.

The sensitivity of the polymerase chain reaction (PCR) in 35 consecutive outpatients with cutaneous leishmaniasis caused by Leishmania (Viannia) guyanensis was evaluated using, as gold standard, the in vitro isolation of the parasite through culture of aspirates of the cutaneous ulcers. All isolates were identified using electrophoretic enzyme analysis. Patients were mainly young males with recent onset disease without prior specific treatment. PCR was performed using DNA extracted from fresh frozen biopsies of cutaneous ulcers. The reaction used a pair of oligonucleotides that amplify the conserved region of the minicircle molecule. PCR showed 100% sensitivity (95% CI from 90.0 to 100.0). These results were similar to the visualization of amastigotes in imprint preparations of cutaneous biopsy tissue and the inoculation of biopsy material in golden hamsters. Despite the high sensitivity of the PCR, in this particular clinical setting of cutaneous leishmaniasis caused by L. (V.) guyanensis in the Brazilian Amazon, it appears that the method of choice for diagnosis should be the direct visualization of amastigotes using imprint preparations and the PCR reserved for those patients with negative imprint results.

Adolescent↗
PubMed2001

Use of DNA-based diagnostic methods for human leishmaniasis in Minas Gerais, Brazil.

DNA hybridisation was used to type 26 samples from lesions of human patients from the Rio Doce Valley (Minas Gerais, Brazil) clinically diagnosed as having cutaneous leishmaniasis, using kinetoplast DNA (kDNA) cloned mini-circle probes specific for the Leishmania mexicana and Leishmania braziliensis complexes. All samples were found to belong to the L. braziliensis complex. When biopsies were pressed directly onto touch blot membranes 38.5% of the samples were positive. The positivity and specificity obtained were both 100% when cultured blotted parasites were used. The results were confirmed by polymerase chain reaction (PCR) analysis using primers specific for the L. mexicana and L. braziliensis complexes.

Animals↗
PubMed2001

Maximum-likelihood divergence date estimates based on rRNA gene sequences suggest two scenarios of Trypanosoma cruzi intraspecific evolution.

The phylogenetic relationships of Trypanosoma cruzi strains were inferred using maximum-likelihood from complete 18S rDNA sequences and D7-24Salpha rDNA regions from 20 representative strains of T. cruzi. For this we sequenced the 18S rDNA of 14 strains and the D7-24Salpha rDNA of four strains and aligned them to previously published sequences. Phylogenies inferred from these data sets identified four groups, named Riboclades 1, 2, 3, and 4, and a basal dichotomy that separated Riboclade 1 from Riboclades 2, 3, and 4. Substitution models and other parameters were optimized by hierarchical likelihood tests, and our analysis of the 18S rDNA molecular clock by the likelihood ratio test suggests that a taxa subset encompassing all 2,150 positions in the alignment supports rate constancy among lineages. The present analysis supports the notion that divergence dates of T. cruzi Riboclades can be estimated from 18S rDNA sequences and therefore, we present alternative evolutionary scenarios based on two different views of T. cruzi intraspecific divergence. The first assumes a faster evolutionary rate, which suggests that the divergence between T. cruzi I and II and the extant strains occurred in the Tertiary period (37-18 MYA). The other, which supports the hypothesis that the divergence between T. cruzi I and II occurred in the Cretaceous period (144-65 MYA) and the divergence of the extant strains occurred in the Tertiary period of the Cenozoic era (65-1.8 MYA), is consistent with our previously proposed hypothesis of divergence by geographical isolation and mammalian host coevolution.

Animals↗
PubMed2001

Parasite persistence in treated chagasic patients revealed by xenodiagnosis and polymerase chain reaction.

Polymerase chain reaction (PCR) was compared with xenodiagnosis performed 20 years after trypanocidal chemotherapy to investigate parasite clearance. Eighty-five seropositive individuals for Chagas disease presenting a positive xenodiagnosis were treated with specific drugs; 37 in the acute phase and 48 in the chronic phase. Fifteen chronic asymptomatic patients received a placebo. Treatment in the acute phase led to PCR negative results in 73% of the cases, while xenodiagnosis was negative in 86%. In the chronic phase, PCR was negative in 65% of the patients and 83% led to xenodiagnosis negative results. Regarding the untreated group (placebo), 73% gave negative results by xenodiagnosis, of which 36% were positive by PCR. Individuals that were considered seronegative (n=10), presented unequivocally negative results in the PCR demonstrating the elimination of parasite DNA. Seventeen individuals had their antibodies titers decreased to such a level that the final results were considered as doubtful and 16 of them presented negative PCR. The molecular method represents a clear advantage over conventional techniques to demonstrate persistent infections in Chagas disease patients that underwent chemotherapy.

Acute Disease↗
PubMed2001

Short report: detection of Leishmania DNA by polymerase chain reaction on blood samples from dogs with visceral leishmaniasis.

Immunological, parasitological, and molecular techniques were applied to blood samples of dogs to diagnose Leishmania infections. In 1997, 644 domestic dogs were studied. Peripheral blood samples were collected for serological diagnosis and detection of Leishmania parasite by polymerase chain reaction (PCR). The indirect immunofluorescence test was positive in 139 (21.6%) of 644 dogs examined. The PCR was performed in 70 blood samples and 3 bone marrow aspirates. A 120-bp fragment specific for Leishmania was present in PCR hybridization analysis of all seropositive samples in the molecular assays. The PCR hybridization test, which used a minicircle of Leishmania chagasi as a probe, was negative in 20 seronegative dogs. These results suggest that a combined PCR-Southern hybridization technique is a highly sensitive approach to diagnose leishmaniasis in dogs, which are a zoonotic reservoir of leishmaniasis for humans.

Animals↗
PubMed2000

Paleoparasitology of Chagas disease revaled by infected tissues from Chilean mummies.

Mummified tissues were sampled from bodies stored at the Museo Arqueologico de San Pedro de Atacama, northern Chile, dated from 2000 years BP-1400 AD, and Trypanosoma cruzi DNA was recovered using polymerase chain reaction (PCR) methodology. Amplification of the conserved region of the minicircle molecule of T. cruzi was achieved in four of the six samples tested. Amplified products corresponding to genetic fragments of the parasite were tested by hybridization experiments with positive results for T. cruzi specific molecular probe. The origin and dispersion of T. cruzi human infection is discussed as well as the molecular paleoparasitological approach, and what it may represent in an evolutionary perspective.

Animals↗
PubMed2000

American tegumentary leishmaniasis (ATL) in Rio de Janeiro State, Brazil: main clinical and epidemiologic characteristics.

BACKGROUND: Rio de Janeiro State in Brazil is an endemic area of American tegumentary leishmaniasis (ATL) induced by Leishmania (Viannia) braziliensis. Objective Our purpose was to describe the main clinical and epidemiologic characteristics of the disease in Rio de Janeiro State. METHODS: Patients from endemic areas of Rio de Janeiro State attending the Evandro Chagas Hospital were included in the study. A general physical, dermatologic, and otorhinolaryngologic examination was performed in all patients, as well as a Leishmanin skin test. Skin biopsy specimens were obtained and utilized for touch preparations (stained with Leishman dye), culture in special media (Nicolle, Nevy and McNeal; NNN), and histopathologic examination after hematoxylin and eosin stain. Positive cultures were identified with regard to species by the isoenzyme technique. Therapy with pentavalent antimonial compounds was employed in all cases. Eco-epidemiologic characteristics were studied through regular field visits to endemic foci. RESULTS: Cutaneous disease was present in 87.2% of patients, and mucosal disease in only 12.7%. A single ulcerative cutaneous lesion was the most common clinical presentation. Demonstration of the parasite was always difficult and culture in special media gave the best results for diagnosis. The species involved in transmission was Leishmania (Viannia) braziliensis. Vectors included phlebotomine sand flies (Diptera: Psychodidae) of the genus Lutzomyia, and the most common species was Lutzomyia intermedia, captured mainly on the external walls of houses. CONCLUSIONS: ATL in Rio de Janeiro is mostly a cutaneous disease. In general, the cases showed great sensitivity to antimony. A pattern of peridomestic transmission seems to be the rule.

Animals↗
PubMed2000

Intraspecific heterogeneity in the mini-exon gene localization of Leishmania (Viannia) panamensis and Leishmania (Viannia) guyanensis from Colombia.

Intraspecific heterogeneity was demonstrated in the mini-exon gene localization from Leishmania (Viannia) panamensis and L. (Viannia) guyanensis. Different karyotypes were detected in human isolates circulating in endemic areas of Colombia. The presence of mini-exon gene sequences on chromosomes of different sizes, ranging from 370 to 800 kb in L. (V.) panamensis and from 500 to 800 kb in L. (V.) guyanensis, was observed and was neither strain nor species specific. In some cases, hybridization with 2 chromosomes in the same strain was observed. The variability of chromosomal localization of mini-exon gene sequences of these 2 species highlights the genetic variability of the Viannia subgenus and the potential utility of the mini-exon gene as a molecular epidemiologic marker.

Animals↗
PubMed2000

Short report: occurrence of Leishmania donovani DNA in donated blood from seroreactive Brazilian blood donors.

Human visceral leishmaniasis (kala-azar) transmitted by blood transfusion has been described in previous reports. Seroprevalence of antibodies to Leishmania donovani was shown to be related to prior blood transfusions in multiply transfused hemodialysis patients in Natal, Rio Grande do Norte, Brazil. In this study, a possible correlation between seroreactivity and the presence of L. donovani DNA was investigated in asymptomatic healthy blood donors. Sera were tested using the fucose mannose ligand (FML) ELISA, which was shown to have a sensitivity of 100%, a specificity of 96-100%, reliability, and diagnostic and prognostic potential for the detection of human and canine kala-azar, respectively. Leishmanial DNA was assessed by the polymerase chain reaction (PCR) and dot-blot hybridization techniques in blood and bone marrow samples. Among 21 FML-seroreactive asymptomatic blood donors, 5 (24%) were positive by the PCR and 9 (43%) were positive in a dot-blot assay of blood samples, showing a significant correlation (chi2 = 14.24, P < 0.01). No Leishmania DNA was detected in 20 FML non-reactive blood donors. Our results point to the need for control of transmission of kala-azar by blood transfusion in areas endemic for this disease.

Animals↗