PubMed Health⌕ Search

Biomedical subjects

R S Pacheco

Publications and source records attributed to R S Pacheco.

At least 19 recordsLinked to original sources
PubMed2005

Mixed infection with Leishmania (Viannia) braziliensis and Leishmania (Leishmania) chagasi in a naturally infected dog from Rio de Janeiro, Brazil.

We report here the first case of co-infection with Leishmania (Viannia) braziliensis and Leishmania (Leishmania) chagasi in a naturally infected dog from Rio de Janeiro, Brazil. Isoenzyme characterisation identified the parasites isolated in culture from the cutaneous lesion as L. (V.) braziliensis and the isolates from blood and lymph node as L. (L.) chagasi. PCR analysis using specific primers followed by molecular hybridisation for direct Leishmania species identification in tissue fragments confirmed the presence of L. (V.) braziliensis DNA in the cutaneous lesion and of L. (L.) chagasi DNA in spleen and popliteal lymph node fragments. This report emphasises the importance of identification of Leishmania species infecting seropositive dogs in endemic areas, and the consequent re-assessment of control and epidemiological surveillance measures for the control of leishmaniasis, as is the case in Brazil.

Animals↗
PubMed2004

Long-term follow-up of co-infected HIV and Trypanosoma cruzi Brazilian patients.

Three cases of Trypanosoma cruzi-HIV co-infected haemophiliacs are described. Parasitological (xenodiagnosis, haemoculture, PCR) and immunological (CD4+ and CD8+ T cell counts, in vitro lymphoproliferative responses) studies were performed. Hybridization of isolated parasites with a specific probe confirmed the T. cruzi aetiology. We observed that despite the high parasitaemia, no clinical or parasitological evidence of T. cruzi reactivation was detected. CD4+ T cells decreased with time in two patients and the lymphocyte proliferative response to T. cruzi was very low in all patients. These data suggest that T. cruzi infection may have a long silent course in immunosuppressed HIV patients. Therefore, this parasitic infection should be investigated in any AIDS patient coming from areas endemic for Chagas' disease.

Adult↗
PubMed2003

RAPD analyses and rDNA intergenic-spacer sequences discriminate Brazilian populations of Triatoma rubrovaria (Reduviidae: Triatominae).

Triatoma rubrovaria, a member of the 'infestans' subgroup, is a potential vector of Trypanosoma cruzi in southern Brazil. Surveillance data indicate a growing domiciliary and peridomiciliary invasion by Tri. rubrovaria in the rural areas of Rio Grande do Sul (RS). In fact, following effective control of Tri. infestans, Tri. rubrovaria, which seems to have pre-adaptative characteristics for anthropic ecotopes, has become the most frequent species of triatomine bug to be collected in these areas. To explore the intraspecific variability and domiciliation of Tri. rubrovaria, the ribosomal DNA (rDNA) of two RS populations of Tri. rubrovaria that were geographically separated by only 220 km was investigated. The RAPD profiles and nucleotide sequences of the intergenic region of the rDNA, including the internal transcribed spacers 1 and 2 (ITS-1 and ITS-2) and the 5.8S gene, were analysed. In the RAPD study, the use of three decameric primers revealed polymorphisms reflecting both genetic differences between the two populations and heterogeneity within each. A phenetic dendrogram of the Tri. rubrovaria specimens, based on the three-primer consensus and a simple-matching coefficient of similarity, showed two clusters, clearly differentiating the bugs from the two localities studied. The rDNA sequencing revealed four different nucleotide sequences, with two different genotypes in each locality. The level of intraspecific variability detected within ITS-1 and ITS-2 of the Tri. rubrovaria, which was remarkably high considering the physical closeness of the two populations sampled, may indicate that the two collection sites are separated by geographical barriers that ensure the reproductive isolation of each population. The ITS sequences, like the RAPD results, clearly distinguished the two populations while showing that there is heterogeneity within each of them. The present study appears to be the first to reveal ITS length differences between populations of the same triatomine species without any associated difference in the number of microsatellite repeats. These results are in agreement with those of earlier studies on iso-enzymes, chromatic patterns, the ecological effects of environmental modification by humans, and bloodmeal sources.

Animals↗
PubMed2002

Epidemiological studies of an outbreak of cutaneous leishmaniasis in the Rio Jequitinhonha Valley, Minas Gerais, Brazil.

We detected an outbreak of American cutaneous leishmaniasis in the Jequitinhonha River Valley, Minas Gerais, Brazil. Clinical and epidemiological aspects were studied for a period of two years. Data include results of physical examinations, Montenegro skin test and serology. In total 72 of the 299 individuals evaluated presented active lesions. Only one case out of these 72 patients showed the mucosal form of the disease. The precarious sanitary conditions, low educational level and low income found in the population studied demonstrated that, as with the other parasitic diseases, cutaneous leishmaniasis occurs with greater frequency in needy populations. A canine serological survey detected 20.3% (30/148) of dogs reactive to the Leishmania antigen. Lutzomyia intermedia was the predominant phlebotomine species and the majority of the specimens (84.9%) were captured in the peridomicile. Four samples from human and three from canine cases were isolated and characterised by PCR and isoenzymes as being Leishmania (Viannia) braziliensis. The peridomiciliary nature of the disease is discussed.

Adolescent↗
PubMed2001

Visceral leishmaniasis in the Metropolitan Region of Belo Horizonte, State of Minas Gerais, Brazil.

In the last few years the number of human cases of American visceral leishmaniasis in the Metropolitan Region of Belo Horizonte (MRBH), Minas Gerais, Brazil has increased, indicating an elevation in the transmission rate of the disease. The total number of notified human cases in the MRBH since 1994, when the first case was identified, up to 1999 was 345 of which 223 (65%) were from the city itself, indicating an urbanization of the disease in this region of Minas Gerais. The age distribution of visceral leishmaniasis cases in the MRBH shows a higher prevalence in children from 0-4 years old, responsible for 28.9% of the notifications. Clinical and immunological findings from dogs infected with Leishmania chagasi are described. The majority of these animals showed no sign of the disease. Sera from all infected dogs showed detectable Leishmania-induced high titles of antibodies based on the results of an indirect fluorescent antibody test. Samples of isolated Leishmania from human and dogs were characterized as L. (L.) chagasi by biochemical and molecular techniques.

Adolescent↗
PubMed2001

Leishmanial antigens in the diagnosis of active lesions and ancient scars of American tegumentary leishmaniasis patients.

Cutaneous biopsies (n = 94) obtained from 88 patients with American tegumentary leishmaniasis were studied by conventional and immunohistochemical techniques. Specimens were distributed as active lesions of cutaneous leishmaniasis (n = 53) (Group I), cicatricial lesions of cutaneous leishmaniasis (n = 35) (Group II) and suggestive scars of healed mucosal leishmaniasis patients (n = 6) (Group III). In addition, active cutaneous lesions of other etiology (n = 24) (Group C1) and cutaneous scars not related to leishmaniasis (n = 10) (Group C2) were also included in the protocol. Amastigotes in Group I biopsies were detected by routine histopathological exam (30.2%), imprint (28.2%), culture (43.4%), immunofluorescence (41.4%) and immunoperoxidase (58.5%) techniques; and by the five methods together (79.3%). In Group II, 5.7% of cultures were positive. Leishmanial antigen was also seen in the cytoplasm of macrophages and giant cells (cellular pattern), vessel walls (vascular pattern) and dermal nerves (neural pattern). Positive reaction was detected in 49 (92.5%), 20 (57%) and 4 (67%) biopsies of Groups I, II and III, respectively. Antigen persistency in cicatricial tissue may be related to immunoprotection or, on the contrary, to the development of late lesions. We suggest that the cellular, vascular and neural patterns could be applied in the immunodiagnosis of active and cicatricial lesions in which leishmaniasis is suspected.

Adult↗
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↗
PubMed1998

N-methylglucamine antimonate (SbV+): intralesional canine tegumentary leishmaniasis therapy.

Twenty five adult dogs of three municipalities of the State of Rio de Janeiro, Brazil, that had been naturally infected by L. (V) braziliensis were treated with N-methylglucamine antimonate (Glucantime). Nine of the animals (36.0%) presented ulcerated skin lesions, twelve (48.0%) had mucosal lesions and four (16.0%) had multiple lesions. In some cases the mucosal lesions were associated to skin lesions. A dose of 85 mg SbV+ or 1 ml of the drug was intralesionally administered to the dogs. The animals were divided into three groups according to the amount of necessary doses (between one and three) for the complete healing of the lesions. The dogs were observed for six months after the third group received the last dose. Within this period two animals perished. Serial antibody evaluation through IFAT has shown that in 14 samples (63.3%) the titers have remained unaltered, in four of them (16%) there has been a decrease in two titers and in five of them (21%) serology was negative. Nineteen of the dogs (86.6%) had their lesions completely healed. The authors suggest intralesional therapy be the first choice of treatment of canine tegumentary leishmaniasis due to its effectiveness.

Animals↗
PubMed1998

Cutaneous scars in American tegumentary leishmaniasis patients: a site of Leishmania (Viannia) braziliensis persistence and viability eleven years after antimonial therapy and clinical cure.

Two former patients treated for the cutaneous form of American tegumentary leishmaniasis were reviewed eight and 11 years, respectively, following clinical cure. We were able to isolate Leishmania parasites in a culture of material from the two scar biopsies, and in one of them the parasite was characterized as Leishmania (Viannia) braziliensis. In both cases, the histopathology revealed discreet hyperceratosis and a slight infiltrate of mononuclear cells surrounding and on the walls of the surface and deep dermal vessels. No amastigotes were seen on immunohistochemical or histopathologic examination. The Montenegro skin test result and the in vitro lymphoproliferative response to Leishmania antigen were positive, but no specific IgG and IgM antibodies were detected. Otorhinolaryngologic examination showed no macroscopic alteration in the mucosae. These findings are important for the evaluation and criteria of post-treatment cure.

Adult↗
PubMed1995

Genotypic polymorphisms in experimental metastatic dermal leishmaniasis.

Molecular karyotype and kDNA restriction analyses were utilized to examine the genetic heterogeneity and plasticity of the Leishmania (Viannia) guyanensis strain WHI/BR/78/M5313, composed of metastatic and non-metastatic populations. Cloning revealed that the strain was constituted by multiple closely related populations that were distinguishable by restriction fragment polymorphisms in kDNA. Size polymorphisms in molecular karyotype were not detected. Passage of clones in hamsters and recovery of parasites from cutaneous metastatic lesions yielded evidence of further genetic heterogeneity among some of the progeny populations. Overall, six kDNA minicircle restriction patterns or schizodemes were observed among clones, subclones and progeny. Although the possibility that population heterogeneity was not resolved by cloning cannot be ruled out, subcloning and kDNA restriction analysis to determine whether the putative clones consisted of homogeneous populations showed the schizodeme of subclones of 3 out of 4 clones to be identical to the clone of origin, while a subclone of the fourth had a co-efficient of similarity of 0.95. Metastasis did not segregate with a particular schizodeme: all six restriction profiles were represented among populations isolated from metastatic lesions and some clones with the same restriction profile did not produce metastatic lesions. The strain from which the clones, subclones and progeny were derived had a kDNA restriction pattern identical to the most prevalent schizodeme (38%) among these subpopulations. This finding together with the reappearance of the repertoire of schizodemes found among clones in the populations recovered from metastatic lesions in hamsters inoculated with a single clone, suggest that sequence polymorphisms in kDNA can emerge during infection.

Animals↗
PubMed1994

Naturally acquired infections with Leishmania enriettii Muniz and Medina 1948 in guinea-pigs from São Paulo, Brazil.

Two domestic guinea-pigs (Cavia porcellus), bought in Pinheros, São Paulo State, Brazil, were taken by their owners to a farm in the rural district of Capão Bonito, close to the Atlantic Forest, São Paulo, where they both developed tumour-like and ulcerating lesions on the ears. The causative agent was identified as Leishmania (L.) enriettii, based on biological characters and isoenzyme profiles. Sources of the parasite in wild mammals, and the possible sandfly vector species are discussed.

Animals↗
PubMed1993

Molecular evidence for the importation of Old World Leishmania into the Americas.

A brief review of work carried out by the laboratory on the systematics of trypanosomatids during the last three years is given. The principal line of research has been on the taxonomy of New World Leishmania and one of the topics studied has been the determination of the autochthonous origin of certain Leishmania species found in the New World. Controversy exists as to whether the etiological agent of American Visceral Leishmaniasis is indigenous. Here, we present evidence from enzyme electrophoresis and schizodeme analysis indicating that L. chagasi has a recent origin and that it is similar to L. infantum. We also describe L. major-like isolates which have been found in the New World and present evidence suggesting that some of these populations may have been imported into the Americas. Reference strains from the subgenus Viannia are examined and compared with other Old World and New World species by enzyme electrophoresis. The results are analyzed numerically and we show that the Viannia species are a group of parasites indigenous to the New World that cluster separately from other Leishmania species. The numerical analyses also indicate that the subgenus forms a monophyletic group in contrast to the subgenus Leishmania which appears to be polyphyletic.

Americas↗
PubMed1990

Population heterogeneity among clones of New World Leishmania species.

Cell cloning techniques and schizodeme analysis were used to detect mixtures of subpopulations in Leishmania parasites, isolated from humans and a reservoir host. Clones were obtained by plating promastigotes, at limited dilution, on solid medium. The resultant colonies were analysed by the restriction profile of kinetoplast DNA minicircles (schizodeme analysis). The efficiency of plating was around 40-60%, and subpopulations of different schizodemes, in stocks of Leishmania isolated from 2 cases of human cutaneous leishmaniasis were detected. The presence of different schizodemes in one isolate from a sylvatic animal suggested the possibility of a mixed natural infection.

Animals↗
PubMed1986

Concurrent human infection with Leishmania donovani and Leishmania braziliensis braziliensis.

In the suburban district of Campo Grande, Municipality of Rio de Janeiro (Brazil) cases of both Leishmania domovani and L. b. braziliensis infections occur. In March 1982 we examined a white male child, five years old, with fever, weight loss and distended abdomen, symptoms said to have started three months before. After the first month a papular lesion appeared on his forehead, rapidly increasing in size and becoming ulcerated: Parasites were isolated from both the ulcer and the bone marrow by culturing in BHI-agar with rabbit blood and a liquid overlay of modified LIT medium. Using the techniques of zymodeme analysis, schizodeme analysis and serodeme analysis using monoclonal antibodies, the parasite from the ulcer was identified as L. b. braziliensis and that from the bone marrow as L. donovani. This seems to be the first described case of concurrent infection of man with a visceral and cutaneous Leishmania. It indicates that, at least in humans, a previous infection with L. donovani does not protect against L. b. braziliensis. This result has important implications for the development of vaccines against leishmaniases.

Animals↗
PubMed1985

Brazilian Leishmania stocks phenotypically similar to Leishmania major.

Screening by enzyme electrophoresis of isolates of New World Leishmania from different geographic areas revealed a number of stocks with enzyme profiles different from those produced by reference strains of described subspecies of L. mexicana, L. braziliensis, and L. donovani. Analysis by six enzymes (aspartate aminotransferase; alanine aminotransferase; malate dehydrogenase; glucose-6-phosphate dehydrogenase; phosphoglucomutase; and glucose-phosphate isomerase) showed that these stocks have identical enzyme profiles and form a distinct zymodeme grouping. These observations were confirmed using the technique of schizodeme analysis and by comparing the k-DNA fingerprints produced by the restriction enzymes MspI, BspRI and AluI. The stocks were further analyzed by monoclonal antibodies and did not react with any of a large panel of L. mexicana, L. braziliensis, and L. donovani species- and/or subspecies-specific monoclonal antibodies using either an indirect radioimmune binding assay or immunofluorescence. These stocks did, however, react with a panel of monoclonal antibodies specific for L. major (formerly L. tropica major). Furthermore, the stocks could not be differentiated from L. major reference strains by enzyme electrophoresis nor could they be distinguished qualitatively from L. major based on their reactivity patterns using 10 Old World cutaneous species- and subspecies-specific monoclonal antibodies. Kinetoplast DNA restriction enzyme profiles, however, were different between these stocks and L. major reference strains. The implications of these results are discussed including the existence of other L. major-like stocks currently misidentified or uncharacterized.

Adult↗
PubMed1984

Schizodeme and zymodeme characterization of Leishmania in the investigation of foci of visceral and cutaneous leishmaniasis.

Leishmania parasites were isolated from humans and canines in foci of cutaneous and visceral leishmaniasis. After in vitro cultivation the parasites were examined by the following biochemical techniques: (i) restriction analysis of kinetoplast DNA (kDNA) also known as schizodeme analysis (Morel et al., 1980); (ii) zymodeme analysis (Barret et al., 1980); by agarose gel electrophoresis and (iii) isoelectricfocusing in polyacrylamide gels. The strains of cutaneous and visceralizing leishmanias studied could be differentiated by schizodeme analysis, using the endonuclease MspI, into three complexes agreeing with those accepted for human New World leishmaniasis. In the municipality of Rio de Janeiro, isolates from a focus of cutaneous leishmaniasis were identified as L. braziliensis braziliensis and from a focus of visceral leishmaniasis were identified as L. donovani by zymodeme characterization. Identical restriction enzyme profiles of kDNA from human and canine isolates indicated that in the cutaneous focus at Jacarepaguá, Rio de Janeiro, the same strain was probably circulating in both the canine and human populations. This suggests a possible role for dogs as a reservoir host for L. braziliensis braziliensis. In addition, our results confirm the importance of dogs as reservoirs in visceral leishmaniasis. The stability of the electrophoretic patterns of restriction digest ("fingerprints") of Leishmania kDNA as well as differences in the sensitivity of the techniques used were demonstrated. Strains from widely different geographical areas as well as strains maintained in vivo and in vitro showed identical kDNA restriction patterns, while strains showing similar banding patterns by enzyme electrophoresis could be differentiated by schizodeme analysis. These results demonstrate the usefulness of an integrated biochemical approach in the identification of Leishmania.

Animals↗