The Leishmania donovani species complex: A new insight into taxonomy☆.

Among the 20 or so Leishmania spp. described as pathogenic for humans, those of the Leishmania donovani complex are the exclusive causative agents of systemic and fatal visceral leishmaniasis. Although well studied, the complex is taxonomically controversial, which hampers clinical and epidemiological research. In this work, we analysed 56 Leishmania strains previously identified as L. donovani, Leishmania archibaldi or Leishmania infantum, isolated from humans, dogs and sandfly vectors throughout their distribution area. The strains were submitted to biochemical and genetic analyses and the resulting data were compared for congruence. Our results show: i) a partial concordance between biochemical and genetic-based data, ii) very limited genetic variability within the L. donovani complex, iii) footprints of frequent genetic exchange along an east-west gradient, marked by a widespread diffusion of alleles across the geographical range, and iv) a large-scale geographical spreading of a few genotypes. From a taxonomic point of view, considering the absence of relevant terminology in existing classes, the L. donovani complex could be treated as a single entity.

Isoenzyme genotyping and phylogenetic analysis of oxacillin-resistance Staphylococcus aureus isolates

Aim: The propagation of S. aureus in hospital and dental environments is considered an important public health problem since resistant strains can cause serious infections in humans. The genetic variability of 99 oxacillin-resistant S. aureus isolates (ORSA) from the dental patients (oral cavity) and environments (air) was studied by isoenzyme genotyping. Methods: S. aureus isolates were studied using isoenzyme markers (alcohol dehydrogenase, sorbitol dehydrogenase, mannitol-1-phosphate dehydrogenase, malate dehydrogenase, glucose dehydrogenase, D-galactose dehydrogenase, glucose-6-phosphate dehydrogenase, catalase and α/ß-esterase) and genetic (Nei's statistics) and cluster analysis (UPGMA algorithm). Results: A highly frequent polyclonal pattern was observed in this population of ORSA isolates, suggesting various sources of contamination or microbial dispersion. Genetic relationship analysis showed a high degree of polymorphism between the strains, and it revealed three taxa (A, B and C) distantly genetically related (0.653≤dij≤1.432) and fifteen clusters (I to XV) moderately related (0.282≤dij<0.653). These clusters harbored two or more highly related strains (0≤dij<0.282), and the existence of microevolutionary processes in the population of ORSA. Conclusion: This research reinforces the hypothesis of the existence of several sources of contamination and/or dispersal of ORSA of clinical and epidemiologically importance, which could be associated with carriers (patients) and dental environmental (air) (AU)

Molecular identification of an old clinical isolate of Indian Kala-azar.

Molecular characterization is an important task for species identification of the isolates belonging to different Leishmania species. Clinical symptoms, tissue tropism, vector preference, reservoir and geographical distribution may act as distinguishing parameters but not always decisive. On the other hand, modern taxonomic tools employed to divulge characteristics of the genome or protein molecules of the parasite would be convincing and for Leishmania sp., they include nuclear and kDNA buoyant density, multi locus enzyme electrophoresis (MLEE), RAPD, RFLP or use of monoclonal antibodies etc. In the present study, we intend to establish the identification of an old clinical isolate of Indian Kala-azar, familiarly known as 'UR6' by MLEE, RAPD, RFLP and species specific monoclonal antibodies. UR6 has been isolated from a confirmed Kala-azar patient admitted in Calcutta School of Tropical Medicine, Kolkata in 1978. From then it is being regularly used for various scientific studies by the Leishmania Research Group of India and abroad. The isozyme profile of UR6 showed similar electrophoretic mobility that of WHO reference strain for Leishmania tropica, K27. Similar findings were obtained in the RAPD and RFLP assays. Screening with species specific monoclonal antibodies showed its strong reactivity towards L. tropica. The Jaccard's Similarity Indices were calculated.

Multilocus enzyme electrophoresis analysis of rapidly-growing mycobacteria: an alternative tool for identification and typing.

OBJECTIVES: Rapidly growing mycobacteria (RGM) have emerged as important pathogens in clinical settings, associated with esthetic procedures and postsurgical infections, pulmonary infections among cystic fibrosis patients, and other structural pulmonary diseases. Microorganisms belonging to Mycobacterium abscessus-Mycobacterium chelonae and to Mycobacterium fortuitum groups have frequently been associated with outbreaks and various epidemics. In the present study, RGM strains were characterized in order to investigate molecular markers based on proteomic analysis. METHODS: Multilocus enzyme electrophoresis (MLEE) was used for species identification and clonal analysis of RGM recovered from postsurgical wound infections during an epidemic. The study included 30M. abscessus subsp. bolletii clinical isolates, most belonging to the BRA100 clone (epidemic in Rio de Janeiro city), as well as 16 RGM ATCC reference strains. RESULTS: Molecular typing allowed the detection of diversity in the studied population and revealed species-specific isoenzymatic patterns. Additionally, the clonal relationship among M. abscessus subsp. bolletii outbreak isolates, as examined using MLEE, was markedly consistent. CONCLUSIONS: Isoenzymatic characterization was found to be a useful molecular tool to identify RGM species and to determine the relatedness among closely related M. abscessus subsp. bolletii isolates. This may be considered a powerful approach for epidemiological studies on RGM.

Evaluation of Leishmania (Leishmania) chagasi strains isolated from dogs originating from two visceral leishmaniasis-endemic areas in Brazil using multilocus enzyme electrophoresis / Avaliação de amostras de Leishmania (Leishmania) chagasi isoladas de cães oriundos de duas áreas endêmicas de leishmaniose visceral no Brasil através da eletroforese de isoenzimas

INTRODUCTION: Domestic dogs are the most important reservoir in the peridomestic transmission cycle of Leishmania (Leishmania) chagasi. The genetic variability of subpopulations of this parasite circulating in dogs has not been thoroughly analyzed in Brazil, even though this knowledge has important implications in the clinical-epidemiological context. METHODS: The objective of this study was to evaluate and compare the phenotypic variability of 153 L. chagasi strains isolated from dogs originating from the municipalities of Rio de Janeiro (n = 57) and Belo Horizonte (n = 96), where the disease is endemic. Strains isolated only from intact skin were selected and analyzed by multilocus enzyme electrophoresis using nine enzyme systems (6PG, GPI, NH1 and NH2, G6P, PGM, MDH, ME, and IDHNADP). RESULTS: The electrophoretic profile was identical for all isolates analyzed and was the same as that of the L. chagasi reference strain (MHOM/BR/74/PP75). Phenetic analysis showed a similarity index of one for all strains, with the isolates sharing 100 percent of the characteristics analyzed. CONCLUSIONS: The results demonstrate that the L. chagasi populations circulating in dogs from Rio de Janeiro and Belo Horizonte belong to a single zymodeme.

INTRODUÇÃO: Cães domésticos são considerados os reservatórios mais importantes no ciclo peridoméstico de transmissão de Leishmania (Leishmania) chagasi. No entanto, a variabilidade genética de sub-populações que circulam neste hospedeiro é ainda pouco explorada no Brasil, sendo tal conhecimento de grande importância no contexto clínico-epidemiológico. MÉTODOS: O objetivo deste estudo foi avaliar e comparar a variabilidade fenotípica de 153 amostras de L. chagasi isoladas de cães oriundos dos municípios do Rio de Janeiro (n = 57) e Belo Horizonte (n = 96), onde a doença é endêmica. Foram selecionadas somente amostras isoladas de pele íntegra e analisadas por eletroforese de isoenzimas (MLEE) empregando nove sistemas enzimáticos (6PG, GPI, NH1 e NH2, G6P, PGM, MDH, ME, IDHNADP). RESULTADOS: Todas as amostras analisadas apresentaram perfil eletroforético idêntico entre si e com a amostra de L. chagasi utilizada como referência neste estudo (MHOM/BR/74/PP75). A análise fenética demonstrou índice de similaridade igual a um para todas as amostras, revelando um compartilhamento de 100 por cento dos caracteres avaliados. CONCLUSÕES: A partir desses resultados, podemos inferir que as populações de L. chagasi que estão circulando nos cães do Rio de Janeiro e Belo Horizonte podem ser agrupadas em um único zimodema.

Typing Candida albicans oral isolates from healthy Brazilian schoolchildren using multilocus enzyme electrophoresis reveals two highly polymorphic taxa

The genetic diversity of C. albicans oral isolates from 75 healthy schoolchildren from eight schools located in different geographic areas of Piracicaba city, São Paulo state, Brazil, was established using isoenzymes marker (Multilocus Enzyme Electrophoresis - MLEE) and cluster analysis. Patterns of monoclonal and polyclonal oral colonization by C. albicans within and between groups of schoolchildren were identified. However, significant divergence between the observed and the expected genotypic frequencies (Hardy-Weinberg equilibrium test) was not detected in the geographically adjacent groups, suggesting the hypothesis that populations of healthy schoolchildren do not correspond to the selection factor (differential survival) of strains. Two highly polymorphic and distantly genetically related taxa (A and B) were identified within the total population of yeasts, each contained subgroups (A1, A2, A3, A4, B1 and B2) and clusters of moderately related strains (from I to X), suggesting the existence of strains restricted or not to certain groups of geographically limited, healthy students. However, the coexistence of identical strains in healthy schoolchildren from the same school (geographically related) reinforces the hypothesis of oral transmission, where the sources of propagation could be explored. Furthermore, this could also be used in current and retrospective analyses of C. albicans isolated from immunocompetent and immunocompromised people, in order to detect commensal or potentially pathogenic yeast groups, predominantly in candidiasis, and in the development of strategies to prevent transmission or human propagation.

Typing Candida albicans oral isolates from healthy brazilian schoolchildren using multilocus enzyme electrophoresis reveals two highly polymorphic taxa.

The genetic diversity of C. albicans oral isolates from 75 healthy schoolchildren from eight schools located in different geographic areas of Piracicaba city, São Paulo state, Brazil, was established using isoenzymes marker (Multilocus Enzyme Electrophoresis - MLEE) and cluster analysis. Patterns of monoclonal and polyclonal oral colonization by C. albicans within and between groups of schoolchildren were identified. However, significant divergence between the observed and the expected genotypic frequencies (Hardy-Weinberg equilibrium test) was not detected in the geographically adjacent groups, suggesting the hypothesis that populations of healthy schoolchildren do not correspond to the selection factor (differential survival) of strains. Two highly polymorphic and distantly genetically related taxa (A and B) were identified within the total population of yeasts, each contained subgroups (A1, A2, A3, A4, B1 and B2) and clusters of moderately related strains (from I to X), suggesting the existence of strains restricted or not to certain groups of geographically limited, healthy students. However, the coexistence of identical strains in healthy schoolchildren from the same school (geographically related) reinforces the hypothesis of oral transmission, where the sources of propagation could be explored. Furthermore, this could also be used in current and retrospective analyses of C. albicans isolated from immunocompetent and immunocompromised people, in order to detect commensal or potentially pathogenic yeast groups, predominantly in candidiasis, and in the development of strategies to prevent transmission or human propagation.

Genetic relationships among rhizopine-producing Rhizobium strains.

Chromosomal and symbiosis-related genotypes of rhizopine-producing and non-producing isolates of Rhizobium meliloti and Rhizobium leguminosarum were examined by multilocus enzyme electrophoresis and RFLP. The distribution of rhizopine production in both species was found to be independent of host genotype. Conversely, rhizopine production was associated with particular symbiotic plasmid types. This association may explain the observed distribution of rhizopine production in R. leguminosarum and R. meliloti. Rhizopine synthesis (mos) genes showed greater sequence divergence than rhizopine catabolism (moc) genes in both R. meliloti and R. leguminosarum. Furthermore, mos and moc genes were less divergent in R. leguminosarum than R. meliloti, suggesting a more recent evolution in the former species.