Utilidad de las herramientas moleculares para la identificación de Leptospira spp. en muestras humanas, animales y ambientales / Usefulness of molecular tools to identify Leptospira spp. in human, animal and environmental samples

Leptospirosis es una enfermedad zoonótica endémica de potencial epidémico que afecta la salud pública y la producción pecuaria alrededor del mundo. Su agente etiológico es una espiroqueta del género Leptospira, con 20 especies reportadas hasta el momento, son las más importantes Leptospira interrogans (patógena) y Leptospirabiflexa (saprófita). Esta bacteria se transmite mediante contacto directo o indirecto en especial con orinade animales infectados, es la transmisión por medio del agua una de las más importantes. En cuanto al diagnóstico se ha evidenciado que diversas pruebas moleculares tienen una alta especificidad y sensibilidad; sin embargo, el conocimiento de la epidemiología de la leptospirosis se ha basado principalmente en estudios serológicos que han utilizado la prueba de aglutinación microscópica que presenta debilidades en sus resultados e interpretación. El objetivo del presente artículo es presentar una revisión actualizada sobre la utilidad de las herramientas moleculares para la identificación de Leptospira spp. en muestras humanas, animales y ambientales. Se llevó a cabo una búsqueda de literaturaen diferentes bases de datos como Pubmed, Science Direct, SciELO, Scopus y Redalyc.Las publicaciones encontradas fueron artículos originales y de revisión, entre otros, publicados entre 1965 y 2014. Se determinó que las herramientas moleculares permiten una identificación directa, rápida, definitivay precisa del agente etiológico, apoyan el diagnóstico, aportan al conocimiento real de laprevalencia e incidencia de la enfermedad. Las herramientas moleculares permiten la identificación de nuevas especies a partir de aislamientos obtenidos de diversas fuentes y ayudan a orientar los programas de prevención y control de esta zoonosis(AU)

Leptospirosis is an endemic and potentially epidemic zoonosis affecting public health and livestock production worldwide. Its etiological agent is a spirochaete of the genus Leptospira, with 20 species reported to date, of which Leptospira interrogans (pathogenic) and Leptospira biflexa (saprophyte) are the most important. This bacterium is transmitted by direct or indirect contact with urine from infected animals, so water is one of the major transmission ways. Regarding diagnosis, many molecular tests have been evinced to have high specificity and sensitivity; however, knowledge on the epidemiology of leptospirosis has been based mainly on serological studies using the microscopic agglutination test, which has weaknesses in its results and interpretation. The aim of this article is to present an update review on the usefulness of molecular tools in the identification of Leptospira spp. in human, animal and environmental samples. A literature search was conducted in different databases such as PubMed, ScienceDirect, SciELO, Scopus and Redalyc. The publications found were original and review articles, among others, published between 1965 and 2014. It was found that the molecular tools allow direct, quick, definitive and precise identification of the etiologic agent, support the diagnosis, and contribute to real knowledge on the disease prevalence and incidence. Molecular tools enable the identification of new species isolates obtained from various sources and help guide prevention programs and control of this zoonosis(AU)

Multi-locus Genotyping Reveals High Occurrence of Mixed Assemblages in Giardia duodenalis within a Limited Geographical Boundary.

Aim: To determine the common genotypes of Giardia duodenalis causing diarrhea in the study region and to assess the extent of genetic polymorphism among them. Study Design: Stool samples were collected from the patients attending IDBG Hospital, Kolkata with diarrheal complaints through a systemic sampling technique and were screened for Giardia duodenalis. The G. duodenalis positive samples were subjected to molecular genotyping through ‘PCR - Direct DNA sequencing’ procedure. All the sequence data obtained were incorporated into MEGA 4 software for multiple alignment and validation followed by phylogenetic analysis. The genotyping data obtained are stored in Excel spreadsheets and incorporated into EpiInfo 3.1 for analyzing possible association of genotype outcome with common physical factors such as age, sex etc. Place and Duration of Study: Department of parasitology, National Institute of Cholera and Enteric Diseases, Kolkata, India from July 2009 to November 2011. Methodology: A total of 68 Giardia duodenalis positive stool samples were identified from the diarrhea patients attending IDBG hospital in the city and were subjected to multi-locus genotyping. Fragments of ß-giardin, Glutamate-dehydrogenase and Triosephosphateisomerase genes of Giardia were amplified from those samples with specific primers and sequenced. All the sequences were analyzed using MEGA 4 software for obtaining the genotyping results. Results: Multi-locus genotyping identified 13 isolates as assemblage A and 41 as assemblage B, whereas 14 of them could not be assigned in a particular group. Detailed phylogenetic analysis revealed that multiple genotypes were observed in those 14 isolates depending upon the marker loci. Conclusion: The study could produce a preliminary idea about the G. duodenalis genotypes found in Kolkata city. High percentage of mixed assemblages in the study population also revealed the presence of genetic diversity among a small population of diarrheal patient within a limited geographical boundary. It has also hypothesized the possibility of inter-assemblage genetic exchange among Giardia.