ABSTRACT
Objective: To determine an algorithm for molecular diagnosis of visceral leishmaniasis (VL) by kinetoplast DNA (kDNA) (RV1/ RV2) and internal transcriber spacer (ITS1) (LITSR/L5.8S) polymerase chain reaction (PCR), complemented by ITS1 PCR restriction fragment length polymorphism (RFLP), using peripheral blood or bone marrow aspirate from patients with suspected VL. Methods: Biological samples were submitted to the gold standard for the diagnosis of VL and molecular diagnosis represented by ITS1 PCR, kDNA PCR, and ITS1 PCR RFLP. The samples were obtained from seven groups: group I, 82 samples from patients with confirmed VL; group H , 16 samples from patients under treatment for VL; groupII, 14 samples from dogs with canine visceral leishmaniasis (CVL); group II, a pool of six experimentally infected sandflies (Lutzomya longipalpis); group IV, 18 samples from patients with confirmed tegumentary leishmaniasis (TL) and groups VI and VII were from control groups without VII. Results: The following gold standard and molecular examination results were obtained for each of the seven groups: group I : parasitologic and immunochromatographic tests showed a sensitivity of 76.3% (61 of 80) and 68.8% (55 of 80), respectively, and a sensitivity of 97.6% (80 of 82) and 92.7% (76 of 82) by ITS1 and kDNA PCR, respectively. After ITS1 PCR RFLP (Hae III) analysis of the 80 positive samples, 52.5% (42 of 80) generated three fragments of 180, 70, and 50 bp, corresponding to the pattern of Leishmania infantum infantum; group Π : negative for the parasitologic methods and positive for IrK39 (100%, 16 of 16), presented 12.5% (2 of 16) of positivity by ITS1 PCR and 25.0% (4 of 16) by kDNA PCR; group III: positive in the parasitologic and serologic tests (100%, 14 of 14), presented 85.7%(12 of 14) of positivity by ITS1 PCR and kDNA PCR. ITS1 PCR RFLP showed that 83.3% (10 of 12) of the canine samples contained parasites with profiles similar to L. infantum; groupIVpresented amplifications by ITS1 PCR and kDNA PCR. ITS1 PCR products were analyzed by RFLP, generating a profile similar to that of L. infantum; group V: positive in the parasitologic examination (100%, 18 of 18), presented 72.2% (13 of 18) of the samples by ITS1 PCR positive. A total of 69.2% (9 of 13) showed profiles corresponding to a Viannia complex by ITS1 PCR RFLP; and group VI and group W were negative by ITS1 and kDNA molecular tests. Comparing the molecular results with the parasitologic and serologic diagnosis from group I, almost perfect agreement was found ( κ both>0.80, P<0.001). ITS1 and RV1/RV2 PCR detected 90.2% (74 of 82) of the samples. Two samples positive by RV1/RV2 were negative by LITSR/L5.8S, and six samples positive by LITSR/L5.8S were negative by RV1/RV2. Therefore, these two systems complemented each other; they diagnosed 100% of the samples as belonging to the Leishmania genus. Conclusions: We suggest an algorithm for the molecular diagnosis of VL, which must consider previous parasitologic and serologic (immunochromatographic) diagnoses, and should combine kDNA and ITS1 to determine the Leishmania subgenus using RFLP as a complement method to define the L. infantum species.
ABSTRACT
Objective: To determine an algorithm for molecular diagnosis of visceral leishmaniasis (VL) by kinetoplast DNA (kDNA) (RV1/ RV2) and internal transcriber spacer (ITS1) (LITSR/L5.8S) polymerase chain reaction (PCR), complemented by ITS1 PCR restriction fragment length polymorphism (RFLP), using peripheral blood or bone marrow aspirate from patients with suspected VL. Methods: Biological samples were submitted to the gold standard for the diagnosis of VL and molecular diagnosis represented by ITS1 PCR, kDNA PCR, and ITS1 PCR RFLP. The samples were obtained from seven groups: group I, 82 samples from patients with confirmed VL; group H , 16 samples from patients under treatment for VL; groupII, 14 samples from dogs with canine visceral leishmaniasis (CVL); group II, a pool of six experimentally infected sandflies (Lutzomya longipalpis); group IV, 18 samples from patients with confirmed tegumentary leishmaniasis (TL) and groups VI and VII were from control groups without VII. Results: The following gold standard and molecular examination results were obtained for each of the seven groups: group I : parasitologic and immunochromatographic tests showed a sensitivity of 76.3% (61 of 80) and 68.8% (55 of 80), respectively, and a sensitivity of 97.6% (80 of 82) and 92.7% (76 of 82) by ITS1 and kDNA PCR, respectively. After ITS1 PCR RFLP (Hae III) analysis of the 80 positive samples, 52.5% (42 of 80) generated three fragments of 180, 70, and 50 bp, corresponding to the pattern of Leishmania infantum infantum; group Π : negative for the parasitologic methods and positive for IrK39 (100%, 16 of 16), presented 12.5% (2 of 16) of positivity by ITS1 PCR and 25.0% (4 of 16) by kDNA PCR; group III: positive in the parasitologic and serologic tests (100%, 14 of 14), presented 85.7%(12 of 14) of positivity by ITS1 PCR and kDNA PCR. ITS1 PCR RFLP showed that 83.3% (10 of 12) of the canine samples contained parasites with profiles similar to L. infantum; groupIVpresented amplifications by ITS1 PCR and kDNA PCR. ITS1 PCR products were analyzed by RFLP, generating a profile similar to that of L. infantum; group V: positive in the parasitologic examination (100%, 18 of 18), presented 72.2% (13 of 18) of the samples by ITS1 PCR positive. A total of 69.2% (9 of 13) showed profiles corresponding to a Viannia complex by ITS1 PCR RFLP; and group VI and group W were negative by ITS1 and kDNA molecular tests. Comparing the molecular results with the parasitologic and serologic diagnosis from group I, almost perfect agreement was found ( κ both>0.80, P<0.001). ITS1 and RV1/RV2 PCR detected 90.2% (74 of 82) of the samples. Two samples positive by RV1/RV2 were negative by LITSR/L5.8S, and six samples positive by LITSR/L5.8S were negative by RV1/RV2. Therefore, these two systems complemented each other; they diagnosed 100% of the samples as belonging to the Leishmania genus. Conclusions: We suggest an algorithm for the molecular diagnosis of VL, which must consider previous parasitologic and serologic (immunochromatographic) diagnoses, and should combine kDNA and ITS1 to determine the Leishmania subgenus using RFLP as a complement method to define the L. infantum species.
ABSTRACT
The objective of this literature review was to investigate the factors that may interfere in the production of breast milk (lactogenesis), making difficult the establishment of lactation and having hypogalactia as a result. Since breast milk is widely known as the most natural and important food for newborns, it is necessary for nurses to understand and acknowledge the physiological mechanisms involved in lactation in order to help mothers to achieve a successful breastfeeding. Methodology: Two computerized databases were assessed in search for articles: Literature Latin American and Caribbean Health Sciences and Medical Literature and Retrieval System On Line, both covering a 10 years (1997-2007) period. Considering the objective of the study, 21 articles were selected; other academic text-books were also consulted for supporting the development of contents and concepts on lactation physiology. Results: Factors related to failure and/or delay in lactogenesis were: placental retention; deficiency and /or resistance to prolactine; presence of ovarian cysts; breasts structural alterations; obesity; primiparity; prolonged labor and delivery; cesarean and hypotension. Conclusion: Lactogenesis is a subject that has not been properly acknowledged by nurses, although being essential for the proper establishment of breastfeeding, which is an important issue in nursing care and research. Relevance to clinical practice: When nurses take care of women who experience difficulties on breastfeeding, it is important to consider factors that may interfere in their lactogenesis; this issue seems to be rarely discussed in nursing literature. Therefore, in order to make womens breastfeeding experience meaningful, enjoyable and effective, nurses should improve their knowledge to investigate factors related to those difficulties and plan interventions...
O objetivo desta revisão de literatura foi investigar os fatores que podem interferir na produção de leite materno (lactogênese), dificultando o estabelecimento da lactação e tendo como resultado a hipogalactia. Uma vez que o leite materno é sabidamente o mais natural e importante alimento para o recém-nascido, é necessário para a enfermeira conhecer os mecanismos fisiológicos envolvidos na lactação, para ajudar a puérpera a amamentar seu bebê com sucesso. Metodologia: Para o desenvolvimento desta revisão foram consultadas duas bases de dados informatizadas em busca de artigos: Literatura Latino-Americana e do Caribe em Ciências da Saúde e Medical Literature and Retrieval System On Line. A consulta abrangeu um período de 10 anos (1997-2007). Considerando o objetivo do estudo, foram selecionados 21 artigos; também foram consultados livros-textos acadêmicos para o desenvolvimento de conteúdos sobre fisiologia e fundamentação dos conceitos. Resultados: Os fatores relacionados à falha e/ou atraso na lactogênese foram: retenção placentária; deficiência e/ou resistência à prolactina; presença de cistos ovarianos; alterações estruturais nas mamas; obesidade; primiparidade; trabalho de parto e parto prolongados; cesárea e hipotensão. Conclusão: A lactogênese é tema pouco abordado por enfermeiros, embora seja um processo imprescindível para o adequado estabelecimento da amamentação, sendo esta última um grande foco de assistência e pesquisa na Enfermagem. Relevância para a prática clínica: Quando o enfermeiro assiste a mulher que vivencia dificuldades no processo de aleitamento materno, é importante que considere fatores que possam interferir na lactogênese, o que parece ainda ser pouco abordado na literatura de Enfermagem. Assim, o enfermeiro que visa tornar esta experiência significativa, prazerosa e eficaz, deve instrumentalizar-se para investigar possíveis fatores relacionados ao insucesso e planejar intervenções...
El objetivo de este trabajo de revisión fue investigar los factores que pueden interferir con la lactogenese, lo que dificulta el establecimiento de la lactancia y tiene como resultado hipogalactia. Dado que la leche materna es la más conocida y importante alimentación natural del recién nacido, las mujeres deben tener todos los mecanismos fisiológicos de la lactancia trabajando bien para amamantar a su bebé correctamente. Metodología: Para el desarrollo de este estudio de revisión de la literatura se utilizaron dos bases de datos: Literatura Latinoamericana y del Caribe en Ciencias de la Salud y Literatura Internacional en Ciencias de la Salud. La consulta abarcó un período de 10 años (1997-2007). Considerando el objetivo del estudio, se seleccionaron 21 artículos. Consultados libros-texto académicos para el desarrollo de contenidos en la fisiología y de los conceptos. Resultados: Los factores relacionados con el fallo y / o el retraso en la lactogenese son: la retención placentária; la discapacidad y / o resistencia a la prolactina; la presencia de quistes ováricos; los cambios estructurales en los senos; la obesidad; primiparidad; trabajo de parto y parto largo; cesárea y la hipotensión. Conclusión: El lactogenese es un tema poco abordado por los enfermeros, apesar de un proceso esencial para el mejor establecimiento de la lactancia materna, siendo esta última una de las principales actividades de la atención y la investigación en enfermería. Importancia para la práctica clínica: Cuando lo enfermero ayuda a la mujer que tiene dificultades en el proceso de la lactancia materna, es importante considerar los factores que pueden interferir con lactogenese, que parece ser poco discutido en la literatura de enfermería. Por lo tanto, para promover una experiencia significativa, agradable y eficaz, lo enfermero debe estudiar para investigar los posibles factores relacionados con este fracaso y planificar intervenciones...