Estudo de Leishmania spp. em amostras provenientes de cães e humanos e suas implicações em saúde pública na região de Bauru, São Paulo.
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Data
2018-07-31
Autores
Ramos-Augusto, Livia Carla
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Universidade Estadual Paulista (Unesp)
Resumo
As leishmanioses são doenças zoonóticas de importância em saúde pública. Os objetivos do trabalho foram comparar as técnicas laboratoriais convencionas utilizadas para o diagnóstico de infecção por Leishmania spp. em amostras de sangue humano e canino com técnicas moleculares e realizar a quantificação da carga parasitária. Além disso, realizar a análise espacial dos casos de leishmaniose visceral humana (LV) e canina (LVC), confirmados laboratorialmente e notificados, baseando-se na incidência da enfermidade, entre 2014 a 2016, no município de Bauru (SP), com intuito de identificar as áreas prioritárias para implementação de medidas de vigilância e de controle. Foram avaliadas amostras de 26 pacientes com suspeita clínica de LV por meio do exame parasitológico direto em aspirado de medula óssea, teste imunocromatográfico rápido (TR), reação de imunofluorescência indireta (RIFI), reação em cadeia da polimerase (PCR) e PCR em tempo real (qPCR). Além disso, foram avaliadas amostras de 22 pacientes humanos com suspeita clínica de leishmaniose tegumentar (LT) para realização da RIFI, PCR e qPCR. As amostras caninas foram obtidas por meio dos inquéritos sorológicos realizados em 4725 cães, pelo Centro de Controle de Zoonoses (CCZ) de Bauru, e, mediante a positividade no TR (TR-DPP®) foi realizado o teste imunoenzimático (ELISA), PCR e qPCR. A análise espacial foi realizada em 39 casos humanos de LV, com diagnóstico obtido a partir do critério clínico-laboratorial, e nos 4725 cães pertencentes aos inquéritos sorológicos realizados entre 2014 a 2016, destacando os casos de LVC. Nas 26 amostras humanas analisadas para LV, a positividade obtida no parasitológico, TR, RIFI, PCR e qPCR foram: 80,8%, 80,8%, 73,1%, 84,6% e 84,6%. A RIFI foi a metodologia que obteve menor positividade e concordância com as demais metodologias e a carga parasitária se concentrou entre 100 e 1.000 parasitas/mL (68,2%). Nas 22 amostras humanas analisadas para LT, a positividade obtida na RIFI, PCR e qPCR foram: 9,1%, 18,2 e 18,2%. A RIFI obteve menor positividade e concordância insignificante com as técnicas moleculares e a carga parasitária das amostras se concentraram entre 100 e 1.000 parasitas/mL (100%). Nas amostras caninas, a positividade de 11,5% (543/4725) foi obtida em ambas as técnicas (TR-DPP® e ELISA). Nos cães TR-DPP® positivos, a positividade no ELISA, PCR e qPCR foram: 64,9%, 63,9% e 65%. A concordância foi excelente entre o ELISA e as técnicas moleculares. A carga parasitária se concentrou entre 1.000 e 10.000 parasitas/mL (73,9%). Foram identificadas cinco áreas com densidade alta de LV, localizadas nas regiões norte e oeste do município. Foram identificadas duas áreas com densidade alta de LVC, ambas localizadas na região norte. Em duas localidades, na região norte, houve sobreposição, compreendendo 68% dos casos de LVC notificados nos inquéritos sorológicos e 38,5% casos de LV humano. As técnicas moleculares em amostras de sangue constituem uma alternativa menos invasiva de diagnóstico tanto para LV quanto para LT em humanos, devido a alta positividade entre as amostras analisadas e/ou excelente concordância com as técnicas convencionais e, em cães, podem ser utilizadas tanto para o diagnóstico quanto para complementar os resultados sorológicos. As análises espaciais são capazes de gerar informações úteis que auxiliam em um melhor planejamento de ações de vigilância e no controle da doença, além da otimização do uso de recursos públicos para a execução dessas ações.
Leishmaniasis is a zoonotic disease of public health significance. The aims of the present study were to compare the conventional laboratory techniques used for the diagnosis of Leishmania spp. in human and canine blood samples with the molecular techniques and carry out the quantification of the parasite load. In addition, to perform a spatial analysis of visceral leishmaniasis in humans (VL) and dogs (CVL), laboratory confirmed and notified, based on the incidence of the disease in Bauru city (SP), during 2014 to 2016, with the objective of identify priority areas for surveillance and control of VL. Twenty-six samples with clinical suspicion of VL were evaluated through direct parasitological in bone marrow aspirate, rapid immunochromatographic test (RT), indirect immunofluorescent antibody test (IFAT), polymerase chain reaction (PCR) and real-time PCR (qPCR). In addition, twenty-two human cases with clinical suspicion of TL were selected for the accomplishment of IFAT, PCR and qPCR. The canine samples were obtained through serological investigations carried out in 4,725 dogs by the Center for Zoonoses Control (CCZ) in Bauru, and by means of the positivity in RT (RT-DPP®) was realized the enzyme-linked immunosorbent assay (ELISA), PCR and qPCR. The spatial analysis was performed in thirty-nine human cases of VL, with a diagnosis obtained from clinical laboratory, and in 4,725 dogs belonging to the serological investigations conducted during 2014 to 2016, highlighting CVL cases. In the twenty-six human samples analyzed for VL, the positivity obtained in parasitological, RT, IFAT, PCR and qPCR were: 80.8%, 80.8%, 73.1%, 84.6% and 84.6%. IFAT was the methodology that obtained less positivity and agreement with other methodologies. The parasite load was concentrated between 100 and 1,000 parasites/mL (68.2%). In the twenty-two human samples analyzed for tegumentary leishmaniasis (TL), the positivity obtained in the IFAT, PCR and qPCR were: 9.1%, 18.2 and 18.2%. The IFAT obtained lower positivity and negligible agreement with the molecular techniques and the parasite load of the samples were concentrated between 100 and 1,000 parasites/mL (100%). In the canine samples, 11.5% (543/4,725) of positivity were obtained for both techniques (RT-DPP® and ELISA). In RT-DPP® positive dogs, positivity in ELISA, PCR and qPCR were: 64.9%, 63.9% and 65%. The agreement was excellent between ELISA and molecular techniques. The parasite load was concentrated between 1,000 and 10,000 parasites/ml (73.9%). Five areas with high VL density were identified, located in the north and west regions of the city. Two areas with high CVL density were identified, both located in the north region. In two locations in the north region was overlap, comprising 68% of cases of CVL notified in serological investigations and 38.5% cases of human VL. Molecular techniques in blood samples consist in a less invasive diagnostic alternative for both VL and TL in humans, due to the higher positivity between the samples analyzed and/or the excellent agreement with the conventional techniques and, in dogs, can be used both for diagnosis and to complement the serological results. Spatial analysis provides useful information and can assist in the better surveillance planning actions and control of the disease, besides optimizing the use of resources for the execution of these actions.
Leishmaniasis is a zoonotic disease of public health significance. The aims of the present study were to compare the conventional laboratory techniques used for the diagnosis of Leishmania spp. in human and canine blood samples with the molecular techniques and carry out the quantification of the parasite load. In addition, to perform a spatial analysis of visceral leishmaniasis in humans (VL) and dogs (CVL), laboratory confirmed and notified, based on the incidence of the disease in Bauru city (SP), during 2014 to 2016, with the objective of identify priority areas for surveillance and control of VL. Twenty-six samples with clinical suspicion of VL were evaluated through direct parasitological in bone marrow aspirate, rapid immunochromatographic test (RT), indirect immunofluorescent antibody test (IFAT), polymerase chain reaction (PCR) and real-time PCR (qPCR). In addition, twenty-two human cases with clinical suspicion of TL were selected for the accomplishment of IFAT, PCR and qPCR. The canine samples were obtained through serological investigations carried out in 4,725 dogs by the Center for Zoonoses Control (CCZ) in Bauru, and by means of the positivity in RT (RT-DPP®) was realized the enzyme-linked immunosorbent assay (ELISA), PCR and qPCR. The spatial analysis was performed in thirty-nine human cases of VL, with a diagnosis obtained from clinical laboratory, and in 4,725 dogs belonging to the serological investigations conducted during 2014 to 2016, highlighting CVL cases. In the twenty-six human samples analyzed for VL, the positivity obtained in parasitological, RT, IFAT, PCR and qPCR were: 80.8%, 80.8%, 73.1%, 84.6% and 84.6%. IFAT was the methodology that obtained less positivity and agreement with other methodologies. The parasite load was concentrated between 100 and 1,000 parasites/mL (68.2%). In the twenty-two human samples analyzed for tegumentary leishmaniasis (TL), the positivity obtained in the IFAT, PCR and qPCR were: 9.1%, 18.2 and 18.2%. The IFAT obtained lower positivity and negligible agreement with the molecular techniques and the parasite load of the samples were concentrated between 100 and 1,000 parasites/mL (100%). In the canine samples, 11.5% (543/4,725) of positivity were obtained for both techniques (RT-DPP® and ELISA). In RT-DPP® positive dogs, positivity in ELISA, PCR and qPCR were: 64.9%, 63.9% and 65%. The agreement was excellent between ELISA and molecular techniques. The parasite load was concentrated between 1,000 and 10,000 parasites/ml (73.9%). Five areas with high VL density were identified, located in the north and west regions of the city. Two areas with high CVL density were identified, both located in the north region. In two locations in the north region was overlap, comprising 68% of cases of CVL notified in serological investigations and 38.5% cases of human VL. Molecular techniques in blood samples consist in a less invasive diagnostic alternative for both VL and TL in humans, due to the higher positivity between the samples analyzed and/or the excellent agreement with the conventional techniques and, in dogs, can be used both for diagnosis and to complement the serological results. Spatial analysis provides useful information and can assist in the better surveillance planning actions and control of the disease, besides optimizing the use of resources for the execution of these actions.
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Palavras-chave
leishmanioses, diagnóstico, análise espacial, leishmaniasis, diagnosis, spatial analysis