Abstract Leptospirosis is important in Uruguay due to the economic loss caused by the diseases of production animals, mainly bovines, and also due to frequent human infection. We decided to study anti-Leptospira antibodies in the sera of dairy workers, rice laborers, veterinarians, suburban slum dwellers and garbage recyclers. Our aims were to estimate the seroprevalence of infection by Leptospira spp. in these people at risk, the relative importance of the known risk factors associated with infection, and the impact of human infections in each setting. Groups at risk were identified and 35 visits to their locations were made, conducting field surveys and exchange talks for information and education. Simple epidemiological questionnaires were administered and sera samples were taken from 308 persons. The microagglutination Technique (MAT) and the IgM Indirect Immunofluorescence (IIF) assay were employed to detect antibodies. Environmental water samples, canine and equine sera were also examined. More than 45% of human sera were reactive and the studied groups were confirmed to be widely exposed to infection. Female sera were frequently reactive, though most illnesses occur in men, and the most severe cases in elderly males; the emergence and evolution of the disease may strongly depend on the host condition and functions. Animal contact and unsafe water usage were the main identified risk factors to be considered in prevention. Fifty per cent of the studied horses showed a positive MAT reaction. The underdiagnosis of the illness and its long-term symptoms require further study, as well as greater health and social attention efforts.

Resumen La leptospirosis es importante en Uruguay por las pérdidas económicas que ocasionan las enfermedades en los animales de producción, principalmente en los bovinos, y a causa de la frecuente infección humana. Decidimos estudiar anticuerpos anti -Leptospira en trabajadores de tambo y de plantaciones de arroz, así como en veterinarios, habitantes de asentamientos y recicladores de residuos. Buscamos evaluar la importancia de distintos factores de riesgo conocidos asociados con la infección por Leptospira spp., y estimar la frecuencia y el impacto de las infecciones humanas en la población expuesta. Se efectuaron 35 visitas a colectivos de riesgo, realizando relevamientos de campo e intercambios educativos, llenando formularios epidemiológicos sencillos y tomando muestras de sangre a 308 personas. Se investigaron anticuerpos séricos con técnicas de microaglutinación (MAT) e inmunofluorescencia indirecta para IgM. Se examinaron también, muestras de agua ambiental, y sueros caninos y equinos. Más del 45% de los sueros humanos fueron reactivos y se confirmó que los grupos estudiados estaban ampliamente expuestos a la infección. Los sueros de mujeres fueron frecuentemente reactivos, aunque la mayoría de las enfermedades ocurren en varones, y los casos más graves en varones añosos. La emergencia y la evolución de la afección pueden depender fuertemente del estado del huésped y su respuesta. El contacto con animales y el uso de agua insegura fueron los principales factores de riesgo identificados para considerar en la prevención. El 50% de los sueros equinos fueron reactivos por MAT. Es necesario incrementar los esfuerzos de atención social y sanitaria en estos sectores, superar el subdiagnóstico y estudiar la evolución y la sintomatología a largo plazo de los pacientes.

ABSTRACT Laboratory diagnosis of human leptospirosis usually relies on indirect methods exploring specific immune response. Isolation and identification of the involved strains are cumbersome, but can provide biological resources for pathogenic studies and relevant information for guiding prevention and control measures. The aim of the research we are hereby reporting was the characterization of Leptospira isolates obtained from humans and the environment in Uruguay. Blood cultures were performed from early samples of 302 Uruguayan patients, mainly rural workers, and from 36 water samples taken from their living or working environments. Eight human isolates and seven environmental isolates were obtained and analyzed by end point Polymerase Chain Reaction (PCR), Multilocus Variable Number of Tandem Repeat Analysis (MLVA) and other molecular methods. Human isolates corresponded to several serogroups and serovars of Leptospira interrogans and Leptospira kirschneri species, probably reflecting the infection with similar involved Leptospira species and serovars of an extended animal reservoir in rural settings of the country, mostly dedicated to meat and dairy production. Culture-positive patients were older than usually affected workers, and presented signs and symptoms of severe illness. A high organic and circulating bacterial burden may explain an easier positive result from these workers’ samples. Environmental isolates were mainly identified as Leptospira biflexa strains, with a single L. meyeri isolate of uncertain significance.

Introducción: la leptospirosis es una enfermedad febril, aguda, que presenta manifestaciones clínicas variadas. Esto dificulta o retarda el diagnóstico clínico, por lo cual es útil disponer de métodos de laboratorio adecuados para orientar el manejo inicial de estos pacientes. Objetivo: evaluar un procedimiento de inmunofluorescencia para detectar IgM (IF-IgM) de elaboración propia aplicado al diagnóstico temprano de leptospirosis. Material y método: se analizaron por IF-IgM y microaglutinación (MAT) (tomada como estándar de referencia) sueros obtenidos de pacientes con sospecha clínica de leptospirosis. La sensibilidad y especificidad de la IF-IgM versus MAT se establecieron utilizando una tabla de doble entrada. La concordancia entre dos observadores se determinó por el test Kappa. Resultados: de 161 muestras precoces analizadas, 97 sueros correspondieron a pacientes con infección aguda confirmada por MAT y 64 sin infección. La sensibilidad y especificidad de la IF-IgM con sueros de fase aguda fueron 79% y 100%, respectivamente. El índice Kappa fue 1. Conclusiones: la IF-IgM aparece como una herramienta útil para el diagnóstico temprano de pacientes con leptospirosis. No requiere el manejo de bacterias viables, puede realizarse en laboratorios que cuenten con microscopio de luz ultravioleta, se necesita una sola muestra de suero y el resultado está listo en tres a cuatro horas. En cuanto a las desventajas, no identifica los serovares involucrados y un resultado negativo no descarta la infección. Teniendo en cuenta esto último es obligatorio analizar por MAT una segunda muestra de suero obtenida a los 10-20 días de la primera para descartarla o confirmarla.

Resumo Introdução: a leptospirose é uma doença febril, aguda, com diferentes manifestações clínicas o que dificulta ou atrasa o diagnóstico clínico. Por essa razão é importante contar com métodos de laboratório adequados para orientar o manejo inicial dos pacientes. Objetivo: avaliar uma técnica de imunofluorescencia para detectar IgM (IF-IgM) para o diagnóstico precoce de leptospirose. Material e método: foram analisados por IF-IgM e microaglutinaçao (MAT) (usada como padrão de referencia) soros de pacientes com suspeita clínica de leptospirose. A sensibilidade e a especificidade da IF-IgM comparada com MAT foi determinada usando uma tabela de dupla entrada. A concordância entre dois observadores foi determinada usando o teste Kappa. Resultados: Cento sessenta e uma amostras da fase precoce foram analisadas, sendo 97 de pacientes com infecção aguda confirmada por MAT e 64 sem infecção. A sensibilidade e a especificidade da IF-IgM com soros obtidos na fase aguda foram 79% e 100%, respectivamente. O índice Kappa foi igual a 1. Conclusões: a técnica IF-IgM que desenvolvemos, pode ser uma ferramenta útil para o diagnóstico precoce de pacientes com leptospirose. Não é necessário manipular bactérias viáveis, pode ser realizada em laboratórios que tenham microscópio com luz ultravioleta, somente é necessária uma amostra de soro e o resultado está disponível em três-quatro horas. Com relação às desvantagens, esta técnica não identifica os serovares e um resultado negativo não descarta a presença de infecção. Considerando estas desvantagens, é obrigatório analisar por MAT uma segunda amostra de soro obtida 10 a 20 dias depois da primeira para descartar ou confirmar a infecção.

Abstract Introduction: leptospirosis is a severe febrile disease, with a variety of clinical presentations. This results in the delay of clinical diagnosis, or in difficulties achieving it, so having the appropriate laboratory tests may deem useful to guide the initial care of these patients. Objective: to evaluate a self-made immunofluorescence technique for the detection of IgM antibodies (IF-IgM) in the early diagnosis of leptospirosis. Method: serum samples obtained from patients with a clinical suspicion of leptospirosis were analysed by IF-IgM antibodies and the microagglutination test (MAT) . Sensitivity and specificity of IF-IgM versus MAT were determined using a double entry table. Agreement between two observers was determined by the Kappa test. Results: out of one hundred and sixty one early samples analysed, 97 serum samples corresponded to patients with a confirmed acute infection by MAT and 64 evidenced no infection. Sensitivity and specificity of IF-IgM assays in the acute phase were 79% and 100% respectively. The Kappa value was 1. Conclusions: IF-IgM seems to be a useful tool for the early diagnosis of patients with leptospirosis. It does not need viable bacteria to be handled; it may be applied in laboratories equipped with ultraviolet light microscopes, a single serum sample is needed and the result is seen three to four hours later. As to its disadvantages, it fails to identify the involved serovars and a negative result does not discard infection. Bearing the latter into account, the MAT test is mandatory in a second serum sample obtained 10-20 days after the first one, to either discard or confirm the disease.