Seroprevalencia de la Leishmaniasis Visceral Canina mediante Elisa con rK39 en Focos Endémicos de Venezuela / Seroprevalence of Canine Visceral Leishmaniasis by rK39-ELISA in Endemic Foci in Venezuela

En el presente trabajo se presenta la seroprevalencia de la leishmaniasis visceral canina (LVC), zoonosis causada por Leishmania infantum/chagasi. Se realizaron pruebas serológicas y examen clínico a 15.822 perros de 13 entidades federales endémicas para la leishmaniasis visceral en Venezuela, en el periodo 2004-2012. Los sueros fueron analizados mediante ELISA con el antígeno recombinante rK39. Los resultados muestran que 14,8% de la población de caninos son positivos para LV. Los estados Lara (19%) y Guárico (18%) mostraron una mayor prevalencia de la enfermedad. Sin embargo, para los años 2010-2012, la prevalencia de la LVC para las entidades federales como Anzoátegui, Aragua, Carabobo, Cojedes, Nueva Esparta y Sucre se mantuvieron entre un 3% y un 31%. Los caninos seropositivos (67,1% machos y 32,9% hembras) tenían edades promedio de 4,8±2,9 años. El 81% de los caninos seropositivos encontrados en estas zonas, no presentó signos clínicos característicos de LVC, mientras que la clínica presentada por el resto fueron ulceraciones cutáneas (8,5%), alopecia (9,4%) y onicogrifosis (19,2%). Este reporte muestra la distribución geográfica (tanto en zonas rurales como urbanas) y características clínicas más resaltantes de perros parasitados en las diferentes regiones endémicas del país, con el fin de tomar medidas estratégicas que fortalezcan los programas de control y prevención de esta zoonosis problema de salud pública.

This study discloses the seroprevalence of canine visceral leishmaniasis (CVL), a zoonotic disease caused by Leishmania infantum/chagasi. In this study, serological tests and clinical examinations were performed in 15,822 dogs from 13 federal entities endemic for visceral leishmaniasis in Venezuela, during the period 2004-2012. Serum samples were analysed by ELISA against the recombinant antigen rK39. Results demonstrated a prevalence of 14.8% of positive dogs for VL. Lara (19%) and Guárico (18%) states showed the highest seroprevalence of the disease. However, for the years 2010-2012, the prevalence of CVL for federal entities as Anzoátegui, Aragua, Carabobo, Cojedes, Nueva Esparta, and Sucre remained between 3% and 31%. The seropositive canines (67.1% males and 32.9% females) average 4.8±2.9 years of age and 81% of the dogs found in these endemic areas did not show clinical signs characteristic of LVC, while clinical symptoms presented by the rest were skin ulceration (8.5%), alopecia (9.4%) and onychogryphosis (19.2%). This report demonstrates the geographical distribution (both rural and urban) and most striking clinical features of parasitized dogs in different endemic regions of the country, in order to take strategic actions to strengthen the control and prevention programs of this public health problem.

Reactividad serológica y celular frente a proteínas micobacterianas en la enfermedad de Hansen / No disponible

Se diseñó un estudio para evaluar la reactividad inmunológica frente a diferentes preparaciones proteicas micobacterianas utilizando pruebas serológicas y de inmunidad celular. Para el estudio fueron incluidos pacientes con manifestaciones clínicas de lepra predominantemente de la forma multibacilar. El 58% correspondía a la forma clínica de Lepra Lepromatosa (LL) n=81, el 29% a la forma Borderline Lepromatosa (BL) n=41 y 10% a Borderline Bordeline (BB) n=14. Sólo el 3% fueron pacientes Borderline Tuberculoide (BT): 74% masculino y 26% femenino. El fenómeno reaccional más frecuente fue de tipo eritema nodoso leproso (ENL). Las proteínas micobacterianas ensayadas fueron: antígenos proteicos crudos totales de Mycobacterium leprae (MISA), Mycobacterium bovis (MbSA y MbSA de excreción), antígeno proteico de excreción parcialmente purificado con una movilidad relativa de 30 kDa (MI30) y proteínas recombinantes de Mycobacterium (Mt70, Mb 65, MI 36, 28, 18 y 10 KDa) encontrándose que las proteínas recombinantes (MI 10KDa, MI36 kDa) a mayor carga bacilar presentaban una mayor reactividad serológica estadísticamente significativa (p=0.0051 y 0.050 respectivamente). La proteína de 30 kDa) a mayor carga bacilar presentaban una mayor reactividad serológica estadísticamente significativa (p=0.0051 y 0.0050 respectivamente). La proteína de 30 kDa fue predominantemente reconocida por anticuerpos de los pacientes multibacilares. Los resultados demuestran que el promedio de los valores de anticuerpos en pacientes no reaccionales fueron superiores en presencia de proteínas completas (MbSA y MbSA de exreción) en comparación con el grupo de pacientes que presentaron fenómenos reaccionales (p=0.000567 y 0.000061 respectivamente). Este mismo comportamiento se observó frente a las proteínas micobacterianas individuales (30kDA, 10kDa y 36 kDa). La respuesta proliferativa de los linfocitos T en los pacientes multibacilares reaccionales y no reaccionales frente a las proteínas micobacterianas (MISA, ML 10kDa, MbSA, MbSA de excreción) fue negativa en ambos grupos (AU)

The study was designed for evaluating immunological reactivity to various mycobacterial protein preparations using serological and cell-mediated immunological test in patients with clinical leprosy signs, predominantly, with the multibacillary forms. Al patients wer adults with ages between 20 and 39 years. Fifty eight (n=81) percent corresponded to Lepromatous Leprsy (LL), 29% (n=41) to Borderline Lepromatuous Leprosy (BL) and 10% (n=14) to Borderline Borderline Leprosy (BB); only 3% were Bordeline Tuberculoid (BT) patients: 74% males and 26% females. The most frequent reactional phenomenon was of the Erythema Nodosum (ENL) type. The mycobacterial proteins tested were: total crude Mycobacterium leprae antigens (MISA); Mycobacterium bovis (MbSA and excretion MbSA); partially purified excretion protein antigen, with a 30KDa relative movility (Ml30); and recombinant M. leprae protein (Mt70, Mb 65, Ml36, 28, 18 and 10kDa). Two of the recombinant proteints (M110 and Ml36 kDA) presented a statiseally significant higher serological reactivity, directly related with a larger bacillary load (p=0.0051 and 0.050 respectively). The 30kDa protein was predominantly recognized by antibodies from multibacillary patients. Results show that mean antibody values were higher in non reactional patients when tested against complete proteins (MbSA and ex MbSA) when compared with the group of patients who presented reactional phenomena (p=0.000567 and 0.000061, respectively). Comparing reactional with non reactional patients, it was seen that mean antibody values against complete proteins (MbSA and ex MbSA) were higher in non reactional individual (p=0.000567 and 0.000061, respectively). This same behavior occurred towards individual mycobacyterial proteins (30, 10 and 36 kDa). The T lymphocyte prolyphetative response in reactional and non reactional patients towards mycobacterial proteins (MISA, Ml 10 kDA, MbSA, ex MbSA) was negative (AU)

[Non invasive diagnostic tools for visceral leishmaniasis: a comparison of the immunoserological tests DAT, rK26 and rK39]. / Herramientas no invasivas en Venezuela: comparación entre las pruebas inmunoserológicas DAT, rK26 y rK39 en el diagnóstico de leishmaniasis visceral.

INTRODUCTION: Human visceral leishmaniasis is a serious public health problem in endemic countries because of its high potential lethality, particularly in children. Rapid diagnosis is essential to early treatment and control of visceral leishmaniasis. OBJECTIVE: The aim was to compare three serodiagnostic tools for human visceral leishmaniasis. MATERIALS AND METHODS: Three methods were compared: the rK39 dipstick (Kalazar detection test, Inbios International Inc.), ELISA rK26 and direct agglutination test (DAT) (KIT Biomedical Research). Fifty serum samples from patients positive for rK39 ELISA were compared from four endemic provinces in Venezuela: Nueva Esparta (Margarita island), Lara, Anzoátegui and Trujillo. Additional serum samples from 17 healthy volunteers and 25 patients with other diseases were included. The rK39 ELISA served as the baseline standard method. Sensitivity, specificity, positive predictive value, negative predictive value and likelihood ratio were calculated for each test. RESULTS: All methods had a positive correlation with rK39 ELISA (p<0.0001). They showed high sensitivity and specificity. The direct agglutination test and the rK39 dipstick showed high sensitivity values, 89.7% (95% CI: 81.34.0-98.2%) and 94.2% (95% CI: 87.7-100%), respectively, and high specificity, 81.0% (95% CI: 80.0-99.5%) and 100%. The rK26 ELISA showed good specificity, 99% (95% CI: 95.2-100%), but a very low sensitivity, 37% (95% CI: 23.4-50.2%). CONCLUSION: Overall results indicated that DAT and rK39 dipstick have the highest specificity and sensitivity. Both are simple, cost-effective and field applicable tests. Therefore, they are recommended for early and accurate diagnosis of visceral leishmaniasis.

Reactividad serológica y celular frente a proteínas micobacterianas en la enfermedad de Hansen / Serological and cellular reactivity to mycobacterial proteins in Hansen´s disease

Se diseñó un estudio para evaluar la reactividad inmunológica frente a diferentes preparaciones proteicas micobacterianas utilizando pruebas serológicas y de inmunidad celular. Para el estudio fueron incluídos pacientes con manifestaciones clínicas de lepra predominantemente de la forma multibacilar. Todos los pacientes fueron adultos con edad comprendida entre 20 y 39 años. El 58 por ciento correspondía a la forma clínica de Lepra Lepromatosa (LL) n= 81, el 29 por ciento a la forma Borderline Lepromatosa (BL) n=41 y 10 por ciento a Borderline Borderline (BB) n=14. Solo el 3 por ciento fueron pacientes Borderline Tuberculoide (BT): 74 por ciento masculino y 26 por ciento femenino. El fenómeno reaccional más frecuente fue del tipo eritema nodoso leproso (ENL). Las proteínas micobacterianas ensayadas fueron: antígenos proteicos crudos totales de Mycobacterium leprae (MlSA), Mycobacterium bovis (MbSA y MbSA de excreción), antígeno proteico de excreción parcialmente purificado con una movilidad relativa de 30 kDa (Ml 30) y proteínas recombinantes de Mycobacterium (Mt70, Mb 65, Ml 36, 28, 18 y 10 kDa) encontrandose que las proteínas recombinantes (Ml10 kDa, Ml 36 kDa) a mayor carga bacilar presentaban una mayor reactividad serológica estadísticamente significativa (p= 0,0051 y 0,050 respectivamente). La proteína de 30 kDa fue predominantemente reconocida por anticuerpos de los pacientes multibacilares. Los resultados demuestran que el promedio de los valores de anticuerpos en pacientes no reaccionales fueron superiores en presencia de proteínas completas (MbSA y MbSA de exc) en comparación con el grupo de pacientes que presentaron fenómenos reaccionales (p=0,000567 y 0,000061 respectivamente) Este mismo comportamiento se observó frente a las proteínas micobacterianas individuales (30 kDa, 10 kDa y 36 kDa). La respuesta proliferativa de los linfocitos T en los pacientes multibacilares reaccionales y no reaccionales frente a las proteínas micobacterianas...

The study was designed for evaluating immunological reactivity to various mycobacterial protein preparations using serological and cell-mediated immunological tests in patients with clinical leprosy signs, predominantly, with the multibacillary forms. All patients were adults with ages between 20 and 30 years. Fifty eight (n= 81) percent corresponded to Lepromatous Leprosy (LL), 29 percent (n= 41) to Borderline Lepromatous Leprosy (BL) and 10 percent (n=41) to Borderline Borderline Leprosy (BB); only 3 percent were Borderline Tuberculoid (BT) patients: 74 percent males and 26 percent females. The most frequent reactional phenomenon was of the Erythema Nodosum (ENL) type. The mycobacterial proteins tested were: total crude Mycobacterium leprae antigens (MISA); Mycobacterium bovis (MbSA and excretion MbSA); partially purified excretion protein antigen, with a 30kDa relative movility (Ml30); and recombinant M. leprae proteins (Mt70, Mb 65, Ml 36, 28, 18 and 10 kDa). Two of the recombinant proteins (Ml10 and Ml 36 kDa) presented a statiscally significant higher serological reactivity, directly related with a larger bacillary load (p= 0.0051 and 0.050 respectively). The 30 kDa protein was predominantly recognized by antibodies from multibacillary patients. Results show that mean antibody values were higher in non reactional patients when tested against complete proteins (MbSA and ex MbSA) when compared with the group of patients who presented reactional phenomena (p= 0.000567 and 0.000061, respectively). Comparing reactional with non reactional patients, it was seen that mean antibody values against complete proteins (MbSA and ex MbSA) were higher in non reactional individuals (p= 0.000567 and 0.000061, respectively). This same behavior occurred towards individual mycobacterial proteins (30, 10 and 36 kDa). The T lymphocyte prolypherative response in reactional and non reactional patients towards mycobacterial proteins (MlSA, Ml 10 kDa, MbSA...

Estudio preliminar de Citocinas en suero de perros con Leishmaniasis visceral en una zona endémica Venezolana / Preliminary Study of Cytokines in Sera from Dogs with Visceral Leishmaniasis from a Venezuelan Endemic Area

Diversos estudios en modelos de ratones han demostrado que la inmunidad protectora frente a Leishmania es mediada por linfocitos T, células presentadoras de antígeno y citocinas. Sin embargo, pocos estudios se han realizado para evaluar citocinas en perros infectados en forma natural con Leishmania infantum/chagasi. El perro doméstico es el principal reservorio del parásito, en tal sentido, el objetivo de este trabajo fue determinar las citocinas en suero de 33 perros con Leishmaniasis Visceral Canina (LVC), provenientes del estado Nueva Esparta, Venezuela (foco endémico). Los perros fueron clasificados en sintomáticos o asintomáticos, de acuerdo al análisis de los signos clínicos de la enfermedad, coincidentes con los títulos de anticuerpos contra las kinesinas recombinantes de Leishmania rK39 y rK26. Otros dos grupos incluyeron: 10 perros de la misma zona endémica como grupo control endémico (CE) y 10 perros de la zona no endémica como control sano (CS). Las concentraciones (pg/mL) de las citocinas solubles IFN-γ, TNF-α, IL-10, IL-6, IL-4 e IL-2 se determinaron por citometría de flujo (Kit CBA Hu Th1/Th2, BD TM). Los resultados mostraron concentraciones estadísticamente mayores (p<0,05) de IFN-γ (69,93±7,46), IL-4 (7,51±2,68), TNF-α (3,86±1,46) e IL-2 (39,85±3,84) en el grupo de perros asintomáticos, con respecto a los perros sintomáticos (60,8±10,6; 5,28±0,80; 2,76±0,72 y 36,04±3,61, respectivamente) y los perros sanos (51±14; 4,65±0,2; 3,21±0,89 y 32,65±5,86, respectivamente). Los perros asintomáticos también presentaron mayor concentración de IL-6 (4,9±0,55) que los CS (4,02±0,64) (p<0,01). Estos resultados demuestran que los perros en estado asintomático exhiben mayor proporción de citocinas de activación celular y proinflamatorias. Los resultados señalan a la medición de citocinas séricas como reflejo del estado inmunológico de los caninos en futuros estudios orientados a vacunación o terapia.

Different experimental murine models have shown that protective immunity against Lesihmania depends upon T cells, cytokines, and antigen presenting cells. However, the role of cytokines in naturally-infected hosts like domestic dogs is controversial. Few studies have evaluated cytokines in dogs naturally-infected with Leishmania infantum/chagasi. Since the domestic dog is the main reservoir of the parasite, a study was conducted to determine cytokines in serum of 33 dogs with Canine Visceral Lesihmaniasis from endemic areas of the State of Nueva Esparta, Venezuela. Dogs were classified as symptomatic (SD) and asymptomatic (AD), according to the expression of three or more clinical signs and levels of  antibodies for rK39 and rK26. Ten non-infected, rK39 negative controls were included from an endemic area (EA) and ten dogs from a non-endemic area were used as healthy controls (HC). The following cytokines (pg/mL) were measured in serum by flow cytometry (CBA Hu Th1/Th 2, BD TM kit): IFN-γ, TNF-α, IL-10, IL-6, IL-4 and IL-2. Results show a higher  concentration (P<0.05) of  IFN-γ (69.93±7.46), IL-4 (7.51±2.68), TNF-α (3.86±1.46), and IL-2 (39.85±3.84) in AD when compared with SD (60.8±10.6; 5.28±0.80; 2.76± 0.72; and 36.04±3.61, respectively); and HC (51±14; 4.65±0.2; 3.21±0.89, and 32.65±5.86, respectively). The AD also showed higher levels (P<0.01) of IL-6 (4.9±0.55) compared with HC (4.02±0.64). Results show that AD exhibit a higher proportion of cellular activation and proinflammatory cytokines. Results indicate that measuring of serum cytokines could reflect the immunological status in dogs in future clinical trials oriented to either vaccination or therapy.

[Serological and cellular reactivity to mycobacterial proteins in Hansen's disease]. / Reactividad serológica y celular frente a proteínas micobacterianas en la enfermedad de Hansen.

The study was designed for evaluating immunological reactivity to various mycobacterial protein preparations using serological and cell-mediated immunological tests in patients with clinical leprosy signs, predominantly, with the multibacillary forms. All patients were adults with ages between 20 and 30 years. Fifty eight (n = 81) percent corresponded to Lepromatous Leprosy (LL), 29% (n = 41) to Borderline Lepromatous Leprosy (BL) and 10% (n = 41) to Borderline Borderline Leprosy (BB); only 3% were Borderline Tuberculoid (BT) patients: 74% males and 26% females. The most frequent reactional phenomenon was of the Erythema Nodosum (ENL) type. The mycobacterial proteins tested were: total crude Mycobacterium leprae antigens (MISA); Mycobacterium bovis (MbSA and excretion MbSA); partially purified excretion protein antigen, with a 30 kDa relative movility (Ml30); and recombinant M. leprae proteins (Mt70, Mb 65, Ml 36, 28, 18 and 10 kDa). Two of the recombinant proteins (Ml10 and Ml 36 kDa) presented a statiscally significant higher serological reactivity, directly related with a larger bacillary load (p = 0.0051 and 0.050 respectively). The 30 kDa protein was predominantly recognized by antibodies from multibacillary patients. Results show that mean antibody values were higher in non reactional patients when tested against complete proteins (MbSA and ex MbSA) when compared with the group of patients who presented reactional phenomena (p = 0.000567 and 0.000061, respectively). Comparing reactional with non reactional patients, it was seen that mean antibody values against complete proteins (MbSA and ex MbSA) were higher in non reactional individuals (p = 0.000567 and 0.000061, respectively). This same behavior occurred towards individual mycobacterial proteins (30, 10 and 36 kDa). The T lymphocyte prolypherative response in reactional and non reactional patients towards mycobacterial proteins (MlSA, Ml 10 kDa, MbSA, ex MbSA) was negative.

Herramientas no invasivas en Venezuela: comparación entre las pruebas inmunoserológicas DAT, rK26 y rK39 en el diagnóstico de leishmaniasis visceral / Non invasive diagnostic tools for visceral leishmaniasis: a comparison of the immunoserological tests DAT, rK26 and rK39

Introducción. La leishmaniasis visceral constituye un problema de salud pública en los países en donde es endémica por ser potencialmente letal, principalmente en niños. El diagnóstico rápido es importante en el control de la enfermedad. Objetivo. Comparar las pruebas inmunocromatográficas rK39 (rK39 dipstick, Kalazar detect test, Inbios Internacional Inc.), ELISA rK26 y la prueba de aglutinación directa (Kit Biomedical Research) en relación con la prueba de ELISA rK39, como herramientas serodiagnósticas para la leishmaniasis visceral en Venezuela.Materiales y métodos. Se estudiaron 50 muestras séricas de pacientes positivos por la prueba ELISA rK39, provenientes de diferentes zonas endémicas: Nueva Esparta, Lara, Anzoátegui y Trujillo; se incluyeron 17 muestras de voluntarios sanos y 25 de pacientes con otras enfermedades. Se utilizó la prueba ELISA rK39 como método de referencia, considerándola como patrón de referencia imperfecto, a partir del cual se determinaron los valores de sensibilidad, especificidad, razón de verosimilitud y valores diagnóstico positivo y negativo en las demás pruebas evaluadas. Resultados. Todas las pruebas mostraron una fuerte correlación (p<0,0001) con la ELISA rK39. La aglutinación directa y la prueba inmunocromatográfica rK39 presentaron altos valores de sensibilidad, 89,74% (IC95% 81,34-98,15) y 94,15% (IC95% 87,65-100), respectivamente, y de especificidad, 81% (IC95% 79,96-99,51) y 100% (IC95% 100-100). La prueba ELISA rK26, a pesar de poseer buena especificidad, 99% (IC95% 95,17-100), tuvo baja sensibilidad, 37% (IC95% 23,41-50,15). Conclusión. Las pruebas de aglutinación directa y la prueba inmunocromatográfica rK39 presentaron los mayores valores de sensibilidad y especificidad. Ambas son simples, económicas y fácilmente aplicables. Por ello, son recomendables para efectuar un diagnóstico de leishmaniasis visceral eficaz y precoz en Venezuela.

Introduction. Human visceral leishmaniasis is a serious public health problem in endemic countries because of its high potential lethality, particularly in children. Rapid diagnosis is essential to early treatment and control of visceral leishmaniasis.Objective. The aim was to compare three serodiagnostic tools for human visceral leishmaniasis. Materials and methods. Three methods were compared: the rK39 dipstick (Kalazar detection test, Inbios International Inc.), ELISA rK26 and direct agglutination test (DAT) (KIT Biomedical Research). Fifty serum samples from patients positive for rK39 ELISA were compared from four endemic provinces in Venezuela: Nueva Esparta (Margarita island), Lara, Anzoátegui and Trujillo. Additional serum samples from 17 healthy volunteers and 25 patients with other diseases were included. The rK39 ELISA served as the baseline standard method. Sensitivity, specificity, positive predictive value, negative predictive value and likelihood ratio were calculated for each test. Results. All methods had a positive correlation with rK39 ELISA (p<0.0001). They showed high sensitivity and specificity. The direct agglutination test and the rK39 dipstick showed high sensitivity values, 89.7% (95% CI: 81.34.0-98.2%) and 94.2% (95% CI: 87.7-100%), respectively, and high specificity, 81.0% (95% CI: 80.0-99.5%) and 100%. The rK26 ELISA showed good specificity, 99% (95% CI: 95.2-100%), but a very low sensitivity, 37% (95% CI: 23.4-50.2%). Conclusion. Overall results indicated that DAT and rK39 dipstick have the highest specificity and sensitivity. Both are simple, cost-effective and field applicable tests. Therefore, they are recommended for early and accurate diagnosis of visceral leishmaniasis.

Evaluación de una prueba de aglutinación directa como método alternativo para el diagnóstico de leishmaniasis visceral canina y humana en Venezuela / The direct agglutination test as an alternative method for the diagnosis of canine and human visceral leishmaniasis

Introduction. Visceral leishmaniasis is the most severe clinical form of leishmaniasis and is often fatal without proper treatment. Therefore, early and accurate diagnosis is important, but often difficult in endemic areas. Objective. The aim was to evaluate a direct agglutination test as a potential visceral leishmaniasis diagnostic method in endemic areas of Venezuela Materials and methods. The performance of the direct agglutination test, based on freezedried Leishmania donovani antigen was evaluated under laboratory conditions using serum samples of humans and dogs from several Venezuelan visceral leishmaniasis endemic areas: Nueva Esparta (Margarita Island), Lara, Anzoátegui and Trujillo Status. The study included confirmed visceral leishmaniasis patients (n=30), visceral leishmaniasis suspected subjects (n=4), healthy controls (n=19) and patients with other confirmed diseases (n=20). In addition, 24 serum samples from dogs with confirmed visceral leishmaniasis and 18 healthy control dogs were tested. Results. All serum samples of visceral leishmaniasis patients, either active or recovered, were positive. They showed anti- L. donovani titers above 1:1600. Three out of four suspected visceral leishmaniasis cases were also positive, while serum samples from endemic controls and patients with other diseases had titers lower than 1:800. A sensitivity of 100 percent was obtained for all threshold levels under consideration and 100 percent specificity at the threshold titer of 1:800 (95 percent confidence interval: 91-100 percent). A 93 percent sensitivity (95 percent confidence interval: 76-99 percent) was observed in dog samples, with 100 percent specificity (95 percent confidence interval: 79-100 percent) at the threshold titer of 1:200. Conclusion. The direct agglutination test seems suitable for use in epidemiological studies and for serological diagnosis of human visceral leishmaniasis and canine visceral leishmaniasis.

Introducción. La leishmaniasis visceral es la forma clínica más grave de la leishmaniasis. Ésta puede ser fatal si no se administra el tratamiento adecuado. Por ello, el diagnostico temprano es importante, pero a menudo difícil, en las áreas endémicas. Objetivo. Evaluar el potencial de la prueba de aglutinación directa como un método para el diagnóstico de leishmaniasis visceral en zonas endémicas de Venezuela. Materiales y métodos. La efectividad de la prueba de aglutinación directa con el antígeno congelación-descongelación de Leishmania donovani fue evaluada bajo condiciones de laboratorio usando muestras de sueros de humanos y perros provenientes de diferentes regiones endémicas de leishmaniasis visceral de Venezuela: Nueva Esparta (Isla de Margarita), Estados Lara, Anzoátegui y Trujillo. Se incluyeron pacientes con diagnóstico confirmado de leishmaniasis visceral (n=30), sospecha de leishmaniasis visceral (n=4), voluntarios sanos (n=19) y pacientes con otras enfermedades (n=20). Además, se evaluaron 24 muestras de suero de perros con leishmaniasis visceral y 18 controles. Resultados. Todas las muestras de los pacientes con leishmaniasis visceral activa o curada fueron positivas. Mostraron títulos anti-L. donovani por encima de 1:1.600. Tres de cuatro casos con sospecha de leishmaniasis visceral también resultaron positivos a la prueba, mientras que los sueros controles y los de los pacientes con otras patologías dieron títulos por debajo de 1:800. Se obtuvo una sensibilidad de 100% a todos los puntos de corte considerados y una especificidad de 100% al punto de corte de 1:800 (intervalo de confianza de 95%, IC95%: 90,97%-100%). Las muestras de perros mostraron una sensibilidad de 92,59% (IC95%: 75,69%- 99,09%) y 100% de especificidad (IC95%: 79,42%-100%) al punto de corte de 1:200. Conclusión. En general, nuestros resultados indican que el uso de la prueba de aglutinación directa es apropiado para la realización de estudios epidemiológicos y para el...

The direct agglutination test as an alternative method for the diagnosis of canine and human visceral leishmaniasis.

INTRODUCTION: Visceral leishmaniasis is the most severe clinical form of leishmaniasis and is often fatal without proper treatment. Therefore, early and accurate diagnosis is important, but often difficult in endemic areas. OBJECTIVE: The aim was to evaluate a direct agglutination test as a potential visceral leishmaniasis diagnostic method in endemic areas of Venezuela. MATERIALS AND METHODS: The performance of the direct agglutination test, based on freeze-dried Leishmania donovani antigen was evaluated under laboratory conditions using serum samples of humans and dogs from several Venezuelan visceral leishmaniasis endemic areas: Nueva Esparta (Margarita Island), Lara, Anzoátegui and Trujillo Status. The study included confirmed visceral leishmaniasis patients (n=30), visceral leishmaniasis suspected subjects (n=4), healthy controls (n=19) and patients with other confirmed diseases (n=20). In addition, 24 serum samples from dogs with confirmed visceral leishmaniasis and 18 healthy control dogs were tested. RESULTS: All serum samples of visceral leishmaniasis patients, either active or recovered, were positive. They showed anti-L. donovani titers above 1:1600. Three out of four suspected visceral leishmaniasis cases were also positive, while serum samples from endemic controls and patients with other diseases had titers lower than 1:800. A sensitivity of 100% was obtained for all threshold levels under consideration and 100% specificity at the threshold titer of 1:800 (95% confidence interval: 91-100%). A 93% sensitivity (95% confidence interval: 76-99%) was observed in dog samples, with 100% specificity (95% confidence interval: 79-100%) at the threshold titer of 1:200. CONCLUSION: The direct agglutination test seems suitable for use in epidemiological studies and for serological diagnosis of human visceral leishmaniasis and canine visceral leishmaniasis.

Epidemiological aspects of human and canine visceral leishmaniasis in Venezuela.

OBJECTIVE: To report recent data on the distribution of human and canine visceral leishmaniasis (VL) in Venezuela, and to highlight problems associated with effective control measures. METHODS: We report the number of cases, incidence rate, age and sex distribution, and mortality rates for human VL (HVL) for the period of 1995 through 2000, based on National Registry of Leishmaniasis data. We carried out serological studies on a total of 3 025 domestic dogs from the 12 states in Venezuela reporting cases of human VL in this 1995-2000 period and also from the state of Yaracuy, where cases were reported earlier during the decade of the 1990s. RESULTS: From 1995 through 2000, 242 cases of HVL were reported from 12 states, in various sections of Venezuela. There was a relatively stable national incidence rate of 0.2 cases per 100 000 persons per year. Of the 242 cases, 26.0% were from Margarita Island, one of the three islands that make up the state of Nueva Esparta (Margarita Island was the only one of the Nueva Esparta islands that had HVL cases). Over the 1995-2000 period, the annual incidence rates for Nueva Esparta ranged from 1.7 to 3.8 cases per 100 000 population. Males in Venezuela were more frequently affected (59.5%) than were females (40.5%). In terms of age, 67.7% of the VL patients were

Epidemiological aspects of human and canine visceral leishmaniasis in Venezuela

OBJECTIVE: To report recent data on the distribution of human and canine visceral leishmaniasis (VL) in Venezuela, and to highlight problems associated with effective control measures. METHODS: We report the number of cases, incidence rate, age and sex distribution, and mortality rates for human VL (HVL) for the period of 1995 through 2000, based on National Registry of Leishmaniasis data. We carried out serological studies on a total of 3 025 domestic dogs from the 12 states in Venezuela reporting cases of human VL in this 1995-2000 period and also from the state of Yaracuy, where cases were reported earlier during the decade of the 1990s. RESULTS: From 1995 through 2000, 242 cases of HVL were reported from 12 states, in various sections of Venezuela. There was a relatively stable national incidence rate of 0.2 cases per 100 000 persons per year. Of the 242 cases, 26.0 percent were from Margarita Island, one of the three islands that make up the state of Nueva Esparta (Margarita Island was the only one of the Nueva Esparta islands that had HVL cases). Over the 1995-2000 period, the annual incidence rates for Nueva Esparta ranged from 1.7 to 3.8 cases per 100000 population. Males in Venezuela were more frequently affected (59.5 percent) than were females (40.5 percent). In terms of age, 67.7 percent of the VL patients were < 4 years of age, and 80.6 percent were younger than 15 years. The mortality rate among the persons with VL was 7.85 percent during the 1995-2000 period. Serological screening with rK39 antigen of 1217 dogs from Margarita Island found a 28.5 percent positivity rate (testing of dogs was not done on the two other islands of Nueva Esparta). In contrast, the rate was 2.8 percent in the 1 808 samples from dogs from 12 states on the mainland. CONCLUSIONS: Human and canine VL are unevenly distributed in Venezuela. The distribution may reflect such factors as differences among the states in human population density, vector density, and the presence or absence of other trypanosomatidae. Particularly high infection rates in very young children as well as in domestic dogs occur in semiurban communities of Nueva Esparta, where other human-infecting trypanosomatidae have not been reported. Control measures related to limiting canine infection might contribute to disease control where VL infections are frequent. Reducing VL mortality requires increased awareness among medical professionals of the possibility of VL in the differential...

Epidemiological and immunological aspects of human visceral leishmaniasis on Margarita Island, Venezuela

Sixty-five patients were diagnosed with visceral leishmaniasis (VL) on Margarita Island in the decade from 1990 to1999; 86.2 percent were <= 3 years old. All were leishmanin-negative at diagnosis. Evaluation of 23 cured patients in 1999 revealed that 22/23 had converted to leishmanin-positive; five had persisting antibodies to rK39 antigen, with no clinical evidence of disease. Leishmanin tests were positive in 20.2 percent of 1,643 healthy individuals from 417 households in endemic areas. Of the positive reactors, 39.8 percent were identified in 35 (8.4 percent) of the households, 15 of which had an antecedent case of VL, a serologically positive dog or both. Weak serological activity to rK39 antigen was detected in 3 of 488 human sera from the endemic areas. The presence of micro-foci of intense peri-urban transmission and the apparent absence of other Trypanosomatidae causing human disease offer a unique opportunity for the study of reservoirs, alternative vectors and evaluation of control measures on the Island

Epidemiological and immunological aspects of human visceral leishmaniasis on Margarita Island, Venezuela.

Sixty-five patients were diagnosed with visceral leishmaniasis (VL) on Margarita Island in the decade from 1990 to1999; 86.2% were <= 3 years old. All were leishmanin-negative at diagnosis. Evaluation of 23 cured patients in 1999 revealed that 22/23 had converted to leishmanin-positive; five had persisting antibodies to rK39 antigen, with no clinical evidence of disease. Leishmanin tests were positive in 20.2% of 1,643 healthy individuals from 417 households in endemic areas. Of the positive reactors, 39.8% were identified in 35 (8.4%) of the households, 15 of which had an antecedent case of VL, a serologically positive dog or both. Weak serological activity to rK39 antigen was detected in 3 of 488 human sera from the endemic areas. The presence of micro-foci of intense peri-urban transmission and the apparent absence of other Trypanosomatidae causing human disease offer a unique opportunity for the study of reservoirs, alternative vectors and evaluation of control measures on the Island.