Malaria remains as a public health problem in Venezuela. In 2015 there were 136,402 cases reported by the Ministry of Popular Power for Health, being the parasite prevalence 73.95% for Plasmodium vivax, 17.6% for Plasmodium falciparum, 0.0095% for Plasmodium malariae and 8.42% mixed infections (P. vivax + P. falciparum). During the period 1999-2002 the number of cases in Venezuela ranged between 21,685 and 29,337, being the Sucre State with highest levels of malaria prevalence, with Plasmodium vivax as the unique specie in this region. In 2002 the Municipality of Cajigal had the highest Annual Parasite Incidence (API) of country, being 260 cases per 1000 inhabitants. In view of the difficulty in controlling malaria in this area, the prevalence of asymptomatic carriers was investigated as one of the epidemiological factors contributing to the persistence of malaria transmission. One hundred fifty people were included in the study, with no history of recent malaria infection, or any symptom and also, not having used antimalarial drugs during the 30 days prior to study entry. To do this, a malaria Rapid Diagnostic Test (mRDTs) was used for the determination of antigenemic (OptiMAL®) and PCR (polymerase chain reaction) in conjunction with the reference "Gold Standard", the conventional thick and thin blood smears (TTBS). It was found a prevalence of infection of 1.33% by mRDTs and TTBS and 8% by PCR which allowed the detection of 10 asymptomatic cases in addition, with a sensitivity and specificity of 100% and 93.4% respectively. The presence of asymptomatic carriers in this area reveals the difficulties that face the Malaria Control Program in the eventual elimination of this specific malaria foci. It is necessary reinforces the maintenance of the epidemiological surveillance using more sensitive diagnostic techniques, as well as to adapt the control measures based on the current findings.

Se planteó identificar antígenos que pudieran ser reconocidos por los anticuerpos IgG1 e IgG3, descritos como protectores en la infección malárica, en personas con respuesta clínica adecuada (RCA) o falla al tratamiento (FT) antimalárico, provenientes de localidades con diferentes grados de endemicidad. Se evaluaron por Immunoblotting muestras de sueros de individuos provenientes de tres localidades del Edo. Amazonas (Venezuela): Puerto Ayacucho (Atures), San Juan de Manapiare (Manapiare) y Platanal (Alto Orinoco). La reactividad de IgG, IgG1 e IgG3 frente a componentes antigénicos del extracto de P. falciparum (FCB2), permitió identificar un mayor número de moléculas específicas en los pacientes con RCA que en los pacientes con FT. La frecuencia de reconocimiento de polipéptidos fue baja en las tres localidades, algunas moléculas con una frecuencia de reconocimiento igual o mayor al 20% pertenecían a sueros de individuos de las localidades de Puerto Ayacucho y Platanal, ambas con exposición permanente a P. falciparum. Dado el reconocimiento de polipéptidos por IgG, IgG1 e IgG3 en sueros de pacientes con RCA, estos podrían ser considerados como posibles blancos relevantes de la respuesta inmunológica protectora que coadyuven con el tratamiento antimalárico. Esto contribuiría al desarrollo y diseño de vacunas más efectivas, que prevengan la infección malárica y/o potencien la eficacia a la quimioterapia.

Here we studied the presence of antigens recognized by IgG1 and IgG3 antibodies, thought as protective, in patients with adequate clinical response (RCA) or treatment failure (FT), living in areas of different degrees of endemicity. Immunoblotting was evaluated from serum samples of individuals from three locations in the State Amazonas (Venezuela): Puerto Ayacucho (Atures), San Juan de Manapiare (Manapiare) and Pantanal (Alto Orinoco). The reactivity of IgG, IgG1 and IgG3 against antigenic components of the extract of P. falciparum (FCB2) identified a greater number of specific molecules in patients with RCA in patients with AFT. The frequency of recognition of polypeptides was low in all three locations, with some molecules having a recognition rate of greater than or equal to 20% sera of individuals belonging to the towns of Puerto Ayacucho and Platanal, both with cases of P. falciparum. Given the recognition of polypeptides by IgG, IgG1 and IgG3 in sera of patients with RCA, they could be considered as possible targets for relevant protective immune responses that contribute to malaria treatment. This would contribute to the development and design of more effective vaccines that prevent malaria infection and/or enhance the efficacy of chemotherapy.

Se evaluó el test rápido OptiMAL® comparándolo con el examen convencional de observación de muestras sanguíneas para la detección de la malaria en dos grupos de individuos procedentes de diferentes áreas endémicas de Venezuela. Uno de los grupos (n = 113) con síntomas sugestivos de malaria, y otro representado por individuos asintomáticos (n = 89). El examen microscópico de las muestras de sangre de estas poblaciones determinó que 67,5% estaban infectados con P.vivax, 31,3% con P. falciparum y 1,2% con infecciones mixtas. La prueba OptiMAL® mostró 96,4% de sensibilidad, 100% de especificidad, con valor predictivo positivo de 100% y valor predictivo negativo de 97,5%. La detección de cualquier infección malárica en la población total presentó una concordancia óptima (kappa = 0,97). Sin embargo, estos parámetros fueron más bajos cuando la parasitemia era £ 300 parásitos/µL. El congelamiento de las muestras no afectó la sensibilidad y especificidad de la prueba. Nosotros concluimos que esta prueba rápida de malaria es sensible y especifica para el diagnóstico rápido de la malaria en el campo y puede complementar a la microcopia convencional.

We evaluated the OptiMAL® rapid dipstick test by comparing it with the conventional standard thick-blood film method, for the detection of malaria in two groups of individuals from different Venezuelan endemic areas. One of the groups consisted of individuals with malaria-like symptoms (n = 113) and the other of asymptomatic individuals (n = 89). The classical microscopy analysis of these populations determined that 67.5% were infected with P. vivax, 31.3% with P. falciparum, and 1.2% with mixed infections. The OptiMAL® test showed 96.4% sensitivity, 100% specificity, 100% positive predictive value, 97.5% negative predictive value and optimal concordance (kappa = 0.97), capable of detecting any malaria infection in the evaluated population. However, these parameters were lower when the parasitaemia was £ 300 parasites/µL. Freezing of the samples did not affect the sensitivity and specificity of the test. We concluded that this rapid malaria test is sensitive and specific for rapid diagnosis of malaria in the field and it is a complement to conventional microscopy.

Se evaluó la prueba Now® malaria ICT P.f/P.v. en personas con síntomas de malaria provenientes de dos áreas endémicas de Venezuela: estado Sucre en la región noreste costera (n=71) y el estado Amazonas en el sur (n=86). 102 muestras resultaron positivas (73 P. vivax, 25 P. falciparum, 2 P. malariae y 2 infecciones mixtas), 55 muestras resultaron negativas mediante el examen microscópico. La sensibilidad, especificidad, VPP y VPN del Now® malaria ICT P.f/P.v. para el diagnóstico de malaria fue 81,4%, 100%, 100% y 74,3%, respectivamente y un Kappa de 0,75. En Sucre la sensibilidad fue de 96,3%, especificidad 100%, VPP 100%, VPN 89,5% y Kappa 0,93 para P. vivax. En Amazonas, la sensibilidad fue 81%, especificidad y VPP 100%, VPN 90,5% y Kappa 0,85 para infecciones activas por P. vivax y P. malariae. Esta sensibilidad para P. vivax aumenta con parasitemias entre 101-500 par/μL. Para P. falciparum la sensibilidad fue 51,9%, especificidad de 100%, VPP de 100%, VPN de 74,5% y un Kappa de 0,56. La sensibilidad del método es de 100% para densidades mayores a 500 par/μL. La concordancia encontrada en este estudio entre la prueba Now® malaria ICT P.f/P.v. y el método parasitológico para las especies P. vivax / P. malariae, aunque no para P. falciparum, aporta un valor adicional a las ya conocidas características de la prueba tales como procedimiento fácil, rapidez e interpretación de los resultados en áreas de difícil acceso, por lo que concluimos que es una herramienta alternativa para el diagnóstico de malaria, sin llegar a sustituir el método parasitológico convencional.

We evaluated the Now® malaria ICT P.f/ P.v. test in symptomatic malaria patients from two different endemic areas of Venezuela: Sucre state in the northeastern costal region (n=71) and the Amazonas state in the south (n=86). One hundred two patients were positive by thick and thin blood smears, 73 with Plasmodium vivax , 25 with P. falciparum , 2 with P. malariae and 2 with mixed infections. Some 55 blood samples were negative. The sensitivity, specificity, PPV, NPV of the Now® malaria ICT P.f/P.v. test, were 81.4%, 100%, 100% and 74.3%, respectively and a Kappa of 0.75. With blood samples from Sucre state, the sensitivity was 96.3%, the specificity 100%, the PPV 100%, NPV 89.5% and a Kappa of 0.93 for P. vivax. In Amazonas State, the sensitivity was 81%, the specificity and PPV 100%, the NPV 90.5% and a Kappa of 0.85 for P. vivax and P. malariae. This sensitivity increased for P. vivax when parasitaemias were above 101 parasites/uL. For P. falciparum , the sensitivity was 51.9%, the specificity 100%, the PPV 100%, NPV was 74.5% and a Kappa of 0.56. The sensitivity was 100%, with more than 500 parasites/mL for this species. The high concordance found in this test with the thick and thin blood smear for all malaria species except P. falciparum gives an additional value to this dipstick, along with other well known characteristics such as the rapid and easy handling and interpretation in areas of difficult access. We concluded that the immunochromatographic test is a valuable additional diagnostic tool for malaria but not a substitute of the classical thin and thick blood smear.

The presence of common antigens between Plasmodium falciparum and Anopheles albimanus was demonstrated. Different groups of rabbits were immunized with: crude extract from female An. albimanus (EAaF), red blood cells infected with Plasmodium falciparum (EPfs), and the SPf66 synthetic malaria vaccine. The rabbit's polyclonal antibodies were evaluated by ELISA, Multiple Antigen Blot Assay (MABA), and immunoblotting. All extracts were immunogenic in rabbits according to these three techniques, when they were evaluated against the homologous antigens. Ten molecules were identified in female mosquitoes and also in P. falciparum antigens by the autologous sera. The electrophoretic pattern by SDS-PAGE was different for the three antigens evaluated. Cross-reactions between An. albimanus and P. falciparum were found by ELISA, MABA, and immunoblotting. Anti-P. falciparum and anti-SPf66 antibodies recognized ten and five components in the EAaF crude extract, respectively. Likewise, immune sera against female An. albimanus identified four molecules in the P. falciparum extract antigen. As far as we know, this is the first work that demonstrates shared antigens between anophelines and malaria parasites. This finding could be useful for diagnosis, vaccines, and the study of physiology of the immune response to malaria.

Epítopes de antígenos compartidos entre Plasmodium falciparum y Anopheles albimanus fueron identificados. Diferentes grupos de conejos fueron inmunizados con: extracto crudo de mosquito hembra de An. albimanus (EAaH), glóbulos rojos infectados con P. falciparum (EPfs) y la vacuna antimalárica sintética SPf66. Los anticuerpos policlonales producidos en conejos fueron evaluados por ELISA, inmunoensayo simultáneo de múltiples antígenos (MABA) e Immunoblotting. Todos los extractos resultaron inmunogénicos cuando se evaluaron por ELISA, MABA e Immunoblotting. Diez moléculas fueron identificadas en los mosquitos hembras y diez en los antígenos de P. falciparum por los sueros autólogos. El patrón electroforético por SDS-EGPA fue diferente para los tres antígenos evaluados. La reactividad cruzada de moléculas entre An. albimanus y P. falciparum fue demostrada por ELISA, MABA e Immunoblotting. Anticuerpos anti-P. falciparum y anti-SPf66 reconocieron diez y cinco componentes respectivamente en el extracto crudo de anofelinos (EAaH). Asimismo, sueros inmunes contra An. albimanus hembra identificaron cuatro moléculas en el extracto del antígeno de P. falciparum. Hasta el presente, este es el primer estudio en el que se demuestra la presencia de antígenos compartidos entre anofelinos y los parásitos de malaria. Este hallazgo podría ser de relevancia para el diagnóstico, vacunas e interpretación de la fisiopatología de la respuesta inmunitaria en malaria.