Leptospira spp y leptospirosis humana

Resumen Introducción. La leptospirosis, enfermedad bacteriana zoonótica y emergente más importante en el mundo, es causada por las especies patógenas de Leptospira spp. Objetivo. Revisar información reciente sobre Leptospira spp. y  leptospirosis humana con énfasis en América y Colombia. Metodología. Revisión de artículos indexados en PubMed, relacionados con la microbiología,  epidemiología, presentación clínica en humanos, diagnostico, tratamiento y prevención  de la enfermedad (vacunas). Resultados. Veinte especies de Leptospira spp han sido descritas;  se ha determinado la secuencia del ADN genómico de algunas cepas patógenas, la función de la mayoría de los genes involucrados en su patogénesis permanece desconocida. La leptospirosis humana presenta un rango de síntomas que van desde una fiebre indiferenciada hasta una ictericia, hemorragia, fallas renales y pulmonares severas. La administración temprana e intravenosa de penicilina G es requerida para reducir las tasas de mortalidad, pero los antibióticos pueden no ser efectivos en la enfermedad pulmonar severa. En las Americas, las areas de alto riesgo son Brasil, Centro América y el Caribe. Pocos estudios han sido  realizados en Colombia. La prueba serológica de oro, la microaglutinación  tiene alta sensibilidad y especificidad cuando se usan baterías de serovariedades locales pero es serogrupo específica. Las vacunas generan respuestas específicas para la serovariedad usada, pero no previenen la infección o trasmisión. Conclusiones. Problemas en el diagnóstico de laboratorio de la leptospirosis conllevan a un sub-registro en el número de casos; altas tasas de mortalidad asociadas a  fallas renal y pulmonar son resultado de las dificultades en el manejo de los casos

Comportamiento de la leptospirosis en el departamento del Atlántico (Colombia) Enero de 1999 a marzo del 2004

Resumen Objetivos: Describir el comportamiento epidemiológico de la leptospirosis en el departamento del Atlántico (Colombia), de enero de 1999 a marzo del 2004. Metodología: Estudio descriptivo. Se analizaron 970 muestras únicas de pacientes sospechosos de infecciones con Leptospira en el Laboratorio Departamental del Atlántico mediante Aglutinación Microscópica (MAT), usando como antígenos los serovares Icterohemorragiae, Pomona, Canícola, Hardjo, Grippotyphosa y Hardjo-bovis de Leptospira interrogans. Información adicional sobre la clínica de pacientes a través de fichas epidemiológicas y visitas a hospitales, además de datos de precipitación anual, fue obtenida. Resultados: El 9,7% de los casos fueron positivos para Leptospira, siendo los serovares Icterohaemor- rhagiae (62%) y Hardjo (12,8%) los más frecuentes. La mayoría de casos (61%), se presentaron en hombres entre 15 y 45 años de edad y la clínica más común se asoció a fiebre (91,7%), mialgias (72,2%), vómito/nausea (70,8%), cefalea (68,1%) e ictericia (63,9%). El 8.6% de los casos asociados a infec- ciones con el serovar Ictherohemorragiae fueron severos; la sintomatología coincidió con el síndrome de Weil, pero no se registraron fatalidades. En los años 2003(23), 2001(21) y 2002(18) se registró la mayor incidencia de casos, en meses de alta precipitación (Agosto-Noviembre). Los municipios con mayor número de casos fueron Barranquilla (46), Soledad (25), Puerto Colombia (6) y Galapa (6). Conclusiones: La leptospirosis debe tenerse en cuenta dentro del diagnóstico diferencial de otras entidades comunes en la región (fiebre de dengue). Un diligenciamiento completo de la ficha epidemio- lógica permitirá un estudio más detallado de esta patología, para desarrollar programas de vigilancia y prevención eficaces. Palabras claves: Leptospira, serología, Atlántico, Colombia. Abstract Objective: This study was performed to describe the epidemiological situation of Leptospira in the Departament of Atlantico (Colombia), from January 1999 to March 2004. Methods: A descriptive study was performed. A total of 970 single serum samples from patients with suspected Leptospira infections, were analyzed using the microscopic agglutination test (MAT). The serovars of Icterohaemorrhagiae, Pomona, Canicola, Hardjo, Grippotyphosa and Hardjo-bovis belonging to L. interrogans, were used as antigens. Information about clinical presentation based on epidemiological sheets, visits to patients and climatological data were obtained. Results: The 9,7% samples were IgM positive for Leptospira and the most prevalent was the serovar Icterohaemorrhagiae (62%), followed by Hardjo (12.8%). Most of the patients were male (61%) between 15 and 45 y.o. The most common presenting features in these patients were (91.7%), myalgia (72%), vomit/nausea (70.8%), headache (68.1%) and icterichia (63.9%). 8.6% of the cases were severe, associated to infections with the serovar Icterohemorragiae and their symptomathology was similar to the Weil ?s syndrome; no fatalities were registered. The highest incidences were recorded during the years 2003 (23), 2001(21) and 2002 (18) especially during the rainy season (August-November). Barranquilla reported the highest number of cases (46) followed by Soledad (25), Puerto Colombia (6) and Galapa (6). Conclusions: Since leptospirosis is an increasing public health problem in the Caribbean Region, dif- ferential diagnosis with other similar pathologies (dengue fever, dengue haemorrhagic fever) has to be performed and surveillance and preventive programmes must be implemented. Key words: Leptospirosis, serology, Atlántico, Colombia

Detection of immune-complex-dissociated nonstructural-1 antigen in patients with acute dengue virus infections

Accurate and timely diagnosis of dengue virus (DEN) infections is essential for the differential diagnosis of patients with febrile illness and hemorrhagic fever. In the present study, the diagnostic value of a newly developed immune-complex dissociated nonstructural-1 (NS-1) antigen dot blot immunoassay (DBI) was compared to a commercially available DEN antigen detection kit (denKEY Blue kit; Globio Co., Beverly, Mass.) and a reverse transcription-PCR (RT-PCR) kit. Serial serum or plasma samples (n = 181) obtained from 55 acute DEN-infected patients were used. In samples obtained from 32 of these 55 DEN-infected patients, viral RNA could be detected by RT-PCR. DEN antigen was detected in only 10 of these 55 patient samples by using the denKEY kit. When these samples were treated with acid to release the immune-complex-associated NS-1 antigen for detection by DBI, 43 of these 55 patients were found to be positive for DEN NS-1 antigen. In nondiss

The NS1 Glycoprotein Can Generate Dramatic Antibody-Enhanced Dengue Viral Replication in Normal Out-Bred Mice Resulting in Lethal Multi-Organ Disease

Antibody-enhanced replication (AER) of dengue type-2 virus (DENV-2) strains and production of antibody-enhanced disease (AED) was tested in out-bred mice. Polyclonal antibodies (PAbs) generated against the nonstructural-1 (NS1) glycoprotein candidate vaccine of the New Guinea-C (NG-C) or NSx strains reacted strongly and weakly with these antigens, respectively. These PAbs contained the IgG2a subclass, which cross-reacted with the virion-associated envelope (E) glycoprotein of the DENV-2 NSx strain, suggesting that they could generate its AER via all mouse Fcγ-receptor classes. Indeed, when these mice were challenged with a low dose (<0.5 LD50) of the DENV-2 NSx strain, but not the NG-C strain, they all generated dramatic and lethal DENV-2 AER/AED. These AER/AED mice developed life-threatening acute respiratory distress syndrome (ARDS), displayed by diffuse alveolar damage (DAD) resulting from i) dramatic interstitial alveolar septa-thickening with mononuclear cells, ii) some hyperplasia of alveolar type-II pneumocytes, iii) copious intra-alveolar protein secretion, iv) some hyaline membrane-covered alveolar walls, and v) DENV-2 antigen-positive alveolar macrophages. These mice also developed meningo-encephalitis, with greater than 90,000-fold DENV-2 AER titers in microglial cells located throughout their brain parenchyma, some of which formed nodules around dead neurons. Their spleens contained infiltrated megakaryocytes with DENV-2 antigen-positive red-pulp macrophages, while their livers displayed extensive necrosis, apoptosis and macro- and micro-steatosis, with DENV-2 antigen-positive Kuppfer cells and hepatocytes. Their infections were confirmed by DENV-2 isolations from their lungs, spleens and livers. These findings accord with those reported in fatal human “severe dengue” cases. This DENV-2 AER/AED was blocked by high concentrations of only the NG-C NS1 glycoprotein. These results imply a potential hazard of DENV NS1 glycoprotein-based vaccines, particularly against DENV strains that contain multiple mutations or genetic recombination within or between their DENV E and NS1 glycoprotein-encoding genes. The model provides potential for assessing DENV strain pathogenicity and anti-DENV therapies in normal mice

Significantly lower anti-Leishmania IgG responses in Sudanese versus Indian visceral leishmaniasis.

BACKGROUND: Visceral leishmaniasis (VL), a widely distributed systemic disease caused by infection with the Leishmania donovani complex (L. donovani and L. infantum), is almost always fatal if symptomatic and untreated. A rapid point-of-care diagnostic test for anti-Leishmania antibodies, the rK39-immunochromatographic test (rK39-ICT), has high sensitivity and specificity in South Asia but is less sensitive in East Africa. One of the underlying reasons may be continent-specific molecular diversity in the rK39 antigen within the L. donovani complex. However, a second reason may be differences in specific IgG anti-Leishmania levels in patients from different geographical regions, either due to variable antigenicity or immunological response. METHODOLOGY/PRINCIPAL FINDINGS: We determined IgG titres of Indian and Sudanese VL patients against whole cell lysates of Indian and Sudanese L. donovani strains. Indian VL patients had significantly higher IgG titres against both L. donovani strains compared to Sudanese VL patients (p<0.0001). Mean reciprocal log10 50% end-point titres (1/log10t50) were i) 3.80 and 3.88 for Indian plasma and ii) 2.13 and 2.09 for Sudanese plasma against Indian and Sudanese antigen respectively (p<0.0001). Overall, the Indian VL patients therefore showed a 46.8-61.7 -fold higher mean ELISA titre than the Sudanese VL patients. The higher IgG titres occurred in children (<16 years old) and adults of either sex from India (mean 1/log10t50: 3.60-4.15) versus Sudan (mean 1/log10t50: 1.88-2.54). The greatest difference in IgG responses was between male Indian and Sudanese VL patients of ≥ 16 years old (mean 1/log10t50: 4.15 versus 1.99 = 144-fold (p<0.0001). CONCLUSIONS/SIGNIFICANCE: Anti-Leishmania IgG responses among VL patients in Sudan were significantly lower than in India; this may be due to chronic malnutrition with Zn(2+) deficiency, or variable antigenicity and capacity to generate IgG responses to Leishmania antigens. Such differential anti-Leishmania IgG levels may contribute to lower sensitivity of the rK39-ICT in East Africa

Manifestaciones clínicas y factores de riesgo asociados a la infección por Cryptosporidium en pacientes de Barranquilla y tres municipios del Atlántico (Colombia)

Resumen Objetivos: Caracterizar manifestaciones clínicas y factores de riesgo asociados a Cryptos- poridiosis. Materiales y métodos: Se realizó un estudio descriptivo transversal en 423 pacientes, con análisis macroscópico y microscópico de muestras fecales, para identificar manifestaciones clínicas y factores de riesgo asociados a Cryptosporidiosis en tres municipios del Atlántico y su capital en un período de 4 meses. Se identificaron ooquistes de Cryptosporidium spp. después de teñirse con Ziehl-Neelsen modificado. Se comparó el método de concentración NaCl saturado con el método en fresco, para la detección de parásitos intestinales en 279/423 (66.0%) pacientes. El análisis estadístico se realizó usando EPI-INFO 6.04. Resultados: La prevalencia de Cryptosporidium spp. fue 1.9% (8/423). Se encontraron asociaciones estadísticas entre cryptosporidiosis y fiebre (p=0.01), sangre en muestras fe- cales (p=0.01) y presencia de animales domiciliarios (p=0.02). La mayoría de los pacientes (267/423:20.3 %) fueron positivos para parásitos intestinales. Los parásitos identificados con mayor frecuencia fueron protozoos no enteropatógenos, Entamoeba coli (118/423: 27.9%) y Endolimax nana (86/423: 20.3%), seguido de Blastocystis hominis (76/423: 18%), Entamoeba histolytica/dispar (28/423: 6.6%) y Giardia lamblia (23/423: 5.4%). Ascaris lumbricoides (6/423: 1.4%) fue el helminto identificado con mayor frecuencia. Una sensibilidad/especificidad de 99.45/95.2% y 87.5%/99.6% se obtuvo para protozoos y helmintos respectivamente usando el método de NaCl saturado. Conclusiones: Los pacientes con cryptosporidosis tuvieron fiebre y muestras fecales san- guinolentas. Probablemente fueron infectados por animales domésticos. Microscópicamente, la utilización de la tinción Ziehl Neelsen modificado fue esencial para la identificación de ooquistes de Cryptosporidium spp. El método de NaCl saturado concentró eficientemente los parásitos. Palabras claves: Cryptosporidium spp., Ziehl-Neelsen modificado, parasitosis in- testinales. Abstract Objectives: To characterize the clinical manifestations and risk factors associated with cryptosporidiosis. Materials and methods: A descriptive study was performed on 423 patients, with mac- roscopic and microscopic faecal sample analyses, to identify the clinical manifestations and risk factors associated with cyptosporidiosis in 3 towns and the principal city in Atlantico (Colombia) over a 4-month period. Cryptosporidium spp oocysts were identified after stain- ing with modified Ziehl-Neelsen. A saturated NaCl parasite-concentration method was also compared with wet-mount method for the detection of all intestinal parasites in 279/423 (66.0%) patients. Statistical analyses were performed using EPI-INFO 6.04. Results: The prevalence of Cryptosporidium spp. was 1.9% (8/423). Statistical associations were found between cryptosporidiosis infections and fever (p=0.01), blood in the faecal samples (p=0.01) and the presence of household animals (p=0.02). Most of the patients (267/423: 63.1%) were positive for intestinal parasites. The most commonly identified parasites were the non-pathogenic protozoa, Entamoeba coli (118/423: 27.9%) and Endoli- max nana (86/423: 20.3%), followed by Blastocystis hominis (76/423: 18%), Entamoeba histolytica/dispar (28/423: 6.6%) and Giardia lamblia (23/423: 5.4%). Ascaris lumbricoi- des (6/423: 1.4%) was the most common helminth identified. Sensitivities/specificities of 99.4%/95.2% and 87.5%/99.6% were obtained for protozoa and helminths respectively using the saturated NaCl method. Conclusions: Patients with cryptosporidiosis had fever and bloody faecal samples, and were probably infected by domestic animals. Microscopy, using the modified Ziehl-Neelsen stain, was essential for Cryptosporidium spp. oocyst identification. The saturated NaCl method efficiently concentrated the parasites. Key words: Cryptosporidium spp., modified Ziehl-Neelsen stain, intestinal parasi- tesis

Development of peptide-based lineage-specific serology for chronic Chagas disease: geographical and clinical distribution of epitope recognition.

BACKGROUND: Chagas disease, caused by infection with the protozoan Trypanosoma cruzi, remains a serious public health issue in Latin America. Genetically diverse, the species is sub-divided into six lineages, known as TcI-TcVI, which have disparate geographical and ecological distributions. TcII, TcV, and TcVI are associated with severe human disease in the Southern Cone countries, whereas TcI is associated with cardiomyopathy north of the Amazon. T. cruzi persists as a chronic infection, with cardiac and/or gastrointestinal symptoms developing years or decades after initial infection. Identifying an individual's history of T. cruzi lineage infection directly by genotyping of the parasite is complicated by the low parasitaemia and sequestration in the host tissues. METHODOLOGY/PRINCIPAL FINDINGS: We have applied here serology against lineage-specific epitopes of the T. cruzi surface antigen TSSA, as an indirect approach to allow identification of infecting lineage. Chagasic sera from chronic patients from a range of endemic countries were tested by ELISA against synthetic peptides representing lineage-specific TSSA epitopes bound to avidin-coated ELISA plates via a biotin labelled polyethylene glycol-glycine spacer to increase rotation and ensure each amino acid side chain could freely interact with their antibodies. 79/113 (70%) of samples from Brazil, Bolivia, and Argentina recognised the TSSA epitope common to lineages TcII/TcV/TcVI. Comparison with clinical information showed that a higher proportion of Brazilian TSSApep-II/V/VI responders had ECG abnormalities than non-responders (38% vs 17%; p<0.0001). Among northern chagasic sera 4/20 (20%) from Ecuador reacted with this peptide; 1/12 Venezuelan and 1/34 Colombian samples reacted with TSSApep-IV. In addition, a proposed TcI-specific epitope, described elsewhere, was demonstrated here to be highly conserved across lineages and therefore not applicable to lineage-specific serology. CONCLUSIONS/SIGNIFICANCE: These results demonstrate the considerable potential for synthetic peptide serology to investigate the infection history of individuals, geographical and clinical associations of T. cruzi lineages

Comparison of Three Commercially Available Dengue NS1 Antigen Capture Assays for Acute Diagnosis of Dengue in Brazil

Dengue is the one of the most prevalent arthropod-borne viral diseases in tropical regions of the world. Manifestations may vary from asymptomatic to potentially fatal complications. Laboratorial diagnosis is essential to diagnose dengue and differentiate it from other diseases. Dengue virus non-structural protein 1 (NS1) may be used as a marker of acute dengue virus infection. Our results, based in the comparison of three NS1 antigen capture assays available, have shown that this approach is reliable for the early diagnosis of dengue infections, especially in the first four days after the onset of the symptoms. A lower sensitivity was observed in DENV-3 cases. Serum positive by virus isolation were more often detected than those positive by RT-PCR by all three assays. Only the Platelia™ NS1 test showed a higher sensitivity in confirming primary infections than secondary ones. In conclusion, NS1 antigen capture commercial kits are useful for diagnosis of acute primary and secondary dengue infections and, in endemic countries where secondary infections are expected to occur, may be used in combination with MAC-ELISA to increase the overall sensitivity of both tests

Distinct Activation Phenotype of a Highly Conserved Novel HLA-B57-Restricted Epitope during Dengue Virus Infection

Variation in the sequence of T cell epitopes between dengue virus (DENV) serotypes is believed to alter memory T cell responses during second heterologous infections. We identified a highly conserved, novel, HLA-B57-restricted epitope on the DENV NS1 protein. We predicted higher frequencies of B57-NS126-34-specific CD8+ T cells in PBMC from individuals undergoing secondary rather than primary DENV infection. However, high tetramer-positive T cell frequencies during acute infection were seen in only 1 of 9 subjects with secondary infection. B57-NS126-34-specific and other DENV epitope-specific CD8+ T cells, as well as total CD8+ T cells, expressed an activated phenotype (CD69+ and/or CD38+) during acute infection. In contrast, expression of CD71 was largely limited to DENV epitope-specific CD8+ T cells. In vitro stimulation of cell lines indicated that CD71 expression was differentially sensitive to stimulation by homologous and heterologous variant peptides. CD71 may represent a useful marker of antigen-specific T cell activation