Dimiconin, a novel coagulation inhibitor from the kissing bug, Triatoma dimidiata, a vector of Chagas disease.

Sequence analysis of a Triatoma dimidiata salivary gland cDNA library resulted in the identification of two transcripts (Td60 and Td101) homologous to triabin, an inhibitor of thrombin in Triatoma pallidipennis saliva. In the present study, a recombinant protein of Td60, designated dimiconin, was expressed in Escherichia coli and its activity was characterized. The resulting protein inhibited the intrinsic but not extrinsic blood coagulation pathway, suggesting that dimiconin is not a thrombin inhibitor. Measurement of the enzymatic activity of coagulation factors using chromogenic substrates revealed that dimiconin efficiently inhibited factor XIIa (FXIIa) activity in a dose-dependent manner. In addition, pre-incubation of dimiconin with FXII effectively inhibited FXIIa activity whereas dimiconin did not affect already activated FXIIa, indicating that dimiconin inhibits the activation of FXII but not the enzymatic activity of FXIIa. These results show that dimiconin is an inhibitor of the contact phase initiated by FXII activation in the blood coagulation cascade, which differs from the bioactivity of triabin.

PCR-RFLP analyses of Leishmania species causing cutaneous and mucocutaneous leishmaniasis revealed distribution of genetically complex strains with hybrid and mito-nuclear discordance in Ecuador.

PCR-Restriction Fragment Length Polymorphism (RFLP) analyses targeting multiple nuclear genes were established for the simple and practical identification of Leishmania species without using expensive equipment. This method was applied to 92 clinical samples collected at 33 sites in 14 provinces of Ecuador, which have been identified at the species level by the kinetoplast cytochrome b (cyt b) gene sequence analysis, and the results obtained by the two analyses were compared. Although most results corresponded between the two analyses, PCR-RFLP analyses revealed distribution of hybrid strains between Leishmania (Viannia) guyanensis and L. (V.) braziliensis and between L. (V.) guyanensis and L. (V.) panamensis, of which the latter was firstly identified in Ecuador. Moreover, unexpected parasite strains having the kinetoplast cyt b gene of L. (V.) braziliensis and nuclear genes of L. (V.) guyanensis, L. (V.) panamensis, or a hybrid between L. (V.) guyanensis and L. (V.) panamensis were identified. This is the first report of the distribution of a protozoan parasite having mismatches between kinetoplast and nuclear genes, known as mito-nuclear discordance. The result demonstrated that genetically complex Leishmania strains are present in Ecuador. Since genetic exchanges such as hybrid formation were suggested to cause higher pathogenicity in Leishmania and may be transmitted by more species of sand flies, further country-wide epidemiological studies on clinical symptoms, as well as transmissible vectors, will be necessary.

Geographic Distribution of Leishmania Species in Ecuador Based on the Cytochrome B Gene Sequence Analysis.

A countrywide epidemiological study was performed to elucidate the current geographic distribution of causative species of cutaneous leishmaniasis (CL) in Ecuador by using FTA card-spotted samples and smear slides as DNA sources. Putative Leishmania in 165 samples collected from patients with CL in 16 provinces of Ecuador were examined at the species level based on the cytochrome b gene sequence analysis. Of these, 125 samples were successfully identified as Leishmania (Viannia) guyanensis, L. (V.) braziliensis, L. (V.) naiffi, L. (V.) lainsoni, and L. (Leishmania) mexicana. Two dominant species, L. (V.) guyanensis and L. (V.) braziliensis, were widely distributed in Pacific coast subtropical and Amazonian tropical areas, respectively. Recently reported L. (V.) naiffi and L. (V.) lainsoni were identified in Amazonian areas, and L. (L.) mexicana was identified in an Andean highland area. Importantly, the present study demonstrated that cases of L. (V.) braziliensis infection are increasing in Pacific coast areas.

Development of a loop-mediated isothermal amplification method for rapid mass-screening of sand flies for Leishmania infection.

Entomological monitoring of Leishmania infection in leishmaniasis endemic areas offers epidemiologic advantages for predicting the risk and expansion of the disease, as well as evaluation of the effectiveness of control programs. In this study, we developed a highly sensitive loop-mediated isothermal amplification (LAMP) method for the mass screening of sand flies for Leishmania infection based on the 18S rRNA gene. The LAMP technique could detect 0.01 parasites, which was more sensitive than classical PCR. The method was robust and could amplify the target DNA within 1h from a crude sand fly template without DNA purification. Amplicon detection could be accomplished by the newly developed colorimetric malachite green (MG)--mediated naked eye visualization. Pre-addition of MG to the LAMP reaction solution did not inhibit amplification efficiency. The field applicability of the colorimetric MG-based LAMP assay was demonstrated with 397 field-caught samples from the endemic areas of Ecuador and eight positive sand flies were detected. The robustness, superior sensitivity, and ability to produce better visual discriminatory reaction products than existing LAMP fluorescence and turbidity assays indicated the field potential usefulness of this new method for surveillance and epidemiological studies of leishmaniasis in developing countries.

Strongylides in Achatina(Lissachatina)fulica (Mollusca, Achatinidae) in Havana, Cuba / Strongylides in Achatina(Lissachatina)fulica (Mollusca, Achatinidae) in Havana, Cuba

ABSTRACT Achatina (Lissachatina) fulica was introduced in Cuba in 2014 for african religious purposes and nowadays, mainly by human activity, it is widespread all over the country and, where besides a general nuisance for people it is a pest and also a public health concern, since it is one of the natural intermediate host of Angiostrongylus cantonensis, ethiological agent of the meningoencephalitis. As in Havana is experiencing the explosive phase of the invasion, LABCEL has been receiving samples of these molluscs for identification and search for Angiostrongylus cantonensis larvae. While examining samples of A. fulica different larvae were obtained, as strongylides found in the interior of the pallial cavity of A. fulica.This is the first report in San Miguel del Padron and Regla municipalities of the development of larvae in A. Fulica evidencing the health importance of this mollusc in the potential transmission of eosinophilic meningoencephalitis. Since the spread of A. fulica is pointed out in the literature as one of the main causative spread of the meningoencephalitis caused by A. cantonensis the authors emphasize the need of sanitary vigilance of snails and rats from vulnerable areas for A. cantonensis introduction as the port side areas like these municipalities.

RESUMEN Achatina (Lissachatina) fulica se introdujo en Cuba en 2014 con fines religiosos africanos y hoy en día, principalmente por actividad humana, está muy extendida en todo el país y, además de una molestia general para las personas, es una plaga y también un problema de salud pública, ya que es uno de los huéspedes intermedios naturales de Angiostrongylus cantonensis, agente etiológico de la meningoencefalitis. Como en La Habana está experimentando la fase explosiva de la invasión, LABCEL ha estado recibiendo muestras de estos moluscos para su identificación y búsqueda de larvas de Angiostrongylus cantonensis. Mientras se examinaban muestras de A. fulica, se obtuvieron diferentes larvas, como strongylides encontrados en el interior de la cavidad paliar de A. fulica. Este es el primer informe en los municipios de San Miguel del Padrón y Regla sobre el desarrollo de larvas en A. Fulica evidenciando La importancia para la salud de este molusco en la transmisión potencial de la meningoencefalitis eosinofílica. Dado que la propagación de A. fulica se señala en la literatura como una de las principales causas de la meningoencefalitis causada por A. cantonensis, los autores enfatizan la necesidad de vigilancia sanitaria de caracoles y ratas de áreas vulnerables para la introducción de A. cantonensis como zonas de babor como estos municipios.

Comparison of major immunoglobulins intrathecal synthesis patterns in Ecuadorian and Cuban patients with angiostrongyliasis.

Angiostrongylus cantonensis meningitis was first reported in Cuba in 1981, and it was recently reported in South America. The aim of this paper is to evaluate the intrathecal immunoglobulin synthesis patterns from Cuba's and Ecuador's patients with angiostrongyliasis; 8 Ecuadorian patients from two different outbreaks and 28 Cuban patients were studied. Simultaneous blood and cerebrospinal fluid samples were taken. Immunoglobulin (Ig) A, IgM, IgG, and albumin were quantified by radial immunodiffusion. Corresponding Reibergrams were applied. A three-Ig pattern was the most frequent in the two groups, but IgM was presented in all Ecuadorian young mature patients; however, in the Cuban children, only 12 of 28 patients had intrathecal IgM, but about 90% had an IgA and IgG synthesis at time of later puncture. This indicates that, with a larger amount of parasites ingested, clinical symptoms are more severe, and a higher frequency of intrathecal IgM synthesis could be observed. This is discussed as a similarity with the intrathecal IgM synthesis in African trypanosomiasis.

Angiostrongylus cantonensis: un parásito emergente en Ecuador / Angiostrongylus cantonensis: an emerging parasite in Ecuador

Introducción: en 2008 se notificó por primera vez la presencia de Angiostrongylus cantonensis en Ecuador, así como los primeros casos de una enfermedad emergente causada por sus larvas, la meningitis eosinofilica. Métodos: se realizó una revisión de la literatura básica y actualizada sobre aspectos generales de Angiostrongylus cantonensis en el mundo y particulares en Ecuador, que incluyó los hallazgos parasitológicos, clínicos y malacológicos relacionados con la enfermedad. Resultados: se informan los hallazgos iniciales acerca de la aparición del parásito en Ecuador, así como la amplia distribución geográfica de sus hospederos intermediarios en el territorio nacional. Además, se notifican los brotes de meningitis eosinofilica por Angiostrongylus cantonensis y un caso de angiostrongyliosis ocular, informados oficialmente por el Ministerio de Salud Pública. Conclusiones: Angiostrongylus cantonensis es un parásito emergente en Ecuador, cuyo diagnóstico en la actualidad es clínico y epidemiológico, de ahí la importancia de contar con métodos de laboratorio que lo oriente. Por otra parte, es importante que se promuevan campañas de promoción y prevención de salud que contribuyan a romper la cadena de transmisión de la enfermedad(AU)

Introduction: the presence of Angiostrongylus cantonensis and the first cases of eosinophilic meningitis, an emerging disease caused by its larvae, were first reported in Ecuador in the year 2008. Methods: a review was conducted of the basic and current bibliography on general aspects of Angiostrongylus cantonensis both worldwide and in Ecuador, including parasitological, clinical and malacological findings. Results: initial findings are reported about the emergence of the parasite in Ecuador, as well as the broad geographic distribution of its intermediate hosts in the national territory. Information is also provided about outbreaks of eosinophilic meningitis due to Angiostrongylus cantonensis and a case of ocular angiostrongylosis, based on official reports by the Ministry of Public Health. Conclusions: Angiostrongylus cantonensis is an emerging parasite in Ecuador whose diagnosis is currently based on clinical and epidemiological findings. Hence the importance of developing relevant laboratory methods. On the other hand, it is important to foster health promotion and prevention campaigns aimed at stopping the transmission of the disease(AU)

Angiostrongylus cantonensis: un parásito emergente en Ecuador / Angiostrongylus cantonensis: an emerging parasite in Ecuador

Introducción: en 2008 se notificó por primera vez la presencia de Angiostrongylus cantonensis en Ecuador, así como los primeros casos de una enfermedad emergente causada por sus larvas, la meningitis eosinofilica. Métodos: se realizó una revisión de la literatura básica y actualizada sobre aspectos generales de Angiostrongylus cantonensis en el mundo y particulares en Ecuador, que incluyó los hallazgos parasitológicos, clínicos y malacológicos relacionados con la enfermedad. Resultados: se informan los hallazgos iniciales acerca de la aparición del parásito en Ecuador, así como la amplia distribución geográfica de sus hospederos intermediarios en el territorio nacional. Además, se notifican los brotes de meningitis eosinofilica por Angiostrongylus cantonensis y un caso de angiostrongyliosis ocular, informados oficialmente por el Ministerio de Salud Pública. Conclusiones: Angiostrongylus cantonensis es un parásito emergente en Ecuador, cuyo diagnóstico en la actualidad es clínico y epidemiológico, de ahí la importancia de contar con métodos de laboratorio que lo oriente. Por otra parte, es importante que se promuevan campañas de promoción y prevención de salud que contribuyan a romper la cadena de transmisión de la enfermedad.

Introduction: the presence of Angiostrongylus cantonensis and the first cases of eosinophilic meningitis, an emerging disease caused by its larvae, were first reported in Ecuador in the year 2008. Methods: a review was conducted of the basic and current bibliography on general aspects of Angiostrongylus cantonensis both worldwide and in Ecuador, including parasitological, clinical and malacological findings. Results: initial findings are reported about the emergence of the parasite in Ecuador, as well as the broad geographic distribution of its intermediate hosts in the national territory. Information is also provided about outbreaks of eosinophilic meningitis due to Angiostrongylus cantonensis and a case of ocular angiostrongylosis, based on official reports by the Ministry of Public Health. Conclusions: Angiostrongylus cantonensis is an emerging parasite in Ecuador whose diagnosis is currently based on clinical and epidemiological findings. Hence the importance of developing relevant laboratory methods. On the other hand, it is important to foster health promotion and prevention campaigns aimed at stopping the transmission of the disease.

Seroprevalence of Trypanosoma cruzi Infection in Schoolchildren and in Pregnant Women from an Amazonian Region in Orellana Province, Ecuador

Chagas disease is a parasitic disease caused by the protozoan parasite Trypanosoma cruzi and about 230,000 persons are estimated to be infected in Ecuador. However, limited studies have been performed in the Amazon region, on the eastern side of the country. We evaluated here the seroprevalence of Trypanosoma cruzi infection in 12 rural villages of the Loreto canton, Orellana Province in schoolchildren aged 5-15 years and in pregnant women. A total of 1,649 blood samples were tested for Trypanosoma cruzi antibodies by enzyme-linked immunosorbent assay and indirect hemaglutination, and discordant samples were tested by indirect immunofluorescence assay. We detected a seroprevalence of anti-Trypanosoma cruzi antibodies of 1.3% in schoolchildren aged 5-15 years, indicating the persistence of a constant and active vectorial transmission in the Loreto County and confirming the need of the implementation of nonconventional vector control. We also observed a seroprevalence of 3.8% in pregnant women, indicating a clear risk of congenital transmission. Further studies should help define this risk more precisely and implement current international guidelines for the diagnosis, treatment, and care of these cases. (AU)