Dicistroviridae: A new viral purification technique / Dicistroviridae: una nueva técnica de purificación viral

The family Dicistroviridae comprises three genera and about twenty species of RNA virus, most of them with health or agricultural importance. The Triatoma virus (TrV) is the only entomopathogenic virus identified in triatomine bugs up to the present. TrV replicates within the intestinal epithelial cells, causing high mortality rate and delayed development of the molt of these bugs. TrV has been proposed as a biological control agent for vectors of Chagas disease. Viral particles were purified from feces of 1, 5 and 10 insects from an experimental colony of Triatoma infestans infected with TrV. Viral concentration and infectivity were corroborated using polyacrylamide gels and RT-PCR, respectively. In this work we report a method of viral purification that allows to reduce necessary reagents and time, using a very small amount of fecal matter.

La familia Dicistroviridae está compuesta por tres géneros y casi una veintena de especies de virus ARN, la mayoría de ellas de importancia sanitaria o agrícola. Triatoma virus (TrV) es el único virus entomopatógeno identificado en triatominos hasta el momento. El TrV se replica en las células del epitelio intestinal; ello provoca una alta tasa de mortalidad y retraso en el desarrollo de la muda del insecto. Se ha propuesto la utilización de TrV como agente de control biológico para vectores de la enfermedad de Chagas. Las partículas virales fueron purificadas a partir de materia fecal de 1, 5 y 10 insectos obtenidos de una colonia experimental infectada con TrV de Triatoma infestans y se corroboró su concentración viral e infectividad mediante geles de poliacrilamida y RT-PCR, respectivamente. En este trabajo se reporta un método de purificación viral que permite la reducción de los reactivos y del tiempo necesario para lograr dicha purificación, partiendo de una mínima cantidad de materia fecal.

Dicistroviridae: A new viral purification technique.

The family Dicistroviridae comprises three genera and about twenty species of RNA virus, most of them with health or agricultural importance. The Triatoma virus (TrV) is the only entomopathogenic virus identified in triatomine bugs up to the present. TrV replicates within the intestinal epithelial cells, causing high mortality rate and delayed development of the molt of these bugs. TrV has been proposed as a biological control agent for vectors of Chagas disease. Viral particles were purified from feces of 1, 5 and 10 insects from an experimental colony of Triatoma infestans infected with TrV. Viral concentration and infectivity were corroborated using polyacrylamide gels and RT-PCR, respectively. In this work we report a method of viral purification that allows to reduce necessary reagents and time, using a very small amount of fecal matter.

Resistencia bacteriana de Escherichia coli uropatogénica en población nativa amerindia Kichwa de Ecuador / Bacterial resistance of uropathogenic Escherichia coli in native Kichwa Amerindian population of Ecuador

Contexto: Escherichia coli uropatógena (ECUP) se presenta como uno de los principales agentes etiológicos en infecciones del tracto urinario (ITUs) no complicadas (70-95%). El objetivo del tratamiento de ITUs no complicadas es obtener curación clínica y microbiológica. Para ello, es de particular importancia el conocimiento de las tasas de resistencia antibiótica local. Objetivo: identificar los perfiles de resistencia a antibióticos de primera línea para ITUs no complicadas en poblaciones nativas amerindias Kichwas ecuatorianas, en donde el tratamiento empírico se basa en trimetoprim/sulfametoxazol, ampicilina, y ciprofloxacina mayoritariamente. Métodos: se analizaron 335 muestras de orina procedentes de las poblaciones de Zumbahua, Colta y Guamote, en un periodo de 4 meses (febrero-mayo 2016). Las muestras fueron incubadas por 24 y 48 horas en agar Eosin Methylene Blue (EMB), para luego ser identificadas en género y especie por pruebas bioquímicas. Para determinar la susceptibilidad antibiótica, se realizó la técnica de difusión en disco de Kirby-Bauer. Para la Concentración Inhibitoria Mínima (CIM), se utilizó la técnica de microdilución en caldo (Vitek 2). El método de doble disco fue la técnica utilizada para la detección de betalactamasas de espectro extendido (BLEE). Resultados: noventa (26,9%) muestras mostraron un recuento significativo de ≥105 (ufc)/ml, compatibles con ITUs. El microorganismo identificado con mayor frecuencia fue E. coli (n=75; 83,3%). La resistencia antibiótica encontrada para los aislados de E. coli fue de 56,7% a trimetoprim/sulfametoxazol, 52,5% a ampicilina, 43.3% a ácido nalidíxico, 32.5% a ciprofloxacina, 28.3% a norfloxacina, 25% a levofloxacina, 15.85% a cefazolina, 17.5% a cefoxitina, 15% a cefuroxima, 15% a ceftazidima, cefotaxima, y ceftriaxona, 15% a cefepima, 7,5% a nitrofurantoina y 1,7% a fosfomicina. Se identificaron 7 aislados productores de betalactamasas de espectro extendido (BLEE). Conclusión: con los resultados obtenidos se recomienda no utilizar ampicilina, trimetoprim/sulfametoxazol, ni quinolonas en la zona estudiada como terapia empírica. Se sugiere instaurar tratamiento empírico con fosfomicina o nitrofurantoina para ITUs no complicadas. (AU)

Seroprevalence of Triatoma virus (Dicistroviridae: Cripaviridae) antibodies in Chagas disease patients.

BACKGROUND: Chagas disease is caused by Trypanosoma cruzi, and humans acquire the parasite by exposure to contaminated feces from hematophagous insect vectors known as triatomines. Triatoma virus (TrV) is the sole viral pathogen of triatomines, and is transmitted among insects through the fecal-oral route and, as it happens with T. cruzi, the infected insects release the virus when defecating during or after blood uptake. METHODS: In this work, we analysed the occurrence of anti-TrV antibodies in human sera from Chagas disease endemic and non-endemic countries, and developed a mathematical model to estimate the transmission probability of TrV from insects to man, which ranged between 0.00053 and 0.0015. RESULTS: Our results confirm that people with Chagas disease living in Bolivia, Argentina and Mexico have been exposed to TrV, and that TrV is unable to replicate in human hosts. CONCLUSIONS: We presented the first experimental evidence of antibodies against TrV structural proteins in human sera.

Prevalence and distribution of parasites and pathogens of triatominae from Argentina, with emphasis on Triatoma infestans and Triatoma virus TrV.

Chagas' disease is the most important endemic arthropod-zoonosis in Argentina with an estimated 1.6 million people infected with the causative agent Trypanosoma cruzi. Triatoma infestans is the main vector of Chagas' disease in Argentina. A survey for parasites and pathogens of Triatominae was conducted from August 2002 to February 2005. Collections of insects were made in domiciles, peridomiciles, and in the natural habitats of the Triatominae. Insects from these collections were dissected and their organs and tissues examined for flagellates. Frass from these insects was collected and examined for detection of the entomopathogenic virus Triatoma virus (TrV) using AC-ELISA and PCR. Triatominae belonging to four species, T. infestans (n=1646), Triatoma guasayana (n=4), Triatoma platensis (n=1) and Triatoma sordida (n=5) were collected from 62 sites located in 13 provinces of Argentina. Triatoma virus and two protozoan species, Blastocrithidia triatomae and T. cruzi, the etiological agent of Chagas disease, were found infecting Triatominae. The total prevalence of TrV in 1646 T. infestans analyzed by ELISA was 9.66% (159/1646) from 7 to 13 provinces where collections were made. Triatoma virus positive triatomines were found in 17 of 62 populations when examined by AC-ELISA but in 38 of 62 populations when PCR was used for detection. The prevalence of B. triatomae in T. infestans was 0.43% (7/1646), while the prevalence of T. cruzi was 1.3% (21/1646). This is the first study on the diversity, distribution and prevalence of flagellated protozoa and TrV of Triatominae in endemic Chagas' disease regions of Argentina.

Equine arteritis virus: a new isolate from the presumable first carrier stallion in Argentina and its genetic relationships among the four reported unique Argentinean strains.

Equine arteritis virus (EAV) was isolated from a testicle of the presumable first stallion infected with EAV in Argentina. This virus isolate (named LT-LP-ARG) was confirmed by GP5-specific PCR and indirect immunofluorescence assays. The PCR product was sequenced, and the phylogenetic analysis revealed that the LT-LP-ARG strain of EAV forms a monophyletic group, together with other strains previously isolated in our laboratory (LP02 group). However, all Argentinean EAV strains belong to a polyphyletic group. We believe that the virus isolate presented in this report could be the origin of EAV infection in our country.

AC-ELISA and RT-PCR assays for the diagnosis of triatoma virus (TrV) in triatomines (Hemiptera: Reduviidae) species.

Triatoma virus (TrV) is the only entomopathogenic virus found in triatomines. TrV replicates in cells of the midgut epithelium of triatomines, causing a high mortality rate and delayed development of the infected insect. In this work, we report an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) and a reverse transcription-polymerase chain reaction (RT-PCR) assay for detection of TrV infection. For antiserum production, rabbits and hens where inoculated with purified TrV. Antiserum reactivity was checked by immunodiffusion, and its specificity was confirmed by western blot and AC-ELISA. Totally 90 fecal samples from T. infestans were analysed. AC-ELISA and RT-PCR results correlated well with transmission electron microscopy (EM) observations, which are considered the gold standard, with Kappa values of 0.73 for AC-ELISA and 0.93 for RT-PCR when compared with EM. Applications and complementary uses of the two techniques reported in this work are discussed.