[Standardization of a method for concentration and detection of enteric viruses from drinking water]. / Estandarización de un método de concentración y detección de virus entéricos en aguas de consumo.

INTRODUCTION: Enteric viruses have been implicated in acute diarrheal disease, food-borne disease, hepatitis A and meningitis outbreaks, in which water was the vehicle of transmission. OBJECTIVE: A concentration method was standardized for the detection of enteric viruses in drinking water. MATERIALS AND METHODS: Twenty liters of water were concentrated to 6 ml by filtration and tangential ultrafiltration. Viral solutions of 20 L each were prepared at 1, 10, 50 and 100 TCID50 of Sabin poliovirus type 1 as positive controls. Viral particles were recovered by tissue culture and detected by conventional polymerase chain reaction (PCR), according to the international standards recommended by the Enterovirus Laboratory at the Centers for Disease Control and Prevention, Atlanta, GA. RESULTS: All positive controls showed cytopathic effect on L20B and RD cells and were amplified by conventional PCR directly from samples. Negative controls did not show any amplification or viral cytopathic effect. CONCLUSIONS: Tangential ultrafiltration for concentrating viruses proved to be a fast, efficient recovery and reproducible. It has the advantage of allowing the detection (at the 1 TCID50 level) and identification of viruses by RT-PCR and the demonstration of viral infectivity by tissue culture.

Estandarización de un método de concentración y detección de virus entéricos en aguas de consumo / Standardization of a method for concentration and detection of enteric viruses from drinking water

Introducción. Los virus entéricos se han visto implicados en brotes de enfermedad diarreica aguda, enfermedades transmitidas por alimentos, hepatitis A y meningitis aséptica, en los que el vehículo de transmisión del agente ha sido el agua.Objetivo. Estandarizar un método de concentración para la detección de virus entéricos en aguas de consumo.Materiales y métodos. Se concentraron 20 litros de agua a un volumen de 6 ml mediante filtración y ultrafiltración tangencial. Como controles positivos se prepararon soluciones de 20 litros a concentraciones virales de 1, 10, 50 y 100 TCID50 (Tissue Culture Infectious Dose 50%) de Poliovirus Sabin de tipo 1. Las partículas virales fueron recuperadas por cultivo en células sensibles a la infección e identificadas por amplificación del genoma viral mediante reacción en cadena de la polimerasa, siguiendo los estándares internacionales de los Centers for Disease Control and Prevention (CDC) de Atlanta. Resultados. Todos los controles positivos causaron efecto citopático en células de rabdomiosarcoma y L20B y fueron detectados por RT- PCR (Real Time- PCR) convencional, directamente de las muestras. Los controles negativos no mostraron efecto citopático ni amplificación viral por RT-PCR.Conclusiones. La ultrafiltración tangencial mostró ser un método rápido y eficaz al recuperar virus desde una TCID50, además de ser reproducible y sencillo. Tiene la ventaja de permitir la detección de su capacidad de contagiosidad viral por el cultivo celular, y la identificación por RT-PCR.

Introduction. Enteric viruses have been implicated in acute diarrheal disease, food-borne disease, hepatitis A and meningitis outbreaks, in which water was the vehicle of transmission.Objective. A concentration method was standardized for the detection of enteric viruses in drinking water. Materials and methods. Twenty liters of water were concentrated to 6 ml by filtration and tangential ultrafiltration. Viral solutions of 20 L each were prepared at 1, 10, 50 and 100 TCID50 of Sabin poliovirus type 1 as positive controls. Viral particles were recovered by tissue culture and detected by conventional polymerase chain reaction (PCR), according to the international standards recommended by the Enterovirus Laboratory at the Centers for Disease Control and Prevention, Atlanta, GA. Results. All positive controls showed cytopathic effect on L20B and RD cells and were amplified by conventional PCR directly from samples. Negative controls did not show any amplification or viral cytopathic effect. Conclusions. Tangential ultrafiltration for concentrating viruses proved to be a fast, efficient recovery and reproducible. It has the advantage of allowing the detection (at the 1 TCID50 level) and identification of viruses by RT-PCR and the demonstration of viral infectivity by tissue culture.