martes, 28 de mayo de 2013

Cell Culture and Electron Microscopy for Identifying Viruses in Diseases of Unknown Cause - Vol. 19 No. 6 - June 2013 - Emerging Infectious Disease journal - CDC

full-text ►
Cell Culture and Electron Microscopy for Identifying Viruses in Diseases of Unknown Cause - Vol. 19 No. 6 - June 2013 - Emerging Infectious Disease journal - CDC

 EID cover artwork EID banner
Table of Contents
Volume 19, Number 6–June 2013

Volume 19, Number 6—June 2013


Cell Culture and Electron Microscopy for Identifying Viruses in Diseases of Unknown Cause

Cynthia S. GoldsmithComments to Author , Thomas G. Ksiazek, Pierre E. Rollin, James A. Comer, William L. Nicholson, Teresa C.T. Peret, Dean D. Erdman, William J. Bellini, Brian H. Harcourt, Paul A. Rota, Julu Bhatnagar, Michael D. Bowen, Bobbie R. Erickson, Laura K. McMullan, Stuart T. Nichol, Wun-Ju Shieh, Christopher D. Paddock, and Sherif R. Zaki
Author affiliations: Centers for Disease Control and Prevention, Atlanta, Georgia, USA (C.S. Goldsmith, P.E. Rollin, J.A. Comer, W.L. Nicholson, T.C.T. Peret, D.D. Erdman, W.J. Bellini, B.H. Harcourt, P.A. Rota, J. Bhatnagar, M.D. Bowen, B.R. Erickson, L.K. McMullan, S.T. Nichol, W.-J. Shieh, C.D. Paddock, S.R. Zaki); University of Texas Medical Branch, Galveston, Texas, USA (T.G. Ksiazek)
Suggested citation for this article


During outbreaks of infectious diseases or in cases of severely ill patients, it is imperative to identify the causative agent. This report describes several events in which virus isolation and identification by electron microscopy were critical to initial recognition of the etiologic agent, which was further analyzed by additional laboratory diagnostic assays. Examples include severe acute respiratory syndrome coronavirus, and Nipah, lymphocytic choriomeningitis, West Nile, Cache Valley, and Heartland viruses. These cases illustrate the importance of the techniques of cell culture and electron microscopy in pathogen identification and recognition of emerging diseases.
Thin section and negative stain electron microscopy (EM) examination of viruses grown in cultured cells have been instrumental in determining an etiologic agent in numerous disease outbreaks caused by previously unknown viruses. Many examples have been reported. In 1976, EM of cell culture isolates identified the causative virus of an outbreak of hemorrhagic fever in Zaire as a member of the family Filoviridae, now known as Zaire ebolavirus (13). Reston ebolavirus was another previously unrecognized virus that was detected by cell culture and EM in 1989; it was isolated from cynomolgus monkeys imported into the United States from the Philippines (4). In Australia in 1994, during an outbreak of fatal respiratory disease in horses and influenza-like illness in humans, a previously unknown virus, Hendra virus, was isolated in culture and recognized as a member of the family Paramyxoviridae by EM (5,6). An outbreak of an unidentified rash illness in humans, associated with sick prairie dogs, occurred in the upper midwestern United States in 2003, and EM detected a poxvirus from a cell culture isolate, which was later characterized as monkeypox virus (7,8). Recently, the etiologic agent of severe fever with thrombocytopenia syndrome in China was isolated and identified by EM as a member of the family Bunyaviridae (9).
Inoculation of patient specimens onto cultured cells or into laboratory animals enables biologic amplification of virus particles to levels where they can be detected by EM and identified to a virus family because, with a few exceptions (10), the morphologic features of all viruses within a given family are the same. Once recognized by EM, the findings can be confirmed by other techniques, including serologic testing, immunohistochemical (IHC) and indirect fluorescence antibody (IFA) assays, and molecular methods that can further characterize the virus to species and strain.
Cell culture methods are relatively unbiased, restricted only by the ability of the virus to grow in a particular cell line. Vero E6 cells, considered one of the most permissive of all cell lines, provide an extremely versatile medium for recovery of unknown pathogens. EM is also an unbiased assay in that there is no need for specific immunologic probes, and has the added advantage of being able to detect and classify the unknown agent. EM observations of cell culture isolates can provide the first clue of an etiologic agent and guide subsequent laboratory and epidemiologic investigations. Detection of a pathogen is critical during outbreaks because identification of an etiologic agent enables public health officials to mount a timely response and limit further spread of the agent involved. In addition, pathogen identification is invaluable in individual cases of severe illness in which an infection is caused by an undetermined agent. We report several cases where cell culture and EM at the Centers for Disease Control and Prevention (CDC) enabled initial recognition and identification of a cause of the viral illness.

No hay comentarios:

Publicar un comentario