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Failure to detect enterovirus in the spinal cord of ALS patients using a sensitive RT-PCR method

From the Respiratory and Enteric Viruses Branch (Drs. Oberste and Pallansch, W.A. Nix), Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, GA; Department of Medicine (Dr. Berger), University of California–Irvine, Orange; Neurology Service (Dr. Brooks), William S. Middleton Memorial Veterans Affairs Medical Center, and Department of Neurology, University of Wisconsin Medical School, Madison; Cecil B. Day Laboratory for Neuromuscular Research (Dr. Brown, D.M. McKenna–Yasek), Massachusetts General Hospital, Charlestown; and Department of Neurology (Dr. Roos), University of Chicago, IL.

Address correspondence and reprint requests to Dr. M.A. Pallansch, Centers for Disease Control and Prevention, Mailstop G-17, Atlanta, GA 30333; e-mail: mpallansch{at}cdc.gov

Objective: To assess the association of enteroviruses (EV) with ALS by applying a sensitive seminested reverse transcription (RT) PCR protocol to the detection of enteroviral RNA in a blinded set of archived tissues from ALS and control cases.

Methods: The specimen set consisted of 24 frozen spinal cord samples from ALS cases, 17 frozen spinal cord samples from negative control (non-ALS) cases, and 5 frozen spinal cord positive control samples. The positive controls were two human spinal cord samples spiked with poliovirus (PV) and three spinal cords from PV-infected transgenic mice. A sensitive, EV-specific, seminested RT-PCR assay was used to detect EV genome in RNA extracted from the specimens and controls.

Results: The assay detected EV RNA in a 10^–5 dilution of infected mouse tissue. EV RNA was not detected in the ALS specimens or in specimens from control cases, despite the presence of amplifiable RNA as assessed by amplification with control primers, whereas all of the positive control specimens yielded the expected PV amplification product.

Conclusion: The reported association between EV infection and ALS was not confirmed by testing this set of specimens with these sensitive methods.

Received August 10, 2003. Accepted in final form December 23, 2003.

See also page 1250

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				W. A. Nix, M. S. Oberste, and M. A. Pallansch Sensitive, Seminested PCR Amplification of VP1 Sequences for Direct Identification of All Enterovirus Serotypes from Original Clinical Specimens. J. Clin. Microbiol., August 1, 2006; 44(8): 2698 - 2704. [Abstract] [Full Text] [PDF]

				B. Jubelt and H. L. Lipton ALS: Persistent scientists do not find persisting enteroviruses Neurology, April 27, 2004; 62(8): 1250 - 1251. [Full Text] [PDF]