Biological and molecular characterization of a persistent influenza type B virus infection.

Abstract

A persistent infection in Madin Darby canine kidney (MDCK) cells by an influenza B virus (B/Tecumseh/63/80) has been characterized. Virus recovered from the persistent state titrates lower in relation to the parental wild-type by hemagglutination, infectivity, and egg and tissue culture infectious dose suggesting that the virus is an attentuated, less cytopathic variant of the original wild-type virus. The persistent virus has decreased cytopathogenicity for both MDCK and primary chicken kidney cell lines, and exhibits a different pattern of genomic RNA and protein electrophoretic migration. Plaques of the persistent virus are smaller and take longer to appear, indicating that this persistent virus is a slower growing variant of the wild-type; it is possible that the small-plaque mutant phenotype is associated with the maintenance of the persistent infection in MDCK cells. Persistently infected cells also exhibit different characteristics in comparison to normal uninfected cells. These cells display a slower growth rate and periodically enter a crisis state in which many cells are destroyed and virus titer is high. About 90% of the cells in the persistent state are infected as determined by immunofluorescence. The persistent cells produce a crystalline deposit between the serosal surface of the cells (basolateral) and the growth substratum. The needle-shaped crystals have been tentatively identified as uric acid by electron microscopy, HPLC and GC/MS. Sequence analysis of the genes coding for the NA and HA external glycoproteins was undertaken in an attempt to provide a molecular explanation for the persistent phenotype. No differences have been found in the region coding for the NB protein, implying that alteration in membrane ion transport is not responsible for the persistent state nor for the formation of the crystalline product. Similarly, no changes have been located in the HA gene that could account for the distinct HI phenotype.