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J Clin Microbiol. 1990 May; 28(5): 965-969

Enhanced recovery of cytomegalovirus in conventional tube cultures with a spin-amplified adsorption.

Department of Pathology and Laboratory Medicine, University of Texas Health Science Center, Houston.

ABSTRACT

Low-speed centrifugation-mediated adsorption was evaluated as an enhancement of infectivity of clinical and laboratory strains of cytomegalovirus (CMV) occurring with cells grown in conventional culture tubes. The time required for reporting of primary isolates of CMV from urine specimens adsorbed onto monolayers of WI-38 cells in culture tubes was calculated. Of 668 specimens adsorbed by the stationary phase (SP) method, 98 were positive by cytopathic effect (CPE) that required an average of 16.8 days for recovery in culture. However, the appearance of CPE required a shorter average time of 11.9 days for 70 CMV strains isolated from 283 specimens adsorbed in tube cultures by the spin-amplified (SA) method. In another phase of clinical CMV recovery, urine specimens were adsorbed by the SA method onto cell cultures grown in both shell vials and test tubes. Of 594 specimens inoculated, a total of 74 were positive by either CPE in test tubes or immunostaining-localized early antigen in shell vials. Approximately one-third of these CMV isolates were recovered only by CPE from specimens adsorbed by the SA method in test-tube cultures. In a related study to further evaluate differences between adsorption methods, the AD-169 laboratory strain of CMV was adsorbed by SP and SA methods onto MRC-5 cells grown in both culture vessels. Early antigen detection by immunomicroscopy was found in the infected cells at least 2 to 4 days prior to the appearance of CPE, regardless of adsorption procedure. In both vessels, the replication of AD-169 virus in cultures adsorbed by the SA method consistently exceeded that of virus adsorbed by the SP procedure. CPE occurred 24 to 48 h earlier and progressed two to four times more extensively; early antigen was expressed two- to fourfold greater within 24 to 48 h postinfection; and foci of infected cells containing late antigen were two to four times greater in number at 1, 2, and 5 days postinfection. Overall, the replication and enhancement of infectivity of laboratory and clinical strains of CMV as determined by CPE and early and late antigen expression occurred most efficiently with specimens adsorbed by the SA method onto cultures grown in conventional tubes or shell vials.