Improved DNA hybridization method for detection of acyclovir-resistant herpes simplex virus.
A simplified DNA hybridization method was developed to detect acyclovir-resistant isolates of herpes simplex virus. Herpes simplex virus-infected cell cultures in microtiter plates were treated with concentrations of acyclovir ranging from 8 to 0.015 micrograms/ml. At 48 h postinfection, infected cells were lysed by a one-step procedure and lysates were absorbed to membranes. Without further treatment, membranes were hybridized by using a herpes simplex virus-specific radioiodinated probe. The membranes were then washed and counted in a gamma counter. The elapsed time for assay performance was 4 h. Parallel plaque reduction assays were performed for comparison. The mean 50% inhibitory dose of in vivo- and in vitro-derived acyclovir-resistant, thymidine kinase-negative isolates was greater than 2 micrograms/ml by DNA hybridization. The 50% inhibitory dose of acyclovir-susceptible, thymidine kinase-positive isolates ranged from 0.01 to 1.1 micrograms/ml. This assay is simple and objective and should facilitate antiviral susceptibility testing in diagnostic laboratories.
Swierkosz
E M
EM
Department of Pediatrics/Adolescent Medicine, St. Louis University School of Medicine, Missouri.
Scholl
D R
DR
Brown
J L
JL
Jollick
J D
JD
Gleaves
C A
CA
D000212:Acyclovir / Q000494:pharmacology*; D000818:Animals; D016026:Bone Marrow Transplantation; D002478:Cells, Cultured; D004279:DNA, Viral / Q000302:isolation & purification*; D004352:Drug Resistance, Microbial; D006801:Humans; D009693:Nucleic Acid Hybridization; D018139:Simplexvirus / Q000187:drug effects*; D013937:Thymidine Kinase / Q000378:metabolism; D014709:Vero Cells; D010948:Viral Plaque Assay
D004279:DNA, Viral; D013937:Thymidine Kinase; D000212:Acyclovir