Improvement in Plaquing Methods for the Enumeration of Anatid Herpesvirus (Duck Plague Virus)

Abstract

The Holland strain of anatid herpesvirus (AHV) was 2- to 10-fold more efficiently plaqued under liquid medium or semisolid medium containing methylcellulose than in media containing agar, agarose, or Nobel agar. Virus adsorption was complete in 45 min, with the maximum virus titers obtained under liquid medium at 48 h postinfection. The AHV dose-response curve was linear. Pekin duck embryo cultures were more efficient than CCL-141 in plaque response and gave the maximum virus yield per cell. Virus titers were independent of avian cell passage number (to 6), fetal or newborn calf sera (to 10% v/v), and the presence of polyions during adsorption. The host range included cells from several species of Anseriformes and Galliformes.

Molecular Characterization of the DNA of Anatid Herpesvirus 1

“…Field isolates of falcon and pidgeon herpesviruses, when ana lyzed by restriction enzymes, were found to be quite similar, suggesting that the host range for these viruses is not restricted to a particular order of avians [60]. Furthermore, the host range of the Holland strain of AHV-ppc3 in culture extends beyong Anseriformes and in cludes many Gallijormes and cultures from em bryos of birds in the Columbiformes and the Passeriformes [12]. The natural host ranges of HO Gardner/Wilkerson/Johnson Anatid Herpesvirus DNA AH V as well as the other avian herpesviruses in this group have not been fully established, and speciation by either antigenic or genetic means of the isolates from the North American conti nent as well as those obtained from China, the Netherlands and England has not been accom plished.…”

“…Embryonated chicken eggs were obtained from pathogen-free flocks (Hyline International). Cell culture preparations from mid-term embryonated eggs and the AHV plaque assay were performed as reported earlier [12], Bacteriophage T4 (ATCC 11303-B4) was grown on Escherichia coli Bb in basal C medium containing 5 ug/ml thymidine and 0.4% glucose [18].…”

“…DVE is caused by the Anatid herpesvirus (AHV), a term proposed and recommended by the Herpesvirus Study Group [9], Biological studies of the virus and its interactions with cells have been aided by the availability of neutralization antisera and a plaque assay ef fective, at least, for some of the AHV isolates [10][11][12], Although the physical description of the virus as obtained by electron microscopy [13,14], the biology of infection as obtained by single-step experiments [15], and initial charac terizations of the virus and viral nucleic acid by nuclease resistance [13,16] indicate that the viral genome is DN A, no reports other than a study of viral multiplication inhibition by phosphono compounds and acyclovir [17] have been pub lished which serve to confirm this finding.…”

Synergistic Inhibition of Anatid Herpesvirus Replication by Acyclovir and Phosphonocompounds

“…Cell cultures were prepared and maintained by published proce dures [12]. The cell maintenance medium (CMM) used consisted of an autoclavable MEM containing 5% heat-inactivated newborn calf serum and 10 mM N -2-hydroxyethylpiperazine -N' -2 -ethanesulfonic acid (HEPES)at pH 7.4.…”

“…An infectious particle assay for AHV that employs a liquid rather than a solid cell maintenance medium overlay has been de scribed [12]. Under the conditions of this as say, one-hit kinetics for AHV plaque forma tion was demonstrated by a linear dose-re sponse curve, assay procedures were simpli fied, and the time required for assay comple tion was decreased.…”