No abstract

Genetic analysis showed that the glycerol non-utilizing isolate gly-u(234) of Neurospora crassa is derived by mutation in a nuclear gene situated in the right arm of linkage group I, about 2.2 cross-over units distal to ad-9 and 11 units proximal to nit-1. Enzymatic testings using a radiochemical method indicate that the mutant is deficient for the enzyme glycerol kinase. The radiochemical testings further indicate that the mutation has inactivated an inducible glycerol kinase, while a low residual activity may be due to a second, basal and non-inducible glycerol kinase, in accordance with a proposal by North (1973, 1974) that Neurospora has two glycerol kinases with these properties.

Herpes simplex virus (HSV) hyperresistant neuroblastoma cells (clone Cl300 RII) were latently infected with HSV-1 if cultured in presence of HSV-neutralizing antibody for one or two passages after infection and then passaged further without antibody. By superinfecting HSV-1 latently infected Cl300 RII cells with HSV-2, progeny virus with HSV-1 characteristics was regularly rescued. Such retrieval of HSV-1 decreased with passage of the latently infected cells.

The virus yields and number of infectious centres of HSV infected mouse neuroblastoma C1300 cells (clone 41 A3) infected at different multiplicities of infection (MOI) were found to vary more than the differences of HSV concentrations of the virus suspensions used for infection of the cells. This suggested that a C1300 cell had to be infected with more than one HSV particle in order to produce progeny virus-multiplicity activation. The greater than expected enhancement of virus production of C1300 cell cultures receiving increasing MOI of HSV was probably not due to improved virus adsorption, nor influenced by non-virus factors in the virus inoculum stimulatory for HSV replication. A hypothesis, that the block in virus replication was promoted by an inhibitor of an HSV specified regulatory protein and could be overcome by the addition of HSV DNA copies in the infected cell, was by the results of two types of experiments. Presence of phosphonoformic acid, an inhibitor of the HSV specified DNA polymerase, in the culture medium of HSV infected permissive GMK cells resulted in non-linear relationships between virus yields and MOI. An HSV temperature sensitive mutant (ts B5), defective in a late structural protein, rescued wild type HSV in C1300 cells.

We developed a method that uses monoclonal antibodies for typing herpes simplex virus type 1 and type 2 strains. Clinical isolates from GMK cells were seeded directly into a monolayer of GMK cells. After incubation overnight, monoclonal antibodies were added to the infected monolayer, and antibody binding was indicated by a peroxidase enzyme-linked immunosorbent assay. Using prototype strains and previously typed patient strains, we verified the specificity of this technique. This method is now used routinely for typing herpes simplex strains in our laboratory. We have also used this technique for specific staining of type 1 plaques in a mixture of type 1 and type 2 plaques. With this method it is possible to find a single type 1 plaque among several hundred type 2 plaques on a single petri dish. Infectious virus can also be recovered from stained, unfixed type 1 plaques.