Attenuation In Mice Research Articles

Insertional inactivation of the vaccinia virus 32-kilodalton gene is associated with attenuation in mice and reduction of viral gene expression in polarized epithelial cells

The mechanism of poxvirus attachment to cells is poorly understood. We have identified a 32-kDa envelope protein of vaccinia virus which binds to the surface of cultured cells. This binding is specific and selective (J.-S. Maa, J. F. Rodriguez, and M. Esteban, J. Biol. Chem. 265:22174-22180, 1990; C. Lai, S. Gong, and M. Esteban, J. Virol. 65:499-504, 1991). In this investigation, we studied the effect of inactivating the 32-kDa gene (32K gene) on the biology of vaccinia virus. We show that inactivation of the 32K gene decreases by 80% the mortality of mice infected with 32K- vaccinia virus. This reduction in mortality correlates with diminished viral gene expression in target tissues. In highly polarized epithelial cells, viral gene expression of 32K- virus was reduced (50 to 60%) at both the apical and basolateral surfaces in comparison with a 32K+ virus. Restriction of virus gene expression in polarized cell surfaces occurs for both intracellular and extracellular forms of infectious 32K- vaccinia virus. The two infectious forms of vaccinia virus 32K+ infect polarized cells preferentially by the basolateral surface. Our findings provide evidence of the importance of the 32-kDa protein in viral pathogenesis.

In vivo behavior of genetically engineered herpes simplex viruses R7017 and R7020: construction and evaluation in rodents.

The herpes simplex virus (HSV) recombinant R7017 was constructed from HSV-1 (strain F) by deleting a portion of the thymidine kinase (tk) gene and by replacing the sequences representing the internal inverted repeats and adjacent genes in the L component with a fragment of the HSV-2 genome encoding the glycoproteins G, D, I, and a portion of E. In addition, the R7020 recombinant contains an HSV-1 DNA fragment encoding the tk gene fused to the alpha 4 gene promoter. The results of studies in mice, guinea pigs, and rabbits were as follows: Both recombinants remained unchanged after nine serial, intracerebral passages in mice; the recombinants could not be differentiated with respect to attenuation in mice injected intracerebrally, in vaginally infected guinea pigs, and in rabbits inoculated on the scarified cornea. Given intradermally or intramuscularly, the recombinants prevented severe infections by virulent challenge viruses, and R7020 established latent infections (at a low frequency) in all species tested, whereas latent R7017 virus was detected in rabbits only.

Repeated electroconvulsive shock attenuates clonidine-induced hypoactivity in rodents

Clonidine administration at low dose produces hypoactivity in both rats and mice, probably by stimulation of presynaptic α 2-adrenoceptors in the brain. This behaviour is antagonised by yohimbine pretreatment but is unaffected by pretreatment with prazosin. An. electroconvulsive shock (ECS) given once daily for 10 days markedly attenuated the hypoactivity in both species when tested 24 h after the final shock. The attenuation in mice was not found after either a single ECS or 10 subconvulsive shocks. Ten ECS did not alter the hypoactivity response produced by (±)-propranolol in mice. Clonidine administration decreased rat brain MOPEG-SO 4 concentrations. A single ECS given daily for 10 days abolished this decrease. These results suggest that repeated ECS causes a subsensitivity of α 2-adrenoceptors in the brain and that these receptors may be located presynaptically.

Attenuation and immunogenicity of ts mutants of Eastern encephalitis virus for mice.

Preliminary experiments were intiated to test the attenuation for mice of chemically induced temperature-sensitive (Ets) mutants of a virulents strain of Eastern encephalitis (E) virus, and the potential of such mutants as live virus vaccines for mice. The reversion frequencies of eight mutants to temperature insensitivity were measured and the defects in their biosynthesis at the non-permissive temperature were studied. All eight mutants were less virulent for mice, but the extent of avirulence varied with the mutant and route of injection. The mutants selected as potential vaccines protected mice against subsequent challenge at 10 to 21 days by the virulent virus of strain E. Neutralizing antibody activity was detected in almost all the mutant-infected mice after 10 days, and was found in all infected mice at 21 days. Immunization by two doses of virus induced a very high protection against intracerebral challenge by virus strain E.

Temperature-Sensitivity Characteristics Distinguishing Substrains of Fixed Rabies Virus: Lack of Correlation with Plaque-Size Markers or Virulence for Mice

Three characteristics of a variety of strains and substrains of cell culture-adapted rabies virus were examined: (1) relative capacity to replicate at 40.5 C (rct/40.5 marker); (2) plaque size in agarose-suspended baby-hamster kidney cells (subclone 13S); and (3) relative pathogenicity for adult mice. Each of four strains of fixed virus was capable of replication at 40.5 C, but quantitative differences in rct/40.5 efficiency, which did not correlate with virulence for mice, were noted. In addition, two substrains of challenge-virus-standard (CVS) cultivated at 23 C25 C and three temperature-sensitive mutants, all incapable of replication at 40.5 C, retained normal virulence for mice. CVS variants tested in the course of continuous passage in snake or lizard cell cultures became incapable of replication at 40.5 C at different passage levels, a phenomenon not related temporally to the acquisition of attenuation for mice or small-plaque markers in the same substrains. The independent variation of these three phenotypic characters suggests that they are controlled by different segments of the viral genome.

Factors affecting bacteriaemia in mice inoculated with Mycoplasma mycoides var. mycoides

Seven fresh strains of M. mycoides produced a high incidence of protracted bacteriaemia in mice inoculated intraperitoneally, but several strains repeatedly subcultured in the laboratory were attenuated. Comparison of the bacteriaemia produced by a freshly isolated strain and the attenuated V5 and KH 3J vaccine strains revealed a parallelism between virulence for mice and cattle and the method may be applicable to the control of virulence of vaccines. Of 4 fresh strains subjected to 53 rapid subcultures, only 1 strain showed evidence of gross attenuation for mice as assessed by blood culture. Definite though not overwhelming differences in susceptibility to M. mycoides were demonstrated in mice obtained from separate colonies. Bacteriaemia was produced more readily by intraperitoneal and intravenous inoculation than by the subcutaneous and intranasal methods.