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S Kit

Publications and source records attributed to S Kit.

At least 37 records · Page 2Linked to original sources
PubMed1990

Safety and efficacy of a thymidine kinase negative equine herpesvirus-1 vaccine in young horses.

A drug induced equine herpesvirus-1 (EHV-1) mutant lacking thymidine kinase inducing activity was developed and evaluated as a vaccine. The safety and effectiveness of the vaccine to protect against experimentally induced EHV-1 respiratory disease were evaluated in weanling horses free of EHV-1 neutralizing antibody. The vaccine was safe when administered either intramuscularly or intravenously, and EHV-1 was not shed intranasally during the 12 days following administration. Intranasal challenge with virulent EHV-1 was used to evaluate vaccine efficacy. Following challenge, there was a significantly (p less than 0.05) greater increase in peak body temperatures and duration of nasal virus shedding in the nonvaccinates, and a significant (p less than 0.05) increase in serum neutralizing antibody titers in the vaccinates.

Animals↗
PubMed1989

Molecular confirmation of an abortigenic strain of equine herpesvirus 1 (subtype 1) in a pregnant mare study.

Four pregnant mares were inoculated intranasally and/or intravenously with equine herpesvirus 1 (EHV-1), subtype 1 during the third trimester of gestation. One mare aborted on postinfection day 15, one mare delivered a sick, weak full term foal, and two mares delivered healthy, full term foals. EHV-1, subtype 1 was isolated from several tissues of the aborted fetus and from the thymus of the sick foal. DNA restriction endonuclease patterns of the recovered EHV-1 viruses were identical to those of the EHV-1 challenge strain, documenting the origin of the abortigenic viruses.

Abortion, Veterinary↗
PubMed1988

Isolation and comparative restriction endonuclease DNA fingerprinting of equine herpesvirus-1 from cattle.

Deoxyribonucleic acid fingerprinting analyses with 4 restriction endonucleases (EcoRI, BamHI, BglII, and HindIII) and serotest results have definitively indicated that 5 herpesviruses isolated from 1974 to 1986 from aborted bovine fetuses and from bovine tissues and nasal secretions were abortigenic subtypes of equine herpesvirus type 1 (EHV-1). The herpesviruses, designated BH1247, 3M20-3, G118, H1753, and 9BSV4, were neutralized by EHV-1-specific antiserum and could be propagated in cultures of either bovine or equine cells. Only minor differences in restriction endonuclease patterns were detected from the pattern of an Army 183 isolate of EHV-1 subtype 1 that had been passaged only in equine cells and from that of an attenuated EHV-1 subtype 1 (RQ) strain that had been passaged several hundred times in non-equine cells. The individual differences in the restriction endonuclease fragments of the 5 bovine isolates and the Army 183 and RQ strains mainly were attributable to alterations in the terminally repeated and the unique short nucleotide sequences of the EHV-1 genomes, which are known to be hot spots for deletions and tandem repeats. The BamHI restriction endonuclease pattern of the 1977 bovine isolate H1753 was identical to that of EHV-1 subtype-1 strains responsible for most of the virus abortions in vaccinated horses since 1981. Abortigenic EHV-1 strains have the ability to infect cattle and cause disease under natural conditions.

Abortion, Veterinary↗
PubMed1987

Phosphorylation of nucleoside analogs by equine herpesvirus type 1 pyrimidine deoxyribonucleoside kinase.

Replication of equine herpesvirus type 1 (EHV-1) was sensitive to 9-(1,3-dihydroxy-2-propoxymethyl)guanine(DHPG) but relatively resistant to E-5-(2-bromovinyl)-2'-deoxyuridine (BVDU). Likewise, plaque formation by EHV-1 was inhibited by DHPG, but not by BVDU. Plaque formation by a thymidine kinase-negative (tk-) mutant of EHV-1 was not inhibited by DHPG. In order to investigate biochemical mechanisms determining the differential sensitivity of EHV-1 to these drugs, the EHV-1-encoded thymidine kinase enzyme activity (TK)1 was partially purified from EHV-1-infected cells and analyzed. The EHV-1-induced enzyme utilized both ATP and CTP as phosphate donors and differed in relative electrophoretic mobility from the TKs of mock-infected and HSV-1-infected cells. Phosphorylation of 3H-dThd by the EHV-1 TK was inhibited by AraT, IdUrd, BVDU, and DHPG. The EHV-1 TK phosphorylated 125I-dCyd and 3H-ACV. The results indicate that EHV-1 encodes a pyrimidine deoxyribonucleoside kinase with broad nucleoside substrate specificity. These observations suggest that the failure of BVDU to inhibit EHV-1 replication is not attributable to an inability of the EHV-1 TK to phosphorylate BVDU, but may result from the incapacity of the viral TK to convert BVDU monophosphate to the triphosphate or from lack of inhibitory effect of BVDU triphosphate on viral DNA polymerase reactions.

Acyclovir↗
PubMed1987

Differential metabolism of (E)-5-(2-iodovinyl)-2'-deoxyuridine (IVDU) by equine herpesvirus type 1- and herpes simplex virus-infected cells.

Thymidine kinase (TK) enzymes encoded by herpes simplex viruses types 1 and 2 (HSV-1, HSV-2), and equine herpesvirus type 1 (EHV-1) catalyze the phosphorylation of thymidine (dThd) and (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU). The replication of HSV-1 is sensitive to BVDU, but the replication of HSV-2 and EHV-1 is not. To investigate the differential sensitivity of the viruses to halogenated vinyldeoxyuridine drugs, the phosphorylation of 125I-labeled (E)-5-(2-iodovinyl)-2'-deoxyuridine (IVDU) was studied. Cytosol enzymes from cells infected by HSV-2 and EHV-1 phosphorylated [125I]IVDU to the monophosphate, IVDUMP, but did not convert IVDUMP to higher di- plus triphosphates (IVDUDP plus IVDUTP) forms. In contrast, enzymes from HSV-1-infected cells converted [125I]IVDU to radioactive IVDUMP and IVDUDP plus IVDUTP. Experiments with mixtures of EHV-1- and HSV-1-induced enzymes showed that the EHV-1 enzyme did not inhibit formation of the IVDUDP plus IVDUTP by the HSV-1 enzyme. With [125I]IVDU as substrate, the Km values for the EHV-1 and HSV-1 TKs were 1.82 and 0.34 microM, respectively, and the Ki (dThd) value for the EHV-1 TK was 0.35 microM. In vivo experiments showed that HSV-1-infected cells converted IVDU to the mono- and the di- plus triphosphate forms. In contrast, EHV-1-infected cells converted IVDU to the monophosphate to a lesser extent than did HSV-1-infected cells, and did not produce the di- plus triphosphates. Thus, inefficient phosphorylation of the monophosphates probably contributes to the insensitivity of EHV-1 replication to IVDU, as it does to the insensitivity of HSV-2 replication to this drug.

Animals↗
PubMed1987

Nucleotide sequence changes in thymidine kinase gene of herpes simplex virus type 2 clones from an isolate of a patient treated with acyclovir.

To identify the nucleotide changes that occur in drug-induced thymidine kinase (TK) mutants of herpes simplex virus type 2 (HSV-2), we compared the nucleotide sequences of the tk genes of two mutant HSV-2 clones isolated from a patient who had been treated with acyclovir [9-(2-hydroxyethoxymethyl)guanine; ACV] with the nucleotide sequence of the parental TK+ HSV-2(8703) strain isolated from the same patient. One of the mutants, TK-altered (TKA) HSV-2(9637), was ACV resistant but induced the incorporation of [14C]thymidine into the DNA of infected rabbit skin cells. The nucleotide sequence of the tk gene of mutant TKA HSV-2(9637) had a single change (G to A) at nucleotide 668, which would cause an arginine-to-histidine substitution at amino acid residue 223 of the TK polypeptide. The second ACV-resistant mutant, TK- HSV-2(8710), did not induce detectable incorporation of [14C]thymidine into the DNA of infected rabbit skin cells. This mutant exhibited a deletion of a single base at nucleotide 217 of its nucleotide sequence. This deletion would cause a frameshift mutation at amino acid residue 73 and chain termination at amino acid residue 86 of the TK polypeptide. The nucleotide sequence of TK+ HSV-2(8703) was the same as that of the laboratory strain, TK+ HSV-2(333). The nucleotide sequence of a bromodeoxyuridine-resistant TK- HSV-2(333) mutant of TK+ HSV-2(333) also exhibited a single-base deletion, but at nucleotide 439. This deletion would cause a frameshift mutation at amino acid residue 147 and chain termination at amino acid residue 182. The frameshift mutations of TK- HSV(8710) and TK- HSV-2(333), respectively, occurred in sequences in which C was repeated three times and G was repeated seven times. The results raise the possibility that TK- frameshift mutations of HSV-2 may be common.

Acyclovir↗
PubMed1987

Second-generation pseudorabies virus vaccine with deletions in thymidine kinase and glycoprotein genes.

A modified-live pseudorabies virus (PRV) vaccine, designated PRV(dlg92/d1tk), with deletions in the thymidine kinase (tk) and glycoprotein-gIII (g92) genes, was derived from the PRV (Bucharest [BUK]-d13) vaccine strain. The vaccine virus also contained a deletion in glycoprotein gI. Despite 3 deletions, PRV(dlg92/d1tk) replicated to high titers in cell culture from 30 C to 39.1 C. Enzyme assays and autoradiography revealed that PRV(dlg92/d1tk) did not induce a functional tk activity in infected tk- RAB(BU) cells (rabbit skin). Rabbit skin cells were infected with PRV(dlg92/d1tk), with vaccine strains derived from BUK or Bartha K strains of PRV or with the virulent Illinois (ILL), Indiana-Funkhauser (IND-F), and Aujeszky (Auj) strains of PRV and were labeled with [3H]mannose from 4 or 5 to 24 hours after infection to investigate whether these viruses induced the synthesis of glycoprotein gIII. Nonionic detergent extracts were prepared and immunoprecipitated with antisera from pigs vaccinated with tk(-)-PRV(BUK-d13) or tk+-Bartha K, pigs vaccinated with tk+-PRV(BUK) strains and then challenge exposed to tk+-PRV(IND-F), naturally infected domestic or feral pigs, and pigs vaccinated with tk-)-PRV(dlg92/d1tk). Mouse monoclonal antibodies against PRV glycoproteins gIII, gp50, and gII were also studied. After immunoprecipitation, labeled PRV-specific proteins were analyzed by sodium dodecylsulfate-polyacrylamide gel electrophoresis and autoradiography. The PRV glycoprotein-gII complex, but not glycoprotein gIII, was synthesized in PRV(dlg92/d1tk)-infected cells. Glycoprotein gII and gIII were made in cells infected with PRV vaccine strains BUK, Bartha K, and BUK-d13 and with virulent PRV strains ILL, IND-F, and Auj. Cells infected with PRV(dlg92/d1tk) and with PRV strains ILL, IND-F, Auj, Bartha K, BUK, and BUK-d13, excreted into the cell culture medium a highly sulfated glycoprotein gX of about 90 kilodaltons. Antibodies to glycoprotein gIII were not detected in the sera of pigs inoculated with PRV(dlg92/d1tk), but were found in all other swine sera.

Animals↗
PubMed1986

Induction of thymidine kinase activity by viruses with group B DNA genomes: bovine cytomegalovirus (bovine herpesvirus 4).

The bovine herpesvirus type 4 (BHV-4) group has a slow replication cycle, a narrow host range, and cytopathogenic effects characteristic of cytomegaloviruses (CMV), but a Group B genome structure similar to that of lymphotropic Herpesvirus saimiri (HVS). Reference BHV-4 strain DN599 and BHV-4 strains N124 and FHV-2 induced in the cytosol fraction of thymidine kinase-negative (TK-) rabbit skin (RAB-BU) cell mutants a novel TK activity. The BHV-4-induced thymidine kinase (TK) differed from the principal cytosol TK of mock-infected cells in PAGE mobility (Rm) under non-denaturing conditions and in the capacity to efficiently substitute CTP for ATP as a phosphate donor. The BHV-4 thymidine phosphorylating activity could also be distinguished from many common herpesvirus-induced TKs because it lacked iododeoxycytidine phosphorylating activity. Iododeoxyuridine, trifluorothymidine and bromovinyldeoxyuridine inhibited [3H]thymidine (0.01 mM) phosphorylation by the BHV-4 enzyme in a dose-dependent manner, but arabinosylthymine and 2'-fluoro-5-methyl-arabinosyluracil (FMAU) were poor inhibitors of [3H]thymidine phosphorylation, and acyclovir and (dihydroxy-2-propoxymethyl)guanine (DHPG) did not inhibit at all at 60 and 40 times the concentrations of [3H]thymidine, respectively.

Animals↗
PubMed1986

Intramuscular and intravaginal vaccination of pregnant cows with thymidine kinase-negative, temperature-resistant infectious bovine rhinotracheitis virus (bovine herpes virus 1).

To test the safety and efficacy of a thymidine kinase-negative (TK-), temperature-resistant (TR) mutant of bovine herpes virus-1 (BHV-1) in pregnant cows, seronegative cows, 2-5 months pregnant, were vaccinated intramuscularly (i.m.) or intravaginally (i.vag.) with this candidate vaccine virus. I.m. vaccinated cows did not shed virus i.vag. or intranasally (i.n.), but i.vag. vaccinated cows replicated virus i.vag. for 8-9 days postvaccination (p.v.) Some of the cows were challenge exposed i.n. at 46 days p.v. with virulent TK+ BHV-1(Cooper). Vaccinated cows showed no clinical disease signs p.v. or postchallenge and responded anamnestically postchallenge. All cows delivered live calves. Pre-colostrum sera of the calves were negative for BHV-1 antibodies.

Animals↗
PubMed1986

Genital reinfection after recovery from initial genital infection with herpes simplex virus type 2 in guinea pigs.

The natural history of genital reinfection with herpes simplex virus type 2 (HSV-2) three weeks to one year after initial infection was explored by using a guinea pig model. Intravaginal reinoculation with HSV-2 of animals that had recovered from initial genital infection produced by wild-type or thymidine kinase-deficient HSV-2 produced asymptomatic genital reinfection characterized by high-titered replication of HSV-2 in the vaginal vault. Reinoculation did not result in acute neural infection, as was seen during initial genital herpes, and no evidence of latent infection by the rechallenge strain could be demonstrated. The frequency of recurrences appeared to be established by the initial infection because reinfection did not alter the pattern of recurrent genital herpes. The present study suggest that asymptomatic reinfection may produce a reservoir of communicable virus and that initial infection with a less virulent mutant of HSV-2 may provide protection against subsequent reexposure to wild-type HSV-2.

Animals↗
PubMed1985

Thymidine kinase-negative bovine herpesvirus type 1 mutant is stable and highly attenuated in calves.

Calves were vaccinated intranasally (IN) or intravenously (IV) with a thymidine kinase-negative (tk-) BHV-1 mutant. Vaccinated calves developed neutralizing antibodies but did not show clinical signs of infectious bovine rhinotracheitis (IBR). Vaccination also prevented clinical signs of IBR disease following IN challenge exposure of the calves to parental Los Angeles (LA) and USDA Cooper strains of tk+ BHV-1. Nasal swabs were collected for 10 days after the vaccination and the challenge exposures to monitor BHV-1 multiplication. At both 91 and 121 days post vaccination (PV), calves were also stressed for 5 days with dexamethasone (DEX) to induce reactivation of BHV-1 and nasal swabs were obtained. tk- BHV-1 multiplied in the nasal mucosa of IN vaccinated calves and was also recovered after DEX treatment. Likewise, tk- BHV-1 was isolated from the buffy coat fraction of IV vaccinated calves, but not from nasal swabs. tk- BHV-1 vaccination reduced the multiplication of tk+ BHV-1 in the nasal mucosa, but did not completely prevent development of a persistent infection by the challenge virus. The phenotypes of viruses isolated from the nasal swabs and buffy coats were analyzed by enzyme assays of extracts from virus-infected cells and by plaque autoradiography. These assays showed that tk- BHV-1 can persist for at least 3 months in vaccinated calves and may also be transmitted from vaccinated to control calves without reverting in vivo to tk+. The results demonstrate that the tk- BHV mutant is attenuated and efficacious as a vaccine.

Animals↗
PubMed1985

Thymidine kinase-deficient herpes simplex virus type 2 genital infection in guinea pigs.

In guinea pigs, thymidine kinase-producing strains of herpes simplex virus type 2 replicated to high titer in the vagina and spinal cord, and animals developed severe clinical disease. Infection with thymidine kinase-deficient virus resulted in similar vaginal virus titers; however, animals exhibited little or no clinical illness and only low titers of virus were detected in spinal cord homogenate cultures. Neural and extraneural latent infection as well as recurrent infection were noted in animals inoculated with either thymidine kinase-producing or -deficient viruses. These data suggest that neural pathways are important in the pathogenesis of genital herpes and that virus-coded thymidine kinase may influence virulence but is not required for latency.

Animals↗
PubMed1985

Attenuated properties of thymidine kinase-negative deletion mutant of pseudorabies virus.

A thymidine kinase (TK)-negative (TK-) deletion mutant of the Bucharest (BUK) strain of pseudorabies virus (PRV) was isolated. The mutant, designated as PRV (BUK d13), did not revert to TK-positive (TK+), even when propagated in medium that selected for TK+ viruses. The mutant also replicated equally well at 39.1 C and 34.5 C, and was easily distinguished from other PRV strains by molecular hybridization experiments, restriction nuclease fingerprints, and plaque autoradiography or other assays for the TK phenotype. The PRV (BUK d13) had greatly reduced virulence for mice and rabbits, compared with parental TK+ strains, PRV (BUK-5) and PRV (BUK-5A-R1), and provided mice with solid protection against the TK+ BUK and Aujeszky strains of PRV. Experiments were done in 5- to 6-week-old pigs to assess the safety and efficacy of PRV (BUK d13) in the natural host. In one experiment, pigs were vaccinated IM with 7.5 X 10(8) plaque-forming units of TK- PRV (BUK d13), and were then challenge exposed intranasally (IN) with 4.3 X 10(8) TCID50 of virulent PRV [Indiana-Funkhauser (IND-F)]. Vaccinated pigs did not have clinical signs of illness after vaccination or after challenge exposure. One nonvaccinated control pig died on postchallenge day 4; a 2nd nonvaccinated control pig became moribund, but eventually recovered. Pigs developed virus-neutralizing antibodies after vaccination, and had a secondary immunologic response after challenge exposure; however, PRV was not isolated from the tonsils or trigeminal ganglia of vaccinated pigs at postchallenge exposure day 11.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Thymidine kinase.

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Amino Acid Sequence↗