Two activities involved in separate pathways for correcting G.T mispairs in DNA have been assayed on duplex substrates containing modified guanine bases. The first, the G.T mismatch incision activity, is specifically involved in short-patch repair of mispairs arising via deamination of 5-methylcytosine. The second activity can be detected by its ability to bind to G.T mispairs and may initiate correction by a long-patch mechanism. 6-Thioguanine and O6-methylguanine paired with thymine were efficiently incised by cell extracts if the modified guanine was in a CpG dinucleotide. Incision was not observed when either purine was paired with cytosine. Extracts of cells that are tolerant both to methylation damage and to 6-thioguanine in DNA also incised 6-thioguanine.T and O6-methylguanine.T base pairs. The data suggest that this activity is unlikely to contribute significantly to the biological effects of O6-methylguanine in DNA. A defect in this pathway is therefore unlikely to explain the cross-resistance of tolerant cells to the two base analogs in DNA. In binding assays, 6-thioguanine.T base pairs were recognized efficiently and to an equivalent extent by the same protein complex as G.T mispairs. O6-Methylguanine.T base pairs were also recognized but with reduced efficiency. No binding was observed to 6-thioguanine.C or O6-methylguanine.C base pairs. Recognition by the binding complex was essentially independent of the base immediately 5' to the mismatched guanine but was somewhat more efficient if O6-methylguanine was preceded by a purine. Extracts of two tolerant lines with a known defect in G.T mismatch binding failed to form complexes with substrates containing the modified bases.(ABSTRACT TRUNCATED AT 250 WORDS)

Summary A randomised, controlled, double‐blind, influenza virus, aerosol challenge of horses was undertaken to determine the efficacy of a cold‐adapted, temperature sensitive, modified‐live virus, intranasal, equine influenza vaccine. Ninety 11‐month‐old influenza‐naïve foals were assigned randomly to 3 groups (20 vaccinates and 10 controls per group) and challenged 5 weeks, 6 and 12 months after a single vaccination. Challenges were performed on Day 0 in a plastic‐lined chamber. Between Days 1 and 10, animals were examined daily for evidence of clinical signs of influenza. Nasal swabs for virus isolation were obtained on Day 1 and Days 1 to 8 and blood samples for serology were collected on Days 1, 7 and 14. There was no adverse response to vaccination in any animal. Following challenge at 5 weeks and 6 months, vaccinates had significantly lower clinical scores (P = 0.0001 and 0.005, respectively), experienced smaller increases in rectal temperature (P = 0.0008 and 0.0007, respectively) and shed less virus (P<0.0001 and P = 0.03, respectively) over fewer days (P<0.0001 and P = 0.002, respectively) than did the controls. After the 12 month challenge, rectal temperatures (P = 0.006) as well as the duration (P = 0.03) and concentration of virus shed (P = 0.04) were significantly reduced among vaccinated animals. The results of this study showed that 6 months after a single dose of vaccine the duration and severity of clinical signs were markedly reduced amongst vaccinated animals exposed to a severe live‐virus challenge. Appropriate use of this vaccine should lead to a marked reduction in the frequency, severity and duration of outbreaks of equine influenza in North America.

No abstract

We have investigated the specificities of G.T mismatch binding proteins and of G.T mismatch cleavage in extracts of mammalian cells. G.T mismatch-specific protein:DNA complex formation by cell extracts was independent of the local sequence context of the mismatch. Cell extracts performed similar levels of protein binding to DNA substrates in which a single G.T mispair was preceded by T, G, A, C, or 5-meC. In contrast, incision by extracts of the T-containing strand of a G.T mismatch exhibited a strong sequence specificity and efficient strand cleavage was only observed when the mismatched G was in a CpG sequence. Thus, oligonucleotides containing either CpgGpT or 5meCpGGpT were efficiently incised, but not those containing GpGCpT, ApGTpT, or TpGApT sequences. Cell lines made resistant to the alkylating agent N-methyl-N-nitrosourea have previously been found to be defective in a G.T mismatch binding reaction. The defect in binding by extracts prepared from these cells extended to G.T mismatches in several sequence contexts. The variant extracts nevertheless incised G.T mismatches normally suggesting that this particular binding activity is not required for incision. The data indicate that incision by this activity is targeted to the CpG sequences in which G.T mismatches are formed by the mutagenic deamination of DNA 5-methylcytosine. In this regard the repair pathway resembles the very short patch (vsp) repair pathway in Escherichia coli.

More than 83 000 patients in England waited more than a year for NHS treatment in July, the highest number since October 2008 and an 81-fold increase from 1032 in July last year, latest figures have shown. 1 The number of people who had been on waiting lists for more than 18 weeks in July was also the highest since records began in 2007, at 2.15 million. Just 46.8% of patients were treated within 18 weeks in July, against the 92% target-the lowest since records began, data from NHS England showed. The average wait for treatment by a hospital consultant rose to 19.6 weeks in July 2020.