Physical Analyses of E. coli Heteroduplex Recombination Products In Vivo: On the Prevalence of 5′ and 3′ Patches

BACKGROUND: Homologous recombination in Escherichia coli creates patches (non-crossovers) or splices (half crossovers), each of which may have associated heteroduplex DNA. Heteroduplex patches have recombinant DNA in one strand of the duplex, with parental flanking markers. Which DNA strand is exchanged in heteroduplex patches reflects the molecular mechanism of recombination. Several models for the mechanism of E. coli RecBCD-mediated recombinational double-strand-end (DSE) repair specify that only the 3'-ending strand invades the homologous DNA, forming heteroduplex in that strand. There is, however, in vivo evidence that patches are found in both strands. METHODOLOGY/PRINCIPLE FINDINGS: This paper re-examines heteroduplex-patch-strand polarity using phage lambda and the lambdadv plasmid as DNA substrates recombined via the E. coli RecBCD system in vivo. These DNAs are mutant for lambda recombination functions, including orf and rap, which were functional in previous studies. Heteroduplexes are isolated, separated on polyacrylamide gels, and quantified using Southern blots for heteroduplex analysis. This method reveals that heteroduplexes are still found in either 5' or 3' DNA strands in approximately equal amounts, even in the absence of orf and rap. Also observed is an independence of the RuvC Holliday-junction endonuclease on patch formation, and a slight but statistically significant alteration of patch polarity by recD mutation. CONCLUSIONS/SIGNIFICANCE: These results indicate that orf and rap did not contribute to the presence of patches, and imply that patches occurring in both DNA strands reflects the molecular mechanism of recombination in E. coli. Most importantly, the lack of a requirement for RuvC implies that endonucleolytic resolution of Holliday junctions is not necessary for heteroduplex-patch formation, contrary to predictions of all of the major previous models. This implies that patches are not an alternative resolution of the same intermediate that produces splices, and do not bear on models for splice formation. We consider two mechanisms that use DNA replication instead of endonucleolytic resolution for formation of heteroduplex patches in either DNA strand: synthesis-dependent-strand annealing and a strand-assimilation mechanism

A review of modelling tools for implementation of the EU Water Framework Directive in handling diffuse water pollution

A numerical catchment-scale model capable of simulating diffuse water pollution is necessary in sustainable environmental management for better implementation of the EU Water Framework Directive. This paper provides critical reviews of most popular and free models for diffuse water modelling, with detailed sources and application potential. Based upon these reviews, further work of selecting and testing the HSPF model was carried out, with a case study in the Upper Bann Catchment, Northern Ireland. The calibrated and validated HSPF model can well represent the characteristics of surface water quantity and quality. Climate change scenario evaluation in 5 years showed that when the annual mean temperature increase 3◦C the mean yearly total runoff volume will decrease by 11.1% and the mean daily river flow 11.4%. If 20% crop and pasture land is converted into forest land in the study area, the mean river concentration of nitrate, nitrite, NH4 and PO4 in 5 years will decrease by 19.4%, 33.3%, 31.3% and 31.3% respectively. When applying filter strip method in 80% crop and pasture land in the area, the reduction of the mean concentration of nitrate, nitrite, NH4 and PO4 in 5 years will be 15.3%, 33.3%, 31.3%, and 5.6% respectively. This study shows that HSPF is a suitable model in handling diffuse source water pollution, which can be introduced into the Programme of Measures in the River Basin Management Plans for better implementation of the EUWFD

Impact of sex on severe asthma: a cross-sectional retrospective analysis of UK primary and specialist care

\ua9 2023 Author(s) (or their employer(s)).Introduction: After puberty, females are more likely to develop asthma and in a more severe form than males. The associations between asthma and sex are complex with multiple intrinsic and external factors. Aim: To evaluate the sex differences in the characteristics and treatment of patients with severe asthma (SA) in a real-world setting. Methods: Demographic, clinical and treatment characteristics for patients with SA in the UK Severe Asthma Registry (UKSAR) and Optimum Patient Care Research Database (OPCRD) were retrospectively analysed by sex using univariable and multivariable logistic regression analyses adjusted for year, age and hospital/practice. Results: 3679 (60.9% female) patients from UKSAR and 18 369 patients (67.9% female) from OPCRD with SA were included. Females were more likely to be symptomatic with increased Asthma Control Questionnaire-6 (UKSAR adjusted OR (aOR) 1.14, 95% CI 1.09 to 1.18) and Royal College of Physicians-3 Question scores (OPCRD aOR 1.29, 95% CI 1.13 to 1.47). However, they had a higher forced expiratory volume in 1 second per cent (FEV1%) predicted (UKSAR 68.7% vs 64.8%, p<0.001) with no significant difference in peak expiratory flow. Type 2 biomarkers IgE (UKSAR 129 IU/mL vs 208 IU/mL, p<0.001) and FeNO (UKSAR 36ppb vs 46ppb, p<0.001) were lower in females with no significant difference in blood eosinophils or biological therapy. Females were less likely to be on maintenance oral corticosteroids (UKSAR aOR 0.86, 95% CI 0.75 to 0.99) but more likely to be obese (UKSAR aOR 1.67, 95% CI 145 to 1.93; OPCRD SA aOR 1.46, 95% CI 1.34 to 1.58). Conclusions: Females had increased symptoms and were more likely to be obese despite higher FEV1% predicted and lower type 2 biomarkers with consistent and clinically important differences across both datasets