Smoking cessation intervention prior to orthopedic surgery: A study protocol to determine patient outcomes and feasibility.

The aim of the study is to analyze the effect of smoking and smoking cessation on the incidence of complications among orthopedic and hand surgery patients, and to determine the feasibility of smoking cessation intervention, as well as factors predicting success in smoking cessation. Orthopedic and hand surgery patients will be invited to participate in the study, which will recruit 550 participants (at least 20% daily smokers). A participant will be defined as a daily smoker if he/she reports daily smoking and/or laboratory tests show active smoking. Data will be collected using a self-reported questionnaire and from medical records. Smokers will receive information about the benefits of smoking cessation and will be encouraged to quit. Medication or nicotine replacement therapy will be prescribed. Laboratory tests will be taken two weeks before and two weeks after surgery. Follow-up phone calls will be made at 3, 6, and 12 months after surgery. The primary outcome is any complication, defined as a prolonged stay in hospital or any additional visit to or measure taken by a health service during the 12 months after surgery. Data on complications are mainly obtained from personal health records and from the information received at the follow-up; the rest of the data will be collected from the register of healthcare-associated infections. Secondary outcomes are the number and types of complications. The sample (n=550) was calculated to observe a 10% difference in complications between smokers and non-smokers (5% alpha level and 80% power), considering a 10% drop-out rate. Logistic regression and log-linear models will be used for data analyses.

An improved RT-qPCR method for direct quantification of enveloped RNA viruses.

Reverse transcription quantitative PCR (RT-qPCR) has emerged as the gold standard for virus detection and quantification, being utilized in numerous diagnostic and research applications. However, the direct detection of viruses has so far posed a challenge as the viral genome is often encapsidated by a proteinaceous layer surrounded by a lipid envelope. This necessitates an additional and undesired RNA extraction step prior to RT-qPCR amplification. To circumvent this limitation, we have developed a direct RT-qPCR method for the detection of RNA viruses. In our method, we provide a proof-of-concept using phage phi6, a safe-to-use proxy for pathogenic enveloped RNA viruses that is commonly utilized in e.g. aerosolization studies. First, the phage phi6 envelope is removed by 1% chloroform treatment and the virus is then directly quantified by RT-qPCR. To identify false negative results, firefly luciferase is included as a synthetic external control. Thanks to the duplex format, our direct RT-qPCR method reduces the reagents needed and provides an easy to implement and broadly applicable, fast, and cost-effective tool for the quantitative analysis of enveloped RNA viruses.•One-step direct RT-qPCR quantification of phage phi6 virus without prior RNA isolation.•Reduced reaction volume for sustainable and cost-effective analysis.

Cold-Active Shewanella glacialimarina TZS-4T nov. Features a Temperature-Dependent Fatty Acid Profile and Putative Sialic Acid Metabolism.

Species of genus Shewanella are among the most frequently identified psychrotrophic bacteria. Here, we have studied the cellular properties, growth dynamics, and stress conditions of cold-active Shewanella strain #4, which was previously isolated from Baltic Sea ice. The cells are rod-shaped of ~2µm in length and 0.5µm in diameter, and they grow between 0 and 25°C, with an optimum at 15°C. The bacterium grows at a wide range of conditions, including 0.5-5.5% w/v NaCl (optimum 0.5-2% w/v NaCl), pH 5.5-10 (optimum pH 7.0), and up to 1mM hydrogen peroxide. In keeping with its adaptation to cold habitats, some polyunsaturated fatty acids, such as stearidonic acid (18:4n-3), eicosatetraenoic acid (20:4n-3), and eicosapentaenoic acid (20:5n-3), are produced at a higher level at low temperature. The genome is 4,456kb in size and has a GC content of 41.12%. Uniquely, strain #4 possesses genes for sialic acid metabolism and utilizes N-acetyl neuraminic acid as a carbon source. Interestingly, it also encodes for cytochrome c3 genes, which are known to facilitate environmental adaptation, including elevated temperatures and exposure to UV radiation. Phylogenetic analysis based on a consensus sequence of the seven 16S rRNA genes indicated that strain #4 belongs to genus Shewanella, closely associated with Shewanella aestuarii with a ~97% similarity, but with a low DNA-DNA hybridization (DDH) level of ~21%. However, average nucleotide identity (ANI) analysis defines strain #4 as a separate Shewanella species (ANI score=76). Further phylogenetic analysis based on the 92 most conserved genes places Shewanella strain #4 into a distinct phylogenetic clade with other cold-active marine Shewanella species. Considering the phylogenetic, phenotypic, and molecular characterization, we conclude that Shewanella strain #4 is a novel species and name it Shewanella glacialimarina sp. nov. TZS-4T, where glacialimarina means sea ice. Consequently, S. glacialimarina TZS-4T constitutes a promising model for studying transcriptional and translational regulation of cold-active metabolism.