One-Step Detection of the 2009 Pandemic Influenza A(H1N1) Virus by the RT-SmartAmp Assay and Its Clinical Validation

<div><h3>Background</h3><p>In 2009, a pandemic (pdm) influenza A(H1N1) virus infection quickly circulated globally resulting in about 18,000 deaths around the world. In Japan, infected patients accounted for 16% of the total population. The possibility of human-to-human transmission of highly pathogenic novel influenza viruses is becoming a fear for human health and society.</p> <h3>Methodology</h3><p>To address the clinical need for rapid diagnosis, we have developed a new method, the “RT-SmartAmp assay”, to rapidly detect the 2009 pandemic influenza A(H1N1) virus from patient swab samples. The RT-SmartAmp assay comprises both reverse transcriptase (RT) and isothermal DNA amplification reactions in one step, where RNA extraction and PCR reaction are not required. We used an exciton-controlled hybridization-sensitive fluorescent primer to specifically detect the HA segment of the 2009 pdm influenza A(H1N1) virus within 40 minutes without cross-reacting with the seasonal A(H1N1), A(H3N2), or B-type (Victoria) viruses.</p> <h3>Results and Conclusions</h3><p>We evaluated the RT-SmartAmp method in clinical research carried out in Japan during a pandemic period of October 2009 to January 2010. A total of 255 swab samples were collected from outpatients with influenza-like illness at three hospitals and eleven clinics located in the Tokyo and Chiba areas in Japan. The 2009 pdm influenza A(H1N1) virus was detected by the RT-SmartAmp assay, and the detection results were subsequently compared with data of current influenza diagnostic tests (lateral flow immuno-chromatographic tests) and viral genome sequence analysis. In conclusion, by the RT-SmartAmp assay we could detect the 2009 pdm influenza A(H1N1) virus in patients' swab samples even in early stages after the initial onset of influenza symptoms. Thus, the RT-SmartAmp assay is considered to provide a simple and practical tool to rapidly detect the 2009 pdm influenza A(H1N1) virus.</p> </div

Association of maternal whole blood fatty acid status during the prenatal period with term birth dimensions : a cross-sectional study

Objective: To investigate selected fatty acid (FA) profiles in maternal whole blood during normal pregnancy and to evaluate their associations with term birth dimensions. Methods: We characterized nine major maternal blood FAs representing four FA families during the second and third trimester of pregnancy, and explored their associations with birth weight, length, and chest or head circumferences by multivariate regression models, using data from 318 mother-newborn pairs of the Hokkaido Study. Results: The absolute and/or relative contents of maternal blood docosahexaenoic acid and arachidonic acid were lowest at 35–41 gestational weeks during pregnancy, as was the essential FA status index. Different from palmitic and stearic acids, palmitoleic and oleic acid contents were higher at 35–41 gestational weeks than those at 23–31 gestational weeks. Three FA components were identified through principal component analysis, and were used in association analysis. Component 3, which was positively and significantly loaded by eicosapentaenoic acid (EPA), was associated with chest circumference [β=0.281, 95% confidence interval (CI): 0.006, 0.556] at 35–41 gestational weeks (P=0.046). No significant associations were observed for Component 1 and 2 loaded by FAs except EPA. Conclusion: Maternal blood EPA content may have an important influence on infant chest circumference

Enhanced antitumor efficacy of fiber‐modified, midkine promoter‐regulated oncolytic adenovirus in human malignant mesothelioma

Oncolytic virotherapy using adenoviruses has potential for therapeutic benefits in malignant mesothelioma. However, the downregulation of coxsackie virus/adenovirus receptor (CAR) expression is frequently a critical rate-limiting factor that impedes the effectiveness of adenovirus serotype 5 (Ad5)-based vectors in many cancer types. We evaluated CAR (Ad5 receptor) and CD46 (adenovirus serotype 35 [Ad35] receptor) expression in six human malignant mesothelioma cell lines. Very low CAR expression was observed in MSTO-211H and NCI-H2052 cells, whereas the other cell lines showed strong expression. In contrast, CD46 was highly expressed in all mesothelioma cell lines. On this basis, we replaced the CAR binding sequence of Ad5 with the CD46 binding sequence of Ad35 in the replication-defective adenoviruses and the tumor-specific midkine promoter-regulated oncolytic adenoviruses. By this fiber modification, the infectivity, virus progeny production, and in vitro cytocidal effects of the adenoviruses were significantly enhanced in low CAR-expressing MSTO-211H and NCI-H2052 cells, also resulting in similar or even higher levels in high CAR-expressing mesothelioma cell lines. In MSTO-211H xenograft models, the fiber-modified oncolytic adenovirus significantly enhanced antitumor effect compared to its equivalent Ad5-based vector. Our data demonstrate that Ad35 fiber modification of binding tropism in a midkine promoter-regulated oncolytic Ad5 vector confers transductional targeting to oncolytic adenoviruses, thereby facilitating more effective treatment of malignant mesothelioma

Enhanced antitumor efficacy of fiber-modified, midkine promoter-regulated oncolytic adenovirus in human malignant mesothelioma

Oncolytic virotherapy using adenoviruses has potential for therapeutic benefits in malignant mesothelioma. However, the downregulation of coxsackie virus/adenovirus receptor (CAR) expression is frequently a critical rate-limiting factor that impedes the effectiveness of adenovirus serotype 5 (Ad5)-based vectors in many cancer types. We evaluated CAR (Ad5 receptor) and CD46 (adenovirus serotype 35 [Ad35] receptor) expression in six human malignant mesothelioma cell lines. Very low CAR expression was observed in MSTO-211H and NCI-H2052 cells, whereas the other cell lines showed strong expression. In contrast, CD46 was highly expressed in all mesothelioma cell lines. On this basis, we replaced the CAR binding sequence of Ad5 with the CD46 binding sequence of Ad35 in the replication-defective adenoviruses and the tumor-specific midkine promoter-regulated oncolytic adenoviruses. By this fiber modification, the infectivity, virus progeny production, and in vitro cytocidal effects of the adenoviruses were significantly enhanced in low CAR-expressing MSTO-211H and NCI-H2052 cells, also resulting in similar or even higher levels in high CAR-expressing mesothelioma cell lines. In MSTO-211H xenograft models, the fiber-modified oncolytic adenovirus significantly enhanced antitumor effect compared to its equivalent Ad5-based vector. Our data demonstrate that Ad35 fiber modification of binding tropism in a midkine promoter-regulated oncolytic Ad5 vector confers transductional targeting to oncolytic adenoviruses, thereby facilitating more effective treatment of malignant mesothelioma

Importance of Controlling Drug-resistant Pseudomonas aeruginosa Infection: Experience from Lung Transplantation in a Cystic Fibrosis Case

Cystic fibrosis (CF) is rare in Japan. We encountered a CF case with drug-resistant Pseudomonas aeruginosa infection and successfully performed lung transplant from living related donors. A combination of betalactams and aminoglycosides for drug-resistant P. aeruginosa infection was administered before lung transplantation. Intravenous colistin was also used immediately before and after transplant surgery. Gram staining of respiratory specimens was performed every day after surgery and it was useful in monitoring infection status. Strict monitoring of infections by the Gram staining and culture of respiratory specimens is considered to be effective in preventing lower respiratory infection in lung transplantation