BACKGROUND: The pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) has prompted a need for mass testing to identify patients with viral infection. The high demand has created a global bottleneck in testing capacity, which prompted us to modify available resources to extract viral RNA and perform reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) to detect SARS-COV-2. OBJECTIVES: Report on the use of a DNA extraction kit, after modifications, to extract viral RNA that could then be detected using an FDA-approved SARS-COV-2 RT-qPCR assay. MATERIALS AND METHODS: Initially, automated RNA extraction was performed using a modified DNA kit on samples from control subjects, a bacteriophage, and an RNA virus. We then verified the automated extraction using the modified kit to detect in-lab propagated SARSCOV-2 titrations using an FDA approved commercial kit (S, N, and ORF1b genes) and an in-house primer-probe based assay (E, RdRp2 and RdRp4 genes). RESULTS: Automated RNA extraction on serial dilutions SARS-COV-2 achieved successful one-step RT-qPCR detection down to 60 copies using the commercial kit assay and less than 30 copies using the in-house primer-probe assay. Moreover, RT-qPCR detection was successful after automated RNA extraction using this modified protocol on 12 patient samples of SARS-COV-2 collected by nasopharyngeal swabs and stored in viral transport media. CONCLUSIONS: We demonstrated the capacity of a modified DNA extraction kit for automated viral RNA extraction and detection using a platform that is suitable for mass testing. LIMITATIONS: Small patient sample size. CONFLICT OF INTEREST: None.

Background: Sever acute respiratory syndrome Coronavirus-2 (SARS-COV-2) spread prompted mass testing. The main method for testing is by any FDA approved kits for RNA extraction followed by One-Step RT-qPCR based on primerprobe assays. Yet, the high demand for these kits created a global bottleneck in the testing capacity. Methods: We developed a Full-In-House Method (FinHM) suitable for automated viral RNA extraction using full in-house solutions utilizing the MagMax TM beads followed by an In-House RT-qPCR based on the CDC/WHO recommended 'primer-probe' assay targeting the following genes; E, RdRp2, and RdRp4. FinHM was validated by an FDA approved kit that targets S, N, and ORF1b genes made by Thermo Fisher Scientific (TF). Results: The sensitivity and specificity of the automated RNA extraction were evaluated on serial dilutions of in-laboratory propagated SARS-COV-2 with a successful detection down to 46 copies in both assays (P>0.05). Moreover, automated FinHM was successful in extraction of SARS-COV-2 RNA in 266 clinical samples, in which the test results replicated the FDA approved test results (>99% similarity, P>0.05). The In-House RT-qPCR assay had low limit of detection (5 RNA templates), with significant negative correlation between the Ct values and RNA titrations as shown by Pearson correlation (-0.8, -0.8 and -0.7 for E, RdRp2 and RdRp4, respectively). Finally, FinHM was also successful in extraction of SARS-COV-2-spiked plasma and patient plasma samples. Conclusion: We report a reliable, reproducible, specific, sensitive and low-cost platform for automated RNA extraction and detection from SARS-COV-2 and other viruses which is suitable for clinical and mass testing.

Purpose:Tiotropium (SPIRIVA®) is used in the treatment of moderate to severe chronic obstructive pulmonary disease (COPD) in patients with persistent dyspnea despite using a short acting bronchodilator (SABD).This paper explores the role of tiotropium in the treatment of mild COPD. Methods:The Cochrane Library, EMBASE, Pubmed, and Clinicaltrials.gov were searched on February 2018.We included randomized controlled trials (RCTs) that evaluated tiotropium in patients with mild COPD.Three authors assessed studies for eligibility. Outcomes included symptoms, quality of life, exercise duration, lung function, COPD exacerbations and hospitalizations, and serious adverse events. Results: Three RCTs were selected as the best available evidence. Based on the results of the main trial, quality of life and symptoms were improved with tiotropium as compared to placebo with a difference between groups at 24 months to be 1.2 (95% CI: 0.5 to 1.9; p=0.0011) using the COPD Assessment test (CAT) score. Frequency of acute exacerbations of COPD (AECOPD) requiring hospitalization was reduced by 10.3% (28.9% with tiotropium vs 39.2% with placebo) in patients receiving tiotropium. One RCT reported no statistically significant difference in exercise duration (27 ± 27 secs) in the tiotropium group vs 50 ± 21 secs in the placebo group; (p=0.4153). Oropharyngeal discomfort was more common with tiotropium (number needed to harm of 12) compared to placebo. Conclusions: Evidence suggests that tiotropium may reduce COPD exacerbations and hospitalizations and improve quality of life in patients with mild COPD.There is an increased risk of oropharyngeal discomfort with tiotropium.

Beta Thalassemia represents a major public health problem in Egypt. The carrier rate varies between 5.5% to > 9%. It is estimated that there are 1000/1.5 million per year live births born with beta thalassemia. (1) β thalassemia occurs when there is a quantitative reduction of β globin chains that are usually structurally normal. (2) They are caused by mutations that nearly all affect the β globin locus and are extremely heterogeneous. Almost every possible defect affecting gene expression at transcription or post-transcriptional level, including translation, have been identified in β thalassemia. (3) These genetic defects lead to a variable reduction in β globin output ranging from a minimal deficit (mild β+ thalassemia alleles) to complete absence (β° thalassemia). Aim of the work:We aimed in this study to assess the molecular changes in transfusion dependent Beta thalassemia patients and the correlation of these molecular changes with their clinical outcomes. Patients & methods: This study will include 40 transfusion dependent β thalassemia patients with age range of 2 -18 years, recruiting the Pediatric Hematology unit in Minia University children hospital. Study procedure: β-Thalassemia mutation identification of samples will be performed by the reverse dot blot hybridization technique (RDB). For RDB, a panel of primers and probes using the beta globin strip assay well be used (β-Globin Strip Assay MED kit, VIENNA lab