Journal of virological methods
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This paper describes the molecular detection of respiratory viruses from nasopharyngeal flocked swabs (flocked swabs) and nasopharyngeal washes (washes) in a clinical setting. Washes and flocked swabs collected from children<3 years old hospitalized with a lower respiratory tract infection were tested for parainfluenza virus 1-3, respiratory syncytial virus, influenza A and B and metapneumovirus (Group 1) and adenovirus, rhinovirus and coronavirus (Group 2) using real-time reverse transcriptase PCR (rRT-PCR). A consensuses standard was used to determine sensitivity and compare cycle thresholds (C(T)) of washes and flocked swabs for each virus and group of viruses. ⋯ When the flocked swabs and washes of Group 1 viruses were collected on the day of admission, the sensitivity of both sample types was 100%. Wash specimens had a lower C(T) value and higher sensitivity than flocked swabs; however there was no statistical difference in the sensitivity of a flocked swab (89%) versus wash (93%) for the detection of Group 1 viruses, particularly when samples were collected on the same day. Flocked swabs may be a useful alternative to washes for detection of respiratory viruses in clinical settings.
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Exhaled breath condensate is a non-invasive method for detecting a wide number of molecules as well as genomic DNA in the airways. No study investigated the detection of viral DNA in exhaled breath condensate, while only one study excluded its usefulness for detection of influenza virus RNA. In this study, the suitability of exhaled breath condensate for detecting herpesviruses infection or reactivation in the respiratory tract of lung transplant recipients was evaluated. ⋯ Eighteen bronchoalveolar lavages (75%), six whole blood samples (25%), and two transbronchial biopsies (8.3%) were positive for at least one herpesvirus. Only one exhaled breath condensate specimen was positive for HCMV DNA (and positive also in the bronchoalveolar lavage, with low viral load in both specimens); while no other patient, irrespective of the viral load in any specimen or the presence of clinical symptoms and signs, had a positive exhaled breath condensate. These findings seem to exclude the suitability of exhaled breath condensate for non-invasive detection of viral DNA in the respiratory tract of lung transplant recipients.
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Ebola virus (EBOV) causes severe hemorrhagic fever in humans and nonhuman primates with high mortality rates. Rapid identification of the virus is required to prevent spread of the infection. In this study, we developed and evaluated a one-step simple reverse transcription-loop mediated isothermal amplification (RT-LAMP) assay for the rapid detection of Zaire ebolavirus (ZEBOV), the most virulent species of EBOV, targeting the trailer region of the viral genome. ⋯ In addition, the assay was highly specific for ZEBOV. The RT-LAMP assay developed in this study is rapid, simple, highly specific, and sensitive for the detection of ZEBOV, and so may be an effective diagnostic tool. Furthermore, as this technique does not require sophisticated instrumentation, it seems very suitable for diagnosis in the field or laboratories in Ebola outbreak areas such as Central Africa.
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The SARS-associated human coronavirus (SARS-HCoV) is a newly described, emerging virus conclusively established as the etiologic agent of the severe acute respiratory syndrome (SARS). This study presents a single-tube RT-PCR assay that can detect with high analytical sensitivity the SARS-HCoV, as well as several other coronaviruses including other known human respiratory coronaviruses (HCoV-OC43 and HCoV-229E). Species identification is provided by sequencing the amplicon, although a rapid screening test by restriction enzyme analysis has proved to be very useful for the analysis of samples obtained during the SARS outbreak in Toronto, Canada.
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The severe acute respiratory syndrome (SARS) epidemic originating from China in 2002 was caused by a previously uncharacterized coronavirus that could be identified by specific RT-PCR amplification. Efforts to control future SARS outbreaks depend on the accurate and early identification of SARS-CoV infected patients. A real-time fluorogenic RT-PCR assay based on the 3'-noncoding region (3'-NCR) of SARS-CoV genome was developed as a quantitative SARS diagnostic tool. ⋯ The assay's detection sensitivity could reach 0.005 pfu or 6-8 GE per assay. It was preliminarily demonstrated that the assay could efficiently detect SARS-CoV from clinical specimens of SARS probable and suspected patients identified in Taiwan. The 3'-NCR based SARS-CoV assay demonstrated 100% diagnostic specificity testing samples of patients with acute respiratory disease from a non-SARS epidemic region.