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Simultaneous Detection of 13 Key Bacterial respiratory pathogens by Combination of Multiplex PCR and Capillary Electrophoresis

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Biomedical and Environmental Sciences 2017年第8期30卷 549-561页

作者： JIANG Lu Xi REN Hong Yu ZHOU Hai Jian ZHAO Si Hong HOU Bo Yan YAN Jian Ping QIN Tian CHEN Yu Department of respiratory Medicine Shengjing Hospital of China Medical University National Institute for Communicable Disease Control and Prevention State Key Laboratory for Infectious Disease Prevention and Control Chinese Center for Disease Control and Prevention Department of respiratory Medicine Zhejiang Provincial People's Hospital

Objective Lower respiratory tract infections continue to pose a significant threat to human health. It is important to accurately and rapidly detect respiratory bacteria. To compensate for the limits of current respiratory bacteria detection methods, we developed a combination of multiplex polymerase chain reaction （PCR） and capillary electrophoresis （MPCE） assay to detect thirteen bacterial pathogens responsible for lower respiratory tract infections, including Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catorrholis, Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus, Mycoplasma pneumoniae, Legionella spp., Bordetella pertussis, Mycobacterium tuberculosis complex, Corynebactefium diphthefiae, and Streptococcus pyogenes. Methods Three multiplex PCR reactions were built, and the products were analyzed by capillary electrophoresis using the high-throughput DNA analyzer. The specificity of the MPCE assay was examined and the detection limit was evaluated using DNA samples from each bacterial strain and the simulative samples of each strain. This assay was further evaluated using 152 clinical specimens and compared with real-time PCR reactions. For this assay, three nested-multiplex-PCRs were used to detect these clinical specimens. Results The detection limits of the MPCE assay for the 13 pathogens were very low and ranged from 10-7 to 10-2 ng/μL. Furthermore, analysis of the 252 clinical specimens yielded a specificity ranging from 96.5%-100.0%, and a sensitivity of 100.0% for the 13 pathogens. Conclusion This study revealed that the MPCE with high specificity and sensitivity. This assay survey of respiratory pathogens. assay is a rapid, reliable, and high-throughput method has great potential in the molecular epidemiological.

关键词： respiratory pathogens Lower respiratory tract infections Multiplex PCR Capillary electrophoresis

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Prevalence and Seroprevalence of Non-SARS Human Coronaviruses 229E and OC43 in East Tyrol/Austria

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Journal of Biosciences and Medicines 2023年第7期11卷 125-139页

作者： Sonnleitner Stefanie Sonnleitner Sissy Eva Hinterbichler Gernot Walder Dr. Gernot Walder GmbH Medical Laboratory Department of Virology Ausservillgraten Austria Division of Hygiene and Medical Microbiology Medical University of Innsbruck Innsbruck Austria

The recent SARS-CoV-2 pandemic renewed interest in other previously discovered human coronaviruses—the non-severe acute respiratory syndrome human coronavirus (non-SARS hCoVs) and this study is a starting point for a closer investigation of those. With the pandemic behind us we believe it to be important to also examine the current and past respiratory pathogen landscape in the patient population in our care. Therefore, 954 nasopharyngeal swabs of patients with respiratory diseases collected between October 2018 and March 2020 were tested against the pathogens Mycoplasma pneumoniae, Bordetella pertussis, Influenza A and virus, Human metapneumovirus, respiratory syncytial virus, Parainfluenza virus, human Adenovirus and Polyoma virus BK/JC. Swabs negative against these pathogens were further tested for OC43 and 229E by RT-qPCR. Human coronaviruses 229E and OC43 were proven as the causative agents in a considerable number of cases, confirmed by PCR. Overall, our results show that those two non-SARS hCoVs were responsible for 13.9% (11 of 79) of infections with flu-like symptoms of unknown etiology in the study area. In the subsequent seroprevalence study, it was shown that the seroprevalence rate of IgG antibodies against 229E and OC43 was over 50%, indicating that a big part of the population in our study area has been in contact with these non-SARS-CoVs and has built the specific humoral immune response accordingly.

关键词： NonSARS Human Coronavirus hCoV hCoV OC43 hCoV 229E respiratory pathogens

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