Investigating the aetiology of respiratory tract infections in children admitted to Tygerberg Children's Hospital using molecular methods and viral culture

[摘要] ENGLISH ABSTRACT: IntroductionAcute respiratory tract infections cause significant morbidity and mortality worldwide, and arethe main reason for the utilisation of health care services. Identifying the aetiological cause of lowerrespiratory tract infections (LRTIs) is difficult at the best of times, and more than 20 viruses and bacteriahave been associated with LRTIs, which cannot be distinguished with clinical examination alone. Virusescan be detected in respiratory samples by a variety of methods, and without exception molecularmethods have proven to be more sensitive than non-molecular-based tests. The increased sensitivity ofmolecular methods may assist in expanding our knowledge of the pathogenesis of severe respiratorytract infections, and could have a positive influence on patient management, infection control,vaccination strategies and public health.Aims and objectives1. Determine the viral causes of lower respiratory tract infections requiring admission in using shellvial culture with immunofluorescent staining and two multiplex PCR assays, the Seeplex® RV15ACE Detection system (Seeplex® RV15 ACE) and the Respiratory Multiplex Real-Time RT-PCRLightMix® Customised Kit (Resp Multiplex RT-PCR).2. Compare the Seeplex® RV15 ACE and the Resp Multiplex RT-PCR with shell vial culture for thedetection of respiratory viruses in routine diagnostic respiratory samples.3. Examine the demographic and clinical characteristics associated with each respiratory viralpathogen.Materials and MethodsOne hundred and thirty-eight paediatric patients, admitted to Tygerberg Children's Hospital fromMay 2010 to August 2010 with a presumptive diagnosis of an acute respiratory tract infection wereincluded in the study. Nasopharyngeal or tracheal aspirates were collected, and all samples were testedby all three diagnostic methods. Clinical, demographic and laboratory data were collected through asystematic review of medical and laboratory records and subsequently anonymisedResultsThirty-seven viruses were detected in 36 samples (26.1%) by shell vial culture withimmunofluorescent staining; 169 viruses in 102 samples (73.9%) with the Seeplex® RV15 ACE; and 90viruses in 73 samples (52.9%) with the Resp Multiplex RT-PCR. Shell vial culture had excellent specificity,but low sensitivity for all of the respiratory viruses. Conversely, the Seeplex® RV15 ACE had excellentsensitivity for all viruses, but slightly lower specificity. This was due to the detection of additional viruses,which may have been true positives due to the increased sensitivity of this assay. The Resp Multiplex RTPCRhad excellent sensitivity and specificity.At least one respiratory pathogen could be identified in 80% of the patients. At least one viruswas detected in 57% of patients, bacterial micro-organisms in 6%, and both viral and bacterial pathogensin 17%. Viral-bacterial co-infections were associated with increased severity compared to otherinfections, as these children were more likely to receive steroids and a blood transfusion (p = 0.002), andmore likely to require mechanical ventilation (p < 0.001) and admission to the intensive care unit (p =0.04).ConclusionsWe confirmed that molecular techniques are significantly more sensitive than shell vial culturefor the detection of respiratory viruses in children. Due to their highly specific nature and the geneticvariability observed in viruses, an excellent, continuous quality control programme is essential to ensurethe continued superiority of these assays. Viral-bacterial co-infection is associated with increasedseverity of LRTIs in children. Further research is needed to elucidate the precise pathogenic andimmunologic mechanism of this interaction.

[发布机构] Stellenbosch University