Experience in developing a new test system for screening and diagnosis of infections that cause acute respiratory diseases, and its use

Abstract

Relevance. Acute respiratory infections (ARI) are a serious health problem not only because of the high frequency of their occurrence, but also because of the economic damage they cause both in the form of direct costs (the cost of diagnosis and treatment) and indirect costs (disability, reduced labor productivity, etc.). Pregnant women and children under 5 years of age are included in the group of patients with risk factors for complications of influenza and other ARI, therefore, an analysis of the etiological structure of ARI and influenza in obstetric hospitals is an urgent task. In recent years, there has been an urgent need to create a national complex diagnostic test system based on molecular genetic methods for detecting infectious agents that cause ARI. Aims. The aim of the study is to analyze the etiological structure of ARI and influenza in patients with clinical symptoms and to develop and implement a new test system for rapid screening and diagnosis of infections that cause ARI. Materials & methods. When studying the etiological structure of ARI and influenza, cultural studies of the nasal and pharyngeal mucosa were carried out, followed by identification of microorganisms using MALDI-TOF mass spectrometry and molecular genetic study (real-time PCR) using an experimental test panel containing primers that allow detecting the following viruses: influenza A, B viruses, parainfluenza viruses of the 1st, 2nd, 3rd and 4th types, coronaviruses OS43, HKU1, NL63, E229, respiratory syncytial virus, metapneumovirus, rhinovirus and adenovirus, as well as bacterial pathogens of ARI: Haemophilus influenzae, Streptococcus pneumoniae, Streptococcus pyogenes, Moraxella catarrhalis, Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa. The test system was developed using the following methods: real-time PCR, a combination of reverse transcription and real-time PCR (RT-PCR) and the next generation sequencing (NGS) method. Results. The etiological structure of ARI and influenza was analyzed in patients with clinical manifestations (cough, tickling/sore throat/hyperemia of the mucous membrane of the palate and the back wall of the pharynx, shortness of breath/difficulty breathing, acute runny nose/nasal congestion). The species spectrum of bacterial and viral pathogens was revealed. A new test system based on PCR, real-time RT-PCR and NGS has been created for complex diagnostics of both viral and bacterial pathogens of ARI, consisting of three separate components: the main test system «ARI», which detects the main viral and bacterial pathogens of ARI, and two additional sets of reagents: «Oseltamivir resistance» and «Oseltamivir/ Zanamivir resistance». Conclusions. The new test system can be used to detect and differentiate nucleic acids of pathogens of ARI of humans. The test system seems to us promising for further use. As a result of the analysis of the etiological structure of acute respiratory infections and influenza, attention is drawn to a significantly smaller variety of identified pathogens in 2020 and a much more pronounced dominance of rhinovirus infection compared to our previous study in 2019.

Keywords

• acute respiratory infection
• acute respiratory viral infection
• influenza
• diagnosis
• test system
• etiological structure