Development of a diagnostic panel for respiratory pathogens using real-time PCR with microfluidics (Biomark HD platform)

• Grant number: 23/14279-9
• Support Opportunities: Scholarships abroad - Research
• Start date: February 16, 2024
• End date: September 30, 2024
• Field of knowledge: Biological Sciences - Microbiology - Applied Microbiology
• Principal Investigator: Ivana Barros de Campos
• Grantee: Ivana Barros de Campos
• Host Investigator: Daniela Mulari Ferreira
• Host Institution: Instituto Adolfo Lutz (IAL). Coordenadoria de Controle de Doenças (CCD). Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Institution abroad: Liverpool School of Tropical Medicine (LSTM), England

Abstract

The composition of the microbiome of the human nasopharynx is very complex and knowing their interactions is important in preventing secondary bacterial infections, which have high mortality. Several factors such as human genetic composition and environmental factors directly affect the microbiome, and knowing its constituents is an important part of controlling pathogens that normally inhabit the human nasopharynx in a commensal manner, but which can cause infections such as otitis, pneumonia, bacteremia or meningitis. The diagnostic technique based on the culture of microorganisms is time-consuming, provides late results, and often presents false-negative results. Therefore, other culture-independent methodologies are important to understand the composition of the nasopharyngeal microbiota, as well as to assist in patient treatment. The PCR (Polymerase chain reaction) technique has high specificity, sensitivity and provides fast results, but is limited to the small number of targets to be detected per reaction. More advanced methodologies, such as next-generation sequencing, are expensive to apply in routine diagnosis, in addition to not being accessible to lower-income countries. Therefore, this project aims to use microfluidic PCR through the Biomark HD platform, based on automated real-time PCR with nanoliters of reagents and samples. As it takes place individually in nanocompartments, with the same volume applied in a single PCR reaction, it is possible to carry out more than 9,000 reactions simultaneously according to the equipment configuration, thus reducing the cost of the technique and the time to obtain results. Therefore, this study intends to apply this methodology for the detection of respiratory pathogens that constitute the microbiota of the human nasopharynx for lower-cost microbiome studies and also perhaps to standardize this methodology for its use in the diagnosis of infectious diseases of the respiratory tract.