Pre-analytics for pandemic preparedness

PI(s)/Head responsible for the resource:

Martin Sundqvist

Host organisation(s):

Dept. of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital

Resource description:

This project will focus on the crucial pre-analytical steps for detection of viruses and bacteria with metagenomics in order to deliver a national standard operating procedure. The main focus will be to compare and evaluate matrices for sample transportation, effect of storage and nucleic acid extraction methods. The main outcome measure will be the effect on the quality (quantity and fragmentation of DNA/cDNA, the amount of microbial reads, bias in relative composition and ability to use sequences for further typing) of fast long-read sequencing using Oxford Nanopore technology and bioinformatics tools. The most likely infection route in coming pandemics is the upper respiratory tract and upper respiratory samples will thus be our primary focus. In the autumn of 2022 the project will focus on methodological issues where exact quantification of SARS-CoV-2, Influenza and RSV using ddPCR will be implemented. Further, samples from the upper airways will be spiked with virus and bacteria, stored in different conditions and then analysed using ddPCR, culture and metagenomic approaches. During the winter of 2022/2023, clinical samples will be collected and analysed to verify these findings. The project aims to expand into more sample matrices and the detection of antibiotic resistance genes and direct typing of viruses and bacteria in 2023. The data from these experiments will render new information regarding the susceptibility of different sample matrices to the environmental conditions.

Research findings:

To be able to evaluate the conditions for storage and transport of respiratory samples we verified the precision of Real time PCR to evaluate the exact concentration of respiratory viruses in frozen respiratory samples.

Impact on prepardness for future pandemics:

Based on our data we can conclude that the use of Ct-values to estimates the viral RNA-load in respiratory samples gives a very good proxy of the amount in the sample. This might be of importance in coming pandemics as the viral load can give an idea on where in the infection period the patient is (early/late) and also if the patient copes with the viral infection (i.e. that the viral levels decreases and disappears over time). During the time of the project several studies has arrived that have investigated the original approach of this project regarding transportation and sample material. This will be useful in a coming laboratory preparedness plan.

Contact information:

Martin Sundqvist
Dept. of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital
Email: martin.sundqvist@regionorebrolan.se