About the project
The problems we are currently facing in connection with the global pandemic situation arose at various levels. The crisis developed not only due to absence or possibly delayed introduction of specific conditions limiting spreading of the pandemic at the political and social level, but also due to the unavailability of systems for express and specific detection at the level of laboratory diagnostics, which would lead to rapid identification and separation of infected individuals not only at the local but also the whole population level. At present, the limitation was not the knowledge about the novel coronavirus SARS-COV-2, which caused the current pandemic, but the global and local availability of rapid and specific high-throughput diagnostic tests or the individual components contained in such diagnostic tests, the limited availability of sufficiently staffed and equipped laboratories, where it would be possible to conduct such tests in a high-throughput mode.
Such a situation can occur unpredictably, but at the same time with a high probability and repeatedly, and therefore it is necessary to eliminate the above-mentioned limitations as much as possible. To this end, it is rational to create the most universal system possible enabling the rapid detection, identification and diagnosis of new infectious diseases with pandemic potential.
The project envisages, in relation to the current status and development of the COVID-19 pandemic, applied research conducted in a retrospective setting. Subsequently, the acquired and standardized procedures will be translated into a system that will be usable in the future in a prospective setting as individual more or less independent components of the pilot system. Using it, it will be possible to ensure rapid response to new, currently unknown threats in the form of new infectious diseases with pandemic potential, which is the main goal of the presented project.
Subject of the project
A system of early and rapid detection, identification and diagnosing of new infectious diseases with pandemic potential, in a pilot phase implemented in direct connection to the current COVID-19 pandemic. The system will consist of several relatively independent components. Their research and development will be a prerequisite for the successful fulfillment of this objective. The individual components are selected and assembled into the system so that the real possibility of achieving the comprehensive goal is maximized. Within the project pilot phase, partial goals will be directly associated with these components.
The project objective is to create a (universal) system of early and rapid detection, identification and diagnosing of new infectious diseases with pandemic potential; in the pilot phase the system is implemented in direct connection with the current pandemic of COVID-19.Procurement
1. Process optimization and development of standard procedures
associated with the primary processing of clinical samples (optimization of sampling and nucleic acid extraction);
2. Design and validation of reagent kit
based on multiplex RT-qPCR for targeted identification of SARS-CoV-2 viral sequences in the sample together with targeted analysis of other types of infectious agents causing diseases with clinical manifestations similar to COVID-19 (molecular diagnostics of SARS-CoV-2 and other infections causing respiratory diseases) with the ambition of certification of the kit for in vitro diagnostic use (CE IVD);
3. Preparation of recombinant enzymes for RNA isolation and RT-qPCR technique
(proteinase K, reverse transcriptase, DNA polymerase). The expression system of Escherichia coli or Pichia pastoris will be used for production. In addition to the fermentation production itself, the isolation and purification of enzymes will be optimized, as well as the long-term storage of enzymes. The ambition is to achieve practically unlimited availability of these components from domestic (national) sources in case of their unavailability at the current (international) commercial level;
4. Introduction and optimization of metagenomic sequencing protocol
for the detection and identification of infectious agents from clinical samples on the basis of information from worldwide available databases enabling the detection of even the previously undescribed infectious agents;
5. Reverse blinded validation study
of the selected clinical samples positive in RT-qPCR analyzes for monitored infectious agents by means of a reference method of metagenomic sequencing (massive parallel sequencing, or MPS) using the so-called hypothesis free de novo protocol created within the project scope;
6. Automation of sample processing procedures
at all levels to ensure the possibility of performing analyzes in a high-throughput mode applicable for the needs of diagnostics at the whole population level.
- universal know-how enabling the detection and identification of infectious agents (including new ones) based methodically on metagenomic sequencing of clinical samples (also available in the form of a new service);
- universal know-how enabling the design of diagnostic kits for the targeted detection of infectious agents based on the analysis of nucleic acids from clinical samples (also available as a new service);
- own and practically unlimited source of key enzymes needed for external source-independent preparation of diagnostic kits (also available in the form of new services and products) – new diagnostic kit for detection of SARS-CoV-2 infection with the possibility of direct differential diagnosis of clinical samples with clinical symptoms similar to COVID-19 (new product).