Development of a microfluidic platform for real-time detection of SARS-CoV-2 virus based on multifunctional silica membrane biosensors

After initial emergence in the Hubei province of China in the last months of 2019, the coronavirus disease 2019 (COVID-19) caused by the new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapidly spread all over the world, thus resulting in a major threat to the global public health security. While aerosols, such as respiratory droplets emitted by the cough and sneeze of symptomatic patients, were identified as the main transmission vectors, asymptomatic patients in incubation period also constitute possible transmitters of the disease. With around 5,000,000 confirmed cases and 330,000 deaths worldwide caused by the COVID-19 pandemic, there is an urgent to develop a set of measures for controlling the spread of the virus, decreasing the severity of the pathology in infected patients and eventually preventing infections by efficient vaccines. The development of accurate, cost-efficient and rapid screening and diagnostic methods is of crucial significance to slow down the circulation of the virus. In particular, the possibility to overtake the present qRT-PCR gold-standard for virus detection by point-of-care (POC) devices which do not rely on specialized personnel and access to biomedical laboratory environment, would bring a real breakthrough to control the spread of COVID-19. Here, we propose an alternative approach to overcome the limitations of the current testing procedures which should eliminate the need for biomedical personnel, specialized laboratory infrastructure, pre-testing sample preparation, additional reagents and bench-top instruments. Based on the expertise of one of the application partners in the design and production of automatized microfluidic devices combining isolation, detection and analysis modules, we propose an interdisciplinary approach for the development of a cheap and ultra-sensitive biosensor, based on porous silica (SiO2) supported active sensing surfaces, for direct viral RNA targeting. The following criteria will be addressed:-Portability: development of a miniaturized, fully-automated microfluidic device integrating all steps of the process within a single system architecture (RNA inactivation and extraction, sensing, readout of the hybridization events); without the need for additional reagents or external instruments;-Sensitivity: the versatility of the functionalization process to engineer porous SiO2 membranes conjugated to nucleic acid probes will allow for the optimization of several parameters determining sensing sensitivity (density and orientation of surface probes, covalent or non-covalent immobilization, discrete or multivalent attachment sites);-Reliability: four target nucleotide sequences will be selected, applying strict filters for probe uniqueness, probe-to-target duplex stability and similarity to non-target sequences;-Screening efficiency and specificity: the design of the microfluidic device will allow for multiple probe sequences to be tested in parallel; -Cost-effectiveness: production of the active sensing surfaces will make use of non-expensive materials and synthetic protocols; the readout step (DC resistance and bioimpedance) does not require additional reagent or labelling agent.These novel biosensors are expected to greatly enlarge the implementation of diagnosis units in non-medical environment such as airports, train stations and ports, where the fast screening of international travelling population would contribute to the early identification of potential vectors and would thus help in containing the disease propagation. Also, the tracking of asymptomatic transmitters will be highly facilitated by the development of cost-effective and portable testing devices. In future perspective, the proposed methodology could be applied to the detection of multiple pathogens within a single sample, thus enlarging the diagnosis scope of such system.