Lab-on-a-chip platforms for pathogen analysis

Rodriguez Mateos, Pablo

Abstract

Infectious diseases caused by pathogenic microorganisms are a big burden in developed and developing countries. The emergence and rapid global spread of virus and antimicrobial resistant bacteria is a significant threat to patients, healthcare systems and the economy of countries. Early pathogen detection is often hampered by low concentrations present in complex matrices such as food and body fluids.
Microfluidic technologies offer new and improved approaches for detection of pathogens on the microscale. Here, two microfluidic platforms for pathogen sorting and molecular identification were investigated: (1) inertial focusing and (2) microscale immiscible filtration. Inertial focusing in two serpentine channel designs etched in glass at different depths was evaluated with different microparticles, bacteria and blood. The shallow design allowed 2.2-fold concentration of Escherichia coli O157 cells, whereas the deep design accomplished recovery of 54% E. coli O157 depleted from 97% red blood cells in 0.81% haematocrit at flowrates of 0.7 mL min-1.
A lab-on-a-chip platform based on microscale immiscible filtration was investigated for capture and detection of nucleic acids and bacteria. For nucleic acids, oligo (dT) functionalised magnetic beads or silica paramagnetic particles in GuHCl were used to capture genomic RNA from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and genomic DNA from Neisseria gonorrhoeae, respectively. On-chip amplification and detection were performed via colorimetric loop-mediated isothermal amplification (LAMP). Results showed sensitive and specific detection of targeted nucleic acids (470 RNA copies mL-1 and 5 × 104 DNA copies mL-1) with no cross-reactivity to other RNAs and DNAs tested. The whole workflow was integrated in a single device and time from sample-in to answer-out was within 1h. The platform only required power for a heat source and showed potential for point of care diagnostics in resource-limited settings. For bacteria detection, anti-E. coli O157 functionalised magnetic beads were used to capture cells with > 90% efficiency and on-chip fluorescence in situ hybridisation and a staining assay were explored for bacteria identification.
A wide variety of microfluidic approaches for pathogen analysis have been devised in the literature with different advantages and drawbacks. Careful evaluation based on their purpose, integrated steps and end user is critical. Input from stakeholders right from the start of a project and throughout is vital to success. The platforms investigated herein have potential for applications such as sample preparation, pathogen concentration and specific molecular detection of E. coli O157, N. gonorrhoeae DNA, and SARS-CoV-2 RNA. With further development and clinical validation, the widespread use of these systems could facilitate early diagnosis of infectious diseases, allowing timely management of outbreaks and treatment and slowing the incidence of antimicrobial resistance.

Citation

Rodriguez Mateos, P. Lab-on-a-chip platforms for pathogen analysis. (Thesis). University of Hull. https://hull-repository.worktribe.com/output/4224144

• Thesis Type: Thesis
• Deposit Date: May 18, 2022
• Publicly Available Date: Feb 24, 2023
• Keywords: Chemistry
• Public URL: https://hull-repository.worktribe.com/output/4224144
• Additional Information: Department of Chemistry and Biochemistry, The University of Hull
• Award Date: Nov 1, 2021