WORLDCOM: Development of new tools for real-time detection of zoonotic bacteria and antimicrobial resistance in veterinary, human and environmental sources
The Project #WORLDCOM
The work to be undertaken in this project will involve the development of real-time diagnostic on-site tools for the detection of zoonotic pathogens focusing on E. coli, Salmonella and Campylobacter together with resistance markers for identification of AMR including CTX-M-15, NDM-5, KPC-2, OXA-48 and MCR-1. Multiplex assays for both pathogen and resistance genes using novel isothermal technology Loop Mediated Endonuclease Cleavage-Loop Mediated Isothermal Amplification (LEC-LAMP) will be developed and evaluated in the first instance for use in a laboratory setting on standard laboratory platforms.
The use of isothermal amplification technologies including LAMP and LEC-LAMP (developed at NUI Galway), with the capacity to detect and differentiate single nucleotide polymorphisms (SNPs), will enable the development of AMR detection assays which will be suitable for use both in a laboratory setting and on-site, since complex instrumentation will not be required for amplification or for product detection. It is planned to incorporate the assays into a nucleic acid lateral flow (NALF) device, which will be evaluated for on-site application, with particular focus in the first instance on pen-side use in a farm setting. Additionally, the tests developed will be evaluated for use in environmental (irrigation, recreational and drinking water) and clinical settings (human samples).
The output of the on-site NALF assays will be communicated via a custom-designed smartphone application, which will link to relevant reference laboratories. This platform will facilitate early warning of potential zoonotic threats and enable follow up epidemiological investigations to be conducted using the full complement of lab-developed tools if warranted, including phenotypic and genotypic characterisation following isolation of the pathogen of interest from the corresponding sample.
A genomics approach will be used to develop a reference database of sequences from currently circulating AMR pathogenic strains, which will complement the IT and diagnostic assay tools in the WorldCom project. This task will involve curating relevant available sequences and targeted de novo sequencing of currently circulating and phenotypically characterised pathogens from the farm, environmental and clinical settings.
The sequence information in the reference database will be used to design the diagnostic assays and to develop and train novel Machine Learning (ML) algorithms for the detection and prediction of antimicrobial resistance from sequence data.
The novel combination of modified isothermal amplification technology, NALF devices capable of being used on-site and mobile technology will be the basis of an innovative, early warning tool-kit for prediction of antimicrobial resistance in various environments.
Delgado-Blas, JF., Ovejero, CM., David, S., Montero, N., Calero-Caceres, W., Garcillan-Barcia, MP., de la Cruz, F., Muniesa, M., Aanensen, DM., Gonzalez-Zorn, B. (2021). Population genomics and antimicrobial resistance dynamics of Escherichia coli in wastewater and river environments. Communications Biology. 4, 457. DOI: https://doi.org/10.1038/s42003-021-01949-x