Rapid Identification of Bacterial isolates Using Microfluidic Adaptive Channels and Multiplexed Fluorescence Microscopy

We demonstrate the rapid capture, enrichment, and identification of bacterial pathogens using Adaptive Channel Bacterial Capture (ACBC) devices. Using controlled tuning of device backpressure in polydimethylsiloxane (PDMS) devices, we enable the controlled formation of capture regions capable of trapping bacteria from low cell density samples with near 100% capture efficiency. The technical demands to prepare such devices are much lower compared to conventional methods for bacterial trapping and can be achieved with simple benchtop fabrication methods. We demonstrate the capture and identification of seven species of bacteria with bacterial concentrations lower than 1000 cells/mL, including common Gram-negative and Gram-positive pathogens such as Escherichia coli and Staphylococcus aureus . We further demonstrate that species identification of the trapped bacteria can be undertaken in the order of one-hour using multiplexed 16S rRNA-FISH with identification accuracies of 73-99% with unsupervised classification methods. ### Competing Interest Statement The work was carried out using a wide-field microscope from Oxford Nanoimaging, a company in which A.N.K. is a co-founder and shareholder. A.N.K. received no payment for this work from Oxford Nanoimaging, and Oxford Nanoimaging was not involved in any aspect of this work. ### Funding Statement This work was supported by an Oxford Martin School (by the establishment of the Oxford Martin School Programme on Antimicrobial Resistance Testing; to ANK, NS, CN), by Wellcome Trust grant 110164/Z/15/Z (to ANK) and by UK Biotechnology and Biological Sciences Research Council grants BB/N018656/1 and BB/S008896/1 (to ANK) and by Oxford's EPSRC/BBSRC Impact Acceleration Account EP/X525777/1 (award 0012379, to ANK, NS, and CN). The research was additionally supported by the National Institute for Health Research (NIHR) Health Protection Research Unit in Healthcare Associated Infections and Antimicrobial Resistance (NIHR200915) at the University of Oxford in partnership with United Kingdom Health Security Agency (UKHSA) and by the NIHR Oxford Biomedical Research Centre. This work was also supported by a John Fell Fund grant to NS (Grant application: 0008776). NS is an NIHR Oxford BRC Senior Research Fellow. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: Ethical approval for the use of clinical samples and isolates processed by the John Radcliffe Hospital microbiology laboratory in the development of diagnostic assays was granted by the UK's Health Research Authority (London - Queen Square Research Ethics Committee [REC reference: 17/LO/1420] I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes The data that support the findings of this study are available from the corresponding authors upon reasonable request.