{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,6,24]],"date-time":"2026-06-24T06:02:42Z","timestamp":1782280962414,"version":"3.54.5"},"posted":{"date-parts":[[2025,1,16]]},"group-title":"Microbiology","reference-count":26,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,1,16]],"date-time":"2025-01-16T00:00:00Z","timestamp":1736985600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,1,16]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Bacteremia is a serious clinical condition in which pathogenic bacteria enter the bloodstream, putting patients at risk of septic shock and necessitating antibiotic treatment. Choosing the most effective antibiotic is crucial not only for resolving the infection but also for minimizing side effects, such as dysbiosis in the healthy microbiome and reducing the selection pressure for antibiotic resistance. This requires prompt identification of the pathogen and antibiotic susceptibility testing, yet these processes are inherently slow in standard clinical microbiology labs due to reliance on growth-based assays. Although alternative methods exist, they are rarely adopted in clinical settings because they involve complex protocols and high costs for training and infrastructure. Here, we present an optimized, simple protocol for rapidly and efficiently isolating bacterial pathogens from blood without altering typical laboratory workflows. Our method is cost-effective and compatible with commonly available laboratory instruments, offering the advantage of isolating bacterial cells directly, which bypasses the delays associated with traditional blood culture methods and enables faster diagnostic results. The protocol achieved over 70% efficiency within 30 minutes, remained effective at low bacterial concentrations (1\u201310 bacteria\/0.3 mL blood), and preserved bacterial viability with no notable change in growth lag times. We validated the protocol on several clinically relevant bacterial strains, including\n                  <jats:italic>Escherichia coli, Klebsiella pneumoniae<\/jats:italic>\n                  , and\n                  <jats:italic>Staphylococcus aureus<\/jats:italic>\n                  . These findings highlight our protocol\u2019s potential utility in clinical and research settings, facilitating timely cultures and minimizing diagnostic delays.\n                <\/jats:p>","DOI":"10.1101\/2025.01.14.633023","type":"posted-content","created":{"date-parts":[[2025,1,16]],"date-time":"2025-01-16T08:50:17Z","timestamp":1737017417000},"source":"Crossref","is-referenced-by-count":0,"title":["A Simple, Low-Cost, and Efficient Protocol for Rapid Isolation of Pathogenic Bacteria from Human Blood"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5537-0867","authenticated-orcid":false,"given":"Fatma S.","family":"Coskun","sequence":"first","affiliation":[{"name":"Department of Pharmacology, The University of Texas Southwestern Medical Center, Dallas, TX 75390, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6376-871X","authenticated-orcid":false,"given":"Joshua","family":"Quick","sequence":"additional","affiliation":[{"name":"School of Biosciences, University of Birmingham, Birmingham, B15 2TT, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4883-7681","authenticated-orcid":false,"given":"Erdal","family":"Toprak","sequence":"additional","affiliation":[{"name":"Department of Pharmacology, The University of Texas Southwestern Medical Center, Dallas, TX 75390, USA"},{"name":"Lyda Hill Department of Bioinformatics, The University of Texas Southwestern Medical Center, Dallas, TX 75390, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026041021401634000_2025.01.14.633023v1.1","doi-asserted-by":"publisher","DOI":"10.1001\/jama.281.1.61"},{"key":"2026041021401634000_2025.01.14.633023v1.2","unstructured":"Centers for Disease Control and Prevention. (2019). Antibiotic Resistance Threats in the United States, 2019."},{"issue":"4","key":"2026041021401634000_2025.01.14.633023v1.3","first-page":"277","article-title":"The Antibiotic Resistance Crisis: Part 1: Causes and Threats","volume":"40","year":"2015","journal-title":"Pharmacy and Therapeutics"},{"issue":"2","key":"2026041021401634000_2025.01.14.633023v1.4","first-page":"159","article-title":"A critical appraisal of 14CO2 blood culture systems","volume":"2","year":"1975","journal-title":"Journal of Clinical Microbiology"},{"issue":"10","key":"2026041021401634000_2025.01.14.633023v1.5","first-page":"2227","article-title":"BacT\/ALERT blood culture system","volume":"28","year":"1990","journal-title":"Journal of Clinical Microbiology"},{"issue":"10","key":"2026041021401634000_2025.01.14.633023v1.6","first-page":"4997","article-title":"BacT\/ALERT microbial detection system: clinical utility and patient management","volume":"41","year":"2003","journal-title":"Journal of Clinical Microbiology"},{"key":"2026041021401634000_2025.01.14.633023v1.7","doi-asserted-by":"publisher","DOI":"10.1016\/j.jpha.2015.11.005"},{"key":"2026041021401634000_2025.01.14.633023v1.8","doi-asserted-by":"publisher","DOI":"10.1093\/ajcp\/45.4_ts.493"},{"key":"2026041021401634000_2025.01.14.633023v1.9","doi-asserted-by":"publisher","DOI":"10.1016\/j.cmi.2015.02.005"},{"issue":"Suppl 1","key":"2026041021401634000_2025.01.14.633023v1.10","first-page":"S26","article-title":"Blood cultures: Key elements for best practices and future directions","volume":"202","year":"2010","journal-title":"Journal of Infectious Diseases"},{"issue":"3","key":"2026041021401634000_2025.01.14.633023v1.11","first-page":"e01836","article-title":"Systematic evaluation of the Accelerate Pheno system for susceptibility testing of Gram-negative bacteria isolated from blood cultures","volume":"9","year":"2021","journal-title":"Microbiology Spectrum"},{"issue":"19","key":"2026041021401634000_2025.01.14.633023v1.12","first-page":"9421","article-title":"Raman spectroscopy for rapid and quantitative detection of pathogens","volume":"88","year":"2016","journal-title":"Analytical Chemistry"},{"key":"2026041021401634000_2025.01.14.633023v1.13","doi-asserted-by":"publisher","DOI":"10.1373\/clinchem.2014.221770"},{"issue":"3","key":"2026041021401634000_2025.01.14.633023v1.14","first-page":"387","article-title":"Applications of MALDI-TOF mass spectrometry in clinical diagnostic microbiology","volume":"41","year":"2017","journal-title":"FEMS Microbiology Reviews"},{"issue":"12","key":"2026041021401634000_2025.01.14.633023v1.15","first-page":"3710","article-title":"MALDI-TOF MS in routine microbial identification: A study on the impact of reference libraries and user expertise","volume":"50","year":"2012","journal-title":"Journal of Clinical Microbiology"},{"issue":"11","key":"2026041021401634000_2025.01.14.633023v1.16","first-page":"e00230","article-title":"MALDI-TOF MS and its applications in clinical microbiology","volume":"58","year":"2020","journal-title":"Journal of Clinical Microbiology"},{"key":"2026041021401634000_2025.01.14.633023v1.17","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2022.858777"},{"key":"2026041021401634000_2025.01.14.633023v1.18","doi-asserted-by":"publisher","DOI":"10.1186\/1471-2180-12-1"},{"key":"2026041021401634000_2025.01.14.633023v1.19","doi-asserted-by":"publisher","DOI":"10.1186\/s13059-014-0572-2"},{"key":"2026041021401634000_2025.01.14.633023v1.20","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.3444"},{"key":"2026041021401634000_2025.01.14.633023v1.21","doi-asserted-by":"crossref","unstructured":"Alonso-Tarr\u00e9s, C. , Benjumea Moreno, C. , Navarro, F. , Habison, A. C. , Gonz\u00e0lez-Bertran, E. , Blanco, F. , \u2026 & Saker, J. (2024). Bacteriuria and phenotypic antimicrobial susceptibility testing in 45 min by point-of-care Sysmex PA-100 System: first clinical evaluation. European Journal of Clinical Microbiology & Infectious Diseases, 1\u201311.","DOI":"10.1007\/s10096-024-04862-3"},{"issue":"5","key":"2026041021401634000_2025.01.14.633023v1.22","first-page":"1461","article-title":"Use of a vacuum filtration system for improved bacterial detection in blood samples","volume":"55","year":"2017","journal-title":"Journal of Clinical Microbiology"},{"issue":"7","key":"2026041021401634000_2025.01.14.633023v1.23","first-page":"4271","article-title":"Microfluidic approaches for detection and characterization of pathogens","volume":"91","year":"2019","journal-title":"Analytical Chemistry"},{"issue":"4","key":"2026041021401634000_2025.01.14.633023v1.24","first-page":"760","article-title":"The use of microfluidics to study the microbiome","volume":"17","year":"2017","journal-title":"Lab on a Chip"},{"key":"2026041021401634000_2025.01.14.633023v1.25","doi-asserted-by":"publisher","DOI":"10.1039\/C8AN00307F"},{"key":"2026041021401634000_2025.01.14.633023v1.26","doi-asserted-by":"publisher","DOI":"10.1016\/j.chom.2024.01.012"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2025.01.14.633023","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,24]],"date-time":"2026-06-24T05:13:20Z","timestamp":1782278000000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2025.01.14.633023"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,1,16]]},"references-count":26,"URL":"https:\/\/doi.org\/10.1101\/2025.01.14.633023","relation":{"is-preprint-of":[{"id-type":"doi","id":"10.3389\/fmicb.2025.1637776","asserted-by":"subject"}]},"subject":[],"published":{"date-parts":[[2025,1,16]]},"subtype":"preprint"}}