New Technologies for the Rapid Identification of Drug-Resistant Bacteria Alexis F. Sauer-Budge * * Exponent, Inc. Polymer Science and Materials Chemistry Practice 9 Strathmore Rd. Natick, MA 01773, asauerbudge@exponent.com ABSTRACT The extensive use of antibiotics to treat infections, without first specifically identifying the underlying cause, has resulted in a strong natural selection for antibiotic resistant organisms. Conservative estimates from the CDC indicate that more than two million people are sickened by antibiotic-resistant infections in the US each year, leading to more than 23,000 deaths. Because it takes days by traditional methods to identify the etiological organism and its drug- susceptibility profile, up to 50% of all antibiotics prescribed are not needed or are not optimally effective. This significant threat to human health is starting to be addressed technologically through the development and commercialization of novel rapid diagnostics. The approaches include phenotypic and genotypic methodologies that target a variety of direct and indirect biomarkers, including protein, lipid, and cell-based phenotypic assays as well as molecular diagnostic approaches. One of the key area of innovation is the development and integration of appropriate and easy-to-use sample preparation that is tailored to the target analyte, the specimen, and the diagnostic modality. A survey of key innovations in the field will be presented. Keywords: microfluidics, antibiotic susceptibility, phenotypic testing, diagnostics 1 INTRODUCTION Antimicrobial resistance (AMR) is a serious and growing problem that impacts the world’s healthcare, veterinary and agricultural industries. In the United States alone, antibiotic- resistant bacteria infect more than 2 million people and cause over 23,000 deaths.[1] The continued spread of AMR is a multi-factorial problem, including inappropriate prescription practices, lack of antimicrobial stewardship programs, insufficient patient education, non-human use of antimicrobials and limitations of existing diagnostics.[2] The criticality of administering appropriate antibiotics has been studied in the case of septic shock, where every hour of delay is associated with an average decrease in survival of 7.6%.[3] However, the standard methods for antibiotic susceptibility determination require multiple days and therefore physicians are left with no choice but to prescribe antibiotics in the absence of this information, which further contributes to proliferation of AMR. To address this threat, the U.S. government developed a five-year National Action Plan in 2015 for Combating Antibiotic Resistant Bacteria, which specifically included a goal to “Advance development and use of rapid and innovative diagnostic tests for identification and characterization of resistant bacteria”.[4] Several other organizations have also recognized the need for improved antibiotic susceptibility testing (AST), and introduced prize- based incentive programs including the Longitude Prize (UK) [5] and the Point-of-Care Diagnostic Challenge Prize (USA) [6]. These incentives intended to hasten the development of novel rapid diagnostics, although significant challenges have yet to be overcome to meet the clinical need. 2 STATE-OF-THE-ART DIAGNOSTICS Traditionally, diagnosis and determination of antibiotic susceptibility has been based on culture methods. Typically, a specimen is taken from the patient and cultured to grow the pathogens to a detectable level and to isolate pure colonies. Once isolated, a second culture step (e.g. disk diffusion, broth dilution or E-test)[7], develops an overall profile of pathogen antibiotic susceptibility, known as an antibiogram. Generation of the antibiogram typically takes days, and, while the methods are accurate and inexpensive, they require strict adherence to standardized protocols.[8] Commercial automated systems are widely used in larger clinical labs (e.g. BD Phoenix, Beckman Coulter MicroScan), which decrease the operator time and overall turn-around-time, but also increase the cost of the analysis. Recently, several new rapid diagnostics have entered the marketplace and are gaining traction in clinical microbiology laboratories. In 2007, the Cepheid Xpert MRSA test became one of the first rapid fully automated diagnostics for detection of antibiotic resistant organisms cleared by the FDA.[9] The test is PCR-based and detects both bacteria- specific and methicillin-resistance genetic markers. While useful for screening for outbreaks, the utility of this test is quite limited as it only detects a single bacterial species and resistance marker. More recent entries to the market, such as the Biomerieux BioFire® FilmArray® and Accelerate Pheno, have broader panels. The BioFire® FilmArray®, based on a multiplexed PCR technology, has FDA cleared panels for respiratory (RP2, 2017), blood culture ID (2016), gastrointestinal (2016), meningitis/encephalitis (2016), and pneumonia (2018) pathogens and resistance markers.[10, 11] While these molecular diagnostic panels are useful, they remain more limited than phenotypic tests because genetic TechConnect Briefs 2019, TechConnect.org, ISBN 978-0-9988782-8-7 310