Rapid Phenotypic Antimicrobial Susceptibility Testing Research Articles

A-273 Rapid, Portable Invasive Fungal Infection Diagnostic and Candida Antimicrobial Susceptibility Testing

Abstract Background Traumatic wounds are associated with a higher risk of invasive fungal infections (IFI), which lead to a significant increase in amputation or death. Actionable treatment decisions are hampered because true infections can be difficult to differentiate from external wound contaminates, and tissue biopsy and quantification of fungal load is required to determine if treatment is necessary, which currently takes several days. GeneCapture is developing the CAPTURE 60-minute, portable infection identification platform to detect infectious levels of common fungi including the genera Aspergillus, Fusarium, and Scedosporium, the family Herpotrichiellaceae, and the order Mucorales. The company is also pursuing rapid identification and antimicrobial susceptibility testing for Candida species from multiple infection types. Methods This study seeks to determine the Limit of Detection (LoD), sensitivity and specificity of the CAPTURE ID assay for fungal pathogens. Since tissue biopsy samples must be used to assess the presence of IFI in wounds, GeneCapture has developed a manual process for the crude homogenization of muscle tissue to allow this matrix to be used as the starting sample for the ID assay. Mock samples were tested for inclusivity and exclusivity on a series of IFI ID panels against multiple strains of the included fungi, against fungi not yet specifically detected on the panel, and against bacteria that commonly infect wounds. While the LoD for yeasts may be determined in CFU/mL, there is no similar, easy-to-define metric for molds. GeneCapture has developed a quantitative PCR method to detect the single copy beta-tubulin gene to roughly correlate fungal dry mass and nuclei number to assay signal for molds cultured in Potato Dextrose Broth. Determining beta-tubulin copy numbers for extracts from mock samples then allowed detection of the progression of IFI over the course of infection. Lastly, antimicrobial susceptibility testing (AST) was performed on two species of Candida to validate utilizing GeneCapture’s rapid phenotypic AST method for fungal infections. Results To date, the CAPTURE ID panel has been tested against 27 included or excluded strains of fungi or bacteria with 97% sensitivity and 100% specificity. The current LoD for mold organisms varies from approximately 1,000 nuclei for Aspergillus species, to 100,000 nuclei for Fusarium species. Additional work to understand this range is underway. Lastly, we have correctly determined the susceptibility of Candida species using our novel approach within 4-6 hours. Conclusions GeneCapture’s ID and AST technology offer a unique solution to address the increasing prevalence of fungal infections. Our platform can identify the most common fungal components in a wound within 60 minutes of receiving a tissue biopsy sample. Candida infections occur across multiple matrices and rapidly identifying the species of Candida and then determining its phenotypic susceptibility profile will have a large impact on treating these difficult pathogens.

A comparative study of a rapid phenotypic antimicrobial susceptibility testing system directly from positive blood cultures to the disk diffusion and VITEK 2 methods

BackgroundSepsis is a life-threatening condition that impacts 49 million people annually and causes 11 million deaths worldwide. Surviving bloodstream infections (BSIs) depends on the rapid administration of effective antimicrobial treatment, underscoring a need for rapid antimicrobial susceptibility testing (AST). AimTo evaluate the performance of Quantamatrix's dRAST v2.5 system (Seoul, South Korea) for AST directly from positive blood cultures as compared to the Disk-Diffusion (DD) and VITEK 2 methods. MethodsThe study included 191 positive blood cultures from clinical samples and spiked blood culture bottles. Following Gram staining and species-level identification, AST was performed by VITEK 2 and standard DD methods using CLSI (2021) interpretation. ResultsdRAST demonstrated very good AST performance for a Gram-negative isolate, and good performance for Gram-positive isolates, meeting CLSI criteria for the acceptance of a new method. Antimicrobials that were not considered verified compared to VITEK 2 and DD were cefazolin, ceftazidime, meropenem, and trimethoprim/sulfamethoxazole for Gram-negatives and clindamycin, erythromycin, penicillin, and oxacillin for Gram-positives. dRAST ESBL detection results were strongly correlated with the ESBL phenotypes obtained with other methods. Additional resistance mechanisms were in concordance with traditional tests. ConclusionsdRAST demonstrated good AST performance, meeting CLSI criteria for most relevant antibiotics. dRAST was associated with a significant reduction in time-to-results, labor, and the subjectivity of result analyses, making it a valuable addition to efforts supporting the treatment of patients with bacteremia.AST (antimicrobial susceptibility test), blood culture, dRAST, rapid methods, sepsis, turnaround time (TAT).

Sedimentation field-flow fractionation for rapid phenotypic antimicrobial susceptibility testing: a pilot study.

The increase in antibiotic resistance is a major public health issue. The development of rapid antimicrobial susceptibility testing (AST) methods is becoming a priority to ensure early and appropriate antibiotic therapy. To evaluate sedimentation field-flow fractionation (SdFFF) as a method for performing AST in less than 3 h. SdFFF is based on the detection of early biophysical changes in bacteria, using a chromatographic-type technology. One hundred clinical Escherichia coli strains were studied. A calibrated bacterial suspension was incubated for 2 h at 37°C in the absence (untreated) or presence (treated) of five antibiotics used at EUCAST breakpoint concentrations. Bacterial suspensions were then injected into the SdFFF machine. For each E. coli isolate, retention times and elution profiles of antibiotic-treated bacteria were compared with retention times and elution profiles of untreated bacteria. Algorithms comparing retention times and elution profiles were used to determine if the strain was susceptible or resistant. Performance evaluation was done according to CLSI and the ISO standard 20776-2:2021 with broth microdilution used as the reference method. AST results from SdFFF were obtained in less than 3 h. SdFFF showed high categorical agreement (99.8%), sensitivity (99.5%) and specificity (100.0%) with broth microdilution. Results for each antimicrobial were also in agreement with the ISO 20776-2 recommendations, with sensitivity and specificity of ≥95.0%. This study showed that SdFFF can be used as a rapid, accurate and reliable phenotypic AST method with a turnaround time of less than 3 h.

Correction for Göransson et al., "Performance of a System for Rapid Phenotypic Antimicrobial Susceptibility Testing of Gram-Negative Bacteria Directly From Positive Blood Culture Bottles".

Correction for Göransson et al., "Performance of a System for Rapid Phenotypic Antimicrobial Susceptibility Testing of Gram-Negative Bacteria Directly From Positive Blood Culture Bottles".

Evaluation of the LifeScale system for rapid phenotypic antimicrobial susceptibility testing from positive blood cultures

Abstract Objective The spread of antimicrobial resistance is a growing threat to public health, thus there is a critical need for prompt and reliable antimicrobial susceptibility testing (AST). Rapid AST may improve patient outcomes by allowing for quicker start of optimal therapy and reducing hospital length of stay, therefore decreasing overall healthcare costs. However, rapid results may not improve patient care if they are not considered equivalent to standard AST. Several platforms are under development that can provide rapid AST results; the objective of this work is to assess the performance of the new LifeScale rapid AST platform (Affinity Biosensors) that uses resonant mass measurements for calculation of antimicrobial susceptibility results. Methods Positive BACTEC blood-cultures collected from patients were included if the gram stain revealed only Gram-negative rods. Standard of care (SOC) AST results, as determined by a combination of BD Phoenix automated, Sensititre broth microdilution, and/or standard disk diffusion methods, were compared to results from direct positive blood culture testing on the LifeScale system. Minimum inhibitory concentrations (MICs) and categorical interpretations based on CLSI and/or FDA AST breakpoints were compared by microorganism and drug to calculate essential and categorical agreement for five species with pending claims on the LifeScale system: Escherichia coli (N=33), Klebsiella pneumoniae (N=10), K. oxytoca (N=5), Pseudomonas aeruginosa (N=11) and Acinetobacter sp. (N=1); tested against 13 relevant antibiotics: Amikacin, ampicillin, aztreonam, cefazolin, cefepime, ceftazidime, ceftazidime/avibactam ertapenem, gentamicin, levofloxacin, meropenem, piperacillin/tazobactam, and trimethoprim/sulfamethoxazole. Results LifeScale had 95.3% essential agreement with the SOC results, and a 93.1% categorical agreement based on CLSI M52 criteria. Of the 665 organism-drug combinations evaluated, agreement was high, with only 1 very major discrepancy, 8 major discrepancies and 37 minor discrepancies, of which 22 (47.8%) were within ±1 doubling dilution. Among these discrepancies, LifeScale MICs averaged 1 dilution higher than SOC MICs, which may reduce the risk of false-susceptible interpretations. Conclusion s: The accuracy and reliability of results from rapid AST is of utmost importance. The LifeScale technology is unique among AST platforms as it measures bacterial cell mass and counts to determine MICs for up to 14 antibiotics in ~4 h. These results suggest this platform is able to deliver rapid, actionable results to guide antimicrobial therapy for patients with bacteremia.

Rapid phenotypic antimicrobial susceptibility testing of Gram-negative rods directly from positive blood cultures using the novel Q-linea ASTar system.

Adequate and timely antibiotic therapy is crucial for the treatment of sepsis. Innovative systems, like the Q-linea ASTar, have been developed to perform rapid antimicrobial susceptibility testing (AST) directly from positive blood cultures (BCs). We conducted a prospective study to evaluate ASTar under real-life conditions with a focus on time-to-result and impact on antimicrobial therapy. Over 2 months, all positive BCs that showed Gram-negative rods upon microscopy were tested with the ASTar and our standard procedure (VITEK 2 from short-term culture). Additionally, we included multidrug-resistant Gram-negative bacteria from our archive. Both methods were compared to broth microdilution. In total, 78 bacterial strains (51 prospective and 27 archived) were tested. ASTar covered 94% of the species encountered. The categorical and essential agreement was 95.6% and 90.7%, respectively. ASTar caused 2.4% minor, 2.0% major, and 2.4% very major errors. The categorical agreement was similar to standard procedure. The average time between BC sampling and the availability of the antibiogram for the attending physician was 28 h 49 min for ASTar and 44 h 18 min for standard procedure. ASTar correctly identified all patients who required an escalation of antimicrobial therapy and 75% of those who were eligible for de-escalation. In conclusion, ASTar provided reliable AST results and significantly shortened the time to obtain an antibiogram. However, the percentage of patients that will profit from ASTar in a low-resistance setting is limited, and it is currently unclear if a change of therapy 29 h after BC sampling will have a significant impact on the patient's prognosis.

Rapid Phenotypic Antimicrobial Susceptibility Testing Using a Coulter Counter and Proliferation Rate Discrepancy.

The rapid determination of antimicrobial susceptibility and evidence-based antimicrobial prescription is necessary to combat widespread antimicrobial resistance and promote effectively treatment for bacterial infections. This study developed a rapid phenotypic antimicrobial susceptibility determination method competent for seamless clinical implementation. A laboratory-friendly Coulter counter-based antimicrobial susceptibility testing (CAST) was developed and integrated with bacterial incubation, population growth monitoring, and result analysis to quantitatively detect differences in bacterial growth between resistant and susceptible strains following a 2 h exposure to antimicrobial agents. The distinct proliferation rates of the different strains enabled the rapid determination of their antimicrobial susceptibility phenotypes. We evaluated the performance efficacy of CAST for 74 clinically isolated Enterobacteriaceae subjected to 15 antimicrobials. The results were consistent with those obtained via the 24 h broth microdilution method, showing 90.18% absolute categorical agreement.

Performance of a System for Rapid Phenotypic Antimicrobial Susceptibility Testing of Gram-Negative Bacteria Directly from Positive Blood Culture Bottles.

The rapid administration of optimal antimicrobial treatment is paramount for the treatment of bloodstream infections (BSIs), and rapid antimicrobial susceptibility testing (AST) results are essential. Q-linea has developed the ASTar system, a rapid phenotypic AST device. Here, we report the performance of the ASTar BC G- (Gram-negative) kit when assessed according to the ISO 20776-2:2007 standard for performance evaluation of in vitro diagnostic AST devices. The evaluated ASTar BC G- kit uses a broad panel of 23 antimicrobials for the treatment of BSIs caused by Gram-negative fastidious and nonfastidious bacteria across a range of 6 to 14 2-fold dilutions, including cefoxitin as a screening agent for AmpC-producing Enterobacterales. The ASTar system processes blood culture samples to generate data on MICs and susceptible, intermediate, or resistant (SIR) category. The automated protocol includes concentration determination and concentration adjustment to enable a controlled inoculum, followed by broth microdilution (BMD) and microscopy performed continuously to generate MIC values within approximately 6 h once the test is run on the ASTar system. The performance of the ASTar system was assessed against the ISO 20776-2:2007 standard BMD reference method. Testing was performed across three sites, with results from 412 contrived blood cultures and 74 fresh clinical blood cultures. The ASTar system was also tested for reproducibility, with triplicate testing of 11 strains. The accuracy study comprised 8,650 data points of bacterium-antimicrobial tests. The ASTar system demonstrated an overall essential agreement (EA) of 95.8% (8,283/8,650) and a categorical agreement (CA) of 97.6% (8,433/8,639) compared to the reference BMD method. The overall rate of major discrepancies (MDs) was 0.9% (62/6,845), and that of very major discrepancies (VMDs) was 2.4% (30/1,239). This study shows that the ASTar system delivers reproducible results with overall EA and CA of >95%.

Under-oil open microfluidic systems for rapid phenotypic antimicrobial susceptibility testing.

Antimicrobial susceptibility testing (AST) remains the cornerstone of effective antimicrobial selection and optimization in patients. Despite recent advances in rapid pathogen identification and resistance marker detection with molecular diagnostics (e.g., qPCR, MALDI-TOF MS), phenotypic (i.e., microbial culture-based) AST methods - the gold standard in hospitals/clinics - remain relatively unchanged over the last few decades. Microfluidics-based phenotypic AST has been growing fast in recent years, aiming for rapid (i.e., turnaround time <8 h), high-throughput, and automated species identification, resistance detection, and antibiotics screening. In this pilot study, we describe the application of a multi-liquid-phase open microfluidic system, named under-oil open microfluidic systems (UOMS), to achieve a rapid phenotypic AST. UOMS provides an open microfluidics-based solution for rapid phenotypic AST (UOMS-AST) by implementing and recording a pathogen's antimicrobial activity in micro-volume testing units under an oil overlay. UOMS-AST allows free physical access (e.g., by standard pipetting) to the system and label-free, single-cell resolution optical access. UOMS-AST can accurately and rapidly determine antimicrobial activities [including susceptibility/resistance breakpoint and minimum inhibitory concentration (MIC)] from nominal sample/bacterial cells in a system aligned with clinical laboratory standards where open systems and optical microscopy are predominantly adopted. Further, we combine UOMS-AST with a cloud lab data analytic technique for real-time image analysis and report generation to provide a rapid (<4 h) sample-to-report turnaround time, shedding light on its utility as a versatile (e.g., low-resource setting and manual laboratory operation, or high-throughput automated system) phenotypic AST platform for hospital/clinic use.

Combinatorial screening SlipChip for rapid phenotypic antimicrobial susceptibility testing.

Antimicrobial resistance (AMR) by bacteria is a serious global threat, and a rapid, high-throughput, and easy-to-use phenotypic antimicrobial susceptibility testing (AST) method is essential for making timely treatment decisions and controlling the spread of antibiotic resistant micro-organisms. Traditional culture-based methods are time-consuming, and their capability to screen against a large number of different conditions is limited; meanwhile genotypic based methods, including sequencing and PCR based methods, are constrained by rarely identified resistance genes and complicated resistance mechanisms. Here, a combinatorial-screening SlipChip (cs-SlipChip) containing 192 nanoliter-sized compartments is developed which can perform high-throughput phenotypic AST within three hours by monitoring the bacterial growth within nanoliter-sized droplets with bright-field imaging and analyzing the changes in bacterial number and morphology. The minimum inhibitory concentration (MIC) of Escherichia coli ATCC 25922 against four antibiotics (ampicillin, ciprofloxacin, ceftazidime, and nitrofurantoin) can be measured in one chip within 3 hours. Furthermore, five antibiotic-resistant E. coli strains were isolated from patients diagnosed with urinary tract infections (UTIs), and an individual isolate was tested using four antibiotics and eleven antibiotic combinations simultaneously with three different concentrations of each. The results from the cs-SlipChip agree with those of a VITEK 2 automated system. This cs-SlipChip provides a practical high-throughput and rapid phenotypic method for AST and can also be used to screen different chemicals and antibiotic combinations for the treatment of multiple antibiotic-resistant bacteria.

Antimicrobial susceptibility testing for Gram positive cocci towards vancomycin using scanning electron microscopy.

The rapid detection of resistant bacteria has become a challenge for microbiologists worldwide. Numerous pathogens that cause nosocomial infections are still being treated empirically and have developed resistance mechanisms against key antibiotics. Thus, one of the challenges for researchers has been to develop rapid antimicrobial susceptibility testing (AST) to detect resistant isolates, ensuring better antibiotic stewardship. In this study, we established a proof-of-concept for a new strategy of phenotypic AST on Gram-positive cocci towards vancomycin using scanning electron microscopy (SEM). Our study evaluated the profiling of Enterococcus faecalis, Enterococcus faecium and Staphylococcus aureus after brief incubation with vancomycin. Sixteen isolates were analysed aiming to detect ultrastructural modifications at set timepoints, comparing bacteria with and without vancomycin. After optimising slides preparation and micrographs acquisition, two analytical strategies were used. The high magnification micrographs served to analyse the division of cocci based on the ratio of septa, along with the bacterial size. Susceptible strains with vancomycin showed a reduced septa percentage and the average surface area was consequently double that of the controls. The resistant bacteria revealed multiple septa occurring at advanced timepoints. Low magnification micrographs made it possible to quantify the pixels at different timepoints, confirming the profiling of cocci towards vancomycin. This new phenotypic AST strategy proved to be a promising tool to discriminate between resistant and susceptible cocci within an hour of contact with vancomycin. The analysis strategies applied here would potentially allow the creation of artificial intelligence algorithms for septa detection and bacterial quantification, subsequently creating a rapid automated SEM-AST assay.

313. Using a Clinical Decision Prediction Tool to Improve Empirical Antimicrobial Therapy in Ceftriaxone-Resistant Enterobacterales Bloodstream Infections

BackgroundThere are several clinical tools for prediction of antimicrobial resistance. However, their utility in management of bloodstream infections (BSI) due to resistant bacteria remains unclear. This quasi-experimental cohort examined the impact of utilization of the extended-spectrum beta-lactamase (ESBL) prediction score on time to initiation of appropriate antimicrobial therapy (AAT) in BSI due to ceftriaxone-resistant (CRO-R) Enterobacterales.MethodsAdults with first episodes of monomicrobial BSI due to CRO-R Enterobacterales at Prisma Health-Midlands Hospitals in South Carolina from January 2010 to December 2017 were included. Antimicrobial stewardship intervention was implemented in January 2014 consisting of real-time alerts for positive blood cultures and rapid diagnostics for identification of bloodstream isolates. The ESBL prediction score was used to stratify risk of resistance prior to conventional antimicrobial susceptibility results. Student’s t-test was used to compare mean time to AAT before and after intervention.ResultsAmong 92 patients with BSI due to CRO-R Enterobacterales, median age was 66 years, 52 (57%) were men, 68 (74%) had community-onset BSI, and 52 (57%) had urinary source of infection. Escherichia coli 52 (57%) and Klebsiella species 27 (29%) were the most common bloodstream isolates and the majority 63 (68%) produced ESBLs. There were no significant differences in demographics or clinical characteristics of patients before (n=45) and after (n=47) antimicrobial stewardship intervention. Mean time to AAT was 3.2 days prior and 1.7 days after utilization of ESBL prediction score (p=0.021). Results were consistent among ESBL-producing Enterobacterales (time to AAT 2.8 before and 1.8 days after intervention, p=0.025).ConclusionUtilization of the ESBL prediction score as part of a real-time antimicrobial stewardship intervention significantly reduced time to AAT in BSI due to CRO-R and ESBL-producing Enterobacterales. The ESBL prediction score represents a viable tool to improve antimicrobial management in clinical settings that lack access to multiplex PCRs for detection of antimicrobial resistance genes and novel methods for rapid phenotypic antimicrobial susceptibility testing.DisclosuresJulie Ann Justo, PharmD, MS, BCPS-AQ ID, bioMerieux (Speaker’s Bureau)TRC Healthcare (Speaker’s Bureau)

Advances in rapid diagnostics for bloodstream infections

Septicemia from bloodstream infections (BSI) is the second largest cause of inpatient mortality and the single most expensive condition for US hospitals to manage. There has been an explosive development of commercial diagnostic systems to accelerate the identification and antimicrobial susceptibility testing (AST) of causative pathogens. Despite adoption of advanced technologies like matrix-assisted laser desorption imaging–time-of-flight mass spectrometry and multiplex polymerase chain reaction for rapid identification, clinical impact has been variable, in part due to the persistent need for conventional AST as well as prescriber understanding of these rapidly evolving platforms. Newer technologies are expanding on rapid detection of genotypic determinants of resistance, but only recently has rapid phenotypic AST been available. Yet, improved outcomes with rapid diagnostic platforms are still most evident in conjunction with active antimicrobial stewardship. This review will outline key advancements in rapid diagnostics for BSI and the role of antimicrobial stewardship in this new era.

Enhanced antimicrobial stewardship based on rapid phenotypic antimicrobial susceptibility testing for bacteraemia in patients with haematological malignancies: a randomized controlled trial

ObjectivesRecently, rapid phenotypic antimicrobial susceptibility testing (AST) based on microscopic imaging analysis has been developed. The aim of this study was to determine whether implementation of antimicrobial stewardship programmes (ASP) based on rapid phenotypic AST can increase the proportion of patients with haematological malignancies who receive optimal targeted antibiotics during early periods of bacteraemia. MethodsThis randomized controlled trial enrolled patients with haematological malignancies and at least one positive blood culture. Patients were randomly assigned 1:1 to conventional (n = 60) or rapid phenotypic (n = 56) AST. The primary outcome was the proportion of patients receiving optimal targeted antibiotics 72 hr after blood collection for culture. ResultsThe percentage receiving optimal targeted antibiotics at 72 hr was significantly higher in the rapid phenotypic AST group (45/56, 80.4%) than in conventional AST group (34/60, 56.7%) (relative risk (RR) 1.42, 95% confidence interval (CI) 1.09–1.83). The percentage receiving unnecessary broad-spectrum antibiotics at 72 hr was significantly lower (7/26, 12.5% vs 18/60, 30.0%; RR 0.42, 95% CI 0.19–0.92) and the mean time to optimal targeted antibiotic treatment was significantly shorter (38.1, standard deviation (SD) 38.2 vs 72.8, SD 93.0 hr; p < 0.001) in the rapid phenotypic AST group. The mean time from blood collection to the AST result was significantly shorter in the rapid phenotypic AST group (48.3, SD 17.6 vs 83.1, SD 22.2 hr). DiscussionASP based on rapid phenotypic AST can rapidly optimize antibiotic treatment for bacteraemia in patients with haematological malignancy. Rapid phenotypic AST can improve antimicrobial stewardship in immunocompromised patients.

Microplate-based surface area assay for rapid phenotypic antibiotic susceptibility testing

Rapid delivery of proper antibiotic therapies to infectious disease patients is essential for improving patient outcomes, decreasing hospital lengths-of-stay, and combating the antibiotic resistance epidemic. Antibiotic stewardship programs are designed to address these issues by coordinating hospital efforts to rapidly deliver the most effective antibiotics for each patient, which requires bacterial identification and antimicrobial susceptibility testing (AST). Despite the clinical need for fast susceptibility testing over a wide range of antibiotics, conventional phenotypic AST requires overnight incubations, and new rapid phenotypic AST platforms restrict the number of antibiotics tested for each patient. Here, we introduce a novel approach to AST based on signal amplification of bacterial surfaces that enables phenotypic AST within 5 hours for non-fastidious bacteria. By binding bacterial surfaces, this novel method allows more accurate measurements of bacterial replication in instances where organisms filament or swell in response to antibiotic exposure. Further, as an endpoint assay performed on standard microplates, this method should enable parallel testing of more antibiotics than is currently possible with available automated systems. This technology has the potential to revolutionize clinical practice by providing rapid and accurate phenotypic AST data for virtually all available antibiotics in a single test.

1513. A New Method for Rapid Phenotypic Antimicrobial Susceptibility Testing Directly from Patient Samples

BackgroundLife-threatening syndromic hospital infections including sepsis, ventilator acquired pneumonia, catheter-associated urinary tract infection (CAUTI), and surgical site infections are often caused by multidrug-resistant pathogens. Implementing the targeted narrow-spectrum antimicrobial therapy as rapidly as possible at the onset of infection is critical for lowering morbidity and mortality for these infections. We present the new MultiPath technology for rapid syndromic infection detection, pathogen identification, and phenotypic antimicrobial susceptibility testing (AST). Our feasibility data demonstrate the technology’s potential application as a rapid CAUTI diagnostic.MethodsThe MultiPath technology detects and counts cells in a 30-minute assay using nonmagnified digital imaging. For identification, target pathogen cells are labeled using fluorescent in situ hybridization (FISH) with rRNA-specific probes, tagged with magnetic nanoparticles, deposited on a surface, imaged, and quantified. For AST, samples are mixed with growth medium, incubated for 4 hours in the presence of serial dilutions of antibiotics, FISH-labeled, magnetically selected, and quantified by digital imaging. The MultiPath assays use a dye-cushion layer to optically sequester the sample and unbound fluorescent probes from the imaging surface, eliminating the need for sample preparation and wash steps.ResultsThe MultiPath ID method specifically detected a range of common CAUTI pathogens including E. coli, K. pneumonia, E. faecium, E. faecalis, and P. aeruginosa. The limit of detection for E. coli was 27 CFU in a 100 µL assay in 10% urine. We present data demonstrating target inclusivity, specificity, and dynamic range. Our AST feasibility study results show excellent correlation with the broth micro dilution reference test for 5 antibiotics. Variable inoculum levels had little impact on MICs in the study.ConclusionThe data presented demonstrate the potential of the rapid ID/AST technology to achieve excellent analytical and clinical performance. This, combined with the method’s simplicity, robustness to sample matrix, and ease-of-use may make the method valuable for rapid syndromic infection diagnostics.Disclosures All authors: No reported disclosures.

2076. Expanded Antibiotic Menu Demonstration for Novel Rapid Phenotypic Antimicrobial Susceptibility Testing Platform

BackgroundFast transitions to targeted therapies for infectious disease patients are paramount for optimal patient care and antibiotic stewardship. A next-generation phenotypic antimicrobial susceptibility test (AST) system that provides rapid results for broad menus of >30 antibiotics per patient sample is required. SeLux has developed a rapid, phenotypic AST platform that utilizes standard 384-well microplates. These consumables provide sufficient wells for simultaneous testing of newly approved antibiotics and broad selections of conventional antibiotics. Here, we demonstrate the platform’s ability to produce fast, accurate results with newly approved and not-yet-approved antibiotics.MethodsThe core of SeLux’s technology is a novel assay for bacterial surface area, which enables delineation of truly resistant bacteria from organisms that filament or swell in antibiotic concentrations above the MIC. AST was performed with the SeLux platform and compared with the CLSI broth microdilution reference method. Testing of 20 representative conventional antibiotics was performed on 1,191 isolates, including 323 FDA-CDC “challenge” strains and comprising 20 species of nonfastidious bacteria. Testing of newly developed antibiotics, generous gifts from the manufacturers, was performed with ~20 to 50 isolates with representative MICs throughout the dilution series and encompassing the breakpoint region.ResultsTesting of conventional antibiotics showed essential agreements (EA) and categorical agreements (CA) ≥90% with the CLSI reference method for all combinations tested (Figures 1 and 2). The platform returned results within 6.5 hours for >98% of the isolates tested to date. The SeLux platform’s EA was ≥90% for all newly developed antibiotics tested to date (Figure 3). For newly approved antibiotics, CAs were similarly ≥90% with no very major errors (Vmj).Figure 1Figure 2Figure 3ConclusionThe SeLux platform’s compatibility with 384-well microplates should transform the rate with which newly approved antibiotics gain use. By speeding the reporting of AST results, SeLux’s platform will further enable hospitals to simultaneously improve patient care, decrease lengths-of-stay, and meet antibiotic stewardship goals.Disclosures E. Stern, SeLux Diagnostics: Board Member, Employee and Shareholder, Salary. A. Vacic, SeLux Diagnostics: Employee and Shareholder, Salary. K. Flentie, SeLux Diagnostics: Employee and Shareholder, Salary. B. Spears, SeLux Diagnostics: Employee and Shareholder, Salary. N. Purmort, SeLux Diagnostics: Employee and Shareholder, Salary. F. Giok, SeLux Diagnostics: Employee and Shareholder, Salary. K. DaPonte, SeLux Diagnostics: Employee and Shareholder, Salary. S. Scott, SeLux Diagnostics: Employee and Shareholder, Salary. D. Puff, SeLux Diagnostics: Employee and Shareholder, Salary. F. Floyd Jr., SeLux Diagnostics: Employee and Shareholder, Salary. Z. Zhang, SeLux Diagnostics: Employee and Shareholder, Salary. P. Reilly, SeLux Diagnostics: Employee, Salary. J. Liu, SeLux Diagnostics: Employee, Salary. E. Viveiros, SeLux Diagnostics: Employee, Salary. N. Phelan, SeLux Diagnostics: Employee, Salary. C. Krebill, SeLux Diagnostics: Employee, Salary. A. Flyer, SeLux Diagnostics: Consultant, Scientific Advisor and Shareholder, Consulting fee. D. Smalley, SeLux Diagnostics: Scientific Advisor and Shareholder, Consulting fee. D. C. Hooper, SeLux Diagnostics: Scientific Advisor, Consulting fee. M. J. Ferraro, SeLux Diagnostics: Scientific Advisor and Shareholder, Consulting fee.

The Slow March toward Rapid Phenotypic Antimicrobial Susceptibility Testing: Are We There Yet?

Antimicrobial susceptibility testing (AST) provides critical information for the management of patients with infections. The gold standard methods for assessing organism susceptibility are still based on growth and require incubation over relatively long periods of time. Until now, little progress has been made in developing rapid, growth-based, phenotypic AST systems. This commentary puts the recently FDA-cleared Accelerate PhenoTest (P. Pancholi et al., J Clin Microbiol 56:e01329-17, 2018, https://doi.org/10.1128/JCM.01329-17) in context by providing a historical perspective on attempts to accelerate phenotypic susceptibility results. In addition, some promising new innovations that promise to shorten the turnaround time for phenotypic AST will be briefly reviewed.

Rapid phenotypic antimicrobial susceptibility testing using nanoliter arrays

Antibiotic resistance is a major global health concern that requires action across all sectors of society. In particular, to allow conservative and effective use of antibiotics clinical settings require better diagnostic tools that provide rapid determination of antimicrobial susceptibility. We present a method for rapid and scalable antimicrobial susceptibility testing using stationary nanoliter droplet arrays that is capable of delivering results in approximately half the time of conventional methods, allowing its results to be used the same working day. In addition, we present an algorithm for automated data analysis and a multiplexing system promoting practicality and translatability for clinical settings. We test the efficacy of our approach on numerous clinical isolates and demonstrate a 2-d reduction in diagnostic time when testing bacteria isolated directly from urine samples.