Rapid antimicrobial susceptibility testing in patients with bacteraemia due to Enterobacterales: an implementation study

AIMS OF THE STUDY: The goal of this descriptive study was to assess the performance as well as the extent of the clinical impact of rapid automated antimicrobial susceptibility testing in patients with bacteraemia due to Enterobacterales. We also aimed to analyse how rapid automated antimicrobial susceptibility testing influences clinical decision-making. METHODS: This single-centre study conducted at the University Hospital of Zurich included data from all consecutive patients with Enterobacterales bacteraemia from November 2019 to October 2020. There was no control group. The primary outcome was the effect of rapid automated antimicrobial susceptibility testing on antibiotic therapy (no adjustment, escalation to a broader-spectrum antibiotic or de-escalation to a narrower-spectrum antibiotic). Rapid automated antimicrobial susceptibility testing results were further compared to susceptibility tests using European Committee on Antimicrobial Susceptibility Testing (EUCAST) standard methods and erroneous results were noted. Additionally, we investigated turnaround times for rapid automated antimicrobial susceptibility testing and routine diagnostic testing. RESULTS: We analysed 106 patients with 116 episodes of bacteraemia due to Enterobacterales, with Escherichia coli and Klebsiella pneumoniae being the most frequent isolates. Almost 8% of pathogens were multidrug resistant. Rapid automated antimicrobial susceptibility testing showed category agreement in 98.4% of all interpretable cases. A significant reduction of more than 20 h in turnaround times could be achieved with rapid automated antimicrobial susceptibility testing compared to the routine diagnostic workflow. In the majority of cases, rapid automated antimicrobial susceptibility testing had no effect, given that the empirical therapy was already correct or circumstances did not allow for de-escalation. In 38.8% of cases, antimicrobial therapy was adjusted, whereas eight cases were de-escalated based on rapid automated antimicrobial susceptibility testing alone. CONCLUSIONS: Rapid automated antimicrobial susceptibility testing may be a valuable and safe way to accelerate diagnosis. In particular, time to suitable therapy can be shortened in cases of incorrect therapy. However, physicians are reluctant to de-escalate antibiotic therapy based on rapid automated antimicrobial susceptibility testing alone, limiting its impact in everyday clinics. To further explore the potential of rapid automated antimicrobial susceptibility testing, a stringent/compulsory antibiotic stewardship programme would be a valuable next step

Fully automated disc diffusion for rapid antibiotic susceptibility test results: a proof-of-principle study

Background Antibiotic resistance poses a significant threat to patients suffering from infectious diseases. Early readings of antibiotic susceptibility test (AST) results could be of critical importance to ensure adequate treatment. Disc diffusion is a well-standardized, established and cost-efficient AST procedure; however, its use in the clinical laboratory is hampered by the many manual steps involved, and an incubation time of 16-18 h, which is required to achieve reliable test results. Methods We have evaluated a fully automated system for its potential for early reading of disc diffusion diameters after 6-12 h of incubation. We assessed availability of results, methodological precision, categorical agreement and interpretation errors as compared with an 18 h standard. In total, 1028 clinical strains (291 Escherichia coli , 272 Klebsiella pneumoniae , 176 Staphylococcus aureus and 289 Staphylococcus epidermidis ) were included in this study. Disc diffusion plates were streaked, incubated and imaged using the WASPLab TM automation system. Results and conclusions Our results demonstrate that: (i) early AST reading is possible for important pathogens; (ii) methodological precision is not hampered at early timepoints; and (iii) species-specific reading times must be selected. As inhibition zone diameters change over time and are phenotype/drug combination dependent, specific cut-offs and expert rules will be essential to ensure reliable interpretation and reporting of early susceptibility testing results

Rapid disc diffusion antibiotic susceptibility testing for Pseudomonas aeruginosa, Acinetobacter baumannii and Enterococcus spp

Background We investigated the feasibility of rapid disc diffusion antibiotic susceptibility testing (rAST) with reading of inhibition zones after 6 and/or 8 h of incubation for Enterococcus faecalis, Enterococcus faecium, Pseudomonas aeruginosa and Acinetobacter baumannii. In addition, we evaluated discrimination of resistant populations from the WT populations at early timepoints and the requirement for clinical breakpoint adaptations for proper interpretation of rAST data. Methods In total, 815 clinical strains [E. faecalis (n = 135), E. faecium (n = 227), P. aeruginosa (n = 295) and A. baumannii (n = 158)] were included in this study. Disc diffusion plates were streaked, incubated and imaged using the WASPLabTM automation system. WT populations and non-WT populations were defined using epidemiological cut-offs. Results and conclusions rAST at 6 and 8 h was possible for A. baumannii and enterococci with readability of inhibition zones >90%. Overall categorical agreement of rAST at 6 h with AST at 18 h was 97.2%, 97.4% and 95.3% for E. faecalis, E. faecium and A. baumannii, respectively. With few exceptions, major categorization error rates were <1% for A. baumannii, and vancomycin-resistant E. faecium were clearly separated from the WT at 6 h. For P. aeruginosa the average readability of inhibition zones was 68.9% at 8 h and we found an overall categorical agreement of 94.8%. Adaptations of clinical breakpoints and/or introduction of technical buffer zones, preferably based on aggregated population data from various epidemiological settings, are required for proper interpretation of rAST

Rapid Detection of PBP2a in Staphylococci from Shortly Incubated Subcultures of Positive Blood Cultures by an Immunochromatographic Assay

Staphylococcus aureus, as well as coagulase-negative staphylococci (CoNS), can cause a wide range of human infections both in nosocomial and community settings. Βeta-lactams are the antibiotics of choice for the treatment of bloodstream infections (BSI) caused by these microorganisms. Resistance to virtually all β-lactams (also referred to as methicillin resistance) primarily results from the production of an alternative penicillin-binding protein (PBP2a) encoded by the mecA gene. While β-lactams are still used as first-line therapy against BSI caused by S. aureus, BSI with CoNS are usually treated with vancomycin due to the high prevalence of methicillin resistance. Rapid detection of methicillin resistance is thus critical for continuation or adjustment of the empirical therapy and therewith to improve the clinical outcome of the patients. The revised version of the immunochromatographic assay PBP2a SA culture colony test (SACCT) is a rapid, inexpensive, and easy method that enables reliable detection of PBP2a in mecA-positive staphylococcal isolates after18 to 24 h of incubation. Here, we evaluated the diagnostic performance of the SACCT using primary subcultures of spiked blood cultures after short incubation (4 to 6 h) and established a modified procedure with an equal analytical performance to that of longer-grown cultures. With the proposed method the SACCT can be employed for PBP2a detection from shortly incubated subcultures of clinically relevant staphylococcal isolates, thereby allowing more rapid and effective management of BSI caused by these organisms. IMPORTANCE Antibiotic resistance poses a major threat to health and incurs high economic costs worldwide. Rapid detection of resistance mechanisms can contribute to improving patient care and preventing the dissemination of antimicrobial resistance. Here, we describe a rapid method to detect the most important beta-lactam resistance mechanism (the plasmid-encoded alternative transpeptidase PBP2a) in staphylococcal isolates causing BSI. We show that, using a modified procedure, PBP2a can be reliably detected from primary subcultures of spiked blood cultures after short incubation (4 to 6 h) with a rapid, inexpensive, and simple immunochromatographic test (SACCT). We provide an accurate, inexpensive, and rapid method to facilitate appropriate management and control of infections in patients suffering from invasive staphylococcal infections

Fully automated disc diffusion for rapid antibiotic susceptibility test results: a proof-of-principle study

Background Antibiotic resistance poses a significant threat to patients suffering from infectious diseases. Early readings of antibiotic susceptibility test (AST) results could be of critical importance to ensure adequate treatment. Disc diffusion is a well-standardized, established and cost-efficient AST procedure; however, its use in the clinical laboratory is hampered by the many manual steps involved, and an incubation time of 16-18 h, which is required to achieve reliable test results. Methods We have evaluated a fully automated system for its potential for early reading of disc diffusion diameters after 6-12 h of incubation. We assessed availability of results, methodological precision, categorical agreement and interpretation errors as compared with an 18 h standard. In total, 1028 clinical strains (291 Escherichia coli , 272 Klebsiella pneumoniae , 176 Staphylococcus aureus and 289 Staphylococcus epidermidis ) were included in this study. Disc diffusion plates were streaked, incubated and imaged using the WASPLab TM automation system. Results and conclusions Our results demonstrate that: (i) early AST reading is possible for important pathogens; (ii) methodological precision is not hampered at early timepoints; and (iii) species-specific reading times must be selected. As inhibition zone diameters change over time and are phenotype/drug combination dependent, specific cut-offs and expert rules will be essential to ensure reliable interpretation and reporting of early susceptibility testing results

Rapid detection of ESBL, carbapenemases, MRSA and other important resistance phenotypes within 6-8 h by automated disc diffusion antibiotic susceptibility testing

Abstract Background In principle, automated systems allow rapid reading of disc diffusion AST (rAST) within 6-8 h. Objectives This study analysed whether rAST can discriminate resistance phenotypes such as ESBL, carbapenemases and MRSA/methicillin-resistant Staphylococcus epidermidis from WT populations. We describe species-drug combinations that may require clinical breakpoint adaptions for early reading due to zone diameter changes during the incubation period. Methods In total, 1852 clinical strains [Escherichia coli (n = 475), Klebsiella pneumoniae (n = 375), Enterobacter cloacae (n = 301), Staphylococcus aureus (n = 407) and S. epidermidis (n = 294)] were included in this study comprising WT populations and important resistance phenotypes, e.g. ESBL, carbapenemases and MRSA. We assessed (i) separation of resistance phenotypes and WT populations after 6, 8 and 12 h as compared with the 18 h standard, and (ii) diameter changes of WT populations and associated putative epidemiological cut-offs during the incubation period. Disc diffusion plates were automatically streaked, incubated and imaged using the WASPLabTM system. Results and conclusions We demonstrated that important resistance phenotypes could reliably be separated from WT populations at early reading times for the most prevalent bacterial pathogens encountered in the clinical laboratory. Current AST expert rules and algorithms for identification of resistance mechanisms can readily be applied for rAST, e.g. EUCAST recommended rules for detection of ESBL, AmpC, carbapenemases and MRSA/methicillin-resistant S. epidermidis. However, several species-drug combinations may require clinical breakpoint adaptations when using rAST as the diameter, and hence the epidemiological cut-off, changes during the incubation period

Rapid disc diffusion antibiotic susceptibility testing for Pseudomonas aeruginosa, Acinetobacter baumannii and Enterococcus spp

Abstract Background We investigated the feasibility of rapid disc diffusion antibiotic susceptibility testing (rAST) with reading of inhibition zones after 6 and/or 8 h of incubation for Enterococcus faecalis, Enterococcus faecium, Pseudomonas aeruginosa and Acinetobacter baumannii. In addition, we evaluated discrimination of resistant populations from the WT populations at early timepoints and the requirement for clinical breakpoint adaptations for proper interpretation of rAST data. Methods In total, 815 clinical strains [E. faecalis (n = 135), E. faecium (n = 227), P. aeruginosa (n = 295) and A. baumannii (n = 158)] were included in this study. Disc diffusion plates were streaked, incubated and imaged using the WASPLabTM automation system. WT populations and non-WT populations were defined using epidemiological cut-offs. Results and conclusions rAST at 6 and 8 h was possible for A. baumannii and enterococci with readability of inhibition zones >90%. Overall categorical agreement of rAST at 6 h with AST at 18 h was 97.2%, 97.4% and 95.3% for E. faecalis, E. faecium and A. baumannii, respectively. With few exceptions, major categorization error rates were <1% for A. baumannii, and vancomycin-resistant E. faecium were clearly separated from the WT at 6 h. For P. aeruginosa the average readability of inhibition zones was 68.9% at 8 h and we found an overall categorical agreement of 94.8%. Adaptations of clinical breakpoints and/or introduction of technical buffer zones, preferably based on aggregated population data from various epidemiological settings, are required for proper interpretation of rAST

Tentative breakpoints and areas of technical uncertainty for early reading automated disc diffusion for Enterobacterales

BACKGROUND Disc diffusion is a reliable, accurate and cost-efficient procedure for antimicrobial susceptibility testing (AST) but requires long (18-24 h) incubation times. Reading of disc diffusion after short incubation times (6-8 h) by automated systems is feasible but should be categorized with time-adapted breakpoints to reduce errors. OBJECTIVES This study systematically compared early readings (6 and 8 h) of disc diffusion using an automated system with that of the standard 18 h EUCAST method. Time-adapted tentative breakpoints were proposed to discriminate susceptible from resistant isolates and areas of technical uncertainty were defined to minimize the risk of errors. METHODS A total of 1106 Enterobacterales isolates with a wide variety of resistance mechanisms and resistance profiles were included. All isolates were analysed for susceptibility to amoxicillin/clavulanic acid, ceftriaxone, cefepime, meropenem, ciprofloxacin and gentamicin using the automated WASPLabTM system. Part of the collection (515 isolates) was also analysed for susceptibility to an additional 10 antibiotics. RESULTS Separation between WT and non-WT populations was poorer at early incubation times than following standard incubation. Editing of rapid automated AST results after 6 and 8 h incubation with time-adapted breakpoints resulted in 84.0% and 88.5% interpretable results with assignment to the resistant or susceptible category. Major error and very major error rates for the 6 h readings were only 0.4% and 0.3%, virtually identical to those of 18 h AST reading. CONCLUSIONS Time-adapted clinical breakpoints in disc diffusion testing for Enterobacterales allow for accurate automated AST interpretation after shortened incubation times for a large number of antibiotics, with the additional possibility of subsequent confirmation after 18 h incubation