Adoption of the updated CLSI fluoroquinolone breakpoints for Gram-negative bacteria in microbiology laboratories
Authors: Maroun M. Sfeir
Journal: Clinical Microbiology and Infection (2020)

In January 2019, the Clinical and Laboratory Standards Institute (CLSI) lowered the clinical breakpoints of fluoroquinolones (FQ) for Enterobacteriaceae other than Salmonella spp. and Pseudomonas aeruginosa after an increase in minimal inhibitory concentrations (MICs) to FQ among Gram-negative bacteria (GNB), application of sophisticated FQ pharmacokinetics and pharmacodynamics technology and the scarcity of clinical data supporting the original FQ breakpoints [[1]Clinical and Laboratory Standards Institute (CLSI)Performance standards for antimicrobial susceptibility testing.29th ed. CLSI supplement M100. CLSI, Wayne, PA2019Google Scholar,[2]Humphries R.M. Hindler J.A. Shaffer K. Campeau S. Evaluation of ciprofloxacin and levofloxacin disk and Etest using the 2019 Enterobacteriaceae CLSI breakpoints.J Clin Microbiol. 2018; 56: 1797-1818Google Scholar]. The revised CLSI breakpoints for Enterobacteriaceae and P. aeruginosa are ≤0.25 and ≤ 0.5 mg/L for ciprofloxacin and ≤0.5 and ≤ 1 mg/L for levofloxacin respectively (Table 1) [[1]Clinical and Laboratory Standards Institute (CLSI)Performance standards for antimicrobial susceptibility testing.29th ed. CLSI supplement M100. CLSI, Wayne, PA2019Google Scholar]. Those new FQ breakpoints are more consistent with the international recommendations from the European Committee on Antimicrobial Susceptibility Testing (AST) [[3]European Committee on Antimicrobial Susceptibility Testing (EUCAST)Clinical breakpoints for bacteria.https://eucast.org/ast_of_bacteria/previous_versions_of_documents/Date: 2019Google Scholar]. The updated CLSI standard was recognized by the US Food and Drug Administration (FDA) on 6 June 2019 [[4]US Food and Drug Administration (FDA)FDA rationale for recognition decision: levofloxacin. Content current as of June 2019.https://www.fda.gov/drugs/development-resources/fda-rationale-recognition-decision-levofloxacinGoogle Scholar]. Similarly, the CLSI breakpoints for Salmonella species to ciprofloxacin and levofloxacin were lowered for susceptible isolates to ≤0.06 and ≤ 0.12 mg/L in 2012 and 2013 respectively after the emergence of plasmid-mediated quinolone resistance called qnrA that was missed by the old FQ MIC breakpoints and has led to clinical failure or delayed response in FQ-treated patients (Table 1) [[1]Clinical and Laboratory Standards Institute (CLSI)Performance standards for antimicrobial susceptibility testing.29th ed. CLSI supplement M100. CLSI, Wayne, PA2019Google Scholar,[4]US Food and Drug Administration (FDA)FDA rationale for recognition decision: levofloxacin. Content current as of June 2019.https://www.fda.gov/drugs/development-resources/fda-rationale-recognition-decision-levofloxacinGoogle Scholar]. While most intestinal Salmonella infections are self-limited and do not need antibiotics or routine susceptibility testing, Salmonella Typhi and Salmonella Paratyphi AC isolated from extraintestinal and intestinal sources as well as extraintestinal Salmonella species infections do require susceptibility testing. Little is known about the number of hospitals that adopted updated FQ breakpoints for GNB.Table 1Historical and updated CLSI ciprofloxacin and levofloxacin breakpoints for Salmonella species, Enterobacteriaceae other than Salmonella species and Pseudomonas aeruginosa.Species and antimicrobial agentHistorical interpretive categories and MIC breakpoints (mg/L)Updated interpretive categories and MIC breakpoints (mg/L)SusceptibleIntermediateResistantSusceptibleIntermediateResistantSalmonella species Ciprofloxacin≤12≥4≤0.060.12–0.5≥1 Levofloxacin≤24≥8≤0.120.25–1≥2Enterobacteriaceae other than Salmonella spp. Ciprofloxacin≤12≥4≤0.250.5≥1 Levofloxacin≤24≥8≤0.51≥2Pseudomonas aeruginosa Ciprofloxacin≤12≥4≤0.51≥2 Levofloxacin≤24≥8≤12≥4CLSI, Clinical and Laboratory Standards Institute Open table in a new tab CLSI, Clinical and Laboratory Standards Institute Accordingly, I conducted a cross-sectional survey querying microbiology laboratories at acute care hospitals and long-term care hospitals in Connecticut, USA, by contacting the microbiology laboratory manager and/or supervisor by phone. I asked about the antimicrobial susceptibility testing (AST) methods and FQ interpretative breakpoints used for Salmonella spp., Enterobacteriaceae and P. aeruginosa. I queried a total of 43 laboratories that serve all the acute care and long-term hospitals in Connecticut. Six laboratories referred AST to a local or national reference laboratory. Thus, I obtained information from 37 (86.1%) microbiology laboratories about the study question verbally by telephone. No microbiology laboratory personnel refused to provide us with information. Characteristics, AST methods for GNB and causes for nonadoption of current FQ breakpoints were summarized in Table 2. Note that updated FQ breakpoints for Salmonella spp. were more often adopted by the laboratories which used Kirby-Bauer (5/5, 100%), gradient diffusion testing (2/2, 100%) and bioMérieux VITEK 2 (5/9, 55.9%) compared to Microscan (0/15) and Phoenix (0/6). On the other hand, laboratories less often adopted the 2019 updated FQ breakpoints for Enterobacteriaceae and P. aeruginosa regardless of the AST method used (Table 2). A bivariate analysis using the Pearson chi-square or Fisher exact test revealed that the AST method used was significantly associated with different level of adoption of updated FQ breakpoints for Salmonella spp. as well as the other Enterobacteriaceae and P. aeruginosa, with p < 0.001 and 0.02 respectively.Table 2Connecticut microbiology laboratories (n = 37) antimicrobial susceptibility testing methods and fluoroquinolone clinical breakpoints used for Gram-negative bacteriaCharacteristicn (%)Affiliation of microbiology laboratory University-affiliated hospital8 (21.6) Community based hospital29 (78.4)AST method for Enterobacteriaceae and Pseudomonas aeruginosa Microscan Beckman coulter15 (40.6) bioMérieux VITEK 213 (35.1) BD phoenix8 (21.6) Sensititre Thermo Fisher Scientific0 Kirby-Bauer disc diffusion0 Gradient diffusion method1 (2.7)AST method for Salmonella spp. Microscan Beckman coulter15 (40.6) bioMérieux VITEK 29 (24.3) BD phoenix6 (16.2) Sensititre Thermo Fisher Scientific0 Kirby-Bauer disc diffusionaSalmonella-specific Kirby-Bauer disc diffusion breakpoints for ciprofloxacin are available, but none have been set for levofloxacin or ofloxacin.5 (13.5) Gradient diffusion method2 (5.4)Implementation of current CLSI breakpoints in microbiology laboratories Updated CLSI breakpoints implemented for Enterobacteriaceae and P. aeruginosa and Salmonella spp.4 (10.8) Updated CLSI breakpoints implemented for Enterobacteriaceae and P. aeruginosa but not for Salmonella spp.5 (13.5) Updated CLSI breakpoints implemented for Salmonella spp. but not for Enterobacteriaceae and P. aeruginosa8 (21.6) None of the updated CLSI breakpoints for Gram-negative bacteria has been implemented20 (54.1)Use of current CLSI breakpoints for Enterobacteriaceae and P. aeruginosa stratified by AST manufacturer among microbiology laboratories Microscan Beckman coulter0 bioMérieux VITEK 25 (13.5) BD phoenix4 (10.8) Sensititre Thermo Fisher Scientific0 Kirby-Bauer disc diffusionaSalmonella-specific Kirby-Bauer disc diffusion breakpoints for ciprofloxacin are available, but none have been set for levofloxacin or ofloxacin.0 Gradient diffusion method0Use of current CLSI breakpoints for Salmonella spp. stratified by AST manufacturer among microbiology laboratories Microscan Beckman coulter0 bioMérieux VITEK 25 (13.5) BD phoenix0 Sensititre Thermo Fisher Scientific0 Kirby-Bauer or disc diffusionaSalmonella-specific Kirby-Bauer disc diffusion breakpoints for ciprofloxacin are available, but none have been set for levofloxacin or ofloxacin.5 (13.5) Gradient diffusion method2 (5.4)Causes of nonadoption of current FQ clinical breakpoints for Enterobacteriaceae and P. aeruginosabEach cause for nonadoption of current fluoroquinolone breakpoints may represent a single cause or may be part of multiple concomitant causes. Lack of knowledge11 (29.7) Lack of updated panels by the AST manufacturers1 (2.7) Lack of knowledge and lack of updated panels by the AST manufacturers12 (32.4) Lack of resources in the microbiology laboratory delaying the verification of the updated panels4 (10.8)Causes of nonadoption of current FQ clinical breakpoints for Salmonella spp.bEach cause for nonadoption of current fluoroquinolone breakpoints may represent a single cause or may be part of multiple concomitant causes. Lack of knowledge11 (29.7)cFive (13.5%) microbiology laboratories adopted for Salmonella spp. the FQ breakpoints of Enterobacteriaceae. Lack of updated panels by the AST manufacturers1 (2.7) Lack of knowledge and lack of updated panels by the AST manufacturers12 (32.4) Lack of resources in the microbiology laboratory delaying the verification of the updated panels1 (2.7)AST, antimicrobial susceptibility test, CLSI, Clinical and Laboratory Standards Institute; FQ, fluoroquinolone.a Salmonella-specific Kirby-Bauer disc diffusion breakpoints for ciprofloxacin are available, but none have been set for levofloxacin or ofloxacin.b Each cause for nonadoption of current fluoroquinolone breakpoints may represent a single cause or may be part of multiple concomitant causes.c Five (13.5%) microbiology laboratories adopted for Salmonella spp. the FQ breakpoints of Enterobacteriaceae. Open table in a new tab AST, antimicrobial susceptibility test, CLSI, Clinical and Laboratory Standards Institute; FQ, fluoroquinolone. FQ have excellent oral bioavailability. Ciprofloxacin, levofloxacin and delafloxacin, if tested susceptible in vitro, represent the only FDA cleared oral option to treat P. aeruginosa infections. Adoption of the updated CLSI breakpoints for GNB to FQ presents several challenges. Revised CLSI breakpoints first have to be approved and released by the FDA so the AST manufacturers, which by US law must use FDA breakpoints, can get FDA approval for their systems with the updated breakpoints. The Generating Antibiotic Incentives Now Act was passed in July 2012 as part of the FDA Safety and Innovation Act to promote rapid diagnostic tests for detection of bacterial infection and AST [5Sfeir M.M. The GAIN Act legislation to combat antimicrobial resistance: where do we stand?.Infect Contr Hosp Epidemiol. 2018; 39: 1499-1500Crossref PubMed Scopus (7) Google Scholar, 6Sfeir M.M. Tigecycline for treatment of multidrug-resistant Gram-negative bacteria in transplant recipient patients: challenges of antimicrobial susceptibility testing.Transpl Infect Dis. 2020; 22e13254Crossref PubMed Scopus (1) Google Scholar, 7Sfeir M.M. Imipenem/relebactam for treatment of carbapenem-resistant Gram-negative bacteria: where do we stand on in vitro susceptibility testing?.Clin Infect Dis. 2020; Crossref Google Scholar]. Likewise, the 21st Century Cures Act was signed into law in December 2016 to help accelerate new medical product development [[8]US Food and Drug Administration (FDA)21st Century Cures Act. 2016.https://www.fda.gov/regulatory-information/selected-amendments-fdc-act/21st-century-cures-actGoogle Scholar,[9]Sfeir M.M. Antimicrobial susceptibility testing for glucose non fermenting Gram-negative bacteria: the tip of the iceberg.Antimicrob Agent Chemother. 2020; 64 (e00011-20)Crossref Scopus (2) Google Scholar]. The Cures Act provides the FDA the help to improve ability to recruit professional experts to review medical products development [[8]US Food and Drug Administration (FDA)21st Century Cures Act. 2016.https://www.fda.gov/regulatory-information/selected-amendments-fdc-act/21st-century-cures-actGoogle Scholar,[9]Sfeir M.M. Antimicrobial susceptibility testing for glucose non fermenting Gram-negative bacteria: the tip of the iceberg.Antimicrob Agent Chemother. 2020; 64 (e00011-20)Crossref Scopus (2) Google Scholar]. In fact, it takes the FDA 6 months on average to approve the updated breakpoints by CLSI [[4]US Food and Drug Administration (FDA)FDA rationale for recognition decision: levofloxacin. Content current as of June 2019.https://www.fda.gov/drugs/development-resources/fda-rationale-recognition-decision-levofloxacinGoogle Scholar]. Additionally, the FDA does not have the legal authority to oblige manufacturers of AST to update breakpoints, despite the noncompliant panels with current FDA regulations. Manufacturers need substantial resources to update the AST devices on the basis of the updated breakpoints, then validate the use of those breakpoints. In such situations, there is often a delay in updating breakpoints by firms, which could jeopardize patients safety. Stricter and more consistent federal and authoritative legislation is needed to shorten the time frame of the FDA review and enforce updating clinical breakpoints by AST manufactures in a timely manner. Another problematic area is the in-house verification study done by the clinical microbiology laboratories to confirm that the updated testing method performs as the manufacturer proposed. That requires that the laboratory personnel be aware of the updates and that the laboratory has adequate resources—including technicians and the ability to test for bacterial isolates—to carry out the verification study. That often represents a burden for microbiology laboratories. Automated AST panels are often limited in the number of antibiotics tested and the lower MIC cutoff for antibiotics that could be attained, requiring the use of additional automated panel that includes the low MIC cutoffs or a manual AST method. Either way is cumbersome and costly to the laboratories, and often leads to slower turnaround time of susceptibility reporting results. Finally, this study showed that the lack of knowledge among the laboratories personnel represents a high barrier to adopting updated CLSI breakpoints. This requires an urgent call to action to achieve consistency in AST reporting across microbiology laboratories. The author reports no conflicts of interest relevant to this letter. I thank the personnel of the microbiology laboratories in Connecticut for their collaboration.