Rapid Antimicrobial Susceptibility Testing Methods for Blood Cultures and Their Clinical Impact.
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ABSTRACT:
SUBMITTER: Banerjee R
PROVIDER: S-EPMC7987685 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:ObjectivesWhen bloodstream infections are caused by resistant bacteria, rapid antimicrobial susceptibility testing (RAST) is important for adjustment of therapy. The EUCAST RAST method, directly from positive blood cultures, was validated in a multi-laboratory study in Europe.MethodsRAST was performed in 40 laboratories in northern Europe (NE) and 15 in southern Europe (SE) from clinical blood cultures positive for Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus or Streptococcus pneumoniae. Categorical results at 4, 6 and 8 h of incubation were compared with results for EUCAST standard 16-20 h disc diffusion. The method, preliminary breakpoints and the performance of the laboratories were evaluated.ResultsThe total number of isolates was 833/318 in NE/SE. The number of zone diameters that could be read (88%, 96% and 99%) and interpreted (70%, 81% and 85%) increased with incubation time (4, 6 and 8 h). The categorical agreement was acceptable, with total error rates in NE/SE of 2.4%/4.9% at 4 h, 1.1%/3.5% at 6 h and 1.1%/3.3% at 8 h. False susceptibility at 4, 6 and 8 h of incubation was below 0.3% and 1.1% in NE and SE, respectively, and the corresponding percentages for false resistance were below 1.9% and 2.8%. After fine-tuning breakpoints, more zones could be interpreted (73%, 89% and 93%), with only marginally affected error rates.ConclusionsThe EUCAST RAST method can be implemented in routine laboratories without major investments. It provides reliable antimicrobial susceptibility testing results for relevant bloodstream infection pathogens after 4-6 h of incubation.
| S-EPMC7566356 | biostudies-literature
Project description:Our objective was to evaluate EUCAST's 'rapid antimicrobial susceptibility testing' (RAST) directly from positive blood culture that delivers antimicrobial results within 6 h for Staphylococcus aureus, Enterococcus spp., Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa, using total lab automation. Zone diameters from RAST were compared with MIC results. Furthermore, its influence on time to report was investigated. RAST was performed to all positive aerobic and anaerobic blood culture bottles by subculturing them, i.e. onto Mueller-Hinton agar and adding six antibiotic discs covering Gram-negative and Gram-positive therapy (cefoxitin, ampicillin, vancomycin, piperacillin/tazobactam, meropenem and ciprofloxacin). RAST was automatically imaged after 6 h. Zone sizes were measured using a TLA software tool and interpreted according to EUCAST clinical breakpoints. Bacteria were identified using MALDI-TOF MS and MIC results were determined using Vitek2 panels. Categorial agreement between agar diffusion and MIC results was investigated. Additionally, time to RAST and time to Vitek were compared for 100 isolates (20 per species). Between November 2018 and April 2019, 3313 positive mono-bacterial blood culture bottles were collected of which 894 bottles with RAST-validated species were investigated. Among these bottles, 2029 individual antibiotic measurements were compared with MIC results from Vitek2 and 14 very major, 28 major and 12 minor errors were found. A median reduction of 17:30 h in time to report was observed. Introduction of RAST with automatic TLA imaging function could reduce time to report by 17:30 h. Excellent accordance between zone diameter and MIC results, particularly for cefoxitin, vancomycin and meropenem, was observed, but drawbacks due to ATU were seen.
| S-EPMC7303068 | biostudies-literature
Project description:Rapid determination of bacterial antibiotic susceptibility is important for proper treatment of infections. The European Committee on Antimicrobial Susceptibility Testing (EUCAST) has recently published guidelines for rapid antimicrobial susceptibility testing (RAST) performed directly from positive blood culture vials. These guidelines, however, were only published for a limited number of common pathogenic bacteria. In this study, we evaluated the applicability of these guidelines to three Tier 1 bioterror agents (Bacillus anthracis, Yersinia pestis and Francisella tularensis) that require prompt antibiotic treatment to mitigate morbidity and mortality. We used spiked-in human blood incubated in a BACTEC™ FX40 system to determine the proper conditions for RAST using disc-diffusion and Etest assays. We found that reliable disc-diffusion inhibition diameters and Etest MIC values could be obtained in remarkably short times. Compared to the EUCAST-recommended disc-diffusion assays that will require adjusted clinical breakpoint tables, Etest-based RAST was advantageous, as the obtained MIC values were similar to the standard MIC values, enabling the use of established category breakpoint tables. Our results demonstrate the promising applicability of the EUCAST RAST for B. anthracis-, Y. pestis- or F. tularensis-positive blood cultures, which can lead to shorter diagnostics and prompt antibiotic treatment of these dangerous pathogens.
| S-EPMC8153291 | biostudies-literature
Project description:Antibiotic resistance is a major public health problem requiring the early optimization of antibiotic prescriptions. Matrix-Assisted Laser Desorption Ionization-Time Of Flight Mass Spectrometry (MALDI-TOF MS) has been shown to accurately identify bacteria from positive blood culture. Here, we developed a rapid detection of Escherichia coli resistance to amoxicillin (AMX) and cefotaxime (CTX) from positive blood culture based on MALDI-TOF MS. Potential sparing of broad-spectrum antibiotics was also evaluated. We tested 103 E. coli-positive blood cultures. Blood cultures were subculture 1-hour in antibiotic-free rich liquid media before further incubation with and without AMX for 2.5 h or CTX for 2 h. Protein extracts associated with an internal control were spotted on a MALDI-TOF target and spectra were analyzed with the MBT-ASTRA prototype software (Bruker Daltonik GmbH, Bremen, Germany). Bacterial growth ratio was calculated from the AUC spectra obtained in the presence and absence of the antibiotic and compared to a threshold which classified E. coli as susceptible or resistant. Results were interpreted with MICs determined using agar dilution method as reference technique. MBT-ASTRA recognized 95% and 84% of the AMX- and CTX-susceptible isolates, respectively. Overall, quantitative analysis of mass spectra allows susceptibility testing within 4 hours after the positivity of blood culture with E. coli. At the first report of positive blood culture, MALDI-TOF MS would then provide the prescribers with the bacterial identification and the susceptibility to AMX and CTX, thus limiting the use of broad-spectrum compounds.
| S-EPMC6181389 | biostudies-literature
Project description:BackgroundAppropriate antibiotic therapy is critical in the management of severe sepsis and septic shock to reduce mortality, morbidity and health costs. New methods for rapid antibiotic susceptibility testing are needed because of increasing resistance rates to standard treatment.AimsThe purpose of this study was to evaluate the performance of a novel microfluidic method and the potential to directly apply this method on positive blood cultures.MethodsMinimum inhibitory concentrations (MICs) of ciprofloxacin, ceftazidime, tigecycline and/or vancomycin for Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae and Staphylococcus aureus were determined using a linear antibiotic concentration gradient in a microfluidic assay. Bacterial growth along the antibiotic gradient was monitored using automated time-lapse photomicrography and growth inhibition was quantified by measuring greyscale intensity changes in the images. In addition to pure culture MICs, vancomycin MICs were determined for S. aureus from spiked and clinical blood cultures following a short centrifugation step. The MICs were compared with those obtained with the Etest and for S. aureus and vancomycin also with macrodilution.ResultsThe MICs obtained with the microfluidic assay showed good agreement internally as well as with the Etest and macrodilution assays, although some minor differences were noted between the methods. The time to possible readout was within the range of 2 to 5 h.ConclusionsThe examined microfluidic assay has the potential to provide rapid and accurate MICs using samples from positive clinical blood cultures and will now be tested using other bacterial species and antibiotics.
| S-EPMC5156554 | biostudies-literature
Project description:For the timely treatment of patients with infections in bloodstream and cerebrospinal fluid, a rapid antimicrobial susceptibility test (AST) is urgently needed. Here, we describe a direct and rapid antimicrobial susceptibility testing (dRAST) system, which can determine the antimicrobial susceptibility of bacteria from a positive blood culture bottle (PBCB) in six hours. The positive blood culture sample is directly mixed with agarose and inoculated into a micropatterned plastic microchip with lyophilized antibiotic agents. Using microscopic detection of bacterial colony formation in agarose, the total time to result from a PBCB for dRAST was only six hours for a wide range of bacterial concentrations in PBCBs. The results from the dRAST system were consistent with the results from a standard AST, broth microdilution test. In tests of clinical isolates (n = 206) composed of 16 Gram-negative species and seven Gram-positive species, the dRAST system was accurate compared to the standard broth microdilution test, with rates of 91.11% (2613/2868) categorical agreement, 6.69% (192/2868) minor error, 2.72% (50/1837) major error and 1.45% (13/896) very major error. Thus, the dRAST system can be used to rapidly identify appropriate antimicrobial agents for the treatment of blood stream infection (BSI) and antibiotic-resistant strain infections.
| S-EPMC5430693 | biostudies-literature
Project description:Briefly incubated agar cultures from positive blood cultures were used for antimicrobial susceptibility testing (AST) by Vitek 2. The cultivation time until inoculation was 3.8 h for Gram-positive cocci and 2.4 h for Gram-negative rods. The error rates were low, providing early and reliable AST without additional time or cost expenditure.
| S-EPMC4313230 | biostudies-literature
Project description: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.
| S-EPMC5530678 | biostudies-literature
Project description:Rapid detection and phenotyping of pathogenic microbes is critical for administration of effective antibiotic therapies and for impeding the spread of antibiotic resistance. Here, we present a novel platform, rapid ultrasensitive detector (RUSD), that utilizes the high reflectance coefficient at high incidence angles when light travels from low- to high-refractive-index media. RUSD leverages a principle that does not require complex manufacturing, labeling, or processing steps. Utilizing RUSD, we can detect extremely low cell densities (optical density [OD] ≥ 5 × 10-7) that correspond to approximately 20 bacterial cells or a single fungal cell in the detection volume, which is nearly 4 orders of magnitude more sensitive than standard OD methods. RUSD can measure minimum inhibitory concentrations (MICs) of commonly used antibiotics against gram-negative and gram-positive bacteria, including Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli, within 2 to 4 h. Here, we demonstrate that antibiotic susceptibility tests for several pathogens can rapidly be performed with RUSD using both small inoculum sizes (500 cells/mL) and larger inoculum sizes (5 × 105 cells/mL) used in standard antibiotic susceptibility tests. We anticipate that the RUSD system will be particularly useful for the cases in which antibiotic susceptibility tests have to be done with a limited number of bacterial cells that are available. Its compatibility with standard antibiotic susceptibility tests, simplicity, and low cost can make RUSD a viable and rapidly deployed diagnostic tool.
| S-EPMC6559665 | biostudies-literature
Project description:Echinocandins are the recommended first-line antifungals for treatment of invasive candidiasis. The increasing number of Candida glabrata strains resistant against echinocandins is an emerging health care concern. The rapid detection of resistant C. glabrata isolates is an urgent requirement for clinical laboratories. In this study, we developed the MALDI Biotyper antibiotic (antifungal) susceptibility test rapid assay (MBT ASTRA) for the rapid detection of anidulafungin-resistant C. glabrata isolates directly from positive blood cultures. Of 100?C. glabrata strains, MBT ASTRA classified 69 as susceptible and 29 as resistant. Microdilution assays performed according to the Clinical and Laboratory Standards Institute (CLSI) guidelines, used as a standard reference, identified 65 susceptible, 9 intermediate, and 26 resistant isolates. Sequencing of hot spot 1 and hot spot 2 regions of the FKS1 and FKS2 genes classified 86 susceptible and 14 resistant isolates. The MBT ASTRA had sensitivity and specificity of 80% and 95%, respectively, compared to the microdilution method. Positive and negative agreement of MBT ASTRA was calculated at 100% and 80%, respectively, compared with the molecular sequencing approach. Together, these results revealed a high accuracy of MBT ASTRA compared to microdilution according to the CLSI and PCR analysis, resulting in a categorical agreement of 90% and 83%, respectively. The validity of MBT ASTRA was 98%. Importantly, MBT ASTRA provided antifungal susceptibility testing (AFST) within 6?h that was both accurate and reliable compared to the other two approaches, which require at least 24?h or are costly. Therefore, this method has the potential to facilitate clinical AFST rapidly at low sample costs for clinical labs already equipped with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS).
| S-EPMC6709499 | biostudies-literature