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Research Article Integrating Bacterial Identification and Susceptibility Hindawi BioMed Research International Volume 2019, Article ID 8041746, 6 pages https://doi.org/10.1155/2019/8041746 Research Article Integrating Bacterial Identification and Susceptibility Testing: A Simple and Rapid Approach to Reduce the Turnaround Time in the Management of Blood Cultures Dariane C. Pereira1 and Luciano Z. Goldani 2 1Microbiology Unit, Laboratory Diagnosis Service, Hospital de Cl´ınicas de Porto Alegre, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil 2InfectiousDiseases Unit, Hospital de Cl´ınicas de Porto Alegre, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil Correspondence should be addressed to Luciano Z. Goldani; [email protected] Received 1 June 2019; Revised 15 August 2019; Accepted 16 September 2019; Published 3 October 2019 Academic Editor: Jiangke Yang Copyright © 2019 Dariane C. Pereira and Luciano Z. Goldani. -is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. We evaluated a rapid bacterial identification (rID) and a rapid antimicrobial susceptibility testing by disk diffusion (rAST) from positive blood culture to overcome the limitations of the conventional methods and reduce the turnaround time in bloodstream infection diagnostics. -e study included hemocultures flagged as positive by bacT/ALERT , identification by MALDI-TOF MS, and rAST. -e results were compared to identification and antimicrobial susceptibility testing (AST)® results by current standard methods, after 24 h incubation. For rAST categorical agreement (CA), very major errors (VME), major errors (ME), and minor errors (mE) were calculated. A total of 524 bacterial samples isolated from blood cultures were obtained, including 246 Gram-negative (GN) and 278 Gram-positive (GP) aerobes. -e overall concordance of rID was 88.6%, and it was highest among GN (96%). A total of 2196 and 1476 antimicrobial agent comparisons were obtained for GN and GP, respectively. Evaluation of rAST, CA, VME, ME, and mE disclosed 97.7, 0.7, 0.5, and 1.1% for GN and 98.0, 0.5, 0.7, and 0.8% for GP, respectively. Meropenem CA, VME, and ME were 98.3, 0.5, and 0.5%, respectively; mE was not observed. Oxacillin CA, ME, and mE were 97.4, 1.6, and 0.6%, respectively; VME was not observed. Overall, kappa scores of the results of the comparisons demonstrated the high agreement between rAST and the standard method. Identification and ASTof aerobic bacteria from positive blood cultures after a short period of incubation on solid blood agar is a fast and reliable method that may improve the management of bloodstream infections. 1. Introduction routine diagnostics are based on bacterial culture, which constitutes the actual gold standard and are precise and -e development of rapid diagnostic assays for the identi- sensitive but rather slow. Traditional identification and fication and analysis of antimicrobial resistance of bacteria antimicrobial susceptibility test results for microorganisms causing bloodstream infections is of utmost importance to causing bloodstream infections can take 48 h or longer to reduce morbidity and mortality. Infectious diseases have a obtain. Today, new methods have been made available to substantial global health impact. Fast diagnosis of pathogens enable faster diagnosis. Recently, matrix-assisted laser de- is critical to guarantee adequate therapy for infections [1]. A sorption/ionization time-of-flight mass spectrometry wide variety of microorganisms can cause bloodstream (MALDI-TOF MS) emerged as a rapid, accurate, cost-ef- infections, commonly E. coli, Klebsiella spp., S. aureus, other fective method and has been effectively used as a fast method bacteria, and yeast. Rapid identification of bloodstream for identifying a wide array of microbial species [3]. In pathogens is a laboratory practice that supports fast tran- MALDI-TOF MS analysis, abundant structural proteins sitions to direct target therapy, supporting timely and ef- such as ribosomal proteins are extracted from an entire fective patient care [2]. Current technologies employed in bacterial colony. -e ionizing laser vaporizes structural 2 BioMed Research International proteins of microorganisms, and unique mass spectra are an appropriate solid agar medium (aerobic Columbia agar generated, having mass-to-charge ratio (m/z) peaks with 5% sheep blood, chocolate agar, MacConkey agar, bio- varying intensities. -e mass spectra of test isolates are Merieux,´ France) following 18–24 h incubation at 35°C and sequentially compared with those in a reference database for 5% CO2 atmosphere. -e colonies grown on overnight agar identification [4]. plate incubation were spotted onto a target slide and pre- In comparison with conventional methods for identifi- pared according to the manufacturer’s instructions for cation of clinical samples by and despite its high-end analysis using VITEK MS system software version 3.0 technology, a MALDI-TOF MS device is simple to use. (bioMerieux).´ -is system® was termed the standard iden- MALDI-TOF MS can provide advantages for a universal tification method (sID). -e colonies grown on overnight procedure of microbial identification. Only a small amount agar plates were also used for the preparation of the in- of an organism, typically a fraction of a single colony from oculum disk diffusion test (Kirby–Bauer disk diffusion primary culture plates, is required for analysis. Compara- susceptibility test protocol), according to CLSI [6]. -is test tively, a larger inoculum and subculture is often required for was termed the standard disk diffusion method (sAST). -e conventional biochemical methods or other automated bacterial identification and antimicrobial resistance de- systems. Furthermore, once the instrument is loaded, tection results obtained using this conventional workflow identifications can typically be performed in less than one were used for comparison in the data analyses. minute, compared with hours to days for conventional methods [4, 5]. 2.4. Rapid Modified Bacterial Identification (rID) from Posi- Even simpler and faster than traditional bacterial tive Blood Culture. -e rapid modified bacterial identifi- identification methods, analysis of blood cultures via cation from positive blood cultures evaluated was previously MALDI-TOF MS and the detection of antimicrobial re- described by Chen et al. [7]. In brief, 3 mL of positive blood sistance require a preliminary bacterial culture in solid culture broth was aspirated from positive blood culture media. In this study, we evaluated a rapid modified bacterial bottles using a sterile syringe and transferred to a 10 mL identification and an early antimicrobial resistance detection serum separator tube. -e aspirate was centrifuged for 5 min from positive blood cultures based on centrifugation and at 3,000 rpm, the supernatant was discarded, and 20 μL of the short-time bacterial incubation to reduce the turnaround bacterial pellet was transferred to the center of a chocolate time in bloodstream infection diagnostics in a routine mi- agar plate (bioMerieux).´ -e inoculum was streaked out to crobiology laboratory. form a 2 by 2 cm, and the dish was incubated at 35°C in 5% CO2 atmosphere, for up to 4–6 h. Growth on the plate was 2. Materials and Methods recovered with a 1 μL inoculating loop, to obtain a sufficient 2.1.StudyDesignandStudyPopulation. Transversal study: all inoculum to be spotted onto a target slide prepared, Gram-negative bacilli and Staphylococcus sp. isolates re- according to the manufacturer’s instructions for analyses covered from bloodstream infection during 2017 and 2018 of using MALDI-TOF VITEK MS® system software version 3.0 patients admitted at a tertiary hospital in southern of Brazil (bioMerieux).´ -e identification process was performed only were included in the study. once for each sample. Criteria for interpretation of the re- sults proposed by the manufacturer were a single organism identification (same genus and species) as successful iden- 2.2. Blood Culture Procedures. Blood culture sets were ob- tification, and more than one species results with the same tained from patients with bloodstream infection. Aerobic genus, as acceptable identification. and anaerobic blood culture bottles (bacT/ALERT culture media FA Plus, PF plus and FN Plus, bioMerieux,´ ® France) 2.5. Rapid Modified Antimicrobial Susceptibility Testing were forwarded to the Microbiology Unit and incubated in (rAST) from Positive Blood Culture. A modification of the the automated blood culture system bacT/ALERT 3D standard disk diffusion method (rAST) was evaluated for the (bioMerieux).´ Negative bottles are automatically resulted® detection of antimicrobial resistance. -e rAST followed and discarded after five days of incubation. -e performance CLSI standards in all aspects, except for the inoculum methods were evaluated for positive blood cultures with preparation. CLSI guideline recommends preparing the monomicrobial bacterial growth. Bottles yielding poly- inoculum for AST with a direct suspension of isolated microbial or yeast growth were excluded. For analyses, we colonies selected from 18–24 h agar plate incubation [6]. In considered only one blood culture series per individual patient. Moreover, the testing only included blood cultures this study, we used colonies selected from a rapid culture from the daily microbiology laboratory routine; no artifi- grown (4–6 h agar plate incubation). -e inoculum was prepared from the rapid culture grown
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