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Quicolor: A novel system for rapid antibacterial susceptibility testing

Quicolor: A novel system for rapid antibacterial susceptibility testing Annals of Microbiology, 57 (1) 131-135 (2007) 1,2,3 4 5 6 1 4 Tanıl KOCAGÖZ *, Serpil ERCIS , Özge DARKA , Siavosh SALMANZADEH-AHRABI , Sesin KOCAGÖZ , Güls ¸en HASÇELIK 1 2 3 Yeditepe University, Faculty of Medicine, Department of Clinical Microbiology and Microbiology, TIBO, and Salubris, Istanbul; 4 5 Hacettepe University, Faculty of Medicine, Department of Clinical Microbiology and Microbiology, Ankara; Ondokuz Mayıs University, Faculty of Medicine, Department of Clinical Microbiology and Microbiology, Samsun, Turkey; Semnan University of Medical Sciences, Department of Microbiology, Semnan, Iran Received 26 September 2006 / Accepted 15 December 2006 Abstract - Early determination of antibacterial susceptibility increases the success of therapy, decreases unnecessary use of antibac- terials and side effects and lowers the overall healthcare costs. We have evaluated a rapid antibacterial susceptibility test, Quicolor (Salubris Inc., Massachusetts, USA), which is based on a rapid culture medium that indicates growth early by changing its colour. Quicolor proved to be a reliable rapid test for determining antibacterial susceptibility, having an overall agreement of 97.6% with the conventional CLSI disk diffusion susceptibility test results. Between two methods overall agreement was 96.7% for Enterobacteriaceae, 96.8% for staphylococci and 94.2% for non-fermentative bacteria. There was only 0.6% major discrepancy in Enterobacteriaceae, 1.7% in staphylococci and 0.9% in non-fermentative bacteria. Since the test provides results in 3.5-6 h, it can provide the means to choose the right treatment regimen the same day the infectious agent is isolated. Key words: disk diffusion, susceptibility, rapid test, Quicolor. INTRODUCTION produced around disks containing antibacterials (Fig. 1A). Quicolor NF agar is used to determine the susceptibility of Early determination of antibacterial susceptibility of bacte- non-fermentative bacteria like Pseudomonas and ria, isolated from cases like meningitis, bacteraemia and Acinetobacter species and this media changes its colour sepsis is very important for the selection of appropriate from yellow to red producing yellow inhibition zones (Fig. therapy as soon as possible. Even when there is no such 1B). urgent need, determination of antibacterial susceptibility on In this study, the susceptibility to various antibacterials the same day the culture results are obtained, it is crucial of 177 clinical isolates grown from the clinical samples sub- for the selection of the right treatment regimen, for increas- mitted to Clinical Pathology Laboratory of Hacettepe ing the success of therapy, for lowering the rate of side- University, Medical School, was determined by disk diffusion effects and mortality and for cutting down the healthcare method using Quicolor and Mueller Hinton agar and the costs (Granato, 1993; Doern et al., 1994; Schifman et al., results were compared. 1997; Barenfanger et al., 1999; Tunney et al., 2004; Kanemitsu et al., 2005). We have developed, produced and tested a media called Quicolor (Salubris Inc., MATERIALS AND METHODS Massachusetts, USA) for increasing efficiency in early deter- mination of antibacterial susceptibility. Quicolor is a media Clinical isolates. For evaluating Quicolor, all the isolates that changes its colour due to metabolic activity of growing grown from clinical samples during a three-month period bacteria. Quicolor is poured in Petri dishes and used to and 34 frozen strains of non-fermentative Gram-negative determine susceptibility to antibacterials by disk diffusion bacteria grown within the previous three months, were method. Quicolor has two types. Quicolor ES agar is used to included in the study. Among the 177 isolates studied, 66 determine the susceptibility of Enterobacteriaceae and belonged to Enterobacteriaceae (45 Escherichia coli, 9 staphylococci. This media changes its colour from red to yel- Klebsiella pneumoniae, 4 Klebsiella oxytoca, 3 Serratia low by bacterial growth and red circular inhibition zones are marcescens, 2 Proteus mirabilis, 1 Enterobacter cloacae, 1 Serratia liquefaciens, 1 Proteus vulgaris), 41 staphylococci (34 Staphylococcus aureus, 6 Staphylococcus epidermidis, 1 Staphylococcus caprae) and 70 non-fermentative Gram- * Corresponding author. Address:Yeditepe University Faculty of negative bacteria (45 Pseudomonas aeruginosa, 14 Medicine, Department of Clinical Microbiology and Microbiology, Acinetobacter baumannii, 5 Acinetobacter lwöffii, 6 Istanbul, Turkey; Phone: +90 532 321 1784, +90 216 578 0535; Fax: +90 216 578 0575; E-mail: tk05-k@tr.net Stenotrophomonas maltophilia). 132 T. Kocagöz et al. FIG. 1 - A: Quicolor ES agar, inoculated with enteric bacteria, turned the colour of the media to yellow, producing red inhibition zones around the antibacterial containing disks that the bacteria are susceptible. B: Quicolor NF agar, inoculated with non-fermenta- tive Gram-negative bacteria, turned the colour of the media to red, producing yellow inhibition zones around the antibacterial containing disks that the bacteria are susceptible. Susceptibility determination by Quicolor. Bacterial sus- RESULTS pensions were prepared from overnight cultures of clinical samples, using the Mueller-Hinton broth (BBL Microbiology The susceptibility results of isolates belonging to Systems, Cockeysville, USA), to produce a turbidity of 0.5 Enterobacteriaceae, with Quicolor ES agar were obtained in Mc Farland. The suspension was spread on the agar surface 4 h for the majority of the strains, the range being from 3.5 of Quicolor ES agar, for Enterobacteriaceae and staphylo- to 5 h. The average incubation time required for obtaining cocci, and Quicolor NF agar for non-fermentative Gram-neg- the susceptibility results of staphylococci and non-fermen- ative bacteria. Disks containing antibacterials (BBL tative bacteria by Quicolor ranged between 4 and 6 h, but Microbiology Systems) were placed and the plates were the majority of strains produced results in 4.5 h. incubated at 35 °C until inhibition zones in colour became The agreement between the susceptibility results apparent (Fig. 1A and 1B). The inhibition zone diameters obtained by Quicolor and CLSI disk diffusion test, for was measured and interpreted according to the “Easy Read Enterobacteriaceae, staphylococci and non-fermentative Chart” (a transparent card with circles that show the break- bacteria (Pseudomonas and Acinetobacter species) are points for susceptibility and resistance for each antibacteri- shown in Tables 1, 2, and 3 respectively. The total agree- al) provided by Salubris Inc. Escherichia coli ATCC 25922, P. ment between two tests was 96.7% for Enterobacteriaceae, aeruginosa ATCC 27853, K. pneumoniae 1951 high ESBL 96.8% for staphylococci and 94.2% for non-fermentative producing strain, K. pneumoniae 1204 low ESBL producing bacteria. Out of 3.3% total discrepancy only 0.6% was clas- strain, S. aureus ATCC 25923, methicillin resistant S. sified as major discrepancy for Enterobacteriaceae. The aureus 27R strain, and COL strain, S. epidermidis glycopep- major discrepancy in staphylococci was 1.7% and in Gram- tides intermediate strain which is one of the challenge negative non-fermentative bacteria 0.9%. strains and numbered as WHO-6 by Tenover et al. (2001) were also evaluated as quality control strains. DISCUSSION Susceptibility determination by CLSI disk diffusion method. From the bacterial suspensions prepared for Clinical and financial benefits of early reporting of antibac- Quicolor, bacteria were spread on the surface of Mueller- terial susceptibility results have been shown in many stud- Hinton agar (BBL Microbiology Systems) and paper disks ies (Granato, 1993; Doern et al., 1994; Schifman et al., containing the same antibacterials tested in Quicolor were 1997; Barenfanger et al., 1999; Tunney et al., 2004; placed. The plates were incubated for 18-24 h at 35 °C. The Kanemitsu et al., 2005). Barenfanger et. al. (1999) report- inhibition zone diameters were measured and interpreted ed that early reporting of antibacterial susceptibility test according to Clinical and Laboratory Standards Institute results decreased the length of stay in the hospital by 2.0 (CLSI) (formerly the NCCLS) criteria (CLSI, 2005). days and the average total cost for patient by $ 2395. In another group of patients Doern et al. (1994) reported a Evaluation of the results. Quicolor was evaluated by com- cost saving of $ 4194 per patient and additionally a statisti- paring the susceptibility results obtained from this and the cally significantly lower mortality rate in rapid antibiotic sus- CLSI disk diffusion test. If the test results were the same, ceptibility test group. In recent years, major technological either susceptible or resistant by both tests, it was defined advances have been made in clinical microbiology that have as “agreement”. If the result was susceptible or resistant by resulted in rapid reporting of antimicrobial susceptibility one test and intermediate with the other, this was called results that many regard as the most important information “minor discrepancy”. If the result was susceptible by one generated by the microbiology laboratory (Granato, 1993). test and resistant by the other it was called “major discrep- Although several automated systems aiming to provide ancy”. early antibacterial susceptibility results became available, Ann. Microbiol., 57 (1), 131-135 (2007) 133 TABLE 1 - The percentage of agreement between Quicolor and CLSI disk diffusion susceptibility results for Enterobacteriaceae (n = 66) Antibacterial Minor discrepancy Major discrepancy Total discrepancy Total agreement (%) (%) (%) (%) Amikacin 0 1.5 1.5 98.5 Ampicillin 4.5 0 4.5 95.5 Cefazolin 4.5 0 4.5 95.5 Cefoperazone 4.5 0 4.5 95.5 Cefotaxime 4.5 0 4.5 95.5 Ceftazidime 3.0 0 3.0 97.0 Cefuroxime 0 0 0 100 Ciprofloxacin 1.5 1.5 3.0 97.0 Gentamicin 1.5 2.0 4.5 95.5 Imipenem 0 0 0 100 Meropenem 0 0 0 100 Piperacillin 1.5 0 1.5 98.5 Ampicilllin/sulbactam 7.5 0 7.5 92.5 Trimethoprim/sulfamethoxazole 4.5 3.0 7.5 92.5 Total 2.7 0.6 3.3 96.7 TABLE 2 - The percentage of agreement between Quicolor and CLSI disk diffusion susceptibility results for staphylococci (n = 41) Antibacterial Minor discrepancy Major discrepancy Total discrepancy Total agreement (%) (%) (%) (%) Cefazolin 0 4.8 4.8 95.2 Ciprofloxacin 4.8 0 4.8 95.2 Clindamycin 4.8 0 4.8 95.2 Erythromycin 7.1 2.4 9.5 90.5 Fusidic acid 0 0 0 100 Oxacillin 0 0 0 100 Penicillin 0 0 0 100 Rifampicin 0 0 0 100 Ampicillin/sulbactam 0 4.8 4.8 95.2 Cefotaxime 2.4 4.8 7.2 92.8 Teicoplanin 0 0 0 100 Trimethoprim/sulfamethoxazole 0 4.8 4.8 95.2 Vancomycin 0 0 0 100 Total 1.5 1.7 3.2 96.8 Table 3 - The percentage of agreement between Quicolor and CLSI disk diffusion susceptibility results for non-fermentative Gram- negative bacteria (n = 70; Pseudomonas, Acinetobacter, Stenotrophomonas species) Antibacterial Minor discrepancy Major discrepancy Total discrepancy Total agreement (%) (%) (%) (%) Amikacin 7.0 1.4 8.4 91.6 Aztreonam 7.0 0 7.0 93.0 Ceftazidime 2.8 1.4 4.2 95.8 Ciprofloxacin 2.8 0 2.8 97.2 Gentamicin 2.8 0 2.8 97.2 Imipenem 9.9 0 9.9 90.1 Meropenem 7.0 1.4 8.4 91.6 Piperacillin 0 2.8 2.8 97.2 Total 4.9 0.9 5.8 94.2 134 T. Kocagöz et al. only limited information about the accuracy and especially mortality by providing the means for choosing the right the speed of these systems can be found in literature. In treatment regimen the same day the infectious agent is comparative evaluations of susceptibility testing proce- grown. In serious cases the susceptibility results may be dures, very major errors should occur in < 1.5% of all tests, confirmed by conventional standard tests. and the overall agreement between tests and the reference Quicolor was previously used successfully in rapid deter- method should be 95% (Nolte et al., 1986). mination of extended spectrum beta-lactamases (ESBL) in Vitek (bioMerieux, NC, USA) and MicroScan Walkaway Enterobacteriaceae using both double disk diffusion and E- (Diamond Diagnostics, MA, USA) are two of the most com- test (Sancak et al., 2005; Kocagöz et al., 2006). monly used automated antimicrobial susceptibility test sys- This is the first study evaluating Quicolor for its speed tems. A study evaluating susceptibility of Gram-negative and efficiency in correctly identifying antibacterial suscepti- bacilli to eleven antibacterials using MicroScan Rapid Neg bility. Further studies are needed in different settings to MIC/Combo panels (Diamond Diagnostics) and autoSCAN- reveal if this novel rapid system can be used as a reliable W/A (Baxter MicroScan, West Sacramento, CA) showed that method. the results were available between 3.5 and 7.0 h in 92.7% of the isolates and overall agreement with the standard test Acknowledgement was 94% with a 3.4% major error rate (Godsey et al., This study was supported by Turkish Scientific and Research 1991). Ribeiro et al. (1999) reported eight Staphylococcus Council. aureus strains initially identified by Vitek GPS-BS or GPS-SA cards as resistant to oxacillin to be found on further testing to be susceptible to oxacillin. REFERENCES Another study evaluating susceptibility of Gram-positive bacteria to 26 antibacterials using MicroScan Rapid Pos Barenfanger J., Drake C., Kacich G. (1999). Clinical and financial MIC/Combo panels and autoSCAN-W/A (Baxter), showed benefits of rapid bacterial identification and antimicrobial sus- that the results were available between 3.5 and 15 h in 98% ceptibility testing. J. Clin. Microbiol., 37: 1415-1418. of the organisms with an overall 96% agreement with stan- Bascomb S., Godsey J.H., Kangas M., Nea L., Tomfohrde K.M. dard MIC panels (Bascomb et al., 1991). McGregor et al. (1991). Rapid antimicrobial susceptibility testing of Gram- (1995) evaluated MicroScan and found out very major or positive cocci using Baxter MicroScan rapid fluorogenic panels and autoSCAN-W/A. Pathol. Biol., 39: 466-470. major discrepancies in 2% and minor discrepancies in 8% of Gram-negative susceptibility tests, the results being CLSI-Clinical and Laboratory Standards Institute (formerly the NCCLS) (2005). Performance Standards for Antimicrobial available in 7 h for 93% of the isolates; in Gram-positive Susceptibility Testing; Fifteenth Informational Supplement: susceptibility tests major and minor discrepancies with the M100-S15. CLSI, Wayne, PA, USA. standard test were 1% and 7% respectively. Comparison of Doern G., Vautour R., Gaudet M., Levy B. (1994). Clinical impact Vitek and Cobas Micro Systems, (Roche Diagnostics, Basel, of rapid in vitro susceptibility testing and bacterial identifica- Switzerland) with a semi automated conventional microsys- tion. J. Clin. Microbiol., 32: 1757-1762. tem MIC2000 (Dynatech, McLean, Va., USA), for suscepti- Godsey J.H., Bascomb S., Bonnette T., Kangas M., Link K., bility testing of Gram-negative bacilli revealed 86% overall Richards K., Tomfohrde K.M. (1991). Rapid antimicrobial sus- agreement with 3% major discrepancies for Vitek and 90% ceptibility testing of Gram-negative bacilli using Baxter overall agreement with 2% major discrepancies for Cobas MicroScan rapid fluorogenic panels and autoSCAN-W/A. Pathol. Biol., 39: 461-465. Micro systems (Simoons-Smit et al., 1994). Evaluation of 500 Gram-negative isolates to determine the number of Granato P.A. (1993). Impact of same day tests versus traditional overnight testing. Diagn. Microbiol. Infect. Dis., 16: 237-243. major susceptibility interpretation discrepancies between the Vitek and MicroScan Walkaway for 9 antimicrobial Kanemitsu K., Kunishima H., Inden K., Hatta M., Saga T., Ueno agents revealed only 1.06% discrepancies between these K., Harigae H., Ishizawa K., Kaku M. (2005). Assessment of RAISUS, a novel system for identification and antimicrobial tests (Rittenhouse et al., 1996). susceptibility testing for enterococci. Diagn. Microbiol. Infect. Ling et al had compared susceptibility testing results of Dis., 53: 23-27. 228 various members of the Enterobacteriaceae, Kocagöz S., Budak F., Gür D. (2006). Evaluation of a chromogenic Pseudomonas aeruginosa and other Gram-negative bacte- medium for rapid detection of extended spectrum β-lacta- ria, obtained with the Vitek 2 AST-No. 12 cards with those mase producing Salmonella spp. Indian J. Med. Res., 124: obtained by the broth microdilution method. They have 465-468. reported 0.5% major errors (resistant with the Vitek 2 sys- Ling T.K.W., Tam P.C., Liu Z.K., Cheng A.F.B. (2001). Evaluation tem but sensitive by the broth microdilution method) and of VITEK 2 Rapid Identification and Susceptibility Testing 0.4% very major errors (sensitive with the Vitek 2 system System against Gram-Negative Clinical Isolates. J. Clin. Microbiol., 39: 2964-2966. but resistant by the broth microdilution method) (Ling et al., 2001). McGregor A., Schio F., Beaton S., Boulton V., Perman M., Gilbert G. (1995). The MicroScan WalkAway diagnostic microbiology In this study, Quicolor proved to be reliable rapid test for system an evaluation. Pathology, 27: 172-176. determining antibacterial susceptibility the former having an overall agreement of 96.7% with CLSI disk diffusion test Nolte F.S., Contestable P.B., Lincalis D., Punsalang A.Jr. (1986). Rapid, direct antibiotic susceptibility testing of blood culture results for Enterobacteriaceae, 96.8% for staphylococci and isolates using the Abbott Advantage System. Am. J. Clin. 94.2% for non-fermentative bacteria. In Quicolor only 0.6% Pathol., 86: 665-669. was classified as major discrepancy for Enterobacteriaceae, Ribeiro J., Vieira F.D., King T., D’Arezzo J.B., Boyce J.M. (1999). 1.7% in staphylococci and 0.9% in non-fermentative bacte- Misclassification of susceptible strains of Staphylococcus ria. Since the test makes the results available between 3.5 aureus as methicillin-resistant S. aureus by a rapid automat- and 6 h, it may have a significant impact on lowering length ed susceptibility testing system. J. Clin. Microbiol., 37: 1619- of stay in the hospital, total cost for patient care and even 1620. Ann. Microbiol., 57 (1), 131-135 (2007) 135 Rittenhouse S.F., Miller L.A., Utrup L.J., Poupard J.A. (1996). Simoons-Smit A.M., MacLaren D.M. (1994). Comparison of Vitek Evaluation of 500 Gram negative isolates to determine the and Cobas Micro systems with semiautomated conventional number of major susceptibility interpretation discrepancies microsystem for identification and susceptibility testing of between the Vitek and MicroScan Walkaway for 9 antimicro- Gram-negative bacilli. J. Clin. Pathol., 47: 71-75. bial agents. Diagn. Microbiol. Infect. Dis., 26: 1-6. Tenover F.C., Mohammed M.J., Stelling J., O’Brien T., Williams R. Sancak B., Ercis S., Kocagöz T., Kocagöz S., Hascelik G., (2001). Ability of laboratories to detect emerging antimicro- Bolmström A. (2005). Rapid 4 to 6 hour detection of extend- bial resistance: proficiency testing and quality control results ed spectrum beta-lactamases (ESBL) using Quicolor Agar from the World Health Organization’s external quality assur- Medium with disk diffusion and Etest. American Society for ance system for antimicrobial susceptibility testing. J. Clin. th Microbiology, 105 General Meeting, Atlanta, Georgia. May, Microbiol., 39: 241-250. 6-9. Tunney M.M., Ramage G., Field T.R., Moriarty T.F., Storey D.G. Schifman R., Pindur A., Bryan J.A. (1997). Laboratory practices (2004). Rapid colorimetric assay for antimicrobial susceptibil- for reporting bacterial susceptibility tests that affect antibiot- ity testing of Pseudomonas aeruginosa. Antimicrob. Agents. ic therapy. Arch. Pathol. Lab. Med., 121: 1168-1170. Chemother., 48: 1879-1881. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Annals of Microbiology Springer Journals

Quicolor: A novel system for rapid antibacterial susceptibility testing

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Publisher
Springer Journals
Copyright
Copyright © 2007 by University of Milan and Springer
Subject
Life Sciences; Microbiology; Microbial Genetics and Genomics; Microbial Ecology; Fungus Genetics; Medical Microbiology; Applied Microbiology
ISSN
1590-4261
eISSN
1869-2044
DOI
10.1007/BF03175062
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Abstract

Annals of Microbiology, 57 (1) 131-135 (2007) 1,2,3 4 5 6 1 4 Tanıl KOCAGÖZ *, Serpil ERCIS , Özge DARKA , Siavosh SALMANZADEH-AHRABI , Sesin KOCAGÖZ , Güls ¸en HASÇELIK 1 2 3 Yeditepe University, Faculty of Medicine, Department of Clinical Microbiology and Microbiology, TIBO, and Salubris, Istanbul; 4 5 Hacettepe University, Faculty of Medicine, Department of Clinical Microbiology and Microbiology, Ankara; Ondokuz Mayıs University, Faculty of Medicine, Department of Clinical Microbiology and Microbiology, Samsun, Turkey; Semnan University of Medical Sciences, Department of Microbiology, Semnan, Iran Received 26 September 2006 / Accepted 15 December 2006 Abstract - Early determination of antibacterial susceptibility increases the success of therapy, decreases unnecessary use of antibac- terials and side effects and lowers the overall healthcare costs. We have evaluated a rapid antibacterial susceptibility test, Quicolor (Salubris Inc., Massachusetts, USA), which is based on a rapid culture medium that indicates growth early by changing its colour. Quicolor proved to be a reliable rapid test for determining antibacterial susceptibility, having an overall agreement of 97.6% with the conventional CLSI disk diffusion susceptibility test results. Between two methods overall agreement was 96.7% for Enterobacteriaceae, 96.8% for staphylococci and 94.2% for non-fermentative bacteria. There was only 0.6% major discrepancy in Enterobacteriaceae, 1.7% in staphylococci and 0.9% in non-fermentative bacteria. Since the test provides results in 3.5-6 h, it can provide the means to choose the right treatment regimen the same day the infectious agent is isolated. Key words: disk diffusion, susceptibility, rapid test, Quicolor. INTRODUCTION produced around disks containing antibacterials (Fig. 1A). Quicolor NF agar is used to determine the susceptibility of Early determination of antibacterial susceptibility of bacte- non-fermentative bacteria like Pseudomonas and ria, isolated from cases like meningitis, bacteraemia and Acinetobacter species and this media changes its colour sepsis is very important for the selection of appropriate from yellow to red producing yellow inhibition zones (Fig. therapy as soon as possible. Even when there is no such 1B). urgent need, determination of antibacterial susceptibility on In this study, the susceptibility to various antibacterials the same day the culture results are obtained, it is crucial of 177 clinical isolates grown from the clinical samples sub- for the selection of the right treatment regimen, for increas- mitted to Clinical Pathology Laboratory of Hacettepe ing the success of therapy, for lowering the rate of side- University, Medical School, was determined by disk diffusion effects and mortality and for cutting down the healthcare method using Quicolor and Mueller Hinton agar and the costs (Granato, 1993; Doern et al., 1994; Schifman et al., results were compared. 1997; Barenfanger et al., 1999; Tunney et al., 2004; Kanemitsu et al., 2005). We have developed, produced and tested a media called Quicolor (Salubris Inc., MATERIALS AND METHODS Massachusetts, USA) for increasing efficiency in early deter- mination of antibacterial susceptibility. Quicolor is a media Clinical isolates. For evaluating Quicolor, all the isolates that changes its colour due to metabolic activity of growing grown from clinical samples during a three-month period bacteria. Quicolor is poured in Petri dishes and used to and 34 frozen strains of non-fermentative Gram-negative determine susceptibility to antibacterials by disk diffusion bacteria grown within the previous three months, were method. Quicolor has two types. Quicolor ES agar is used to included in the study. Among the 177 isolates studied, 66 determine the susceptibility of Enterobacteriaceae and belonged to Enterobacteriaceae (45 Escherichia coli, 9 staphylococci. This media changes its colour from red to yel- Klebsiella pneumoniae, 4 Klebsiella oxytoca, 3 Serratia low by bacterial growth and red circular inhibition zones are marcescens, 2 Proteus mirabilis, 1 Enterobacter cloacae, 1 Serratia liquefaciens, 1 Proteus vulgaris), 41 staphylococci (34 Staphylococcus aureus, 6 Staphylococcus epidermidis, 1 Staphylococcus caprae) and 70 non-fermentative Gram- * Corresponding author. Address:Yeditepe University Faculty of negative bacteria (45 Pseudomonas aeruginosa, 14 Medicine, Department of Clinical Microbiology and Microbiology, Acinetobacter baumannii, 5 Acinetobacter lwöffii, 6 Istanbul, Turkey; Phone: +90 532 321 1784, +90 216 578 0535; Fax: +90 216 578 0575; E-mail: tk05-k@tr.net Stenotrophomonas maltophilia). 132 T. Kocagöz et al. FIG. 1 - A: Quicolor ES agar, inoculated with enteric bacteria, turned the colour of the media to yellow, producing red inhibition zones around the antibacterial containing disks that the bacteria are susceptible. B: Quicolor NF agar, inoculated with non-fermenta- tive Gram-negative bacteria, turned the colour of the media to red, producing yellow inhibition zones around the antibacterial containing disks that the bacteria are susceptible. Susceptibility determination by Quicolor. Bacterial sus- RESULTS pensions were prepared from overnight cultures of clinical samples, using the Mueller-Hinton broth (BBL Microbiology The susceptibility results of isolates belonging to Systems, Cockeysville, USA), to produce a turbidity of 0.5 Enterobacteriaceae, with Quicolor ES agar were obtained in Mc Farland. The suspension was spread on the agar surface 4 h for the majority of the strains, the range being from 3.5 of Quicolor ES agar, for Enterobacteriaceae and staphylo- to 5 h. The average incubation time required for obtaining cocci, and Quicolor NF agar for non-fermentative Gram-neg- the susceptibility results of staphylococci and non-fermen- ative bacteria. Disks containing antibacterials (BBL tative bacteria by Quicolor ranged between 4 and 6 h, but Microbiology Systems) were placed and the plates were the majority of strains produced results in 4.5 h. incubated at 35 °C until inhibition zones in colour became The agreement between the susceptibility results apparent (Fig. 1A and 1B). The inhibition zone diameters obtained by Quicolor and CLSI disk diffusion test, for was measured and interpreted according to the “Easy Read Enterobacteriaceae, staphylococci and non-fermentative Chart” (a transparent card with circles that show the break- bacteria (Pseudomonas and Acinetobacter species) are points for susceptibility and resistance for each antibacteri- shown in Tables 1, 2, and 3 respectively. The total agree- al) provided by Salubris Inc. Escherichia coli ATCC 25922, P. ment between two tests was 96.7% for Enterobacteriaceae, aeruginosa ATCC 27853, K. pneumoniae 1951 high ESBL 96.8% for staphylococci and 94.2% for non-fermentative producing strain, K. pneumoniae 1204 low ESBL producing bacteria. Out of 3.3% total discrepancy only 0.6% was clas- strain, S. aureus ATCC 25923, methicillin resistant S. sified as major discrepancy for Enterobacteriaceae. The aureus 27R strain, and COL strain, S. epidermidis glycopep- major discrepancy in staphylococci was 1.7% and in Gram- tides intermediate strain which is one of the challenge negative non-fermentative bacteria 0.9%. strains and numbered as WHO-6 by Tenover et al. (2001) were also evaluated as quality control strains. DISCUSSION Susceptibility determination by CLSI disk diffusion method. From the bacterial suspensions prepared for Clinical and financial benefits of early reporting of antibac- Quicolor, bacteria were spread on the surface of Mueller- terial susceptibility results have been shown in many stud- Hinton agar (BBL Microbiology Systems) and paper disks ies (Granato, 1993; Doern et al., 1994; Schifman et al., containing the same antibacterials tested in Quicolor were 1997; Barenfanger et al., 1999; Tunney et al., 2004; placed. The plates were incubated for 18-24 h at 35 °C. The Kanemitsu et al., 2005). Barenfanger et. al. (1999) report- inhibition zone diameters were measured and interpreted ed that early reporting of antibacterial susceptibility test according to Clinical and Laboratory Standards Institute results decreased the length of stay in the hospital by 2.0 (CLSI) (formerly the NCCLS) criteria (CLSI, 2005). days and the average total cost for patient by $ 2395. In another group of patients Doern et al. (1994) reported a Evaluation of the results. Quicolor was evaluated by com- cost saving of $ 4194 per patient and additionally a statisti- paring the susceptibility results obtained from this and the cally significantly lower mortality rate in rapid antibiotic sus- CLSI disk diffusion test. If the test results were the same, ceptibility test group. In recent years, major technological either susceptible or resistant by both tests, it was defined advances have been made in clinical microbiology that have as “agreement”. If the result was susceptible or resistant by resulted in rapid reporting of antimicrobial susceptibility one test and intermediate with the other, this was called results that many regard as the most important information “minor discrepancy”. If the result was susceptible by one generated by the microbiology laboratory (Granato, 1993). test and resistant by the other it was called “major discrep- Although several automated systems aiming to provide ancy”. early antibacterial susceptibility results became available, Ann. Microbiol., 57 (1), 131-135 (2007) 133 TABLE 1 - The percentage of agreement between Quicolor and CLSI disk diffusion susceptibility results for Enterobacteriaceae (n = 66) Antibacterial Minor discrepancy Major discrepancy Total discrepancy Total agreement (%) (%) (%) (%) Amikacin 0 1.5 1.5 98.5 Ampicillin 4.5 0 4.5 95.5 Cefazolin 4.5 0 4.5 95.5 Cefoperazone 4.5 0 4.5 95.5 Cefotaxime 4.5 0 4.5 95.5 Ceftazidime 3.0 0 3.0 97.0 Cefuroxime 0 0 0 100 Ciprofloxacin 1.5 1.5 3.0 97.0 Gentamicin 1.5 2.0 4.5 95.5 Imipenem 0 0 0 100 Meropenem 0 0 0 100 Piperacillin 1.5 0 1.5 98.5 Ampicilllin/sulbactam 7.5 0 7.5 92.5 Trimethoprim/sulfamethoxazole 4.5 3.0 7.5 92.5 Total 2.7 0.6 3.3 96.7 TABLE 2 - The percentage of agreement between Quicolor and CLSI disk diffusion susceptibility results for staphylococci (n = 41) Antibacterial Minor discrepancy Major discrepancy Total discrepancy Total agreement (%) (%) (%) (%) Cefazolin 0 4.8 4.8 95.2 Ciprofloxacin 4.8 0 4.8 95.2 Clindamycin 4.8 0 4.8 95.2 Erythromycin 7.1 2.4 9.5 90.5 Fusidic acid 0 0 0 100 Oxacillin 0 0 0 100 Penicillin 0 0 0 100 Rifampicin 0 0 0 100 Ampicillin/sulbactam 0 4.8 4.8 95.2 Cefotaxime 2.4 4.8 7.2 92.8 Teicoplanin 0 0 0 100 Trimethoprim/sulfamethoxazole 0 4.8 4.8 95.2 Vancomycin 0 0 0 100 Total 1.5 1.7 3.2 96.8 Table 3 - The percentage of agreement between Quicolor and CLSI disk diffusion susceptibility results for non-fermentative Gram- negative bacteria (n = 70; Pseudomonas, Acinetobacter, Stenotrophomonas species) Antibacterial Minor discrepancy Major discrepancy Total discrepancy Total agreement (%) (%) (%) (%) Amikacin 7.0 1.4 8.4 91.6 Aztreonam 7.0 0 7.0 93.0 Ceftazidime 2.8 1.4 4.2 95.8 Ciprofloxacin 2.8 0 2.8 97.2 Gentamicin 2.8 0 2.8 97.2 Imipenem 9.9 0 9.9 90.1 Meropenem 7.0 1.4 8.4 91.6 Piperacillin 0 2.8 2.8 97.2 Total 4.9 0.9 5.8 94.2 134 T. Kocagöz et al. only limited information about the accuracy and especially mortality by providing the means for choosing the right the speed of these systems can be found in literature. In treatment regimen the same day the infectious agent is comparative evaluations of susceptibility testing proce- grown. In serious cases the susceptibility results may be dures, very major errors should occur in < 1.5% of all tests, confirmed by conventional standard tests. and the overall agreement between tests and the reference Quicolor was previously used successfully in rapid deter- method should be 95% (Nolte et al., 1986). mination of extended spectrum beta-lactamases (ESBL) in Vitek (bioMerieux, NC, USA) and MicroScan Walkaway Enterobacteriaceae using both double disk diffusion and E- (Diamond Diagnostics, MA, USA) are two of the most com- test (Sancak et al., 2005; Kocagöz et al., 2006). monly used automated antimicrobial susceptibility test sys- This is the first study evaluating Quicolor for its speed tems. A study evaluating susceptibility of Gram-negative and efficiency in correctly identifying antibacterial suscepti- bacilli to eleven antibacterials using MicroScan Rapid Neg bility. Further studies are needed in different settings to MIC/Combo panels (Diamond Diagnostics) and autoSCAN- reveal if this novel rapid system can be used as a reliable W/A (Baxter MicroScan, West Sacramento, CA) showed that method. the results were available between 3.5 and 7.0 h in 92.7% of the isolates and overall agreement with the standard test Acknowledgement was 94% with a 3.4% major error rate (Godsey et al., This study was supported by Turkish Scientific and Research 1991). Ribeiro et al. (1999) reported eight Staphylococcus Council. aureus strains initially identified by Vitek GPS-BS or GPS-SA cards as resistant to oxacillin to be found on further testing to be susceptible to oxacillin. REFERENCES Another study evaluating susceptibility of Gram-positive bacteria to 26 antibacterials using MicroScan Rapid Pos Barenfanger J., Drake C., Kacich G. (1999). 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Annals of MicrobiologySpringer Journals

Published: Nov 20, 2009

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