Performance of VITEK® 2 AST-GN Ceftazidime/Avibactam (S < 8/4 µg/mL and R ≥16/4 µg/mL) for Antimicrobial Susceptibility Testing of Enterobacteriaceae and Pseudomonas aeruginosa S. FRANKLIN¹, H.P. DWIVEDI1, V. LABOMBARDI2,T. DAVIS3, R. HUMPHRIES4, J. HINDLER4, S. CAMPEAU4, P. DEOL1, 1bioMérieux, Inc., Hazelwood, MO; 2New York Presbyterian Hospital, Flushing, NY; 3Indiana University School of Medicine, Indianapolis, IN, 4University of California Los Angeles (UCLA), Los Angeles, CA. ASM 2018 ● Atlanta, GA Friday, June 8, 2018 Poster # 232
[email protected] 314.731.8754
Background: Ceftazidime/Avibactam (CZA) is a cephalosporin and beta-lactamase inhibitor combination drug with activity against extended-spectrum β-lactamase (ESBL)producing Enterobacteriaceae and multidrug-resistant (MDR) Pseudomonas aeruginosa (P. aeruginosa). In this study, the performance of VITEK2 CZA, an in vitro technique for determining the antimicrobial susceptibility of Enterobacteriaceae and P. aeruginosa, was evaluated against the CLSI broth microdilution (BMD) reference method Method: A total of 1073 isolates (866 Enterobacteriaceae and 207 P. aeruginosa) were tested by VITEK2 CZA method and BMD reference procedure (CLSI M07-A10). The results were analyzed for essential agreement (EA), category agreement (CA), major and very major error rates following FDA/ISO performance criteria using the FDA/EUCAST breakpoints for Ceftazidime/Avibactam (Enterobacteriaceae and P. aeruginosa S < 8/4 µg/mL and R ≥16/4 µg/mL). Results: VITEK2 CZA overall performance for Enterobacteriaceae and P. aeruginosa met the performance criteria for EA and CA, major errors and very major errors. The overall categorical major error rate for Enterobacteriaceae and P. aeruginosa combined was 1.4% (14/998). Nine (9) major errors were one dilution apart from the reference method and as such fall within EA. Based on the EA, and the lack of an intermediate breakpoint for Ceftazidime/Avibactam, the adjusted FDA categorical major error rate was 0.4% (3/829) for Enterobacteriaceae , 1.2% (2/169) for P. aeruginosa, and 0.5% (5/998) for all organisms combined. Table 1: Summary of VITEK2 CZA performance
Claimed Species Enterobacteriaceae (FDA) P. aeruginosa (FDA) Overall-FDA Overall-EUCAST
EA
Evaluate VITEK2 CZA performance when used with the VITEK®2 and VITEK®2 Compact Systems in a clinical setting, and to show the equivalency of the product to the CLSI BMD technique. Data obtained was used for submission to the FDA and CE marking. VITEK2 CZA is FDA 510 (k) approved and CE marked.
MATERIALS AND METHODS Fresh clinical and stock isolates were evaluated at three clinical trial sites in the United States. The range of species tested included Enterobacteriaceae and P. aeruginosa. Each isolate was first subcultured on TSA with sheep blood agar. From an 18-24 hour culture, a DensiCHEK Plus instrument was used to create a suspension with a 0.5-0.63 McFarland range. Clinical, Challenge, Reproducibility, and Quality Control (QC) isolates were tested on the VITEK®2 System with the auto dilution option. In addition, the VITEK®2 System with manual for inoculum preparation was evaluated for QC, Reproducibility and Challenge strains. The VITEK®2 Compact System was also evaluated during the clinical trial. Testing consisted of QC, Reproducibility and Challenge strains. One initial McFarland suspension was prepared for each isolate for inoculation of all the cards, and the reference when applicable. Table 2 : Sample type and testing method
1st card
CA
Table 5 : Frequency table of Overall Enterobacteriaceae and P. aeruginosa
OBJECTIVE
ABSTRACT
CA ME Rate
CA VME Rate
94.2% (816/866)
99.2% (859/866)
0.8% (7/829)
0.0% (0/37)
95.7% (198/207)
96.6% (200/207)
4.1% (7/169)
0.0% (0/38)
94.5% (1014/1073)
98.7% (1059/1073)
1.4% (14/998)
0.0% (0/75)
94.5% (1014/1073)
98.9%* (1061/1073)
1.2%* (12/998)
0.0% (0/75)
* Resolved data shown Conclusion: When compared to the BMD reference method, VITEK2 CZA proved to be a suitable test for susceptibility testing of Enterobacteriaceae and P. aeruginosa.
Total
EA
%EA
CA
%CA
#R
#ME
#VME
Citrobacter freundii
57
49
86.0%
57
100.0%
5
0
0
Citrobacter koseri
44
43
97.7%
44
100.0%
0
0
0
1
1
100.0%
1
100.0%
0
0
0
Enterobacter aerogenes
47
43
91.5%
46
97.9%
1
1
0
Enterobacter cloacae
43
43
100.0%
43
100.0%
9
0
0
Enterobacter cloacae cloacae
24
20
83.3%
24
100.0%
0
0
0
Enterobacter cloacae complex
19
19
100.0%
19
100.0%
0
0
0
169
163
96.4%
168
99.4%
7
1
0
48
46
95.8%
48
100.0%
3
0
0
103
97
94.2%
103
100.0%
1
0
0
Edwardsiella tarda
Escherichia coli Klebsiella oxytoca
Clinical and QC
Challenge and QC
Reproducibility
Klebsiella pneumoniae
65
63
96.9%
64
98.5%
7
1
0
VITEK®2 automatic dilution
VITEK®2 automatic dilution
VITEK®2 automatic dilution
Morganella morg. morganii
18
18
100.0%
18
100.0%
0
0
0
Morganella morganii
12
11
91.7%
12
100.0%
0
0
0
Proteus mirabilis
59
57
96.6%
59
100.0%
0
0
0
Proteus vulgaris
29
25
86.2%
29
100.0%
0
0
0
25
23
92.0%
23
92.0%
3
2
0
207
198
95.7%
200
96.6%
38
7
0
Salmonella enteritidis
7
7
100.0%
7
100.0%
0
0
0
Salmonella typhi
4
3
75.0%
4
100.0%
0
0
0
76
69
90.8%
74
97.4%
1
2
0
Shigella flexneri
1
1
100.0%
1
100.0%
0
0
0
Shigella sonnei
15
15
100.0%
15
100.0%
0
0
0
1073
1014
94.5%
1059
98.7%
75
14
0
VITEK®2 manual dilution
VITEK®2 manual dilution
3rd card
VITEK®2
VITEK®2
Yes
Compact manual
Compact manual
dilution
dilution
Providencia stuartii
Yes
No
P. aeruginosa
RESULTS
Serratia marcescens
Four gram-negative QC organisms were tested throughout comparative testing at each study site by both the VITEK2 CZA and BMD reference method containing CZA. The testing was performed a minimum of twenty times by both card and reference method at each site. Overall performance is shown in table 3 below. Table 3 : Quality Control
INTRODUCTION VITEK2 AST GN-CZA is an in vitro quantitative test device for determining the antimicrobial susceptibility of Enterobacteriaceae and P. aeruginosa against Ceftazidime / Avibactam (CZA). It determines the Minimum Inhibitory Concentration (MIC) for Ceftazidime / Avibactam. The VITEK2 Susceptibility components are antimicrobials in combination with growth medium, which are incorporated into VITEK2 card products for use with the VITEK2 system for performing antimicrobial susceptibility testing of microorganisms in clinical laboratory settings. The VITEK2 AST card consists of a maximum of 64 wells. Each well contains an aliquot of a specific concentration of antimicrobial agent. Once the cards are placed in the VITEK®2 System along with the appropriate organism suspension, no additional handling is required. Dilutions (if required) are made automatically and the cards are inoculated using a vacuum filling process. The cards are sealed and placed into the reader / incubator automatically. The VITEK®2 Compact System will require the user to manually prepare the AST dilution and transfer the AST cards between filling and loading stations. For both the VITEK®2 and VITEK®2 Compact Systems, the computer determines growth based on the attenuation of light measured by an optical scanner. The data is used to determine the MIC values for the antimicrobial agents.
Organism
Klebsiella pneum. pneumoniae
2nd card
BMD
Table 6 : Frequency table of Overall Enterobacteriaceae and P. aeruginosa
To determine the reproducibility of the VITEK2 CZA test, ten on-scale gramnegative isolates were tested in triplicate each day for three days at three clinical trial sites. Overall performance is shown in table 4 below. Table 4 :Overall Combined Reproducibility
All species combined
Although within EA when compared to the reference method, errors (major and/or very major) were observed due to the lack of intermediate category. The overall categorical major error rate for Enterobacteriaceae and P. aeruginosa combined was 1.4% (14/998). Nine (9) major errors were one dilution apart from the reference method and as such fall within EA. Based on the EA, and the lack of an intermediate breakpoint for CZA, the adjusted categorical major error rate was 0.5% (5/998). Testing should be repeated using an alternative testing/reference method prior to reporting results for P. aeruginosa when the VITEK®2 MIC is ≥16 μg/mL due to observed major errors (4.1%). Due to a low overall EA, an alternate method of testing should also be used prior to reporting results for Providencia rettgeri isolates.
CONCLUSIONS When compared to the broth microdilution reference method, VITEK2 AST GN-CZA was found to be an accurate and reliable method for susceptibility testing of CZA providing MIC results for Enterobacteriaceae and P. aeruginosa. The addition of the VITEK2 CZA test to the current VITEK2 antimicrobial susceptibility test menu increases the options available to clinical laboratories for testing ESBL-producing Enterobacteriaceae and MDR P. aeruginosa isolates.
