1
Rapid Detection of Gram-Positive Organisms Using the Verigene Gram-Positive Blood 1
Culture Nucleic Acid Test and the BacT/ALERT Pediatric FAN System in a 2
Multi-Centered Pediatric Evaluation 3
4
K. V. Sullivan1,2,# 5
N. N. Turner1 6
S. S. Roundtree1 7
S. Young3 8
C. A. Brock-Haag3 9
D. Lacey3 10
S. Abuzaid3 11
D. L. Blecker-Shelly1 12
C. D. Doern3,4 13
14 1 The Children’s Hospital of Philadelphia, Philadelphia, USA 15 2 Perelman School of Medicine, University of Pennsylvania, Philadelphia, USA 16 3 Children’s Medical Center of Dallas, Dallas, Texas, USA 17 4 University of Texas Southwestern, Dallas, Texas, USA 18 19 Running title: Rapid Blood Culture Testing in Pediatrics 20 21 Keywords: Pediatric, Rapid Testing, Bacteremia, Diagnosis 22 23
#Corresponding Author: 24
Dr. Kaede V. Sullivan, M.D., M.Sc. 25 The Children’s Hospital of Philadelphia 26 34th Street and Civic Center Boulevard 27 Main Building, Room 5112A 28 Philadelphia, Pennsylvania, USA 19104-4399 29 Tel: 215-983-6792 30 Fax: 215-590-2171 31 E-mail: [email protected] 32 33
JCM Accepts, published online ahead of print on 21 August 2013J. Clin. Microbiol. doi:10.1128/JCM.01224-13Copyright © 2013, American Society for Microbiology. All Rights Reserved.
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Abstract 34
Assays that expedite reporting of organism identification and antibiotic susceptibility status in 35
positive blood cultures can fast-track interventions that improve clinical outcomes. We evaluated 36
the Verigene Gram-Positive Blood Culture Nucleic Acid Test (BC-GP) in two pediatric 37
hospitals. Positive BacT/ALERT Pediatric FAN blood cultures with Gram-positive organisms 38
were tested using BC-GP in tandem with routine laboratory procedures. To test organisms 39
underrepresented in the clinical blood culture evaluation, blood culture bottles were spiked with 40
diluted organism suspensions at concentrations of 10-100 colony forming units per milliliter. 249 41
Gram-positive bacterial isolates were recovered from 242 blood cultures. BC-GP detected S. 42
aureus, methicillin-susceptible S. aureus, and methicillin-resistant S. aureus with sensitivity of 43
100%, 99% and 100%; and specificity of 100%, 100% and 99.5% respectively. BC-GP detected 44
S. epidermidis, methicillin-susceptible S. epidermidis and methicillin-resistant S. epidermidis 45
with sensitivity of 95%, 80% and 96% respectively and 100% specificity. BC-GP correctly 46
identified 14/15 cases of E. faecalis and E. faecium bacteremia and 9 cases with S. pneumoniae. 47
It misidentified 5/15 clinical blood cultures with S. mitis/oralis and 1/3 blood cultures spiked 48
with Streptococcus anginosus Group as S. pneumoniae. BC-GP detected a case of S. pyogenes 49
bacteremia but failed to detect 2/3 clinical blood cultures with S. agalactiae. BC-GP’s rapid and 50
accurate detection of Staphylococcus spp., E. faecium and E. faecalis, and its ability to ascertain 51
mecA, vanA and vanB status may expedite clinical decisions pertaining to optimal antibiotic use. 52
False-positive S. pneumoniae results may warrant reporting of “Streptococcus spp.” only when 53
this organism is reported by BC-GP. 54
55
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Background 58
Laboratories typically require 24-48 hours to determine the identity and antimicrobial 59
susceptibility of an agent of bacteremia following the signal of a positive blood culture in a 60
continuous monitoring system. Rapid testing that expedites this process can fast-track 61
interventions that improve clinical outcomes. 62
As in adults, S. aureus bacteremia is associated with high risk of mortality and embolic 63
complications in children (1-3). Timely removal of central venous catheters (1) and expedited 64
initiation of optimized antibiotics (ex. nafcillin, oxacillin or cefazolin in methicillin-susceptible 65
S. aureus bacteremia) has been associated with a reduction in treatment failures and hospital 66
costs (4-6). 67
Similarly, delayed initiation of targeted enterococcal therapy has been associated with 68
increased length of hospital stay and mortality, regardless of vancomycin susceptibility status (7 69
-9). Rapid detection of Enterococcus spp. from positive blood cultures may expedite therapeutic 70
modifications required to optimally treat enterococcal bacteremia (9). These include the addition 71
of an aminoglycoside agent to beta-lactam or glycopeptide therapy for synergy to compensate for 72
the inherently poor response of Enterococcus spp. when they are administered alone. Moreover, 73
treatment of vancomycin-resistant enterococcal (VRE) bacteremia requires initiation of linezolid 74
or quinupristin-dalfopristin. 75
By contrast, literature derived from implementation of rapid molecular assays have 76
suggested that expedited identification of coagulase-negative Staphylococcus spp. (CoNS) from 77
positive blood cultures coupled with pharmacy intervention can shorten length of hospital stay 78
and lower hospital costs by mitigating initiation of unnecessary antibiotics (10). 79
The Verigene Gram-Positive Blood Culture Nucleic Acid Test (BC-GP; Nanosphere, 80
Northbrook, IL) is a rapid, FDA-cleared, microarray assay that detects clinically relevant Gram-81
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positive organisms directly from positive blood cultures. It uses an automated, 2.5-hour format. 82
This study examined the diagnostic performance of BC-GP in two tertiary, pediatric populations 83
using the BacT/ALERT Pediatric FAN blood culture system (bioMérieux, Durham, NC). 84
Results were presented at the 113th American Society for Microbiology General Meeting in 85
Denver, Colorado. 86
Methods 87
Clinical blood culture specimens 88
Positive clinical BacT/ALERT Pediatric FAN blood culture bottles with Gram-positive 89
organisms on Gram stain in the microbiology laboratories at The Children’s Hospital of 90
Philadelphia (CHOP) and at Children’s Medical Center of Dallas (CMCD) were included in this 91
study. The study period was September, 2012 - March, 2013. 92
Verigene BC-GP 93
BC-GP was performed on all qualifying blood cultures according to manufacturer’s 94
instructions. This assay uses a microarray format with probes that detect tuf gene targets for 95
genus-level detection of Staphylococcus spp., Streptococcus spp., and Listeria spp., and various 96
gene targets for species-level detection of the following: gyrB for S. aureus, S. anginosus Group, 97
and S. pneumoniae; hsp60 for S. epidermidis, E. faecalis, E. faecium, S. agalactiae, and S. 98
pyogenes; and sodA for detection of S. lugdunensis. Additional probes detect mecA in S. aureus 99
and S. epidermidis; and vanA and vanB in E. faecium and E. faecalis. 100
Conventional identification and susceptibility testing 101
Reference standard genus/species identification of organisms isolated from clinical blood 102
cultures was performed using conventional identification tests and the Vitek 2 system 103
(bioMérieux, Durham, NC) at CHOP and Matrix-assisted laser desorption-ionization time-of-104
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flight mass spectrometry (MALDI-TOF MS, BioTyper 3.0, Bruker Daltonics, Billerica, MA) at 105
CMCD. An internal MALDI-TOF MS validation study at CMCD included all species present in 106
the BC-GP panel. In the validation study, MALDI-TOF MS was found to accurately identify all 107
Gram-positive species except Streptococcus pneumoniae and the Streptococcus mitis/oralis 108
group. S. pneumoniae and viridans group streptococci were therefore identified using colonial 109
morphology, Gram stain, catalase, optochin susceptibility and bile solubility status at both sites. 110
Vitek 2 and the MicroScan WalkAway system (Siemens Healthcare Diagnostics, Deerfield, IL) 111
were used for species-level identification of streptococcal isolates at CHOP and CMCD 112
respectively. Methicillin susceptibility in Staphylococcus spp. and vancomycin susceptibility in 113
E. faecium and E. faecalis were determined using Vitek 2 at CHOP and the MicroScan 114
WalkAway system at CMCD. At CMCD, organism identification and susceptibility testing is not 115
routinely pursued when coagulase-negative Staphylococcus spp. (CoNS) is isolated in a single 116
occurrence. BC-GP was performed on these blood cultures but they were analyzed separately. 117
Spiked blood culture specimens 118
Spiked BacT/ALERT Pediatric FAN blood culture bottles were prepared to evaluate BC-119
GP’s ability to detect organisms on the BC-GP microarray that were underrepresented in the 120
clinical blood cultures evaluated. Starting with a 1 MacFarland Standard suspension of organism, 121
three successive 100-fold dilutions were performed by placing 10 µL of suspension into 990 µL 122
of diluent. This was followed by a 30-fold dilution by placement of 100 µL of suspension into a 123
blood culture bottle with 3 mL of banked blood to produce a final organism concentration 124
between 10-100 colony forming units/milliliter. Colony counts were performed prior to the final 125
1:30 dilution to verify organism concentration (11). Clinical and ATCC strains were used. 126
Clinical strains included: S. lugdunensis (1), vancomycin-resistant E. faecium (6), vancomycin-127
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susceptible E. faecium (1), L. monocytogenes (2), S. agalactiae (10), S. pyogenes (10), S. 128
constellatus (2), S. intermedius (1), S. mitis/oralis (2), S. sanguis (1), and S. parasanguinis (1). 129
ATCC strains included: Enterococcus faecalis ATCC 51299 (VRE), Enterococcus faecium 130
ATCC 51559, Enterococcus avium ATCC 14025, Streptococcus mutans ATCC 35668, 131
Streptococcus bovis ATCC 9809, Streptococcus agalactiae ATCC 12386, and Streptococcus 132
pyogenes ATCC 19615. 133
We compared the proportion of tests with inaccurate BC-GP results at the two centers 134
and also, the proportion of tests with false-negative results using the Chi-Square Test for 135
proportions and SAS statistical software. 136
Results 137
Results from clinical and spiked blood culture testing are summarized in Tables 1 and 2, 138
respectively. Bacterial isolates recovered from 242 clinical blood cultures (166 from CHOP and 139
76 from CMCD) underwent organism identification and susceptibility testing. 249 Gram-positive 140
bacterial isolates were recovered. 106 S. aureus isolates (85 methicillin-susceptible, 21 141
methicillin-resistant) and 61 S. epidermidis isolates (5 methicillin-susceptible, 56 methicillin-142
resistant) were identified. BC-GP detected S. aureus, MSSA, and methicillin-resistant S. aureus 143
(MRSA) with sensitivity of 100%, 99% and 100%; and specificity of 100%, 100% and 99.5% 144
respectively. BC-GP detected S. epidermidis, methicillin-susceptible S. epidermidis and 145
methicillin-resistant S. epidermidis with sensitivity of 95%, 80% and 96% respectively and 100% 146
specificity. At CMCD, BC-GP appropriately reported “Staphylococcus spp.” or “S. 147
epidermidis” for all 34 “single-positive” CoNS isolates not identified to the species level. 148
BC-GP correctly detected E. faecalis or E. faecium to the species level in 19 of 20 149
clinical blood cultures that grew these organisms but failed to detect E. faecalis in a case of 150
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mixed E. faecalis and E. faecium bacteremia. A repeat test also failed to detect E. faecalis. BC-151
GP also correctly detected vanA in the only clinical case of VRE bacteremia encountered in the 152
study. BC-GP also detected 9/9 blood cultures spiked with E. faecalis or E. faecium, and 153
reported vanA or vanB positivity in 7/7 blood cultures bottles inoculated with VRE. One blood 154
culture spiked with E. avium (ATCC 14025) was reported by BC-GP as E. faecium. 155
BC-GP correctly identified 9 cases of S. pneumoniae bacteremia, but misidentified 5/15 156
clinical blood cultures with S. mitis/oralis and 1/3 blood cultures spiked with S. anginosus Group 157
as S. pneumoniae. BC-GP detected a case of S. pyogenes bacteremia but failed to detect 2/3 158
clinical blood cultures with S. agalactiae. In one case, BC-GP failed to detect both the tuf and 159
hsp60 targets. Repeat testing yielded positive results for both targets. The second case was a 160
mixed blood culture that grew S. agalactiae and MSSA. BC-GP detected both the tuf and gyrB 161
targets required to report MSSA and Streptococcus spp. but it failed to detect the hsp60 target 162
needed to report the presence of S. agalactiae. Repeat testing yielded the same result. BC-GP 163
detected 11/11 blood cultures spiked with S. pyogenes and 11/11 cultures spiked with S. 164
agalactiae. Ten blood cultures grew more than one organism and are summarized in Table 3. 165
Three cultures grew pathogens that should have been detected by the BC-GP microarray but 166
were falsely negative (CMC036, CMC041 and CMC088). Also, a culture with MSSA, S. 167
saprophyticus and S. hominis was reported as mecA-positive S. aureus, implying MRSA 168
bacteremia. 169
BC-GP reported invalid (“No-call”) results in 7 of the 249 (2.8%) clinical blood cultures 170
analyzed. Two were due to internal control failure (Micrococcus spp., S. epidermidis) and four 171
were the result of variation in the target-specific signals (Rothia mucilaginosa (3), Micrococcus 172
sp. (1), and S. warneri (1)). 173
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BC-GP reported incorrect results in 4/166 (2%) at CHOP and 4/83 (5%) at CMCD 174
(p=0.45), and failed to detect a pathogen represented on the microarray in 6/166 (4%) blood 175
cultures from CHOP and 4/83 (5%) from CMCD (p=0.74). Post hoc power analysis yielded 39% 176
power for these analyses. 177
Discussion 178
The Verigene BC-GP assay detects a wide spectrum of Gram-positive organisms that are 179
highly relevant in the management of pediatric bacteremia. By using a rapid, automated format, 180
BC-GP is designed to expedite provision of actionable results to clinicians. This multi-centered 181
study is the first to evaluate its performance in conjunction with an automated pediatric blood 182
culture system. 183
In our pediatric cohort, BC-GP detected MSSA bacteremia from positive blood cultures 184
with sensitivity of 99% and specificity of 100% suggesting that de-escalation of vancomycin 185
(commonly used in various empiric antimicrobial therapy regimens) based on BC-GP to a 186
narrower anti-staphylococcal agent, as recommended by various national guidelines, could 187
proceed safely (12, 13). This is consistent with published data from adult populations (14, 15). 188
In one mixed culture with MSSA, S. saprophyticus, and S. hominis, BC-GP appropriately 189
detected the Staphylococcus spp., and S. aureus targets but also reported detection of mecA. 190
Because the BC-GP algorithm only reports mecA status in S. aureus and S. epidermidis, for 191
analysis, the mecA result was interpreted as a false-positive mecA result. 192
We tested a combined total of 110 “single-positive” (n=34) and fully characterized 193
(n=76) pediatric blood cultures that grew CoNS. BC-GP detected CoNS with sensitivity of 96% 194
and specificity of 100%. Among the 58 blood cultures that BC-GP reported S. epidermidis, BC-195
GP correctly assigned mecA status in all 58 cases. Therefore, in instances where S. epidermidis 196
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bacteremia is considered significant (ex. central line associated bloodstream infections), use of 197
BC-GP mecA results to guide therapeutic decisions appears safe and appropriate. 198
In our cohort, BC-GP detected E. faecium and E. faecalis in 19 of the 20 instances in 199
which either pathogen was recovered from clinical blood cultures. BC-GP also detected vanA or 200
vanB in all clinical and spiked blood cultures with VRE, supporting the use of BC-GP for this 201
indication. 202
Three clinical blood cultures with S. mitis/oralis and a blood culture spiked with S. 203
constellatus tested positive for S. pneumoniae, suggesting that the gyrB target used to detect S. 204
pneumoniae may lack specificity. This phenomenon has been reported in the literature (14, 15). 205
An option is to report BC-GP S. pneumoniae results as “S. pneumoniae/viridans group 206
Streptococcus spp. Further identification to follow” while awaiting the results of routine testing 207
(14). 208
In two cases of S. agalactiae bacteremia, BC-GP failed to detect the hsp60 target, and in 209
one case, it also failed to detect the tuf target, suggesting insufficient sensitivity of BC-GP to 210
detect this pathogen. Additional data evaluating the sensitivity of BC-GP in the detection of S. 211
agalactiae would be beneficial. Because specificity of BC-GP appeared satisfactory for the 212
detection of S. agalactiae and S. pyogenes, it seems appropriate to report positive S. pyogenes 213
and S. agalactiae results as is. 214
We recovered 17 clinical isolates that were not represented on the BC-GP microarray. 215
Three blood cultures with Rothia spp. and one culture with Micrococcus spp. were reported by 216
BC-GP as “no call – variation”. Each target is spotted three times on the BC-GP microarray. 217
Target signals, when positive, are expected to be within a consistent range of intensity across the 218
three spots. The background of the slide is also expected to elicit a uniform, albeit weaker, signal 219
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across the surface of the slide. Variability in the target and/or background signal elicits a “no-call 220
variation” report. While we are uncertain about the cause of these results, we hypothesize that 221
the microarray slides may not have been adequately rinsed after removal from the Processor and 222
prior to insertion into the Reader instrument. However, if this was the case, we would have 223
expected more systematic reports of “no-call variation”. Another explanation may have been 224
partial, but irregular binding of nucleic acid from these organisms to the microarray targets. 225
Mixed blood cultures have been reported to be associated with higher rates of discordant 226
results between BC-GP and reference methods (14, 15). Mixed cultures accounted for 4% 227
(10/242) of tests analyzed. Discordance between reference and BC-GP testing occurred in 30% 228
(3/10) of polymicrobial cultures and in 6% (14/232) of monomicrobial blood cultures. It is 229
difficult to make definitive conclusions based on the small number of mixed blood cultures 230
encountered in this study. 231
An aspect of the BC-GP algorithm that complicates reporting of mixed cultures with S. 232
aureus and coagulase-negative Staphylococcus spp. is the suppression of the mecA result (14, 15) 233
in species other than S. aureus and S. epidermidis. BC-GP reports of mixed cultures with MSSA 234
and methicillin-resistant S. epidermidis are ambiguous as it is impossible to ascertain whether the 235
positive mecA result originated from S. aureus or S. epidermidis. This ambiguity may lead to 236
missed opportunities to initiate narrow-spectrum anti-staphylococcal therapy to treat MSSA 237
bacteremia. Secondly, BC-GP will report a mixed culture with MSSA and S. lugdunensis with 238
negative mecA status regardless of the methicillin susceptibility status of the latter. Suppression 239
of the mecA status could therefore result in the erroneous assumption that both pathogens are 240
mecA-negative. Finally, in any blood culture with S. aureus, S. lugdunensis, or S. epidermidis, 241
BC-GP reports detection of both Staphylococcus spp. and the species-level target. For example, 242
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a culture with S. aureus will be reported by BC-GP as positive for Staphylococcus spp. and S. 243
aureus. In these cases, the presence of a mixed infection with a Staphylococcus spp. other than 244
the three aforementioned species would be impossible to discern. One way to mitigate confusion 245
would be to report all blood cultures with Staphylococcus spp. with the following disclaimer: 246
“Cannot exclude mixed culture with coagulase-negative Staphylococcus spp.” 247
Other assays with the capability of detecting pathogens directly from positive blood 248
cultures have received approval from the Food and Drug Administration. The BD GeneOhm 249
Staph SR (BD GeneOhm, San Diego, CA) and the currently unavailable Cepheid Xpert 250
MRSA/SA BC (Cepheid Diagnostics, Inc., Sunnyvale CA) are examples. However, clinical data 251
generated after FDA approval revealed cases of MRSA bacteremia testing falsely as MSSA by 252
both assays. This phenomenon was attributed to failure of the assay primers to detect variants of 253
SCCmec (16). Instances of MSSA testing positive for MRSA have also been reported. This 254
observation has been attributed to amplification of the SCCmec target in the absence of the mecA 255
gene (the “empty cassette” or “mecA dropout” phenomenon) and supports the inclusion of a 256
mecA target in the design of assays that detect MSSA and MRSA as Nanosphere has done with 257
their BC-GP assay (17). Matrix-assisted laser desorption ionization time-of-flight mass 258
spectrometry (MALDI-TOF MS) may also accurately detect various pathogens directly from 259
positive blood cultures (18, 19). However, MALDI-TOF MS is not FDA-cleared nor capable of 260
reliably performing antibiotic susceptibility testing at this time. 261
This study had several limitations. Consistent with other studies in the literature, the 262
majority (73%) of the organisms recovered in our clinical blood cultures were Staphylococcus 263
spp. (14, 15). We attempted to address this through the preparation of spiked blood cultures 264
inoculated with Enterococcus and Streptococcus spp. Second, the charcoal present in 265
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BacT/ALERT PF bottles could have theoretically affected the performance of BC-BG by 266
interfering with nucleic acid binding to magnetic beads during DNA extraction, resulting in 267
false-negative BC-GP results. Comparison of false-negative rates in charcoal-containing bottles 268
with those in bottles containing antibiotic binding resins, or no antibiotic removal device, would 269
be valuable future avenues of investigation. 270
Finally, the two study centers used different automated systems for “reference standard” 271
organism identification and antibiotic susceptibility testing. CHOP relied on the Vitek 2 system 272
while CMCD used MALDI-TOF MS for organism identification and the Microscan Walkaway 273
system for antibiotic susceptibility testing. Given that the Vitek 2 and Microscan Walkaway 274
instruments have met FDA approval, we felt confident in their ability to serve as reference 275
standards in this study. The MALDI-TOF MS instrument at CMCD was extensively validated 276
prior to use with both clinical and reference strains (20). There was no statistically significant 277
difference in “incorrect” BC-GP identifications between the centers although this study was 278
somewhat underpowered for this post-hoc analysis. 279
In summary, there is considerable need for accurate, reliable, and rapid assays that 280
provide actionable organism identifications and susceptibility testing results from positive blood 281
cultures. These data show that the Verigene Gram-Positive Blood Culture Nucleic Acid Test is a 282
viable option for testing of positive pediatric blood cultures. BC-BP accurately determined mecA 283
status in S. aureus and S. epidermidis; and vancomycin resistance in E. faecium and E. faecalis. 284
Further research and development aimed at reducing false-positive S. pneumoniae and false-285
negative S. agalactiae results would be beneficial. 286
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Acknowledgements 289
We thank the microbiology laboratory staff at The Children’s Hospital of Philadelphia and 290
Children’s Medical Center of Dallas for their assistance with blood culture processing. We also 291
thank Dr. Laura Chandler, Alpa Shah and Maria Dirkes for their assistance with the blood 292
culture spiking. Verigene Gram-Positive Blood Culture Nucleic Acid Test consumables were 293
donated in kind by Nanosphere, Inc. 294
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Table 1. Performance of Verigene BC-GP Assay Compared to Standard Procedures in Positive Clinical Blood Cultures
No. organisms on BC-GP panel isolated
No. (%) isolates correctly identified1
No. (%) isolates incorrectly identified
No. (%) isolates not detected2
CHOP3 CMCD4 Total CHOP CMCD Total CHOP CMCD Total CHOP CMCD Total S. aureus
methicillin-susceptible 61 24 85 61/61 (100%)
23/245 (96%)
84/85 (99%) 0 (0%) 1/24
(4%) 0 (0%) 0 (0%) 0 (0%) 0 (0%)
methicillin-resistant 10 11 21 10/10 (100%)
11/11 (100%)
21/21 (100%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%)
S. epidermidis
methicillin-susceptible 4 1 5 4/4 (100%)
0/1 (0%)
4/5 (80%) 0 (0%) - 0 (0%) 0 (0%) 1/1
(100%) 1/5
(20%)
methicillin-resistant 50 6 56 48/506 (96%)
6/6 (100%)
54/56 (96%)
1/50 (2%) 0 (0%) 1/56
(2%) 1/50
(2%) 0 (0%) 1/56 (2%)
Other CoNS 13 2 15 12/137 (92%)
2/2 (100%)
14/15 (93%) 0 (0%) - 0 (0%) 1/13
(8%) 0 (0%) 1/15 (7%)
E. faecium
vancomycin-susceptible 1 3 4 1/1 (100%)
3/3 (100%)
4/4 (100%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%)
vancomycin-resistant 1 - 1 1/1 (100%) - 1/1
(100%) 0 (0%) - 0 (0%) 0 (0%) - 0 (0%)
E. faecalis
vancomycin-susceptible 7 8 15 7/7 (100%)
7/88 (88%)
14/15 (93%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 1/8
(13%) 1/15 (7%)
S. pneumoniae 3 6 9 3/3 (100%)
6/6 (100%)
9/9 (100%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%)
S. anginosus Group - 1 1 - 1/1 (100%)
1/1 (100%) - 0 (0%) 0 (0%) - 0 (0%) 0 (0%)
Other viridans group streptococci 5 10 15 2/5
(40%) 8/10
(80%) 10/159 (66%) 3/5 (60%) 2/10
(20%) 5/15
(33%) 0 (0%) 0 (0%) 0 (0%)
S. pyogenes - 1 1 - 1/1 (100%)
1/1 (100%) 0 (0%) 0 (0%) 0 (0%) - 0 (0%) 0 (0%)
S. agalactiae - 3 3 - 1/3 (66%)
1/3 (66%) - 1/3
(33%) 1/3
(33%) - 1/3 (33%)
1/3 (33%)
Listeria monocytogenes - 1 1 - 1/1 (100%)
1/1 (100%) - 0 (0%) 0 (0%) - 0 (0%) 0 (0%)
Other Gram-positive organisms10 11 6 17 7/11
(36%) 5/6
(83%) 12/17 (69%) 0 (0%) 0 (0%) 0 (0%) 4/11
(36%) 1/5
(20%) 5/17
(29%)
Total 166 83 249 156/166 (94%)
75/83 (90%)
231/249 (93%)
4/166 (2%)
4/83 (5%)
8/249 (3%)
6/166 (4%)
4/83 (5%)
10/249 (4%)
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1. “Corrected identified” by BC-GP was defined as concordance between standard procedures and BC-GP result for genus, species and mecA or vanA/vanB status for S. aureus, S. epidermidis, S. lugdunensis, E. faecium, and E. faecalis; genus and species for S. pneumoniae, S. anginosus Group, S. pyogenes, S. agalactiae; “Staphylococcus spp.” for other coagulase-negative Staphylococcus spp., “Streptococcus spp.” for other viridians group Streptococcus spp., and “Listeria spp.” for L. monocytogenes.
2. “Not detected” by BC-GP was defined as a BC-GP result that was negative or a “no-call” due to internal control failure or signal variation.
3. CHOP = The Children’s Hospital of Philadelphia
4. CMCD = The Children’s Medical Center of Dallas
5. Standard procedures yielded methicillin-susceptible S. aureus, S. saprophyticus and S. hominis. BC-GP reported S. aureus, positive mecA.
6. Standard procedures yielded methicillin-resistant S. epidermidis. BC-GP reported “Staphylococcus” (coagulase-negative Staphylococcus) in one case, a “No-call” due to internal control failure in the other.
7. Standard procedures yielded identification of S. warneri. BC-GP reported “No-call” due to signal variation.
8. Standard procedures yielded a mixed culture with E. faecalis and E. faecium (both vancomycin-susceptible). BC-GP reported E. faecium but not E. faecalis.
9. Standard procedures yielded identification of S. mitis/oralis. BC-GP reported S. pneumoniae.
10. “Other Gram-positive organisms” included Bacillus spp. (1), Corynebacterium spp. (2), Micrococcus spp. (9), and Rothia spp. (3) and E. gallinarum (1).
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Table 2. Performance of Verigene BC-GP assay compared to standard procedure in spiked blood cultures
1BC-GP identified one isolate as S. pneumoniae and reported the second as a “no-call” due to internal control failure twice.
3BC-GP reported one isolate with a report of “no-call” due to internal control failure. Repeat testing resulted in a correct result.
3BC-GP reported a blood culture spiked with E. avium as E. faecium.
Organism No. isolates tested
No. isolates correctly identified
No. isolates incorrectly identified
No. isolates not detected
S. lugdunensis 1 1 0 0
E. faecium
Vancomycin-susceptible
2 2 0 0
Vancomycin-resistant
6 6 0 0
E. faecalis
Vancomycin-resistant
1 1 0 0
S. anginosus Group1 3 1 1 1
Other viridians group streptococci
6 6 0 0
S. pyogenes 11 11 0 0
S. agalactiae 11 11 0 0
Listeria monocytogenes
2 2 0 0
Other Gram-positive organisms1
1 0 1 0
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Table 3. Performance of Verigene BC-GP in Mixed Cultures
Blood culture
Organisms isolated BC-GP result BC-GP Errors
CHOP093 S. aureus (methicillin-susceptible) K. oxytoca
S. aureus, mecA-negative None.
CHOP148 S. epidermidis (methicillin-resistant) S. hominis (methicillin-resistant)
S. epidermidis, mecA-positive None.
CHOP149 S. epidermidis (methicillin-resistant) S. hominis (methicillin-resistant)
S. epidermidis, mecA-positive None.
CMC027 S. bovis Salmonella spp.
Streptococcus spp. None.
CMC036 S. agalactiae S. aureus (methicillin-susceptible)
S. aureus, mecA-negative S. agalactiae not detected.
CMC041 E. faecalis, E. faecium E. faecium E. faecalis not detected.
CMC073 S. aureus (methicillin-susceptible) S. saprophyticus (methicillin-resistant) S. hominis (methicillin-susceptible)
S. aureus, mecA-positive False positive mecA result.
CMC082 S. mitis/oralis Abiotrophia
Streptococcus spp. None
CMC088 S. aureus (methicillin-susceptible) S. epidermidis (methicillin-susceptible)
S. aureus, mecA-negative S. epidermidis not detected.
CMC108 E. faecium S. lutentiensis E. coli
E. faecium, Streptococcus spp. None
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