Evaluation of a novel diagnostic test for the rapid
negative exclusion of blood cultures
Andrew J Rogers1, Rachel Parker1, Matthew Dryden2, Kordo Saeed2, David Thomas2 1 Momentum Bioscience Ltd., 10
Blenheim Office Park, Long Hanborough OX29 8LN, UK.2 Hampshire Hospitals NHS Foundation Trust, Aldermaston
Rd, Basingstoke RG24 9NA, UK.
Approximately 10% of blood culture samples submitted for microbiological
testing are found to be positive. The remaining 90% of samples are
generally reported as negative after five days of incubation, during
which time clinicians commonly continue to prescribe broad spectrum
antibiotics to patients with suspected infection. This overuse of antibiotics
leads to increased pharmacy costs, extra bed space being used on
clinical wards, the promotion of antibiotic resistance and an increased risk
of antibiotic-associated disease. To minimise these collateral damages,
novel technologies are required to exclude bacterial or fungal infection in
a timely manner.
Enzymatic Template Generation and Amplification (ETGA®) is a novel
technology for the universal and rapid phenotypic detection of viable
micro-organisms by detecting the presence of bacterial or fungal nucleic
acid-modifying enzymes such as DNA polymerase (Figure 1).
Results and discussion
In total, 1,497 blood culture specimens were tested, with 1,366 specimens
being considered valid samples post-arbitration. More than half of the
specimens tested (54%) were either paediatric patients (0-5 years) or
more than 70 years of age (Figure 2).
Age distribution of tested specimens (n=1,366)
600
500
400
Count
Background
300
200
100
0
0-5
6-10
11-20 21-30 31-40 41-50 51-50 61-70
Age
70+
Figure 2. Age distribution of the blood culture specimens tested during
the study.
Specimen
Sample
Clean-up
Microbial
Lysis
Substrate
Modification
PCR
Template
Figure 1. Overview of ETGA® methodology, used in Cognitor® Minus
Study Objectives
The aim of this study was to evaluate the use of Cognitor® Minus, a
diagnostic test using ETGA® technology, for the rapid confirmation and
exclusion of negative blood culture specimens, with the aim of reducing
the time taken to report negative results.
The study was carried out at two hospital sites within the Hampshire
Hospitals NHS Foundation Trust, between December 2014 and March
2015, and compared Cognitor Minus® results to results from automated
blood culture after 5 days incubation.
Prior to clinical arbitration, the study results had a FPR of approximately
10% when compared to blood culture. However, clinical arbitration
suggested that approximately 70% of these FPs should be considered
TPs. The FPR [FP/(FP+TN)] for the dataset was re-calculated [40/
(40+1,291)] (Figure 3).
The false positive rate was calculated to 3%.
After clinical arbitration of the data, the NPV [TN/(TN+FN)] for the study
was also calculated: [1,291/(1,291+7)] (Figure 3).
The NPV was calculated to be 99.5%.
Ct cut-off >43.5 units
Study methods
This study summarises the clinical application and validation of a
diagnostic test using ETGA® technology for the rapid confirmation and
exclusion of negative blood culture specimens.
Surplus samples were obtained from both the aerobic (BacT/ALERT®
SA, bioMérieux #259789) and anaerobic (BacT/ALERT® SN, bioMérieux
#259790) adult blood culture bottle (0.5 mL/ bottle), or from a paediatric
(BacT/ALERT® PF Plus, bioMérieux #410853) blood culture bottle (1 mL),
sent routinely to the microbiology laboratory. Samples were taken from
blood culture bottles incubated on a blood culture machine (BacT/ALERT®
3D system, bioMérieux) for 12 hours or more and that were negative at the
time of sample collection.
During the study, each valid specimen was categorised as either ‘true
negative’ (TN), ‘true positive’ (TP), ‘false negative’ (FN) or ‘false positive’
(FP) based on the eventual blood culture result after 5 days.
All discrepant results (i.e. FN or FP) were arbitrated by the Clinical
Investigators for a final clinical judgement as to whether to include or
exclude the specimen. After review, Negative Predictive Value (NPV) and
false positive rate (FPR) for the study were calculated.
Cognitor result interpretation
Results from the Cognitor Minus® qPCR reaction were obtained in the
form of a Ct value automatically generated by the SmartCycler (Cepheid)
software. For the purpose of the Cognitor Minus® test, a Ct value is defined
as the point in the qPCR reaction where amplification is seen. Ct values
below a defined cut-off value are considered positive or ‘Not Determined’;
those that exceeded this Ct cut-off value (or did not amplify) were
considered ‘Negative’, and unlikely to contain a living micro-organism.
TP
28
FP
40
NPV 99.5%
[1,291/(1,291+7)]
FN
7
TN
1,291
FPR 3%
[40/(40+1,291)]
Figure 3. Final, clinically arbitrated data set showing a very high NPV
(99.5%), and a correspondingly low FPR (3%).
Conclusion
Cognitor Minus®, using ETGA® technology, was found to be an excellent
diagnostic predictor and suitable for rapidly identifying negative blood
culture specimens with an NPV of 99.5%, and thereby assisting in the
clinical decision making.
Using Cognitor Minus® in the clinical setting may:
•
•
•
•
Promote better antibiotic stewardship;
Allow cost savings from a reduction in antibiotic usage;
Reduce pressure on antibiotic resistance;
Minimise antibiotic associated disease for the patient.
Acknowledgements
Momentum Bioscience Ltd., acknowledges the support for this study from
the clinical and laboratory teams at the Department of Microbiology, Royal
Hampshire County Hospital, Winchester, the Department of Microbiology,
Basingstoke and North Hampshire Hospital, and the Hampshire Hospitals
NHS Foundation Trust.