streptococcus equi field trial data

Maine Biotechnology Services, Inc. 2014

Streptococcus equi “Strangles” Rapid

Stall Side Test

Release of Field Trial Data


Development of Stall Side Test

Resulted from multi-directional collaborationsUMO, U-Kentucky, MBS, Diagnostic Innovations

Address need for rapid identification of Strangles (+) horsesEnabling Barn Manager to take appropriate

actions days before PCR results available. Follow up with confirmatory PCR expected.


Prototype Rapid Test Developed

Sensitivity Quantitative PCR done on Strep equi culture Enabled assessment of sensitivity Artificial swabs with known bacterial counts

tested Limit of sensitivity ~ 50 CFU (colony forming

units) per strip Clinical target: 1000 organisms, as recommended

by Dr. John Timoney University of Kentucky


Sensitivity:


Prototype Rapid Test Developed

SpecificityBank of Strep equi and Strep zoo clinical

isolates provided by Dr. Causey Isolates were tested and compared the signal

to the CF32 Strep equi reference strain at 16 cells per strip.


The data on clinical isolates supports specificity for Strep equi with no cross reactivity to Strep zoo isolates.

Test against known CFU of Strep equi. UMO provided known concentrations of

Strep equi via the CFU metric Provided both bacteria in water and in saline

To evaluate possible effect of saline or water nasal flushes

Results show detection limit of 50-500 CFULimit of detection needed for clinical application

reported to be 1000 CFU Dr. John Timoney, University of Kentucky


Summary:Specificity metrics show no cross reactivity to the Strep zoo ancestor of Strep equi subsp. equiSensitivity as determined by

Quantitative PCR CFU Indicates that the prototype can detect 50-100 CFU


Nasal Swabs taken from 10 healthy horses.

UMO performed nasal swabs (2 per horse) on 10 horses documented as healthy

Prototype Strep equi test run to validate no cross reactivity with equine nasal flora

Negative swabs were spiked to assess interference with nasal natural flora


Results

Diagnostic Innovations


Summary Monoclonal antibodies to Strep equi Prototype rapid assay designed Strep equi CF32 reference strain Q-PCR Strep equi and zoo clinical isolates

specificity. Nasal swabs from healthy horses tested Spiked nasals swabs tested Ready to take the rapid test to the field.


Test Components

Diagnostic Innovations


Clinical Trial, 23 horses in Maine December 2013, barn in Cumberland

Maine was stricken with a highly contagious respiratory disease

May have been started via purchase of a yearling in Texas that showed signs of a mild respiratory illness.

Attending Veterinarian, Dr. Martha Smith


Clinical Trial: Compare PCR to rapid stall side assay 23 horses dual nasal swabs

1 sent to UC Davis for Quantitative PCR1 run on Rapid Stall side assay

Health histories during outbreak recorded. Random selection of samples also tested

for Streptococcus zooepidemicus. Ancestor of Streptococcus equi. subsp. equi


Diagnostic Test Metrics:• Specificity

•refers to the cross reactivity profile of the test

• Sensitivity

•refers to the lowest detectable load of bacteria

• Positive Predictive Value (PPV)

•is the proportion of patients with positive results, who are correctly diagnosed

• Negative Predictive Value (NPV)

•is the proportion of patients with negative results, who are correctly diagnosed.


Results:Disease + Disease - Total

Test + 5 (TP) 2 (FP) 7 (PPV Denominator)Test - 0 (FN) 16 (TN) 16 (NPV Denominator)

5 18 23

Positive Predictive Value: TP / (TP + FP) x 100 = 72%

Negative Predictive Value: TN / (TN + FN) x 100 = 100%

Sensitivity: TP / (TP + FN ) x 100 = 100 %

Specificity : TN / (TN + FP) x 100 = 88%

Take away message from this small clinical trial is: The test may have false positives, but it does not show the propensity for false negatives. It errs on

the side of safety for the barn managers with a 100% NPV and 100% Sensitivity metric.

Both (–) Strep zoo


Questions for End User How might you incorporate this test in your

practice What rapid test attributes are most

important to you? (cost, ease of use, time, et al..)

Can you identify any weaknesses for the commercialization of this test?

Questions, concerns?

streptococcus equi field trial data

Science