Rapid Diagnosis of Babesia gibsoni by Point‐of‐Need Testing by Insulated Isothermal PCR in Dogs at High Risk of Infection

Abstract

BackgroundDogs seized by law enforcement agencies during dogfighting investigations are at increased risk of Babesia gibsoni infection. A rapid and cost‐effective diagnostic test would increase the feasibility of mass screening of dogs for infection and monitoring treatment efficacy in B. gibsoni‐infected dogs.ObjectiveTo determine the performance of a point‐of‐need insulated isothermal PCR (iiPCR) test for diagnosis of B. gibsoni in dogs rescued in dogfighting investigations.AnimalsTwo hundred and thirty‐three dogs seized in dogfighting investigations.MethodsCross‐sectional study. Whole blood samples were tested for B. gibsoni and Babesia spp. by iiPCR. Results were compared to a reference standard comprised of concordant results from real‐time PCR in a commercial diagnostic laboratory and antibody titers.ResultsThe iiPCR system was quick to learn, portable, and had a short processing time of <2 hours. Sensitivity and specificity of the iiPCR assay for B. gibsoni were 90% (95% confidence interval [CI] 81–95%) and 99% (CI, 95–100%), respectively. Sensitivity and specificity of the iiPCR assay for Babesia spp. were 87% (CI, 78–93%) and 98% (CI, 0.94–99%), respectively.Conclusions and Clinical ImportanceThe iiPCR system produced few false‐positive results, indicating that positive results are likely to represent true infections when used in high‐risk animals. The iiPCR system can fail to identify 10–15% of truly infected dogs. However, the portability, speed, and economy of the iiPCR system compared to testing through a reference laboratory can allow rescue groups to screen and identify infection in more dogs.

An RT-PCR panel for rapid serotyping of dengue virus serotypes 1 to 4 in human serum and mosquito on a field-deployable PCR system

“…In this system, insulated isothermal PCR (iiPCR) is achieved consistently in a capillary tube (R-tube, GeneReach) in a simple, specially designed insulated heater and relies on fluorescent probe hydrolysis for signal detection [23,24]. Various iiPCR/RT-iiPCR-based reagents, available commercially in a lyophilized format to facilitate long-term storage and easy shipping, have been validated to have analytical and clinical performance comparable to reference methods (real time PCR, nested PCR, virus isolation) for different important microbial pathogen hosts [25][26][27][28][29][30][31]. The POCKIT device has been bundled with easy field-deployable methods for nucleic acid extraction, with potential to serve as a flexible mobile PON tool for rapid DENV detection in human and mosquito.…”