Development of a low-cost copro-LAMP assay for simultaneous copro-detection of Toxocara canis and Toxocara cati

• Published in: Parasitology
• Main Authors: Avila, Héctor Gabriel, Risso, Marikena Guadalupe, Ruybal, Paula, Repetto, Silvia Analía, Butti, Marcos Javier, Trangoni, Marcos David, Grune Löffler, Sylvia, Pérez, Verónica Mirtha, Periago, María Victoria
• Format: Article in Journal/Newspaper
• Language: English
• Published: Cambridge University Press (CUP) 2021
• Subjects: Canis lupus
• Online Access: http://dx.doi.org/10.1017/s0031182021000342; https://www.cambridge.org/core/services/aop-cambridge-core/content/view/S0031182021000342

Abstract Toxocariasis is a zoonotic disease caused mainly by Toxocara canis and Toxocara cati and diagnosis in dogs and cats is an important tool for its control. For this reason, a new coprological loop-mediated isothermal amplification (LAMP) assay was developed for the simultaneous detection of these species. The primer set was designed on a region of the mitochondrial cox -1 gene. Amplification conditions were evaluated using a temperature gradient (52°C to 68°C), different incubation times (15–120 min), and different concentrations of malachite green dye (0.004–0.4% w/v). The analytical sensitivity was evaluated with serial dilutions of genomic DNA from T. canis and T. cati adult worms, and with serial dilutions of DNA extracted from feces using a low-cost in-house method. The specificity was evaluated using genomic DNA from Canis lupus familiaris , Felis catus , Escherichia coli , Toxascaris leonina , Ancylostoma caninum , Echinococcus granulosus sensu stricto and Taenia hydatigena . The LAMP assay applied to environmental fecal samples from an endemic area showed an analytical sensitivity of 10–100 fg of genomic DNA and 10 −5 serial dilutions of DNA extracted from feces using the low-cost in-house method; with a specificity of 100%. Additionally, the total development of the assay was carried out in a basic laboratory and per-reaction reagent cost decreased by ~80%. This new, low-cost tool can help identify the most common agents of toxocariasis in endemic areas in order to manage prevention strategies without having to rely on a laboratory with sophisticated equipment.