Rapid and Visually Specific Detection of Sarcocystis miescheriana and Sarcocystis suihominis Infections in Domestic Pigs (Sus scrofa domesticus) Using Loop-Mediated Isothermal Amplification

The domestic pig (<i>Sus scrofa domesticus</i>) serves as an intermediate host for two <i>Sarcocystis</i> species: the non-zoonotic <i>S. miescheriana</i> and the zoonotic <i>S. suihominis</i>, both of which threaten animal and human health and contribute to economic losses in swine production. Existing diagnostic methods, such as microscopy and PCR, suffer from limitations regarding their sensitivity, cost, and field applicability, especially in resource-constrained settings. To address these challenges, we developed a highly specific and ultrasensitive loop-mediated isothermal amplification (LAMP) assay targeting the mitochondrial cytochrome c oxidase subunit I (<i>cox</i>1) gene for rapid detection. The optimized protocol (62 °C for 40 min) showed absolute specificity, with no cross-reactivity to related coccidia. The assay exhibited remarkable sensitivity, detecting as little as 6.7 × 10^-6 ng/μL (<i>S. miescheriana</i>) and 5.4 × 10^-7 ng/μL (<i>S. suihominis</i>), representing 10-fold and 10,000-fold improvements over conventional PCR, respectively. With a simple visual readout, the proposed LAMP assay eliminates the need for sophisticated equipment, making it an ideal field-deployable diagnostic tool for basic laboratories and under-resourced regions.