One-pot nanoflower-based sensitive colorimetric biosensor for multihost detection of zoonotic clonorchiasis.

Clonorchis sinensis, a significant but frequently overlooked foodborne zoonotic helminth, is responsible for infecting approximately 15 million individuals globally, poses an infection risk to nearly 200 million people, and represents a threat to public health while imposing considerable economic burdens. The broad diversity and widespread distribution of definitive hosts significantly contribute to the sustained prevalence of clonorchiasis, while the diagnostic challenges are further exacerbated by the intricate variability in host species and their geographical dispersion, alongside the absence of accurate multihost detection techniques, ultimately driving increasing disease incidence. In this study, we fabricated a one-pot nanoflower-based sensitive competitive ELISA (nano-cELISA) for the multihost detection of clonorchiasis. The functional organic-inorganic hybrid nanoflowers retain the ability to specifically bind to the C. sinensis tandem repeat sequence 1 (CSTR1) antigen and leverage the catalytic function of horseradish peroxidase, thereby facilitating signal amplification while ensuring specificity. The detection performance of the colorimetric biosensor was systematically assessed via multihost serum samples (humans, dogs and mice). Seroconversion in artificially infected mice and dogs occurred at 21 days post-infection (dpi), indicating high specificity without cross-reactivity with sera from other parasitic infections. The nano-cELISA demonstrated a sensitivity of 100% and a specificity of 91.67% in clinical serum samples from dogs and 95.49% sensitivity and 96.72% specificity in human samples. The developed nano-cELISA method has straightforward preparation and operational simplicity, representing a useful tool for epidemic containment and offering a robust technical strategy for the prevention and control of C. sinensis-infection.