Development and application of DNA-aptamer-coupled magnetic beads and aptasensors for the detection of Cryptosporidium parvum oocysts in drinking and recreational water resources

Abstract

Environmentally stable and disinfectant-resistant oocysts of Cryptosporidium spp. shed in the feces of infected humans and animals frequently contaminate water resources and are subsequently spread via potable and recreational waters. The current monoclonal-antibody-based methods for detecting them in water are slow, labor-intensive, and demand skills to interpret the results. We have developed DNA-aptamer-based aptasensors, coupled with magnetic beads, to detect and identify the oocysts of C. parvum for monitoring recreational and drinking water sources. A sensitive and specific electrochemical aptasensor (3′-biotinylated R4–6 aptamer) was used as a secondary ligand to bind the streptavidin-coated magnetic beads. This was incorporated into a probe using gold nanoparticle modified screen-printed carbon electrodes. Square wave voltammetry allowed for specific recognition of C. parvum oocysts. The aptamer-coated probes had an oocyst detection limit of 50. It did not bind to the cysts of Giardia duodenalis, another common waterborne pathogen, thus indicating its high specificity for the target pathogen. The system could successfully detect C. parvum oocysts in spiked samples of the raw lake and river waters. Therefore, the combined use of the aptasensor and magnetic beads has the potential to monitor water quality for C. parvum oocysts in field samples without relying on monoclonal antibodies and skill-demanding microscopy.