A Pumpless and Tubeless Microfluidic Device Enables Extended In Vitro Development of Cryptosporidium parvum

Background. The enteric parasite Cryptosporidium remains a treatment challenge for drinking water utilities globally due to its resistance to chlorine disinfection. However, the lack of an in vitro culture system for Cryptosporidium that is both cost-effective and reliable remains a key bottleneck in Cryptosporidium research. Methods. Here we report that the microfluidic culture of human ileocecal colorectal adenocarcinoma (HCT-8) cells under fluid shear stress enables the extended development of Cryptosporidium parvum. Specifically, the growth of C. parvum in a user-friendly pumpless microfluidic device was assessed using immunofluorescence assays, scanning electron microscopy, and quantitative polymerase chain reaction, which revealed that development continued for 10 days in total. Results. Oocysts produced within the microfluidic device were infective to fresh HCT-8 monolayers; however, these oocysts were only present at low levels. Conclusions. We anticipate that such microfluidic approaches will facilitate a wide range of in vitro studies on Cryptosporidium and may have the potential to be further developed as a routine infectivity assessment tool for the water industry.