Microfluidics for Waterborne Pathogen Detection

Cryptosproidium parvum is a deadly enteric waterborne pathogen which requires regular monitoring and detection. Dielectrophoresis is a label-free separation method which uses the dielectric properties of oocysts for separation in a microfluidic device. We propose a two-region electrode structure with a focusing region which deviates viable oocysts to the central part of the channel and a second finger-pair separation region which deviates the viable oocysts to the opposite end of the channel from its entry location. From a 50% mixed sample, 85.1% of all detected events found to be focused were viable oocysts, when combined with excystation results. Nanoparticles are known for their lysing and anti-microbial properties. We present a simplified lysis protocol for the efficient extraction of molecular material from protozoan pathogens such as Cryptosporidium via the use of nanoparticles. The lysis of Cryptosporidium by zinc oxide and silver nanoparticles (NPs) was investigated at concentrations ranging from 125 to 1000 µg/ml, and exposure times up to 120 minutes, and compared to freeze-thaw. To confirm the lysis of cells, qPCR and confocal microscopy were used. The effectiveness of the different lysis approaches can be ranked: ZnO NPs=freeze thawing>Ag NPs though all methods worked across the whole range of oocyst concentrations tested. The NP approach to oocyst lysis is faster and requires less equipment so provides a useful alternative to freeze thaw and has the benefit that it offers easier miniaturisation capabilities, and the possibility of developing portable methods to detect Cryptosporidium via molecular approaches.