Permeabilization of Cryptosporidium spp. Oocysts in Water, Apple and Carrot Juice by Pulsed Electric Field Technology

<i>Cryptosporidium</i> spp. oocysts are highly resistant to conventional disinfection methods and have been associated with foodborne outbreaks linked to unpasteurized fruit and vegetable juices. This study aimed to evaluate the effectiveness of Pulsed Electric Fields (PEF) in permeabilizing <i>Cryptosporidium</i> oocysts in water, apple juice, and carrot juice. Oocysts were exposed to monopolar square-wave pulses (3 µs) at electric field strengths ranging from 15 to 35 kV/cm, with treatment times up to 180 µs, and application temperatures between 25 °C and 60 °C. Membrane permeabilization was assessed using propidium iodide uptake via fluorescence microscopy and flow cytometry. Results showed that oocyst permeabilization increased with electric field strength, treatment time, and temperature, with up to 90% permeabilization achieved at 35 kV/cm and 45 °C. Carrot juice treatments yielded higher permeabilization levels than apple juice, attributed to greater electrical conductivity and energy input. Temperatures below 60 °C alone had negligible effects, but synergistically enhanced PEF efficacy. These findings demonstrate that PEF, particularly when combined with mild heat, is a promising non-thermal technology for reducing <i>Cryptosporidium</i> viability in beverages, offering an effective alternative for improving the microbiological safety of minimally processed juices while preserving sensory and nutritional quality.