The Use of an Atmospheric Cold Plasma Jet to Inactivate Cryptosporidium parvum Oocysts on Cilantro

Introduction: In 2015, the CDC reported a rise in outbreaks linked to parasites like Cryptosporidium. Cryptosporidium parvum outbreaks have been associated with contaminated drinking or recreational water; however, there is growing concern that oocysts may become a more common contaminant in foods, especially in produce and herbs, which have been identified as transmission vehicles of protozoan illness.

Purpose: This study examined the effects of a cold plasma jet on viability of C. parvum oocysts on fresh cilantro.

Methods: Oocysts (6.25 x 10⁵ oocysts/per 500 µl PBS) were applied to glass slides (controls) or fresh cilantro, and treated in duplicate. The 90 min air-dried samples were treated for 0, 30, 90, and 180 s with atmospheric cold plasma jet and placed into 50 ml conical tubes containing 15 ml PBS. Samples were placed on a rotating shaker plate for 30 min and inverted at 15 min, then centrifuged for 20 min at 2,000 x g, treated with 2% bleach, and triple rinsed. Recovered oocysts were subjected to excystation, in duplicate per sample. Oocysts were treated with a 0.75% taurocholic acid/0.25% trypsin solution, incubated at 37°C for 45 mins, and observed. Percent excystation was determined using differential interference contrast microscopy of 100 fields, counting oocyst ghosts, and observing moving sporozoites compared to non-viable oocysts. Data were analyzed using one-way ANOVA.

Results: Data analysis indicated a decrease in percent excystation of oocysts that was statistically significant between the untreated oocysts with >85% excystation and treated (180 s) samples on glass (P=0.0060). Oocysts recovered from cilantro showed significant differences in excystation, between untreated and treated (90 s, 180 s) samples (P<0.01). Treated oocysts showed varying percent excystation at <67%.

Significance: Oocyst excystation is the initial step in infection. Data suggests cold plasma treatment effects C. parvum oocyst viability. These results indicate potential inactivation and warrant use of cell culture infectivity assays coupled with variation in oocyst inoculum levels and treatment parameters.