A reduced-cost approach for analyzing fresh produce for contamination with Cryptosporidium oocysts and/or Giardia cysts

• Fresh produce can be contaminated by Giardia and/or Cryptosporidium (oo)cysts.
• As the Standard ISO Method is costly, we investigated cost-saving modifications.
• Immunomagnetic separation was adapted by using less beads and modified buffers.
• Recovery efficiencies were tested in-house and in a blinded inter-laboratory trial.
• The modification is a cheaper supplementary method to the Standard ISO Method.

Fresh produce is a recognized vehicle for transmission of various protozoan parasites, including Toxoplasma gondii, Cyclospora cayetanensis, Giardia duodenalis, and Cryptosporidium spp. For Giardia and Cryptosporidium, a Standard ISO Method for analyzing fresh produce is being developed, based on the standard methods for analyzing water. Although it is undoubtedly of value to have a Standard Method available, if the Method is very expensive or difficult to perform this may hamper routine surveys, particularly in settings where resources are restrained, although arguably such settings may produce the results of most importance. Here we present a modified method for analyzing green leafy vegetables such as lettuce or spinach for Cryptosporidium oocysts and Giardia cysts.The modified method is considerably cheaper than the Standard Method; by using a smaller volume of magnetic beads in the immunomagnetic separation (IMS) step and buffers that are complementary to those provided in the IMS kit, the cost per analysis is reduced significantly.In-house seeding trials resulted in acceptable levels of recovery. The modified method has also been trialed in 10 different microbiology analysis labs with experience of detecting protozoa, and results have been shown to be satisfactory; recovery rates ranged from 4% to 88% with a mean of 53% for Cryptosporidium and 33% for Giardia. Generally poor results were associated with problems in shipping reagents. This modified method is not proposed as an alternative to the Standard Method, but as a complementary approach providing a cheaper option for projects on limited budgets or for laboratories performing analyses in situations or countries where application of the ISO Standard Method is too expensive.