Long-Term Preservation and Storage of Faecal Samples in Whatman® Cards for PCR Detection and Genotyping of Giardia duodenalis and Cryptosporidium hominis.

Abstract

Simple SummaryPreservation and storage of biological samples prior to testing and analysis is a pressing issue in the epidemiological field studies conducted in remote or poor-resource areas with limited or no access to electricity where the cold chain cannot be maintained. This is particularly true for faecal specimens of human and animal origin exposed to high degradation rates under environmental conditions characterised by high temperatures and humidity, such as those present in tropical and subtropical regions. Under this scenario, simple, safe, and cost-effective methods are highly needed to allow the collection and transportation of well-preserved faecal samples intended for pathogen detection without compromising the performance, reliability, and accuracy of molecular procedures methods used for detection and genotyping purposes. This study assessed the suitability of three commercially available filter cards for the preservation of faecal samples containing common diarrhoea-causing enteric protozoan parasites at different storage periods and temperature conditions. Obtained results demonstrated that filter cards impregnated with faecal matrices containing these pathogens are fully compatible with downstream molecular methods for up to six months at room temperature. Therefore, filter cards can be used for the safe transportation, preservation, and storage of faecal samples without the need of the cold chain.Preservation and conservation of biological specimens, including faecal samples, is a challenge in remote areas or poor-resource settings where the cold chain cannot be maintained. This study aims at evaluating the suitability of filter cards for long-term storage of faecal samples of animal and human origin positive to the diarrhoea-causing protozoan parasites, Giardia duodenalis and Cryptosporidium hominis. Three commercially available Whatman® Filter Cards were comparatively assessed: the FTA® Classic Card, the FTA® Elute Micro Card, and the 903 Protein Saver Card. Human faecal samples positive to G. duodenalis (n = 5) and C. hominis (n = 5) were used to impregnate the selected cards at given storage (1 month, 3 months, and 6 months) periods and temperature (−20 °C, 4 °C, and room temperature) conditions. Parasite DNA was detected by PCR-based methods. Sensitivity assays and quality control procedures to assess suitability for genotyping purposes were conducted. Overall, all three Whatman® cards were proven useful for the detection and molecular characterisation of G. duodenalis and C. hominis under the evaluated conditions. Whatman® cards represent a simple, safe, and cost-effective option for the transportation, preservation, and storage of faecal samples without the need of the cold chain.