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

Abstract

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.

Performance of Sewage Treatment Technologies for the Removal of Cryptosporidium Sp. And Giardia Sp.: Toward Water Circularity

“…In addition, more expensive equipment and qualified personnel are needed; therefore, this clinical diagnosis approach has not achieved widespread implementation (Ahmed and Karanis et al, 2018). The genetic analysis of parasitic protozoa has also presented problems, both in the preservation and extraction of DNA and the search for genes that allow the identification of species or subspecies (Lalonde and Gajadhar, 2009;Wilke and Robertson, 2009;Abdelsalam et al, 2016;Costa et al, 2021;Köster et al, 2021).…”

Investigating the Role of Animal Behaviour in the Transmission and Spread of Zoonotic Diseases

“…It is important to study the various causes of emerging zoonoses, as these diseases account for more than 60% of infectious diseases encountered by humans and can create worldwide devastation, as seen during the COVID-19 pandemic. As mankind becomes more aware of the threats of pandemics, such as the plague, Spanish flu, and SARS-CoV-2, it is of great interest to better characterize ways to minimize activities that increase reservoirs or human-animal contact (Köster et al, 2021). While it is seemingly impossible to completely stop new infections from spreading from animals to humans, it may be possible to reduce the severity of risks to the human population via quicker or more efficient methods of detection, early warning systems, and proper control or prevention policies if we better understand the activities that influence or drive these zoonotic transmissions (Kelly et al, 2017).…”

Zoonoses and Wildlife: One Health Approach

“…More than 50 different species—including wild and domestic ungulates (e.g., red deer, roe deer, fallow deer, chamois, mouflon, European bison, wild boar, sheep, goat, cattle), wild carnivores (e.g., wolf, Eurasian lynx, Eurasian badger, coypu, beech marten, golden jackal), micromammals (e.g., yellow-necked field mouse, long-tailed field mouse, European water vole, white-toothed shrew, garden dormouse, common vole, house mouse, western Mediterranean mouse, black rat, Eurasian red squirrel), non-human primates (the genera Cebuella, Cercocebus, Cercopithecus, Eulemur, Hylobates, Lemur, Macaca, Mandrillus, Saimiri, and Varecia ), turtles (e.g., Testudo hermanni , T. h. boettgeri , T. graeca , and T. marginata ), bats (the families Pteropodidae, Emballonuridae, Rhinolophidae, Hipposideridae, and Vespertilionidae), and ticks ( Ixodes ricinus , Dermacentor marginatus , Hyalomma marginatum )—are included. Regarding the zoonotic pathogens represented in this issue, the presence of or exposure to 17 different pathogens—including viruses [ 4 ] (West Nile virus), bacteria [ 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 ] ( Anaplasma phagocytophilum, Coxiella burnetii, Helicobacter pylori, H. suis, Mycobacterium tuberculosis Complex, Salmonella sp., and Leptospira interrogans sensu stricto), and parasitic protists [ 14 , 15 ] (e.g., Cryptosporidium spp., Giardia duodenalis , Blastocystis sp., Enterocytozoon bieneusi , Entamoeba histolytica , Entamoeba dispar , Balantioides coli , Troglodytella spp., Leishmania spp. )—are presented.…”

“…In addition, the study of parasites involves searching for them, or their DNA—mainly in the feces of animals. In remote areas or resource-poor settings where the cold chain cannot be maintained, preservation and conservation of biological specimens—including fecal samples—is a challenge; for this reason, Köster et al [ 15 ] evaluated the suitability of filter cards for the long-term storage of fecal samples of animal and human origin that were positive for the diarrhea-causing protozoan parasites Giardia duodenalis and Cryptosporidium hominis . For this purpose, three commercially available Whatman ® filter cards were comparatively evaluated: the FTA ® Classic card, the FTA ® Elute Micro card, and the 903 Protein Saver card.…”

“…Giardia duodenalis ( n = 5)- and C. hominis ( n = 5)-positive human stool samples were used to impregnate the selected cards at selected storage times (1 month, 3 months, and 6 months) and temperatures (−20 °C, 4 °C, and room temperature). Data presented by Köster et al [ 15 ] demonstrate that Whatman ® cards are a cost-effective option for the preservation and long-term storage (up to six months) of fecal samples under a wide range of temperatures (from −20 °C to room temperature), without compromising their biospecimen stability and suitability for molecular-based diagnostic methods. Indeed, Whatman ® cards enable the molecular detection and genotyping of common diarrhea-causing enteric protozoan parasites, including C. hominis and G. duodenalis .…”