Evaluation of the Increased Genetic Resolution and Utility for Source Tracking of a Recently Developed Method for Genotyping Cyclospora cayetanensis

• Published in: Microorganisms (Basel) Vol. 12; no. 5; p. 848
• Main Authors: Leonard, Susan R, Mammel, Mark K, Almeria, Sonia, Gebru, Solomon T, Jacobson, David K, Peterson, Anna C, Barratt, Joel L N, Musser, Steven M
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 24.04.2024
• Subjects: Assaying; Clusters; Cyclospora cayetanensis; Cyclosporiasis; Disease control; Epidemics; epidemiology; Feces; Food; Food contamination; Food contamination & poisoning; foodborne parasite; Gene loci; Genomes; Genotyping; Haplotypes; Medical laboratories; Parasites; Phylogeny; Raspberries; targeted amplicon sequencing; traceback; United States > US; epidemiology; genetic relatedness; genotyping; cyclosporiasis; foodborne parasite; targeted amplicon sequencing; traceback
• ISSN: 2076-2607; 2076-2607
• DOI: 10.3390/microorganisms12050848

is a foodborne parasite that causes cyclosporiasis, an enteric illness in humans. Genotyping methods are used to genetically discriminate between specimens from cyclosporiasis cases and can complement source attribution investigations if the method is sufficiently sensitive for application to food items. A very sensitive targeted amplicon sequencing (TAS) assay for genotyping encompassing 52 loci was recently designed. In this study, we analyzed 66 genetically diverse clinical specimens to assess the change in phylogenetic resolution between the TAS assay and a currently employed eight-marker scheme. Of the 52 markers, ≥50 were successfully haplotyped for all specimens, and these results were used to generate a hierarchical cluster dendrogram. Using a previously described statistical approach to dissect hierarchical trees, the 66 specimens resolved into 24 and 27 distinct genetic clusters for the TAS and an 8-loci scheme, respectively. Although the specimen composition of 15 clusters was identical, there were substantial differences between the two dendrograms, highlighting the importance of both inclusion of additional genome coverage and choice of loci to target for genotyping. To evaluate the ability to genetically link contaminated food samples with clinical specimens, was genotyped from DNA extracted from raspberries inoculated with fecal specimens. The contaminated raspberry samples were assigned to clusters with the corresponding clinical specimen, demonstrating the utility of the TAS assay for traceback efforts.