Genotyping Cyclospora cayetanensis From Multiple Outbreak Clusters With An Emphasis on a Cluster Linked to Bagged Salad Mix-United States, 2020

Abstract

Cyclosporiasis is a diarrheal illness caused by the foodborne parasite Cyclospora cayetanensis. Annually reported cases have been increasing in the United States prompting development of genotyping tools to aid cluster detection. A recently developed Cyclospora genotyping system based on 8 genetic markers was applied to clinical samples collected during the cyclosporiasis peak period of 2020, facilitating assessment of its epidemiologic utility. While the system performed well and helped inform epidemiologic investigations, inclusion of additional markers to improve cluster detection was supported. Consequently, investigations have commenced to identify additional markers to enhance performance.

Keywords: Cyclospora cayetanensis; cyclosporiasis; epidemiology; genotyping; outbreaks.

Figure 1.

Epidemiologic curve of cases linked to the bagged salad outbreak investigation (n = 705) by the end of 2020, by illness onset date (A), and MicrobeTrace network generated from a distance matrix containing genotypes analyzed between 24 June and 9 November 2020 (B). A, Cases linked by genotyping to clusters 2020_001 (n = 116) and 2020_003 (n = 73) and that were also epidemiologically linked to the bagged salad outbreak are highlighted in blue and orange, respectively. Gray represents cases linked to the bagged salad mix outbreak without a specimen provided for genotyping (n = 516). B, The matrix used to generate the network shown was calculated using the Barratt-Plucinski ensemble of algorithms, as described elsewhere [2, 3]. Links were filtered in MicrobeTrace [5] to link specimens separated by a distance of <0.15 for visualization purposes. Color coding reflects specimens belonging to each temporal-genetic cluster (TGC), determined from the same distance matrix using Ward’s clustering method, as described elsewhere [2]. Some outlying specimens (dots distal from their same-colored partners in 2-dimensional space) appear in this network, reflecting minor discordance between Ward’s clustering method and clusters detected using MicrobeTrace. Specimens assigned to clusters 001 (blue) and 003 (orange) are partitioned into 2 hemispheres of the same larger cluster, based on their mitochondrial genotype. This network includes 778 genotyped specimens submitted to the Centers for Disease Control and Prevention (CDC) by the state health department and 38 submitted by the Public Health Agency of Canada by 9 November 2020, for which sequence data of sufficient quality were obtained (all colors except light gray; n=816), analyzed alongside specimens submitted in previous years and/or those collected from outside the United States or Canada (light gray; n=710). Notably, some genetic clusters (dots positioned closely together in space) are marked with different colors indicating membership in different TGCs despite possessing a similar genotype. This is a consequence of using specimen collection dates to identify outbreaks that may have been epidemiologically unrelated yet were genetically similar based on the CDC’s current 8-marker genotyping system.