Molecular characterization of Tubifex populations in Alberta, Canada, and their role in the spread of whirling disease

Abstract

Whirling disease is a parasitic infection caused by Myxobolus cerebralis Hofer (Cnidaria: Myxozoa) and is debilitating to the salmonid fish that act as hosts. As of 2016, this parasite has been confirmed in Alberta, Canada. This thesis focused on estimating how long M. cerebralis has been present in the province of Alberta, Canada, and the potential for this parasite to spread in the province through the oligochaete host, Tubifex tubifex (Müller) (Annelida: Naididae). To discern the introduction date of M. cerebralis into the province I used a combination of eDNA detections utilizing quantitative polymerase chain reactions (qPCR) and soil radioisotope dating. Sediment samples from Johnson Lake in Banff National Park, the location of the first whirling disease-positive fish detected in Alberta, were taken as well as two other waterbodies in the same watershed. These sediment samples were sectioned by depth and assessed for age and M. cerebralis DNA. These methods agreed with the introduction of the parasite to Johnson Lake, AB, around 2016. As a control, Johnson Lake core samples were also assessed for Brook Trout and Rainbow Trout DNA through qPCR, as introduction dates are known to begin after 1900 for this waterbody. Trout DNA was not detected in sediment prior to their introduction, with low DNA detections as early as 1972.9, confirming the reliability of these molecular methods. To increase our knowledge of the distribution of Tubifex spp. and M. cerebralis in Alberta, I used CoxI barcoding of worms combined with qPCR detections for M. cerebralis DNA and Whole genome sequencing (WGS). Oligochaete samples from southern watersheds in the Crowsnest region of Alberta were taken in 2019. A total of 409 worm samples were sequenced and barcoded, with 156 returning positive qPCR results for M. cerebralis DNA. All positive M. cerebralis detections with >1000 DNA copies belonged to Tubifex sp. 3, corresponding to individuals suspected of shedding triactinomyxons (TAM), the parasite life stage responsible for infecting new fish. Whole genome sequencing was completed for each Tubifex spp. described from Alberta, which allowed mitochondrial genomes to be assembled and compared phylogenetically, expanding on the current understanding of Tubifex spp. and indicating new species in need of descriptions are present in Alberta, Canada. Observations of these Tubifex spp. suggest they differ in susceptibility to M. cerebralis.