Introduction
Amoebic Gill Disease (AGD) is one of the most significant health constraints affecting Atlantic salmon aquaculture caused by the marine amoeba Neoparamoeba perurans. Despite its economic importance, tools for high-resolution epidemiological surveillance remain limited, restricting the ability to distinguish local persistence from reinvasion, identify transmission pathways, or detect emerging lineages. Genomic analysis of N. perurans has been hindered by difficulties in axenic culture and contamination from associated microbial communities. We therefore developed a targeted AmpSeq panel for simultaneous genotyping of both N. perurans and its kinetoplastid endosymbiont, Perkinsela, directly from biological material.
Methods
An AmpSeq panel targeting nuclear and mitochondrial loci from both partners was designed from a draft N. perurans genome assembly. This panel was applied to 58 isolates collected from 12 Atlantic salmon aquaculture sites in Scotland and Norway. Sequence reads were processed to generate high-confidence SNP datasets for population genomic analysis. Population structure, clonal diversity, linkage disequilibrium, isolation by distance, and host-symbiont evolutionary congruence were assessed using standard multivariate and phylogenetic approaches.
Results and Discussion
The panel resolved 50 host and 44 symbiont high-confidence SNPs with mean coverage exceeding 150x. The N. perurans host population showed weak geographic structuring (Fst = 0.008; p = 0.123) and comparatively high genotype richness (47 MLGs), consistent with substantial regional connectivity. In contrast, the Perkinsela showed lower diversity (21 MLGs), stronger clonality and significant geographic structure (Fst = 0.035; p < 0.001). Significant isolation-by-distance was detected in both partners, indicating that dispersal is extensive but not fully homogenised.
Global cophylogenetic analyses supported predominantly vertical transmission between host and symbiont. However, frequent genotype mismatches were observed with 10 of 11 host MLG genotypes represented by multiple isolates harbouring different Perkinsela lineages. This indicates that strict co-inheritance is not absolute and that reassortment or lineage switching may occur.
These findings demonstrate that AmpSeq can be a scalable tool to provide actionable genomic surveillance data for Amoebic Gill Disease by differentiating persistence from reinfection, identifying transmission connectivity between farms or regions, and detecting emerging host or symbiont lineages relevant to management and biosecurity.
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