The diversity and pathogenicity of amoebae on the gills of Atlantic salmon (Salmo salar) with amoebic gill disease (AGD)

• Main Author: English, Chloe
• Format: Thesis
• Language: English
• Published: The University of Queensland, School of Biological Sciences 2020
• Subjects: Amoebozoa; Salmo salar; gill disease; aquaculture; Neoparamoeba; Nolandella; Pseudoparamoeba; aetiology; dysbiosis; 0602 Ecology; 0605 Microbiology; 0707 Veterinary Sciences; Atlantic salmon
• Online Access: https://espace.library.uq.edu.au/view/UQ:f3ffca5/thumbnail_s41732975_phd_final_thesis_t.jpg; https://espace.library.uq.edu.au/view/UQ:f3ffca5/s41732975_phd_final_thesis.pdf; https://espace.library.uq.edu.au/view/UQ:f3ffca5

Amoebic gill disease (AGD) is an ectoparasitic condition affecting many teleost fish globally, and it is one of the main health issues impacting farmed Atlantic salmon, Salmo salar in Tasmania’s expanding aquaculture industry. To date, Neoparamoeba perurans is considered the only aetiological agent of AGD, based on laboratory trials that confirmed its pathogenicity, and its frequent presence on the gills of farmed Atlantic salmon with branchitis. However, the development of gill disease in salmonid aquaculture is complex and multifactorial and is not always closely associated with the presence and abundance of N. perurans. Moreover, multiple other amoeba species colonise the gills and their role in AGD is unknown.In this study we profiled the Amoebozoa community on the gills of AGD-affected and healthy farmed Atlantic salmon and performed in vivo challenge trials to investigate the possible role these accompanying amoebae play alongside N. perurans in AGD onset and severity. Significantly, it was shown that despite N. perurans being the primary aetiological agent, it is possible AGD has a multi-amoeba aetiology.Specifically, the diversity of amoebae colonising the gills of AGD-affected farmed Atlantic salmon was documented by culturing the gill community in vitro, then identifying amoebae using a combination of morphological and sequence-based taxonomic methods. In addition to N. perurans, 11 other Amoebozoa were isolated from the gills, and were classified within the genera Neoparamoeba, Paramoeba, Vexillifera, Pseudoparamoeba, Vannella and Nolandella. This work highlighted that there is a far greater diversity of amoebae colonising AGD-affected gills than previously established.Drawing on this culture-based study, five new TaqMan quantitative PCR (qPCR) assays were developed and applied to more accurately determine the prevalence and abundance of multiple amoeba species colonising the gills of Atlantic salmon held at two Tasmanian farm sites over a one-year period. The presence of N. perurans was also assessed in parallel using a previously established qPCR method. N. perurans was the dominant species in the Amoebozoa community on gills, and its abundance positively correlated with the progression of gross gill pathology. Only sporadic detections of Pseudoparamoeba sp. and Vannellida species were observed across the sampling period at either farm site. Nolandella spp., however, were highly prevalent at one site at one sample time when N. perurans were not detected on gills presenting low levels of gross gill pathology.To investigate the pathogenic potential of Nolandella sp. and a more closely related amoeba to N. perurans, Pseudoparamoeba sp., in vivo challenges of naïve Atlantic salmon were performed. Additionally, to elucidate how Nolandella sp. and Pseudoparamoeba sp. influence the onset or severity of N. perurans-induced AGD, the gill condition of fish challenged with N. perurans alone was compared to fish challenged with a mix of all three amoeba strains. Immersion challenge of all three species resulted in minor gill lesions, with the most severe epithelial hyperplasia documented in the N. perurans treatments, while lesions with infiltrating lymphocytes were the predominate pathology observed in fish challenged by Nolandella sp. and Pseudoparamoeba sp. The presence of individual Nolandella or Pseudoparamoeba cells were not linked with lesion sites, so the precise cause of pathology remains inconclusive. Moreover, the presence of these non-N. perurans species did not significantly increase the severity of N. perurans-induced branchitis.Overall this investigation supports N. perurans being the primary agent of AGD, yet also shows that other species of amoebae which colonise the gills of Tasmanian Atlantic salmon can dominate the gill community and may be capable of causing some gill pathology under specific conditions. Thus, the involvement of non-N. perurans amoebae in AGD should not yet be discounted. The increasing list of putative gill pathogens, and the complexity of disease expression, provides supportive rationale to the consideration of gill disease in the context of dysbiosis of microbial community structure, rather than a pathological response to a single agent.