A molecular assessment of infectious agents carried by Atlantic salmon at sea and in three eastern Canadian rivers, including aquaculture escapees and North American and European origin wild stocks
Abstract
Introduction
Methods
Sample collection, preservation, and transfer
Source location | Continent of origin | Year | Tissue type(s) | N | Fork length, mm (mean ± SD) |
---|---|---|---|---|---|
Labrador Sea near Greenland | North America | 2016 | Kidney | 10 | 654 ± 24 |
2017 | Multi-tissuea | 26 | 672 ± 55 | ||
European | 2016 | Kidney | 33 | 618 ± 25 | |
2017 | Multi-tissuea | 4 | 668 ± 20 | ||
Restigouche River | North America | 2017 | Multi-tissuea | 30 | 836 ± 67 |
St. John River | North America | 2017 | Gill | 30 | 572 ± 31 |
Magaguadavic River | North America | 2017 | Multi-tissuea | 17 | 653 ± 47 |
Laboratory protocols
Agent | Type | Abbreviation | Limits of detection | Primer and probe sequences | Accession no. | Assay reference |
---|---|---|---|---|---|---|
Atlantic salmon calicivirusa | Virus | ascv | 27.14 | F: ACCGACTGCCCGGTTGT R: CTCCGATTGCCTGTGATAATACC P: CTTAGGGTTAAAGCAGTCG | — | Gideon Mordecai |
Infectious hematopoietic necrosis virus | Virus | ihnv | 27.64 | F: AGAGCCAAGGCACTGTGCG R: TTCTTTGCGGCTTGGTTGA P: TGAGACTGAGCGGGACA | NC_001652 | |
Infectious pancreatic necrosis virus | Virus | ipnv | 27.63 | F: GCAACTTACTTGAGATCCATTATGCT R: GAGACCTCTAAGTTGTATGACGAGGTCTCT P: CGAGAATGGGCCAGCAAGCA | — | |
Infectious salmon anemia virus | Virus | isav | 26.12 | F: TGGGCAATGGTGTATGGTATGA R: GAAGTCGATGAACTGCAGCGA P: CAGGATGCAGATGTATGC | EU118822 | |
Salmonid herpesvirus | Virus | omv | 26.59 | F: GCCTGGACCACAATCTCAATG R: CGAGACAGTGTGGCAAGACAAC P: CCAACAGGATGGTCATTA | — | |
Piscine myocarditis virus | Virus | pmcv | 26.29 | F: AGGGAACAGGAGGAAGCAGAA R: CGTAATCCGACATCATTTTGTGA P: TGGTGGAGCGTTCAA | HQ339954 | |
Piscine reovirus | Virus | prv | 26.11 | F: TGCTAACACTCCAGGAGTCATTG R: TGAATCCGCTGCAGATGAGTA P: CGCCGGTAGCTCT | — | |
Salmon alphavirus | Virus | sav | 26.28 | F: CCGGCCCTGAACCAGTT R: GTAGCCAAGTGGGAGAAAGCT P: TCGAAGTGGTGGCCAG | AY604235 | |
Salmonid gill poxvirus | Virus | sgpv | 25.15 | F: ATCCAAAATACGGAACATAAGCAAT R: CAACGACAAGGAGATCAACGC P: CTCAGAAACTTCAAAGGA | — | |
Putative Totiviridae | Virus | toti | 25.87 | F: TCTGCGCGCTGCACCTA R: ATGCGGAGGAACTCACACACT P: CAAGTGCTACACTGCG | — | Gideon Mordecai |
Viral erythrocytic necrosis virus | Virus | ven | 24.85 | F: CGTAGGGCCCCAATAGTTTCT R: GGAGGAAATGCAGACAAGATTTG P: TCTTGCCGTTATTTCCAGCACCCG | — | |
Viral encephalopathy and retinopathy virus | Virus | venv | 26.21 | F: TTCCAGCGATACGCTGTTGA R: CACCGCCCGTGTTTGC P: AAATTCAGCCAATGTGCCCC | AJ245641 | |
Viral hemorrhagic septicemia virus | Virus | vhsv | 26.86 | F: AAACTCGCAGGATGTGTGCGTCC R: TCTGCGATCTCAGTCAGGATGAA P: TAGAGGGCCTTGGTGATCTTCTG | Z93412 | |
Aeromonas hydrophila | Bacterium | ae_hy | 28.67 | F: ACCGCTGCTCATTACTCTGATG R: CCAACCCAGACGGGAAGAA P: TGATGGTGAGCTGGTTG | AY165026 | |
Aeromonas salmonicida | Bacterium | ae_sal | 25.61 | F: TAAAGCACTGTCTGTTACC R: GCTACTTCACCCTGATTGG P: ACATCAGCAGGCTTCAGAGTCACTG | M64655 | Keeling et al. 2013 (modified) |
Flavobacterium psychrophilum | Bacterium | fl_psy | 29.46 | F: GATCCTTATTCTCACAGTACCGTCAA R: TGTAAACTGCTTTTGCACAGGAA P: AAACACTCGGTCGTGACC | — | |
Ca. Piscichlamydia salmonisa | Bacterium | pch_sal | 30.72 | F: TCACCCCCAGGCTGCTT R: GAATTCCATTTCCCCCTCTTG P: CAAAACTGCTAGACTAGAGT | EU326495 | |
Piscirickettsia salmonis | Bacterium | pisck_sal | 23.32 | F: TCTGGGAAGTGTGGCGATAGA R: TCCCGACCTACTCTTGTTTCATC P: TGATAGCCCCGTACACGAAACGGCATA | U36943 | |
Renibacterium salmoninarum | Bacterium | re_sal | 25.91 | F: CAACAGGGTGGTTATTCTGCTTTC R: CTATAAGAGCCACCAGCTGCAA P: CTCCAGCGCCGCAGGAGGAC | AF123890 | |
Rickettsia-like organism | Bacterium | rlo | 25.23 | F: GGCTCAACCCAAGAACTGCTT R: GTGCAACAGCGTCAGTGACT P: CCCAGATAACCGCCTTCGCCTCCG | EU555284 | |
Ca. Syngnamydia salmonisa | Bacterium | sch | 27.9 | F: GGGTAGCCCGATATCTTCAAAGT R: CCCATGAGCCGCTCTCTCT P: TCCTTCGGGACCTTAC | FJ897519 | |
Tenacibaculum maritimum | Bacterium | te_mar | 26.71 | F: TGCCTTCTACAGAGGGATAGCC R: CTATCGTTGCCATGGTAAGCCG P: CACTTTGGAATGGCATCG | — | |
Vibrio anguillarum | Bacterium | vi_ang | 26.41 | F: CCGTCATGCTATCTAGAGATGTATTTGA R: CCATACGCAGCCAAAAATCA P: TCATTTCGACGAGCGTCTTGTTCAGC | L08012 | |
Aliivibrio salmonicida | Bacterium | vi_sal | 25.84 | F: GTGTGATGACCGTTCCATATTT R: GCTATTGTCATCACTCTGTTTCTT P: TCGCTTCATGTTGTGTAATTAGGAGCGA | AF452135 | |
Yersinia ruckeri | Bacterium | ye_ruc | 28.13 | F: TCCAGCACCAAATACGAAGG R: ACATGGCAGAACGCAGAT P: AAGGCGGTTACTTCCCGGTTCCC | — | |
Paramoeba perurans | Amoeba | ne_per | 25.39 | F: GTTCTTTCGGGAGCTGGGAG R: GAACTATCGCCGGCACAAAAG P: CAATGCCATTCTTTTCGGA | EF216905 | |
Ichthyophthirius multifiliis | Ciliate | ic_mul | 23.7 | F: AAATGGGCATACGTTTGCAAA R: AACCTGCCTGAAACACTCTAATTTTT P: ACTCGGCCTTCACTGGTTCGACTTGG | IMU17354 | |
Gyrodactylus salaris | Fluke | gy_sal | 26.42 | F: CGATCGTCACTCGGAATCG R: GGTGGCGCACCTATTCTACA P: TCTTATTAACCAGTTCTGC | — | |
Spironucleus salmonicida | Flagellate | sp_sal | 26.05 | F: GCAGCCGCGGTAATTCC R: CGAACTTTTTAACTGCAGCAACA P: ACACGGAGAGTATTCT | AY677182 | |
Nanophyetus salmincola | Fluke | na_sal | 24.3 | F: CGATCTGCATTTGGTTCTGTAACA R: CCAACGCCACAATGATAGCTATAC P: TGAGGCGTGTTTTATG | AY269674 | |
Sphaerothecum destruens | Mesomycetozoea | sp_des | 26.5 | F: GGGTATCCTTCCTCTCGAAATTG R: CCCAAACTCGACGCACACT P: CGTGTGCGCTTAAT | AY267346 | |
Facilispora margolisi | Microsporidian | fa_mar | 30.55 | F: AGGAAGGAGCACGCAAGAAC R: CGCGTGCAGCCCAGTAC P: TCAGTGATGCCCTCAGA | HM800849 | |
Loma salmonae | Microsporidian | lo_sal | 25.42 | F: GGAGTCGCAGCGAAGATAGC R: CTTTTCCTCCCTTTACTCATATGCTT P: TGCCTGAAATCACGAGAGTGAGACTACCC | HM626243 | |
Paranucleospora theridiona | Microsporidian | pa_ther | 28.16 | F: CGGACAGGGAGCATGGTATAG R: GGTCCAGGTTGGGTCTTGAG P: TTGGCGAAGAATGAAA | FJ59481 | |
Ceratonova shasta | Myxozoan | ce_sha | 28.5 | F: CCAGCTTGAGATTAGCTCGGTAA R: CCCCGGAACCCGAAAG P: CGAGCCAAGTTGGTCTCTCCGTGAAAAC | AF001579 | |
Myxobolus arcticus | Myxozoan | my_arc | 26.8 | F: TGGTAGATACTGAATATCCGGGTTT R: AACTGCGCGGTCAAAGTTG P: CGTTGATTGTGAGGTTGG | HQ113227 | |
Myxobolus insidiosus | Myxozoan | my_ins | 26.43 | F: CCAATTTGGGAGCGTCAAA R: CGATCGGCAAAGTTATCTAGATTCA P: CTCTCAAGGCATTTAT | EU346375 | |
Parvicapsula kabatai | Myxozoan | pa_kab | 25.58 | F: CGACCATCTGCACGGTACTG R: ACACCACAACTCTGCCTTCCA P: CTTCGGGTAGGTCCGG | DQ515821 | |
Parvicapsula minibicornis | Myxozoan | pa_min | 29.62 | F: AATAGTTGTTTGTCGTGCACTCTGT R: CCGATAGGCTATCCAGTACCTAGTAAG P: TGTCCACCTAGTAAGGC | AF201375 | |
Parvicapsula pseudobranchicolaa | Myxozoan | pa_pse | 25.16 | F: CAGCTCCAGTAGTGTATTTCA R: TTGAGCACTCTGCTTTATTCAA P: CGTATTGCTGTCTTTGACATGCAGT | AY308481 | |
Tetracapsuloides bryosalmonae | Myxozoan | te_bry | 24.98 | F: GCGAGATTTGTTGCATTTAAAAAG R: GCACATGCAGTGTCCAATCG P: CAAAATTGTGGAACCGTCCGACTACGA | AF190669 | |
Trypanoplasma salmositica | Protozoan | cr_sal | 24.34 | F: TCAGTGCCTTTCAGGACATC R: GAGGCATCCACTCCAATAGAC P: AGGAGGACATGGCAGCCTTTGTAT | — | |
Dermocystidium salmonis | Protozoan | de_sal | 25.49 | F: CAGCCAATCCTTTCGCTTCT R: GACGGACGCACACCACAGT P: AAGCGGCGTGTGCC | U21337 | |
Ichthyophonus hoferi | Protozoan | ic_hof | 24.17 | F: GTCTGTACTGGTACGGCAGTTTC R: TCCCGAACTCAGTAGACACTCAA P: TAAGAGCACCCACTGCCTTCGAGAAGA | AF467793 | |
Si:dkey-78d16.1 protein | Host reference | hkg | 45 | F: GTCAAGACTGGAGGCTCAGAG R: GATCAAGCCCCAGAAGTGTTTG P: AAGGTGATTCCCTCGCCGTCCGA | — |
Note: F, forward primer; R, reverse primer, P, probe
Sequencing
Statistical methodology
Results
Genotyping of marine-collected Atlantic salmon and annual infection differences
Prevalence and load differences among collection locations and strains
Marine: N. American origin (N = 36) | Marine: European origin (N = 37) | Magaguadavic River (N = 17) | Restigouche River (N = 30) | St. John River (N = 30) | Total | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Agent | N | Mean | SD | N | Mean | SD | N | Mean | SD | N | Mean | SD | N | Mean | SD | N | |
Parasites | |||||||||||||||||
P. pseudobranchicola | 21 | 6463 | 15 914 | 4 | 538 | 384 | 7 | 1927 | 2716 | 14 | 1674 | 3924 | 3 | 1087 | 837 | 49 | |
T. bryosalmonae | 12 | 31 547 | 71 331 | 18 | 351 773 | 656 993 | 5 | 71 395 | 99 215 | 0 | — | — | 0 | — | — | 35 | |
P. theridion | 12 | 3395 | 5968 | 7 | 3697 | 5195 | 1 | 181 | — | 10 | 2807 | 6367 | 4 | 3527 | 6171 | 34 | |
I. hoferi | 2 | 385 | 482 | 0 | — | — | 0 | — | — | 15 | 1 608 948 | 3 587 305 | 10 | 338 474 | 433 839 | 27 | |
S. destruens | 1 | 1485 | — | 0 | — | — | 0 | — | — | 0 | — | — | 0 | — | — | 1 | |
Bacteria | |||||||||||||||||
Ca. P. salmonis | 3 | 83 350 | 130 482 | 2 | 248 725 | 194 166 | 9 | 148 466 | 255 231 | 11 | 1 989 479 | 4 397 990 | 9 | 109 302 | 221 325 | 34 | |
F. psychrophilum | 0 | — | — | 0 | — | — | 0 | — | — | 11 | 3614 | 5175 | 19 | 2284 | 4426 | 30 | |
A. salmonicida | 0 | — | — | 0 | — | — | 0 | — | — | 7 | 2 805 611 | 7 186 680 | 3 | 139 877 | 241 677 | 10 | |
Ca. S. salmonis | 0 | — | — | 0 | — | — | 0 | — | — | 4 | 85 | 53 | 0 | — | — | 4 | |
Viruses | |||||||||||||||||
PRV | 2 | 5980 | 8363 | 1 | 18 799 | — | 13 | 24 | 33 | 0 | — | — | 0 | — | — | 16 | |
ASCV | 0 | — | — | 0 | — | — | 10 | 188 248 | 383 841 | 1 | 10 | — | 0 | — | — | 11 | |
ISAV | 2 | 218 | 264 | 0 | — | — | 0 | — | — | 0 | — | — | 0 | — | — | 2 | |
SGPV | 0 | — | — | 0 | — | — | 2 | 227 | 81 | 0 | — | — | 0 | — | — | 2 | |
VERV | 1 | 22 | — | 0 | — | — | 0 | — | — | 0 | — | — | 0 | — | — | 1 | |
Richness | — | 1.6 | 1.1 | — | 0.9 | 0.9 | — | 2.8 | 1.3 | — | 2.4 | 1.3 | 1.6 | 1.3 | — | ||
RIB | — | 0.23 | 0.5 | — | 0.14 | 0.3 | — | 0.21 | 0.34 | — | 0.36 | 0.4 | 0.12 | 0.23 | — |
Note: Total positive detections (N) and the mean and standard deviation (SD) of agent copy numbers, richness (total unique agents per host), and relative infection burden (RIB) are shown for each agent relative to the sampling location. RIB is a composite metric that incorporates richness and load information from all agents detected in each host; PRV, piscine orthoreovirus; ASCV, Atlantic salmon calicivirus; ISAV, infectious salmon anemia virus; SGPV, salmonid gill poxvirus; VERV, encephalopathy and retinopathy virus.
PRV and ISAV sequence analysis
Discussion
Marine transmission potential between continental stocks
Infection profiles of wild and escaped farm salmon in rivers
Conclusions
Acknowledgements
References
Supplementary materials
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