Dataset Information

Catching the fish with the worm: a case study on eDNA detection of the monogenean parasite Gyrodactylus salaris and two of its hosts, Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss).

ABSTRACT: BACKGROUND:Environmental DNA (eDNA) monitoring is growing increasingly popular in aquatic systems as a valuable complementary method to conventional monitoring. However, such tools have not yet been extensively applied for metazoan fish parasite monitoring. The fish ectoparasite Gyrodactylus salaris, introduced into Norway in 1975, has caused severe damage to Atlantic salmon populations and fisheries. Successful eradication of the parasite has been carried out in several river systems in Norway, and Atlantic salmon remain infected in only seven rivers, including three in the Drammen region. In this particular infection region, a prerequisite for treatment is to establish whether G. salaris is also present on rainbow trout upstream of the salmon migration barrier. Here, we developed and tested eDNA approaches to complement conventional surveillance methods. METHODS:Water samples (2 × 5 l) were filtered on-site through glass fibre filters from nine locations in the Drammen watercourse, and DNA was extracted with a CTAB protocol. We developed a qPCR assay for G. salaris targeting the nuclear ribosomal ITS1 region, and we implemented published assays targeting the mitochondrial cytochrome-b and NADH-regions for Atlantic salmon and rainbow trout, respectively. All assays were transferred successfully to droplet digital PCR (ddPCR). RESULTS:All qPCR/ddPCR assays performed well both on tissue samples and on field samples, demonstrating the applicability of eDNA detection for G. salaris, rainbow trout and Atlantic salmon in natural water systems. With ddPCR we eliminated a low cross-amplification of Gyrodactylus derjavinoides observed using qPCR, thus increasing specificity and sensitivity substantially. Duplex ddPCR for G. salaris and Atlantic salmon was successfully implemented and can be used as a method in future surveillance programs. The presence of G. salaris eDNA in the infected River Lierelva was documented, while not elsewhere. Rainbow trout eDNA was only detected at localities where the positives could be attributed to eDNA release from upstream land-based rainbow trout farms. Electrofishing supported the absence of rainbow trout in all of the localities. CONCLUSIONS:We provide a reliable field and laboratory protocol for eDNA detection of G. salaris, Atlantic salmon and rainbow trout, that can complement conventional surveillance programs and substantially reduce the sacrifice of live fish. We also show that ddPCR outperforms qPCR with respect to the specific detection of G. salaris.

SUBMITTER: Rusch JC

PROVIDER: S-EPMC5987472 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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Catching the fish with the worm: a case study on eDNA detection of the monogenean parasite Gyrodactylus salaris and two of its hosts, Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss).

Rusch Johannes C JC Hansen Haakon H Strand David A DA Markussen Turhan T Hytterød Sigurd S Vrålstad Trude T

Parasites & vectors 20180604 1

<h4>Background</h4>Environmental DNA (eDNA) monitoring is growing increasingly popular in aquatic systems as a valuable complementary method to conventional monitoring. However, such tools have not yet been extensively applied for metazoan fish parasite monitoring. The fish ectoparasite Gyrodactylus salaris, introduced into Norway in 1975, has caused severe damage to Atlantic salmon populations and fisheries. Successful eradication of the parasite has been carried out in several river systems in ...[more]

PMID: 29866158

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Project description:Gyrodactylus salaris (Monogenea, Platyhelminthes) is a notifiable freshwater pathogen responsible for causing catastrophic damage to wild Atlantic salmon stocks, most notably in Norway. In some strains of Baltic salmon (e.g., from the river Neva) however, the impact is greatly reduced due to some form of innate resistance that regulates parasite numbers, resulting in fewer host mortalities. Gyrodactylus salaris is known from 17 European states; its status in a further 35 states remains unknown; the UK, the Republic of Ireland and certain watersheds in Finland are free of the parasite. Thus, the parasite poses a serious threat if it emerges in Atlantic salmon rearing regions throughout Europe. At present, infections are generally controlled via extreme measures such as the treatment of entire river catchments with the biocide rotenone, in order to remove all hosts, before restocking with the original genetic stock. The use of rotenone in this way in EU countries is unlikely as it would be in contravention of the Water Framework Directive. Not only are such treatments economically and environmentally costly, they also eradicate the potential for any host/parasite evolutionary process to occur. Based on previous studies, UK salmon stocks have been shown to be highly susceptible to infection, analogous to Norwegian stocks. The present study investigates the impact of a G. salaris outbreak within a naïve salmon population in order to determine long-term consequences of infection and the likelihood of coexistence. Simulation of the salmon/ G. salaris system was carried out via a deterministic mathematical modelling approach to examine the dynamics of host-pathogen interactions. Results indicated that in order for highly susceptible Atlantic strains to evolve a resistance, both a moderate-strong deceleratingly costly trade-off on birth rate and a lower overall cost of the immune response are required. The present study provides insights into the potential long term impact of G. salaris if introduced into G. salaris-free territories and suggests that in the absence of external controls salmon populations are likely to recover to high densities nearing 90% of that observed pre-infection.

Project description:BackgroundAquaculture is a globally important and rapidly growing industry. It contributes positively to the economy and sustainability of coastal communities, but it is not without regulatory challenges. These challenges are diverse, and may include identification of fish discarded in an illegal manner, biological discharge from fish ensilage tanks, and partially destroyed or processed tissues. Robust genetic tools are required by management authorities to address these challenges. In this paper, we describe nine species-specific primer sets amplifying very short DNA fragments within the mitochondrial DNA cytochrome c oxidase (COI) gene, which were designed to permit diagnostic identification of degraded DNA from two of the most commonly farmed salmonids in Europe and North America.ResultsOf the nine designed primer sets, six were found to be species-specific (four Atlantic salmon, two rainbow trout), whereas the remaining three sets (two Atlantic salmon, one rainbow trout) also amplified a product from other, closely related, salmonid DNA templates. Screening of DNA templates from 11 other non-salmonid native fish species did not produce PCR products with any of the primer sets. Specific tests confirmed the ability of these markers to identify Atlantic salmon and rainbow trout tissues in treated food products, chemically treated ensilage waste and fillets left to degrade in saltwater for up to 31 days at 15°C. Importantly, these markers provided diagnostic identification in cases where other genetic methods failed because of degraded DNA quality.ConclusionsResults from this study demonstrate that amplification of very short DNA fragments using species-specific primers represents a robust and versatile method to create cheap and efficient genetic tests that can be implemented in a range of forensic applications. These markers will provide fishery, aquaculture and food regulatory authorities with a method to investigate and enforce regulations within these industries.

Project description:BACKGROUND: Describing and evaluating miRNA inventories with Next Generation Sequencing is a goal of scientists from a wide range of fields. It requires high purity, high quality, and high yield RNA extractions that do not only contain abundant ribosomal RNAs but are also enriched in miRNAs. Here we compare 6 disparate and commercially available totalRNA extraction kits for their suitability for miRNA-preparations from Gyrodactylus salaris, an important but small (500 ?m in length) monogenean pathogen of Norwegian Atlantic salmon (Salmo salar). FINDINGS: We evaluated 1 salt precipitation method (MasterPure™ Complete RNA Purification Kit, Epicentre), 2 Phenol based extraction methods (mirVana Kit, Ambion, and Trizol Plus Kit, Invitrogen), 1 paramagnetic bead extraction method (RNA Tissue kit, GeneMole) and 2 purification methods based on spin column chromatography using a proprietary resin as separation matrix (Phenol-free Total RNA Purification Kit, Amresco, and ZR MicroPrep Kit, Zymo Research). The quality of the extractions from 1, 10 and 100 individuals, respectively, was assessed in terms of totalRNA yield, RNA integrity, and smallRNA and miRNA yield. The 6 RNA extraction methods yielded considerably different total RNA extracts, with striking differences in low molecular weight RNA yield. The Phenol-free Total RNA Purification Kit (Amresco) showed the highest totalRNA yield, but the best miRNA/totalRNA ratio was obtained with the ZR MicroPrep Kit (Zymo Research). It was not possible to extract electrophoretically detectable miRNAs from Gyrodactylus salaris with the RNA Tissue Kit (GeneMole) or the Trizol Plus Kit (Invitrogen). CONCLUSIONS: We present an optimized extraction protocol for single and small numbers of Gyrodactylus salaris from infected Atlantic salmon that delivers a totalRNA yield suitable for downstream next generation sequencing analyses of miRNA. Two of the six tested totalRNA kits/methods were not suitable for the extraction of miRNAs from Gyrodactylus salaris.

Project description:BackgroundGyrodactylus salaris Malmberg, 1957 is an OIE (Office International des Epizooties)-listed parasitic pathogen and had until the current study been reported from 19 countries across Europe, although many of these records require confirmation. The last comprehensive evaluation regarding the distribution of G. salaris, however, was made in 2007, although some of the states identified as being G. salaris-positive were ascribed this status based on misidentifications, on partial data resulting from either morphological or molecular tests, or from records that have not been revisited since their early reporting. It is thus important to go through the reports on G. salaris to obtain a status for each country.MethodsTo provide a revised update of the G. salaris distribution, a literature review was necessary. This literature, however, was not always readily accessible and, in certain cases, the article only made superficial reference to the parasite without providing details or data to support the identification. In most cases, the original specimens were not deposited in a national collection. Additional Gyrodactylus material for the current study was obtained from selected salmonid populations with the aim to contribute to current understanding regarding the distribution of G. salaris. Additional parasite material collected for this study was processed following standard procedures for species identification in Gyrodactylus [1].ResultsFrom the work conducted in the current study, G. salaris is reported from a further three regions in Italy, alongside three other species, and appears to occur extensively throughout central Italy without causing significant mortalities to its rainbow trout, Oncorhynchus mykiss (Walbaum), host. The analysis of archive material from G. salaris-positive farms would suggest that G. salaris has been in this country since at least 2000. Material obtained from rainbow trout from Finland and Germany are confirmed as G. salaris, supporting existing data for these countries. No specimens of G. salaris, however, were found in the additional Gyrodactylus material obtained from rainbow trout reared in Portugal and Spain. A morphologically similar species, Gyrodactylus teuchis Lautraite, Blanc, Thiery, Daniel et Vigneulle, 1999, however, was found.ConclusionsFollowing the present review, Gyrodactylus salaris is reported from 23 out of 50 recognised states throughout Europe; only records from 14 of these states have been confirmed by either morphology and/or by an appropriate molecular test and are considered valid, while only nine of these records have been confirmed by a combination of both methods.

Dataset Information

Catching the fish with the worm: a case study on eDNA detection of the monogenean parasite Gyrodactylus salaris and two of its hosts, Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss).

Publications

Catching the fish with the worm: a case study on eDNA detection of the monogenean parasite Gyrodactylus salaris and two of its hosts, Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss).

Similar Datasets

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