Going to Court to Stop Spread of Disease (Part 2 of 2)

[rockdog]

Actually, for some reason CK seems pretty focused on Morton. She is just 1 of many, many scientists around the world who are against salmon farming as it is currently. All you need to do is read the research papers that Englishman posted previously, and you can see it's not just her.
I tend to read the credible research papers, and respect the information held within them. Having some guy who works at a feedlot, paid to put spins on things on a sport fishing website, doesn't quite cut it for me, sorry.

 

[cuttlefish]

Wouldn't mind getting more info about these outbreaks; http://www.inspection.gc.ca/animals...icaemia-2014-/eng/1396452410303/1396452411272

Know anything about those reports, CK? If not the herring then how about the Atlantics? Can you ask the BCSFA to find out which farm in BC and share with the public?

I asked a simple question. Still hoping to get an honest answer...

 

[seadna]

seadna, I suspect you are ignoring my question regarding genome sequencing of Atlantic salmon. A man I respect, Dr. Brian Riddell, had good things to say about this and I had hoped you with a background in this science would chime in.
I have been accused of $hit disturbing on this site, and rightfully so, but that is not the case now. You have stated you have expertise in this stuff; please share your thoughts on how this will impact salmon farming here in BC and Washington, and the rest of the world.

Sorry Dave, I just missed this. Genome sequencing is something I've done a good bit of (from viruses to bacteria and some larger genomes like apple). In general, genome sequencing means that one genome of a hopefully representative individual has been sequenced. Typically the between individual variation is small (<1-2%) so that the one genome that is sequenced is a fairly good estimate of the sequence of any other individual (for example in humans the sequence difference is about 1 base pair in 1000).

As far as the impact of genome sequencing one huge impact is that the sequence alone is a useful "reagent" for further study. That is, with a technique called PCR (polymerase chain reaction), one can amplify a given region of DNA from any sample provided you know the approximate sequence of that region. In practice this is done by synthesizing two short pieces of DNA (known as "oligos") that are outside of a region of interest based on the known genome sequence. If you put these oligos, the 4 nucleotide building blocks, and the enzyme that copies DNA (polymerase) into a tube with DNA isolated from a given fish and heat that tube up and down about 30 times, the PCR process will make about 2 to the 30th power (about 1,000,000,000) copies of the region of DNA between the two primer sequences. This allows one to easily extract that DNA from the rest of the DNA (e.g. the entire genome) and sequence it to look at variation in a given region across many individuals. This can be very useful to look for variants that give rise to some useful trait (say viral resistance or increased growth rate or fat content). So the immediate impact is that it gives researchers tools to study the fish far more efficiently.

As far as the impact on farming, that's a bit harder to say. One possible impact is that researchers can use the genome sequence and PCR to more rapidly follow pieces of the genome around in selective breeding experiments. This should make it quicker to breed farmed fish with desirable traits.

 

[Dave]

Sorry Dave, I just missed this. Genome sequencing is something I've done a good bit of (from viruses to bacteria and some larger genomes like apple). In general, genome sequencing means that one genome of a hopefully representative individual has been sequenced. Typically the between individual variation is small (<1-2%) so that the one genome that is sequenced is a fairly good estimate of the sequence of any other individual (for example in humans the sequence difference is about 1 base pair in 1000).

As far as the impact of genome sequencing one huge impact is that the sequence alone is a useful "reagent" for further study. That is, with a technique called PCR (polymerase chain reaction), one can amplify a given region of DNA from any sample provided you know the approximate sequence of that region. In practice this is done by synthesizing two short pieces of DNA (known as "oligos") that are outside of a region of interest based on the known genome sequence. If you put these oligos, the 4 nucleotide building blocks, and the enzyme that copies DNA (polymerase) into a tube with DNA isolated from a given fish and heat that tube up and down about 30 times, the PCR process will make about 2 to the 30th power (about 1,000,000,000) copies of the region of DNA between the two primer sequences. This allows one to easily extract that DNA from the rest of the DNA (e.g. the entire genome) and sequence it to look at variation in a given region across many individuals. This can be very useful to look for variants that give rise to some useful trait (say viral resistance or increased growth rate or fat content). So the immediate impact is that it gives researchers tools to study the fish far more efficiently.

As far as the impact on farming, that's a bit harder to say. One possible impact is that researchers can use the genome sequence and PCR to more rapidly follow pieces of the genome around in selective breeding experiments. This should make it quicker to breed farmed fish with desirable traits.

Well, I would be lying if I said I understood all of the above, lol!
Thanks for the reply.

 

[agentaqua]

Seadna - thanks so much for your post. I understood most of what you said. Just wondering? - what is your opinion on the strengths/weaknesses of the genome methodology verses the current CFIA methodology when looking for diseases in fish?

My current and limited understanding is that CFIA uses DFO labs that have been certified through an ISO scheme - but are not "approved" by the OIE - YET - they are the only ones CFIA uses or pays attention to. In fact they were so rattled by the positives generated by Fred Kibenges lab in Charlottetown - they complained to the OIE and had his lab decertified.

My current understanding of the CFIA methodology is that they try to get fresh samples preserved in RNALater and ship them off for PCR amplification and then analysis. If they get a "unconfirmed" positive - they then try to grow the virus directly in cell culture looking for cytopathic abnormalities. That is the "confirmed" part of the positive. I know there are problems with being able to use the PCR method - as I understand it - you need to be able to know what segment to amplify. There are over 11 different strains of ISA - for example. Then there is the type of tissue - from what species - and the length of time that you need to incubate the virus - to get a positive. The non-pathogenic strain of ISA HPR0 - cannot be grown in cell culture, for example. Then you need to have an appropriately-trained tech to read the results. All issues - are far as I know - wrt getting a "confirmed" positive, along with degradation and inactivation of the virus when getting and transporting the samples. An example was that Rick Routledge with SFU used previously frozen samples for the ISA/PCR test - and still got weak positives. CFIA claimed that the test was a false negative and meaningless w/o looking at a statistical analysis for probabilities of a weak positive verses a false negative. I was wondering what your take is on this method?

I believe it is likely that Rick sequenced the non-pathogenic strain - which can not be grown in cell culture - and that strain readily mutates - and that strain may be the one in Cultus Lake and Oweekeno stocks - both lakes have had "unconfirmed" positive results for ISA - and both have seen dramatic stock crashes and limited recovery.

I thought the genome method also looked at the RNA that was turned-on in the cell?

 

[ClayoquotKid]

I asked a simple question. Still hoping to get an honest answer...

CFIA shares information about salmon aquaculture equal to, or at a greater level, than every other type of food production industry they are responsible for - I have no role in this.

I am not the company's veterinarian, nor am I employed in the fish health department in any way - I don't even know if those reports are related to my company.

 

[ClayoquotKid]

Actually, for some reason CK seems pretty focused on Morton. She is just 1 of many, many scientists around the world who are against salmon farming as it is currently. All you need to do is read the research papers that Englishman posted previously, and you can see it's not just her.
I tend to read the credible research papers, and respect the information held within them. Having some guy who works at a feedlot, paid to put spins on things on a sport fishing website, doesn't quite cut it for me, sorry.

I have read the papers, discussed them with peers, looked at their strengths and weaknesses, and considered their conclusions given observations and other known data.

Many are based on incredibly small sample sizes, with issues around chain-of-custody and collection methods, or simply come to conclusions which are highly speculative and unsupported by empirical evidence.

Others are simply alarmist, containing a wealth of "Could's" and "May's" and serve no purpose other than to propose a theory using "peer -review" as a vehicle for the purpose of creating fear, uncertainty and doubt about the aquaculture industry.

Others contain valid concerns which are being followed by the scientific community in the effort to identify and quantify impacts that may exist regarding salmon aquaculture.

All have failed to definitively identify a direct link between the presence of aquaculture and a related, abnormal decline in wild salmon stocks - this is especially true on the BC coast.

After nearly 40 years of management invoking the, "Precautionary Principle" in a variety of ways (some found to be acceptable, others not), the salmon farm industry (and I speak to my area of experience - BC) has proven to have elevated its practices to a level which pushes the limits of the world standard.

Being dismissive of anonymous posters whining about character assassination in one breath and calling people sociopaths in the next is something that I take no issue with.

People demanding peer-reviewed rebuttals to every wad of scientific spaghetti thrown at the wall will be disappointed to find that there is no, and not likely ever will there be, a paper outlining exactly how salmon farms have zero impact on wild populations. (Believe me, I would love to see it.)

Science is left looking at links, correlations, and trends - which have to date not shown evidence that gives reason to move away from current practices regarding net-pen aquaculture, choosing to rather employ high levels of monitoring and selective breeding in the farms (and things like genomic sequencing: http://www.fis.com/fis/worldnews/worldnews.asp?monthyear=&day=20&id=69378&l=e&special=&ndb=1 target= ) , and related (increasing) monitoring outside, to ensure that farmed fish are kept healthy and their wild counterparts are not put at risk.

Alarmism, catastrophism and hostility from the anti-aquaculture side does nothing to further the debate, and the continued reliance on conspiracies and cries of collusion to fill in the spots where evidence, or the lack thereof, falsifies their hypotheses serves to even further weaken their position.

There are plenty of people working on both sides of the issue to ensure that both wild and farmed salmon are abundant on the BC coast into the future - unfortunately, people like Morton, (And to a lesser degree Volpe, Krkosek et al.) are not part of this effort, and their contributions (in many cases) rely on misinformation, half-truths, suppression of evidence, and what would be considered poor scientific practice.

They are not out to make salmon farming better, they are out to end the practice.

There is a distinct difference, and their material is recieved accordingly.

 

[Dave]

I have read the papers, discussed them with peers, looked at their strengths and weaknesses, and considered their conclusions given observations and other known data.

Many are based on incredibly small sample sizes, with issues around chain-of-custody and collection methods, or simply come to conclusions which are highly speculative and unsupported by empirical evidence.

Others are simply alarmist, containing a wealth of "Could's" and "May's" and serve no purpose other than to propose a theory using "peer -review" as a vehicle for the purpose of creating fear, uncertainty and doubt about the aquaculture industry.

Others contain valid concerns which are being followed by the scientific community in the effort to identify and quantify impacts that may exist regarding salmon aquaculture.

All have failed to definitively identify a direct link between the presence of aquaculture and a related, abnormal decline in wild salmon stocks - this is especially true on the BC coast.

After nearly 40 years of management invoking the, "Precautionary Principle" in a variety of ways (some found to be acceptable, others not), the salmon farm industry (and I speak to my area of experience - BC) has proven to have elevated its practices to a level which pushes the limits of the world standard.

Being dismissive of anonymous posters whining about character assassination in one breath and calling people sociopaths in the next is something that I take no issue with.

People demanding peer-reviewed rebuttals to every wad of scientific spaghetti thrown at the wall will be disappointed to find that there is no, and not likely ever will there be, a paper outlining exactly how salmon farms have zero impact on wild populations. (Believe me, I would love to see it.)

Science is left looking at links, correlations, and trends - which have to date not shown evidence that gives reason to move away from current practices regarding net-pen aquaculture, choosing to rather employ high levels of monitoring and selective breeding in the farms (and things like genomic sequencing: http://www.fis.com/fis/worldnews/worldnews.asp?monthyear=&day=20&id=69378&l=e&special=&ndb=1 target= ) , and related (increasing) monitoring outside, to ensure that farmed fish are kept healthy and their wild counterparts are not put at risk.

Alarmism, catastrophism and hostility from the anti-aquaculture side does nothing to further the debate, and the continued reliance on conspiracies and cries of collusion to fill in the spots where evidence, or the lack thereof, falsifies their hypotheses serves to even further weaken their position.

There are plenty of people working on both sides of the issue to ensure that both wild and farmed salmon are abundant on the BC coast into the future - unfortunately, people like Morton, (And to a lesser degree Volpe, Krkosek et al.) are not part of this effort, and their contributions (in many cases) rely on misinformation, half-truths, suppression of evidence, and what would be considered poor scientific practice.

They are not out to make salmon farming better, they are out to end the practice.

There is a distinct difference, and their material is recieved accordingly.

View attachment 12247

Excellent link ck. That is what Riddell was meaning I believe. As I understand it, some Atlantic stocks are also more resistant to sea lice.
Seadna, if IPN can be coded as in the link provided, is it not a matter of time before ISA is also??

 

[agentaqua]

BC) has proven to have elevated its practices to a level which pushes the limits of the world standard.

No - absolutely not CK> Scotland uses a particle tracking model - to model sea lice plumes in the lochs. That model was developed by DFO on this side of the Atlantic - where it is not used. Similarly, fish health reporting in other countries - US, Norway, and Scotland - is open, public, and transparent - but not here. Not even a nice try "try" CK. Read the UVic Law report on the link I posted - even though you claim you have read all the stuff we post - it is obvious this fact escaped your attention - somehow...unless poor or bad is also a "limit" of a standard in your definition.

 

[agentaqua]

Finstad et al. Veterinary Research 2014, 45:35
http://www.veterinaryresearch.org/content/45/1/35

Piscine orthoreovirus (PRV) infects Atlantic salmon erythrocytes
Øystein Wessel Finstad1*, Maria Krudtaa Dahle2, Tone Hæg Lindholm1, Ingvild Berg Nyman1, Marie Løvoll3, Christian Wallace3, Christel Moræus Olsen1, Anne K Storset1 and Espen Rimstad1

1Department of Food Safety and Infection Biology, Norwegian University of Life Sciences, Oslo, Norway


Abstract
Piscine orthoreovirus (PRV) belongs to the Reoviridae family and is the only known fish virus related to the Orthoreovirus genus. The virus is the causative agent of heart and skeletal muscle inflammation (HSMI), an emerging disease in farmed Atlantic salmon (Salmo salar L.). PRV is ubiquitous in farmed Atlantic salmon and high loads of PRV in the heart are consistent findings in HSMI. The mechanism by which PRV infection causes disease remains largely unknown. In this study we investigated the presence of PRV in blood and erythrocytes using an experimental cohabitation challenge model. We found that in the early phases of infection, the PRV loads in blood were significantly higher than in any other organ. Most virus was found in the erythrocyte fraction, and in individual fish more than 50% of erythrocytes were PRV-positive, as determined by flow cytometry. PRV was condensed into large cytoplasmic inclusions resembling viral factories, as demonstrated by immunofluorescence and confocal microscopy. By electron microscopy we showed that these inclusions contained reovirus-like particles. The PRV particles and inclusions also had a striking resemblance to previously reported viral inclusions described as Erythrocytic inclusion body syndrome (EIBS). We conclude that the erythrocyte is a major target cell for PRV infection. These findings provide new information about HSMI pathogenesis, and show that PRV is an important factor of viral erythrocytic inclusions.

Apparently CK has not only discussed this article with his buds around the pens and they really don't believe the science. After all - since they sling feed to the cages - they should know the science - right? They rather believe a press release from a company PR firm that stated that in the courts - third hand verbal evidence - that they heard that somebody told them that Gary Marty had an "opinion" - with no science behind it. BUT that's good enough for CK - and don't bother asking any inconvenient questions - as only those people like Marty are special enough to ask questions about such annoying, inconvenient, and inconsequential things such as risk assessment and management, siting criteria, open and transparent fish health reporting, scoping - or determining the boundaries of effects, particle modelling, agent-based modelling, due diligence, the precautionary approach - or the democratic process. Nothing to see there folks - move along. BC salmon farmers are doing more than anyone else. CK says so - and you should just take his word for it - or he will post a cute picture and ignore your questions.

 

[cuttlefish]

CFIA shares information about salmon aquaculture equal to, or at a greater level, than every other type of food production industry they are responsible for - I have no role in this.

I am not the company's veterinarian, nor am I employed in the fish health department in any way - I don't even know if those reports are related to my company.

Okay, well thanks anyway. I thought you might be able, in the very least to find out if those reports are related to your company. Or if your industry association might want to provide an example of the transparency they seem to now be espousing.
http://www.theglobeandmail.com/news/...ticle19192479/
Jeremy Dunn, executive director of the B.C. Salmon Farmers Association, said his group endorses the call to make disease data public.

“We support the release of fish health information. We’ve asked our regulator to release that information and we understand that DFO’s working on releasing a greater detail of fish health information,” he said. “Obviously the CFIA has their own release [standards] with respect to just certain diseases and we support being transparent and putting up the information for the public.”

 

[seadna]

Seadna - thanks so much for your post. I understood most of what you said. Just wondering? - what is your opinion on the strengths/weaknesses of the genome methodology verses the current CFIA methodology when looking for diseases in fish?

My current and limited understanding is that CFIA uses DFO labs that have been certified through an ISO scheme - but are not "approved" by the OIE - YET - they are the only ones CFIA uses or pays attention to. In fact they were so rattled by the positives generated by Fred Kibenges lab in Charlottetown - they complained to the OIE and had his lab decertified.

My current understanding of the CFIA methodology is that they try to get fresh samples preserved in RNALater and ship them off for PCR amplification and then analysis. If they get a "unconfirmed" positive - they then try to grow the virus directly in cell culture looking for cytopathic abnormalities. That is the "confirmed" part of the positive. I know there are problems with being able to use the PCR method - as I understand it - you need to be able to know what segment to amplify. There are over 11 different strains of ISA - for example. Then there is the type of tissue - from what species - and the length of time that you need to incubate the virus - to get a positive. The non-pathogenic strain of ISA HPR0 - cannot be grown in cell culture, for example. Then you need to have an appropriately-trained tech to read the results. All issues - are far as I know - wrt getting a "confirmed" positive, along with degradation and inactivation of the virus when getting and transporting the samples. An example was that Rick Routledge with SFU used previously frozen samples for the ISA/PCR test - and still got weak positives. CFIA claimed that the test was a false negative and meaningless w/o looking at a statistical analysis for probabilities of a weak positive verses a false negative. I was wondering what your take is on this method?

I believe it is likely that Rick sequenced the non-pathogenic strain - which can not be grown in cell culture - and that strain readily mutates - and that strain may be the one in Cultus Lake and Oweekeno stocks - both lakes have had "unconfirmed" positive results for ISA - and both have seen dramatic stock crashes and limited recovery.

I thought the genome method also looked at the RNA that was turned-on in the cell?

AgentAqua - I need to do some searching to find out what "confirmation methods" are used by CFIA but if that involves culture, I'd be worried as that is IMHO, a great way to increase the rate of false negatives. In brief, PCR is VERY sensitive and can detect false positives but the false negative rate is generally pretty low. Culture based methods are much more complicated and they suffer from the opposite problem, high false negatives but low false positive rates. Some things don't culture very efficiently (or at all) in the lab so requiring a culture positive test to get a "confirmed" positive means that possible infections will be missed. Depending on the virus and sample handling, it's conceivable that one would miss the majority of infections by requiring a culture positive result. In addition, culture positive results are sensitive to both the length of time one does the culturing, the amount of "live" virus in the initial sample and the rate of replication in the culture system. Samples with low viral (or other pathogen) load have to be cultured for longer to produce enough pathogen for it (or its effects on other cell) to be visible in a culture dish. Usually culture based approaches culture the samples for a finite period of time (say 1 week) and only call things positive if they see as positives at or prior to this time threshold. This is another source of false negative results. Choosing an assay that has a high false negative rate is effectively slanting results to favor the industry.

On the other hand, PCR may detect levels of virus that are not biologically relevant AND without proper technique in the lab one can also get false positives due to contamination with PCR products from previous assays (e.g. an aerosol of a previous positive test can contaminate surfaces in the area which can in principle contaminate future samples). The latter can be mostly solved by appropriate lab setup and cleaning. For example we do all our PCR in a room that is physically separated from the area in which we setup PCR reactions and we set up PCR reactions in a clean hood to limit contamination. Nonetheless using any PCR positive in and of itself as an indicator of presence of virus (or other pathogen) can result in false positives. With careful experimentation, one can determine the rate of false positives. Also, one can use a method called "Quantitative PCR" to determine the level at which the virus is present in the sample. False positives due to contamination are easier to separate out quantitative PCR since they are generally from DNA present at low levels. So it is correct to say that a PCR positive result does not necessarily indicate the presence of an infectious agent. In fact for DNA based viruses or bacterial pathogens, the DNA of the pathogen may persist in the animal after the infection is cleared. So I would agree with the CFIA claim "that the test MAY (my emphasis added) be meaningless w/o looking at a statistical analysis for probabilities of a weak positive versus a false negative". That said, I would note that almost all human virological testing these days is done by molecular methods (mostly PCR) and the false positive issue is addressed with negative controls and other appropriate methodologies. Also, if there is a concern about infections being transmitted from fish farms to wild salmon (and that is a legitimate concern), false positives cost the industry while false negatives cost the wild salmon and everyone else that depends upon them. So IMHO, I'd rather select an assay to err on the side of false positives rather than one the errs on the side of false negatives.

As for "I thought the genome method also looked at the RNA that was turned-on in the cell?" - Generally when one refers to genome sequencing they are referring to sequencing the DNA. RNA (which is transcribed from the DNA genome) can also be sequenced. Since different genes are needed in differing amounts and those amount vary depending on conditions, the level of RNA present varies as a function of gene and condition. You can think of DNA as the computer code that provides the instructions for making all the genes. As different genes are turned on and off (or up and down) in different conditions, measuring the RNA levels tells you what genes are in operation (what subroutines are running in the computer analogy). One way to measure which RNA's are present, is to sequence them. The term used to describe the collection of RNA's present in a given sample is the "transcriptome" and sequencing the transcriptome is usually referred to as RNA-Seq. One reason why one may want to sequence RNA in the case of virus infected samples is that some viruses (like ISAV) are RNA viruses. Hence it's necessary to measure the RNA to detect the virus. In practice, sequencing (and PCR) require converting RNA into DNA. This is done with an enzyme called reverse transcriptase. So if one wants to sequence the RNA, the process is a) Isolate RNA from the cell b) (optional) selectively remove high abundance RNA's that are not of interest or select a subset of RNA's that are likely to be of interest c) convert the RNA into DNA using reverse transcriptase. This DNA made from RNA is usually referred to as cDNA for copy or complementary DNA. My username SeaDNA is a play on that term. d) Further process and sequence the cDNA (how this is done depends on the sequencing method).

 

[agentaqua]

Thanks so much for the info cDNA! Here's an interesting blog with articles and chronological info: http://www.homolog.us/blogs/blog/2013/08/28/salmon-next-gen-sequencing/

 

[agentaqua]

have you seen this one, cDNA?

http://people.landfood.ubc.ca/anthony.farrell/pubs/p310-Miller_et_al_2011.pdf

Genomic Signatures Predict Migration and Spawning Failure in Wild Canadian Salmon

Kristina M. Miller,1,2* Shaorong Li,1 Karia H. Kaukinen,1 Norma Ginther,1 Edd Hammill,3 Janelle M. R. Curtis,3 David A. Patterson,4 Thomas Sierocinski,5 Louise Donnison,5 Paul Pavlidis,5 Scott G. Hinch,2 Kimberly A. Hruska,2 Steven J. Cooke,6 Karl K. English,7 Anthony P. Farrell8

Long-term population viability of Fraser River sockeye salmon (Oncorhynchus nerka) is threatened by unusually high levels of mortality as they swim to their spawning areas before they spawn. Functional genomic studies on biopsied gill tissue from tagged wild adults that were tracked through ocean and river environments revealed physiological profiles predictive of successful migration and spawning. We identified a common genomic profile that was correlated with survival in each study. In ocean-tagged fish, a mortality-related genomic signature was associated with a 13.5-fold greater chance of dying en route. In river-tagged fish, the same genomic signature was associated with a 50% increase in mortality before reaching the spawning grounds in one of three stocks tested. At the spawning grounds, the same signature was associated with 3.7-fold greater odds of dying without spawning. Functional analysis raises the possibility that the mortality-related signature reflects a viral infection.

 

[agentaqua]

Another good one:

http://onlinelibrary.wiley.com/doi/10.1111/eva.12164/pdf

REVIEWS AND SYNTHESIS
Infectious disease, shifting climates, and opportunistic predators: cumulative factors potentially impacting wild salmon declines

Kristina M. Miller,1,2 Amy Teffer,3 Strahan Tucker,1 Shaorong Li,1 Angela D. Schulze,1 Marc Trudel,1,3 Francis Juanes,3 Amy Tabata,1Karia H. Kaukinen,1 Norma G. Ginther,1 Tobi J. Ming,1 Steven J. Cooke,6 J. Mark Hipfner,5 David A. Patterson4 and Scott G. Hinch2
1 Pacific Biological Station, Fisheries and Oceans Canada, Nanaimo, BC, Canada
2 Forest and Conservation Sciences, University of British Columbia, Vancouver, BC, Canada
3 Biology Department, University of Victoria, Victoria, BC, Canada
4 Fisheries and Oceans Canada, School of Resource and Environmental Management, Simon Fraser University, Science Branch, Burnaby, BC, Canada
5 Environment Canada, Wildlife Research Division, Delta, BC, Canada
6 Fish Ecology and Conservation Physiology Laboratory, Department of Biology, Carleton Univerisy, Ottawa, ON, Canada

Abstract

Emerging diseases are impacting animals under high-density culture, yet few studies assess their importance to wild populations. Microparasites selected for enhanced virulence in culture settings should be less successful maintaining infectivity in wild populations, as once the host dies, there are limited opportunities to infect new individuals. Instead, moderately virulent microparasites persisting for long periods across multiple environments are of greatest concern. Evolved resistance to endemic microparasites may reduce susceptibilities, but as barriers to microparasite distributions are weakened, and environments become more stressful, unexposed populations may be impacted and pathogenicity enhanced. We provide an overview of the evolutionary and ecological impacts of infectious diseases in wild salmon and suggest ways in which modern technologies can elucidate the microparasites of greatest potential import. We present four case studies that resolve microparasite impacts on adult salmon migration success, impact of river warming on microparasite replication, and infection status on susceptibility to predation. Future health of wild salmon must be considered in a holistic context that includes the cumulative or synergistic impacts of multiple stressors. These approaches will identify populations at greatest risk, critically needed to manage and potentially ameliorate the shifts in current or future trajectories of wild populations.

 

[agentaqua]

Molecular Genetic Analysis of Stomach Contents Reveals Wild Atlantic Cod Feeding on Piscine Reovirus (PRV) Infected Atlantic Salmon Originating from a Commercial Fish Farm
Kevin Alan Glover1*, Anne Grete Eide Sørvik1, Egil Karlsbakk1, Zhiwei Zhang2, Øystein Skaala1
1 Institute of Marine Research, Bergen, Norway, 2 Jiangsu Institute of Marine Fisheries, NanTong City, P. R. China

Abstract
In March 2012, fishermen operating in a fjord in Northern Norway reported catching Atlantic cod, a native fish forming an economically important marine fishery in this region, with unusual prey in their stomachs. It was speculated that these could be Atlantic salmon, which is not typical prey for cod at this time of the year in the coastal zone. These observations were therefore reported to the Norwegian Directorate of Fisheries as a suspected interaction between a local fish farm and this commercial fishery. Statistical analyses of genetic data from 17 microsatellite markers genotyped on 36 partially degraded prey, samples of salmon from a local fish farm, and samples from the nearest wild population permitted the following conclusions: 1. The prey were Atlantic salmon, 2. These salmon did not originate from the local wild population, and 3. The local farm was the most probable source of these prey. Additional tests demonstrated that 21 of the 36 prey were infected with piscine reovirus. While the potential link between piscine reovirus and the disease heart and skeletal muscle inflammation is still under scientific debate, this disease had caused mortality of large numbers of salmon in the farm in the month prior to the fishermen’s observations. These analyses provide new insights into interactions between domesticated and wild fish.

p.6: "Four main conclusions can be drawn from these analyses: 1. The partially digested and morphologically difficult to identify
prey were revealed to be Atlantic salmon, 2. Based upon several independent genetic parameters, these salmon prey were identified
as farmed and not from the local wild population, thus demonstrating this to be a human induced, as opposed to natural
phenomena, 3. Despite partial digestion, the majority of the prey, including the single escapee, carried detectable levels of PRV.
PRV is associated with the disease HSMI [32,33]. This disease had caused significant mortality of salmon on the local farm in the
immediate time-period prior to the prey being captured in the wild cod, 4."

 

[agentaqua]

Here's one pertinent to the PCR/cell culture discussion about false positives/negatives: http://www.tandfonline.com/doi/pdf/10.1577/H09-028.1

Journal of Aquatic Animal Health

Comparison of Quantitative RT-PCR with Cell Culture to Detect Viral Hemorrhagic Septicemia Virus (VHSV) IVb Infections in the Great Lakes
Kristine M. Hope a , Rufina N. Casey a , Geoffrey H. Groocock a , Rodman G. Getchell a , Paul R. Bowser a & James W. Casey a
a Aquatic Animal Health Program, Department of Microbiology and Immunology , College of Veterinary Medicine, Cornell University, Ithaca, New York, 14853, USA
Published online: 09 Jan 2011.

Abstract.—Viral hemorrhagic septicemia virus (VHSV) is an important pathogen of cultured and wild fish in marine and freshwater environments. A new genotype, VHSV IVb, was isolated from a fish collected from the Great Lakes in 2003. Since the first isolation, VHSV IVb has been confirmed in 28 species, signaling the early invasion and continued spread of this Office International des Epizooties–reportable agent. For surveillance of this virus in both wild and experimental settings, we have developed a rapid and sensitive onestep quantitative real-time polymerase chain reaction (qRT-PCR) assay that amplifies a 100-base-pair conserved segment from both the genomic negative strand and the mRNA positive strand of the nucleoprotein (N) gene of VHSV IVb. This assay is linear over seven orders of magnitude, with an analytical capability of detecting a single copy of viral RNA and reproducibility at 100 copies. The assay is approximately linear with RNA input from 50 to 1,000 ng per assay and works equally well with RNA prepared from a column-based or phenol-chloroform-based method. In wild-caught fish, 97% of the cases were found to be more than three orders of magnitude more sensitive using qRT-PCR than using cell culture. Of the 1,428 fish from the Great Lakes region tested in 2006 and 2007, 24% were positive by qRT-PCR whereas only 5% were positive by cell culture. All of the fish that were positive by cell culture were also positive by qRT-PCR. Importantly, qRTPCR sensitivity is comparable to that of cell culture detection when comparing VHSV viral RNA levels with
viral titer stocks, confirming that the high qRT-PCR signals obtained with diagnostic samples are due to the accumulation of N gene mRNA by transcriptional attenuation. The qRT-PCR assay is particularly valuable for rapid and high-throughput prescreening of fish before confirmatory testing by cell culture or sequencing tissue-derived amplicons and especially in detecting infection in fish that do not show clinical signs of VHS.

 

[agentaqua]

http://onlinelibrary.wiley.com/doi/10.1111/jfd.12228/abstract

Potential disease interaction reinforced: double-virus-infected escaped farmed Atlantic salmon, Salmo salar L., recaptured in a nearby river.

A S Madhun, E Karlsbakk, C H Isachsen, L M Omdal, A G Eide Sørvik, O Skaala, B T Barlaup, K A Glover
Journal of Fish Diseases (Impact Factor: 1.59). 01/2014; DOI:10.1111/jfd.12228
Source: PubMed
ABSTRACT The role of escaped farmed salmon in spreading infectious agents from aquaculture to wild salmonid populations is largely unknown. This is a case study of potential disease interaction between escaped farmed and wild fish populations. In summer 2012, significant numbers of farmed Atlantic salmon were captured in the Hardangerfjord and in a local river. Genetic analyses of 59 of the escaped salmon and samples collected from six local salmon farms pointed out the most likely source farm, but two other farms had an overlapping genetic profile. The escapees were also analysed for three viruses that are prevalent in fish farming in Norway. Almost all the escaped salmon were infected with salmon alphavirus (SAV) and piscine reovirus (PRV). To use the infection profile to assist genetic methods in identifying the likely farm of origin, samples from the farms were also tested for these viruses. However, in the current case, all the three farms had an infection profile that was similar to that of the escapees. We have shown that double-virus-infected escaped salmon ascend a river close to the likely source farms, reinforcing the potential for spread of viruses to wild salmonids.

 

[agentaqua]

http://onlinelibrary.wiley.com/doi/10.1111/jfd.12228/abstract

Health monitoring of wild anadromous salmonids in seawater in Norway

Abdullah S. Madhun, Cecilie Helen Isachsen, Linn Maren Omdal, Ann Cathrine Bårdsgjære
Einen, Vidar Wennevik, Terje Svåsand and Egil Karlsbakk

The Institute of Marine Research

 

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http://www.plosone.org/article/fetc....1371/journal.pone.0082202&representation=PDF

Phylogenetic Evidence of Long Distance Dispersal and Transmission of Piscine Reovirus (PRV) between Farmed and Wild Atlantic Salmon
A˚se Helen Garseth1,2*, Torbjørn Ekrem2, Eirik Biering1
1Department of Health Surveillance, Norwegian Veterinary Institute, Trondheim, Norway, 2Department of Natural History, Norwegian University of Science and Technology University Museum, Trondheim, Norway

Abstract
The extent and effect of disease interaction and pathogen exchange between wild and farmed fish populations is an ongoing debate and an area of research that is difficult to explore. The objective of this study was to investigate pathogen transmission between farmed and wild Atlantic salmon (Salmo salar L.) populations in Norway by means of molecular epidemiology. Piscine reovirus (PRV) was selected as the model organism as it is widely distributed in both farmed and wild Atlantic salmon in Norway, and because infection not necessarily will lead to mortality through development of disease. A matrix comprised of PRV protein coding sequences S1, S2 and S4 from wild, hatchery-reared and farmed Atlantic salmon in addition to one sea-trout (Salmo trutta L.) was examined. Phylogenetic analyses based on maximum likelihood and Bayesian inference indicate long distance transport of PRV and exchange of virus between populations. The results are discussed in the context of Atlantic salmon ecology and the structure of the Norwegian salmon industry. We conclude that the lack of a geographical pattern in the phylogenetic trees is caused by extensive exchange of PRV. In addition, the detailed topography of the trees indicates long distance transportation of PRV. Through its size, structure and infection status, the Atlantic salmon farming industry has the capacity to play a central role in both long distance transportation and transmission of pathogens. Despite extensive migration, wild salmon probably play a minor role as they are fewer in numbers, appear at lower densities and are less likely to be infected. An open question is the relationship between the PRV sequences found in marine fish and those originating from salmon.