DFO still says NO ISA???

[Charlie]

From: Fred Kibenge
To: Simon Jones@dfo-mpo.gc.ca
Date: 14/11/201111:28 AM
Subject: RE: Re: Paper
CC: Laura Richards; Mark Higgins; Mark Saunders; Molly Kibenge; Stewart ...

Dear Simon,

Thank you for your input on the publication question and for the information about the Cohen Commission.

For your information, this data (with your qualifiers below) is being relayed to CFIA as part of their on-going investigation. Our lab is currently making preparations for participation in this process and will disclose this work notwithstanding its age. I think that this historical data may also clarify some of the issues around recent ISAV testing in BC.

As you likely know, the OlE Manual of Diagnostic Tests for Aquatic Animals 2010 (http://www.cabi.org/ac/default.aspx?site= 162&page=332S) in the chapter on ISAV (http://www.oie.int/fileadmin/Home/eng/Health_standards/aahm/2010/2.3.0S_ISA .pdf) lists both criteria for suspicion and for confirmation of ISAV infection. This manuscript, which describes a targeted surveillance with significant prevalence rates of suspected infection in specific Pacific salmon species, accompanied by virus sequence data, may assist the CFlA.

Thanks.
Fred.


»> "Jones, Simon" <SimonJones@dfo-mpo.gc.ca> 11/4/2011 8:35 PM »>
Dear Molly,

Thanks for the message and for asking permission to submit the draft manuscript for publication.

Your email is timely. Recent events in BC concerning the alleged detection of ISA virus in wild Pacific salmon brought to mind the research you conducted while you were an NSERC Visiting Fellow at the Pacific Biological Station, and some of the questions it raised.

You may recall that Fish Health staff at DFO disagreed that your data supported the conclusion that ISA virus, whether asymptomatic or otherwise, occurred in the salmon you examined. For example, all attempts to isolate the virus into cell culture failed. As you are aware, the OlE case definition for ISAV infection requires either the isolation and identification of the virus into cell culture from two independent samples taken on two separate occasions, or the isolation and identification of ISAV in cell culture plus RT-PCR or serologic confirmation from tissue preparations. You may further recall that an independent laboratory was unable, on more than one occasion, to reproduce your RT-PCR results on the same samples. In my opinion, it will be very important to better understand the disagreement in laboratory results and to better test the hypothesis of "Asymptomatic ISAV" before moving towards publication.

On the related issue, CFIA is now conducting confirmatory testing of more recent samples from Pacific salmon in which initial positive RT-PCR results for ISAV were obtained. In addition, the Cohen Commission will reconvene for two days in December to hear evidence on ISA virus in British Columbia. I will wait to hear the outcome of these processes before further discussion on a 7-year-old manuscript. Consequently, I do not give permission to submit this work, whether in this manuscript or any other, for publication.

With my best wishes,

Simon
Dr. Simon R.M. Jones
Aquatic Animal Health Section
Pacific Biological Station
Fisheries and Oceans Canada
3190 Hammond Bay Road
Nanaimo, British Columbia
V9T 6N7, Canada
Tel: 2507298351
Fax: 250756 7053
E-mail: simon.jones@dfo-mpo.gc.ca


From: Molly Kibenge [mailto:Mkibenge@upeLca]
Sent: November 4,2011 7:19 AM
To: Jones, Simon
Cc: Fred Kibenge
Subject: Fwd: Re: Paper

Dear Simon,

I hope all is well in Nanaimo. It has been long since we last corresponded.

I am writing to follow up on the work I did in your lab during my post-doc at DFO-PBS, Nanaimo. As you recall we prepared a draft manuscript (attached). I am writing to ask your permission to submit it for publication as soon as possible. I would like to submit it to Diseases of Aquatic Organisms or Journal of Fish Diseases. What do you think?

Please let me know. If you have additional changes, please send them along and I will then proceed from there.

Please say hello to the family.

Best regards,
Molly.

 

[Charlie]

Asymptomatic infectious salmon anaemia in juvenile
Oncorhychus species from the North West Pacific Ocean

Kibenge Molly. T 1*, Jones Simon1, Traxler Garth1, Kibenge Frederick. S.B2

1Pacific Biological Station, Fisheries and Oceans Canada, Nanaimo, British Columbia, V9T 6N7, Canada,
2 Department of Pathology and Microbiology, Atlantic Veterinary College, University of Prince Edward Island, Charlottetown, PE, Canada CIA 4P3.

Running title: Surveillance of wild pacific salmon for viruses.

*Corresponding Author: Dr. Molly Kibenge;
E-mail: KibengeM@pac.dfo-mpo.gc.ca; Tel: (250) 729-8380;
Fax: (250) 756-7053

Abstract
Juvenile chinook (Oncorhynchus tshawytscha), chum (O. keta), coho (O. kisutch), pink (O. gorbuscha), and sockeye salmon (O. nerka) from the West Coast of Vancouver Island, Southeast Alaska, and the Bering Sea were surveyed between August 2002 and April 2003 for infectious salmon anaemia virus (ISAV). Spawning sockeye from the Cultus Lake and Kokanee from Lois Lake, BC population was also sampled. Pooled or individual tissues were tested by RTPCR, nucleotide sequencing and virus isolation. ISAV segment 8 was amplified from 34 of 121 (28%) chinook and 15 of 88 (17%) pink salmon caught off the west Coast Vancouver Island and southeast Alaska. ISAV segment 8 was also amplified from all 64 spawning sockeye and one cultured Atlantic salmon. The 220bp RT-PCR products were 94% to 98% homologous with Canadian ISAV isolates and 92% to 93% with European ISAV isolates. A product of 377 bp was obtained with Segment 7 ORF1 products were obtained in 5 chinook fish and the nucleotide sequence corresponded to ISAV segment 7 ORF2 products and was 95.7% identical to NBISA01 control isolate (Canadian isolate) and 99.7 % identity to an ISAV isolate 810/9/99 from Norway. ISAV segments 2, 6 and full opening frame for segment 8 were not amplified nor was ISAV isolated onto SHK or CHSE and ASK-2 cells. These results lead us to conclude that an asymptomatic form of ISA occurs among some species of wild Pacific salmon in the north Pacific.

KEYWORDS: Oncorhynchus, wild Pacific salmon, ISAV, Surveillance

Introduction
Viruses are important pathogens in marine-farmed fish however their importance as pathogens in wild stocks and the roles played by wild fish as reservoirs of infections are poorly understood. Infectious salmon anemia (ISA) is a severe disease of marine farmed Atlantic salmon, Salmo salar caused by an Orthomyxovirus-like virus, (Mjaaland et al., 1997) The disease is characterized by severe anemia, leucopenia, ascites and hemorrhagic liver necrosis and kidney necrosis. The disease has affected marine farmed Atlantic salmon in Norway since 1984 (Thorud and Djupvik, 1988). More recently the disease has been diagnosed in marine farmed Atlantic salmon in Eastern Canada (New Brunswick and Nova Scotia), Scotland, eastern USA (Maine), and the Faroe Islands (1998, Mullin et al., Ritchie et al., 2001, 1998, Rogers et al., 1998, Bouchard et al., 2001, Anonymous 2000). ISAV was isolated from farmed coho salmon in Chile (Kibenge et al., 2001). The virus has also been found in health farmed rainbow trout (Anon 2002). Although, Oncorhynchus species have not been shown to be affected by the natural disease and experimental infection, virus was recovered from chum, coho and steelhead trout after inoculation with a very high virus concentration of 107 TCID50 (Rolland and Winton, 2003). Also experimental infection with high concentration (TCID50 over 107 /200μl) of selected ISAV isolates caused mortality in rainbow trout (Oncorhynchus mykiss) (McWilliams et al., 2003, Kibenge unpublished data) indicating that some of Oncorhynchus species could be affected by ISAV and also serve as reservoirs of the virus. Natural and experimentally infected sea trout (Salmo trutta) and brown trout (Salmo trutto) become asymptomatic ISAV carriers and transmit the virus to Atlantic salmon by cohabitation (Raynard et al., 2001, Nylund and Jakobosen, 1995; Nylund et al., 1995; Nylund et al., 1997). Also ISA virus was isolated from asymptomatic wild Atlantic salmon (Olivier G, 2002, Raynard et al., 2001) Arctic char (Salvelinus alpinus) (Snow et al., 2001), pollock (Pollachius virens) from ISA infected cages and cod (MacLean et al., 2002). Thus wild marine fish stock may act as the virus reservoir and transmit the virus to farmed Atlantic salmon.Nylund et al., (2003) hypothesized that wild salmonids species are the source of benign ISAV isolates which become virulent after mutation either in the wild or in the farmed Atlantic salmon after transmission based on the analysis of the highly polymorphic region (HPR) of ISAV haemagglutinin gene. (Nylund et at., 2003). The Northwest Pacific coast of Canada and the United States are the only regions where marine Atlantic salmon farming is practiced that are still free of ISA disease. Because routine screening for the virus has been mostly done by virus isolation on cell culture, there is a possibility that low levels of virus in fish tissues or presence of non-pathogenic virus strains could go undetected. Although there is increased pathogen surveillance studies they are directed to culture fin fish. Therefore for this study, molecular biology technique of reverse transcription-polymerase chain reaction (RT-PCR) was added to diagnostic methods for investigation of ISAV in wild pacific salmon species. In this study five Pacific salmon species were examined for the presence of infectious salmon anemia (ISAV) virus.

Materials and methods
Fish, tissue collection and storage.
Juvenile chinook (Oncorhynchus tshawytscha), chum (O. keta), coho (O. kisutch), pink (O. gorbuscha), and sockeye salmon (O. nerka) from the West Coast of Vancouver Island, Southeast Alaska, and the Bering Sea were surveyed between August 2002 and April 2003 for infectious salmon anaemia virus (ISAV). Spawning sockeye from the Cultus Lake and Kokanee salmon from the Lois Lake were also sampled (Table 1). Organ tissues (head kidney, heart, spleen and liver) were harvested from partially thawed high sea pacific salmon and from freshly sacrificed sockeye, Kokanee and Atlantic salmon from Cultus Lake, Lois Lake and PBS respectively. Two of the 8 Atlantic salmon sampled had been on
salt water for a while the remaining 6 were only exposed to salt water for 2 weeks for the purpose of fungal treatment. Location and the weights of fish are shown in Table .1 in the results section. The samples were stored and -80C°individually or pooled until they were analyzed for virus by RT-PCR and cell culture.

Cells and Virus propagation.
Chinook salmon embryonic cells (CHSE-214) cells were grown in 24 or 48 well plates using Eagle’s minimum essential medium supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (Invitrogen Life Technologies) while salmon head kidney (SHK-1) and Atlantic salmon kidney (ASK-2) cells monolayers were grown using Leibowitz’s L-15 medium (Dannevig et al., 1997) The tissues were macerated and homogenised in presence of HBSS supplemented with 10% antibiotic/mycotic solution. The homogenate was then clarified by centrifugation at 3000 rpm for 15 minutes was inoculate on cell monolayer and incubated for 1hr at 15 °C to allow virus adsorption. After which MEM or L-15 supplemented with 4% FBS, 1% antibiotic/mycotic solution (Invitrogen Life Technologies) and HEPES (Sigma) was added to the wells. Wells of uninfected cells were left as negative controls and the plates were returned to the 15°C incubator. The cell monolayers were checked for development of CPE using inverted microscope. Normally ISAV causes CPE in CHSE-214 cells by 15 days while it may take up to 21 days for CPE to develop in SHK-1 cells.

 

[Charlie]

RNA extraction and reverse transcription-polymerase chain reaction (RTPCR).
Samples of kidney, liver, spleen and heart were harvested from individual fish and were either stored and analysed separately or pooled. The tissues were then macerated in presence of Hank’s balanced salt solution (HBSS) with 10% antibiotic solution. Total RNA was extracted from 300 μl of tissue homogenate or cell culture lysate in case of the control IHNV and VHSV using the Trizol LS Reagent (Invitrogen, Canada) following the manufacturer’s protocol. The RNA pellet was dissolved in 50 μl RNase free dionized water. The one step reverse transcription-polymerase chain reaction (RT-PCR) was performed using 1 μl (1 μg-2 μg) of RNA in a 25 μl volume reaction using one step RT-PCR kit (Qiagen). RT-PCR was performed in PTC-200DNA engine Peltier thermal cycler (MJ Research Inc.) Cycling conditions for all viruses consisted of once cycle of cDNA synthesis at 50°C for 30 minutes followed by 15 minutes at 95°C to activate the Hot Star Taq DNA polymerase and inactivate the reverse transcriptase. This was followed by 40 cycles, each consisting of denaturation at 95 °C for 30s, annealing at 61 °C for 45s in case of for ISAV and extension at 72 °C for 90s with final extension at 72 °C for 10 minutes. The primer sequence used in the RT-PCR for ISAV amplification of 220 bp fragment of segment 8 of the virus (Devold et al., 2000, Kibenge et al., 2001), and the nucleotide sequence for both the sense (342-363 nt) and antisense (532-552 nt) are (5’GAA GAG TCA GGA TGC CAA GAC G 3’, 5’GAA GTC GAT GAT TCG CAG CGA 3’) respectively. The samples sent to AVC were also analyzed with segment 7 ORF1 and segment 7OR2 primers. The two opening frames share the reverse primer and the primer nucleotide sequence and were designed using ISAV Accession # AF328627 (Ritchie et al., 2002). Here are the sequences for both sense and the antisense primers are: ISAV SEG7 (ORF1) FOR (23 mer from 1-23nt) 5'-ATG GAT TTC ACC AAA GTG TAT GG-3' , and ISAV SEG7 (ORF1) REV (23 mer from 881-903) 5'- TCA CAT TCT GAA GTG AAG TCCAG-3' and for the segment 7 ORF2 the sense primer is (82 mer from 1-63/590-608 nt) 5'-ATG GAT TTC ACC AAA GTG TAT GGT GTG CTG GTT GAC CAA CTA AAA CTT CAC GGA AAA GAC AAG GTG GCT TCT TTC CTG TCG G-3'. The PCR products were resolved by electrophoresis on 1.5% agarose gel with 0.01 mg ethidium bromide and visualized using the imaging system ( ). In samples that had a PCR product of 220 bp, amplification of the other ISAV gene segments (2, 6 and ORF1 and 2 for segment 8 proteins) was attempted. In the beginning of the study we included ISAV positive control to make sure that the reactions were working properly. We obtained positive RNA control from Atlantic Veterinary College (AVC) Virology Research laboratory (Charlottetown, PEI).

Cloning, sequencing and sequence analysis
The RT-PCR products were purified from the agarose gel using either Roche or Qiagen PCR purification kit and then cloned into pCR®II vector using a TOPO TA cloning kit (Invitrogen Life Technologies) in preparation for sequencing. Clones were screened for inserts by PCR and then positive ones were sequenced using big dye terminator sequencing kit (Applied Biosystems). The nucleotide sequences were trimmed and assembled using the Sequencer program (Gene Codes Corporation, Ann Arbor, MI) Nucleotide sequence pairwise comparison was performed using the BLAST program and Multialignment with the known segment 8 nucleotide sequences was carried out using multiple sequence alignment program (Corpet, 1988).

Results
Virus Isolation: No virus was isolated from any sample using SHK-1, CHSE-214 and ASK-2 cell lines.

RT-PCR and sequence analysis:
A 220 base-pair product was produced in the ISAV segment 8 RT-PCR assay of 116 of 520 (22.3%) samples. Table 1 shows the source and number fish samples tested with ISAV segment 8 primer. The number in the brackets denotes fish that had the expected PCR product of 220 bp. Alt least 2 clones for
each PCR product was sequenced.

Control ISAV NBISA01.
The RT-PCR product (220bp) was clone and sequenced and nucleotide sequence of our product (193 bp) was 99% identical to the original sequence of the same isolate in the GenBank.

Chinook: Of 121 samples analyzed with segment 8 primers, 31 had a 220 bp PCR product. The AVC Virology lab identified an additional 3 positive samples bringing the positive to 34(28%). DNA sequence from two samples was not that of ISAV (Table 2). Those that were sent to AVC that turned out positive were also tested with segment 7 primers and all positive products were sequenced. The Pacific salmon DNA inserts had an identity of 94% identity with NBISA01 +ve while among there was a homology ranging between 97% to 100%. All the samples that showed a positive product with segment 8 primers were analyzed with other ISAV genes (segment 2(PB1), segment 6 (HA) segment 7 both open reading frames and segment both open reading frames 1 and 2). The ones we tried here were all negative but from AVC virology lab samples (Table 2) had PCR product of 377 bp. This product corresponded with ISAV segment 7 ORF2 products and had an identity of 99.7% and 95.8% with ISAV isolates 810/9/99 from Norway and NBISA01 from New Brunswick, respectively.

Pink: Out of 89 fish and all the tissues organs were pooled, 15 (17%) were RTPCR positive with ISAV segment 8 primers. DNA sequence of 213 bp from one fish was had 97% identity to NBISA01 and most other North American Isolates and 92 % identity to European Isolates.

Atlantic salmon (salt water): RNA extracted from the heart of one fish was positive with segment 8 primer and PCR product of 211bp had 98% identity to most Canadian ISAV isolates including NBISA01 isolate in the 202 bp that were overlapping and 93% identical to European isolates in 197 nucleotides.

Sockeye: Although all Cultus lake sockeye samples had correct size PCR product, most of the products could not be clone, however fish # 7890C product was successfully cloned and sequenced and all three clones had a product of 241 bp. The nucleotide sequence of these inserts had identity to ISAV only in the primer sequence. However when all DNA inserts were aligned, using hierarchial clustering of nucleotides sequence (Corpet, 1988), they fell into three groups, with the inserts from Atlantic salmon, one from chinook (sample HS 2002 38 IVI02-124-019P), pink and the control NBISAV01 in one group, the other chinook samples ((HS200238-ISEA124-005L and (HS200238-IVI07-124-001h) fell into another group and the sockeye inserts had their own group although this group is close to the second group with the chinook samples.

Chum and coho salmon samples were all negative. ISAV segments 2 (PB1), 6 (HA) and full opening frame for segment 8 (NS1 and NS2) were not amplified nor was ISAV isolated onto SHK or CHSE cells. The expected RT-PCR products with the other ISAV genes used were obtained but they were obtained
Segment 7 ORF1 and ORF2. Five samples (Table 2) from Chinook salmon had PCR product of approximately 400bp with segment 7 ORF1 primers, while the positive control NBISA01 had an insert of 903 bp corresponding to segment 7 ORF1. These products were cloned and sequenced, the sequence obtained (377bp) corresponded with that of segment 7ORF2 nucleotide sequence. All chinook salmon samples the DNA inserts were 99.7% identical and corresponded to the ISAV Segment 7 ORF2 product. They had a 95.8% identity with the NBISA01 +ve control, and 99.7% identity with ISAV strain 810/9/99 from Norway. Segment 7 products were not amplified in any other fish samples.

Discussion.
Infectious salmon anaemia or ISA virus has not been detected in Pacific North west coast of Canada or USA. In this investigation we were able to detect ISAV segment 8 and segment 7 PCR products in juvenile chinook, pink, spawning sockeye and culture Atlantic salmon using RT-PCR. Sequence analysis of some of the product reveal a homology of 94-99% and 92-93% to the North American and European ISAV isolates, respectively. The fact that segment 8 and segment 7ORF2 sequences amplified in some samples were not 100% identical to the control NBISAV01 rules out contamination during sample processing. These results are not unique since Raynard et al. (2001) also reported detection of ISAV by RT-PCR in wild fish in Scotland without isolating the virus by cell culture even though they were able to amplify the full segment 8 and 2. Thus our results may imply that wild pacific salmon species have a nonpathogenic ISAV isolate or just non-infective particle of ISAV. It has been speculated from analysis of ISAV haemagglutinin gene that the wild salmon may harbour a non pathogenic ISAV isolate with no mutation in this gene and that after this ISAV isolate mutates by deletions it becomes pathogenic to Atlantic salmon (Nylund et al., 2003). Thus pacific salmon species could harbour non pathogenic ISAV that has not yet experienced any mutations thus is unable to infect cultured Atlantic salmon in the area. The fact that were able to amplify ISAV segment 8 sequences in salt water cultured Atlantic salmon which did not have ISA symptoms may attest to that fact and could also indicate that ISAV nucleotide are present in the sea water.

 

[Charlie]

In experimental infection of Oncorhynchus species, steelhead trout, (O. mykiss), chum (O.keta), chinook (O. tshawytscha) and coho (O. kitsutch), virus was recovered 13-15 d p.i only in fish that were given a very high dose although no disease was produced in any of these fish while the same viral doses cause mortality in Atlantic salmon show that the virus might be able to survive in these Pacific salmon species, therefore could act as carriers of ISAV (Rolland and Winton, 2003). In natural environment, the viral titre may be very low thus making it un detectable. We were not able to detect any nucleotide sequences in chum and coho in this study. Other diagnostic methods such as IFAT and should be used along with RT-PCR (Snow et al., 2003).

From these results, failure to isolate ISAV on cell culture combined with failure to amplify other genes of ISAV from juvenile pacific salmon species leads to the conclusion that there is a non-pathological ISAV isolate in the Pacific North west coast. In future use fresh material and in vivo experiments and use of serological methods may provide a different perspective.

It will be interesting to see how DFO tries to get out of this one!

 

[Whole in the Water]

Thanks for posting this info Charlie - good stuff!

I am wondering if you could try to put some of the scientific, biological, testing information in more laymans terms so the majority of us can understand what this information is saying. I think I speak for all lot of us on this issue we I say many of us can get confused/overwhelemd with a lot of this info since the majority of us our do not formal scientific backgrounds. Just a thought -if you can't/don't have time, no worries.

 

[Klob]

Good to see your on the case Charlie. But what is the "Coles Notes" on that data?

 

[agentaqua]

scary scenario

Wanna know my scary scenario?

1/ - Back a few years ago when the province was in charge - If the prov fish vet says it isn't ISA - it isn't. If he says it isn't and doesn't test - it isn't. If they have all the clinical signs and he doesn't test using RNA & PCR - it isn't. If it "isn't", then it goes unreported. In the farm fish health database, there are over 1100 fish with clinical signs of ISA - but the government can state that "no ISA has ever been found in BC". In the case of the River's Inlet samples - if the frozen samples get tested and they don’t show ISA at that time - the government can state that "no ISA has ever been found in BC" even though they reused frozen samples. Freezing juveniles is a practice common to preserve samples for many uses - but not normally used for DNA/RNA analysis. If they stay well frozen (below -20C) and not unthawed, but only ONCE – then the DNA/RNA is reasonably preserved. If not - then it is extremely difficult to get intact DNA/RNA from frozen/refrozen samples.

2/ ISA may have come-in with a shipment of Atlantic eggs destined for a BC fish farm from an uncertified hatchery in Iceland. DFOs Laura Richards successfully asks the DFO fisheries minister to override the fish health importation guidelines due to "low risk" and that the poor fish farmers will be at a "competitive disadvantage" if they don't. Laura wins. Eggs come in (this is all in the Cohen commission emails). I'd like to know why Laura is so sympathetic to fish farms. She also tried to muzzle DFOs Kristi Miller on the salmon anemia issue, as you all may know..

3/ there are at least 2 species of fish normally nearby fish farms - juvie coho salmon and juvie herring - both of which are susceptable to ISA and are often in close contact with each other in the salt chuck. You may all know this already.

the ISA virus can be carried by other salmonid species such as brown trout (Salmo trutta), sea trout (S.trutta) (Raynard et al. 2001), rainbow trout (Oncorhynchus mykiss) and, to a lesser extent Arctic charr (Salvelinus alpinus) (Snow et al. 2001), without the fish showing clinical signs of the disease. Unconfirmed positive tests for the virus have also been reported in Coho salmon (Oncorhynchus kisutch) (Kibenge et al. 2001), the European eel (Anguilla Anguilla) (Raynard 2000), wild plaice (Hippoglossoidesplatessoides ) (Mjaaland et al. in press) and possibly Atlantic herring (Clupea harengus) (Mullins et al. 1999) and haddock (Melanogrammus aeglefinus) (Mjaaland et al.).

4/ Coho can be asymptomatic - meaning they don't have to die and can stay carriers. The virus can also rapidly mutate like our common flu. coho show it when they come back at spawning time when they are stressed-out. They may take it back into the creeks where charr (Dolly Varden) and rainbow trout and 1st yr old coho juvies would then get it.

Horizontal transmission of ISAv in fresh water has been achieved experimentally (Brown et al. 1998) and occurs rapidly between infected and naive smolts in fresh water. Even under these conditions, asymptomatic smolts may remain infective to naïve parr for 18 months after the original challenge (Melville and Griffiths 1999)

5/ 2nd yr old coho often live on lakeshores, close to rearing sockeye. The juvie coho and resident charr & trout then transfer it to sockeye.

6/ River's inlet low productivity has been a concern for some time, and the variability has greatly increased in River's Inlet and other systems (e.g. Lakelse, Babine) in the past few years. The smolts had it LEAVING the system. This virus doesn't have to kill by itself - it lowers the immune response to other diseases.

7/ Herring then got it, and travelled up and down the coast, and mix with lots of juvie coho outside of many creeks. This coho/herring combo could then infect systems coast-wide.

8/ What happens if we test other systems showing the same population-level fluxes now? Will we find ISA?

THATS my worst-case nightmare...

 

[searun]

Great post "A" Scary set of circumstances, and from what you post it would appear we have not seen the end of this issue even though the Salmon Farmers and DFO want this thing buried away. Hard to imagine why DFO would not aggressively pursue a testing program to get to the bottom of this issue.

 

[searun]

Holmes, that is an interesting twist. Would be sweet justice. Salmon Farming brought to an end by a disease wild Pacific Salmon are immune to....love it.

 

[Charlie]

Wanna know my scary scenario?

1/ - Back a few years ago when the province was in charge - If the prov fish vet says it isn't ISA - it isn't. If he says it isn't and doesn't test - it isn't. If they have all the clinical signs and he doesn't test using RNA & PCR - it isn't. If it "isn't", then it goes unreported. In the farm fish health database, there are over 1100 fish with clinical signs of ISA - but the government can state that "no ISA has ever been found in BC". In the case of the River's Inlet samples - if the frozen samples get tested and they don’t show ISA at that time - the government can state that "no ISA has ever been found in BC" even though they reused frozen samples. Freezing juveniles is a practice common to preserve samples for many uses - but not normally used for DNA/RNA analysis. If they stay well frozen (below -20C) and not unthawed, but only ONCE – then the DNA/RNA is reasonably preserved. If not - then it is extremely difficult to get intact DNA/RNA from frozen/refrozen samples.

Not so sure about your scary scenario? I believe IMHO - That is already FACT!
Yep! As the government of Canada states, "there are no confirmed cases of ISAv ever been found in BC.

2/ ISA may have come-in with a shipment of Atlantic eggs destined for a BC fish farm from an uncertified hatchery in Iceland. DFOs Laura Richards successfully asks the DFO fisheries minister to override the fish health importation guidelines due to "low risk" and that the poor fish farmers will be at a "competitive disadvantage" if they don't. Laura wins. Eggs come in (this is all in the Cohen commission emails). I'd like to know why Laura is so sympathetic to fish farms. She also tried to muzzle DFOs Kristi Miller on the salmon anemia issue, as you all may know..

Yep, here also! However, if you really start digging you will find DFO has known BC has had the ISAv since the the early 90's. If you get down to it, they confirmed ISAv in 1998, right before they dropped the research program back then. IMHO Dr Laura Richards is well aware of this fact, so why ban any egg imports from anywhere? ISAv is already in BC and DFO knows it!

3/ there are at least 2 species of fish normally nearby fish farms - juvie coho salmon and juvie herring - both of which are susceptable to ISA and are often in close contact with each other in the salt chuck. You may all know this already.

the ISA virus can be carried by other salmonid species such as brown trout (Salmo trutta), sea trout (S.trutta) (Raynard et al. 2001), rainbow trout (Oncorhynchus mykiss) and, to a lesser extent Arctic charr (Salvelinus alpinus) (Snow et al. 2001), without the fish showing clinical signs of the disease. Unconfirmed positive tests for the virus have also been reported in Coho salmon (Oncorhynchus kisutch) (Kibenge et al. 2001), the European eel (Anguilla Anguilla) (Raynard 2000), wild plaice (Hippoglossoidesplatessoides ) (Mjaaland et al. in press) and possibly Atlantic herring (Clupea harengus) (Mullins et al. 1999) and haddock (Melanogrammus aeglefinus) (Mjaaland et al.).

"ALL" salmon can carry the ISAv. That doesn't mean they will die of ISAv, it just means they will probably die of someting. Start looking at (and why) all those farmed Chinook have been dying since the early 90's. DFO even decided to gave it a new name!

4/ Coho can be asymptomatic - meaning they don't have to die and can stay carriers. The virus can also rapidly mutate like our common flu. coho show it when they come back at spawning time when they are stressed-out. They may take it back into the creeks where charr (Dolly Varden) and rainbow trout and 1st yr old coho juvies would then get it.

Horizontal transmission of ISAv in fresh water has been achieved experimentally (Brown et al. 1998) and occurs rapidly between infected and naive smolts in fresh water. Even under these conditions, asymptomatic smolts may remain infective to naïve parr for 18 months after the original challenge (Melville and Griffiths 1999

5/ 2nd yr old coho often live on lakeshores, close to rearing sockeye. The juvie coho and resident charr & trout then transfer it to sockeye.

6/ River's inlet low productivity has been a concern for some time, and the variability has greatly increased in River's Inlet and other systems (e.g. Lakelse, Babine) in the past few years. The smolts had it LEAVING the system. This virus doesn't have to kill by itself - it lowers the immune response to other diseases.

7/ Herring then got it, and travelled up and down the coast, and mix with lots of juvie coho outside of many creeks. This coho/herring combo could then infect systems coast-wide.

8/ What happens if we test other systems showing the same population-level fluxes now? Will we find ISA?

THATS my worst-case nightmare...

Everything in the above has already happened! Yep, nothing worry about!

DFO already knows ISAv exists in BC, which is now coming to light. Yes, ISAv is killing some of the wild salmon as we speak - there is no doubt about that in my mind. I recently posted, it will be true justice will when the "wild" salmon give this disease right back to the farmed and wipes the entire "farmed" salmon industry out and they leave BC as they did everywhere else. IMHO that day is coming sooner than later.

Here is your kicker! Canada (DFO) knows ISAv exists in BC. Canada (DFO) is not doing anything about it. Canada (DFO) knows it is currently killing wild salmon, see number 6) above. Canada (DFO) knows the biggest threat is to those penned up Atlantic salmon. Canada (DFO) knows it is not harmful to humans - Let me add to that, YET!

Here is the real threat that Canada seems to be forgetting - This is a VIRUS! This virus truely has the ability to mutate and kill every salmon on the West Coast. Then dealing with a virus, first look up what is referred when the word "mutate" is used! A virus needs a host. You want the real "scary senario"? When the host of a virus is finally eradicated, the virus will search for another host. A virus can change sybiotic sorces in a matter of hours/days, hence comes the word "mutation." Which means the virus is just "adapting" to another host. Will it be deadly to that new host - who knows. I assure, DFO sure doesn't! Let me give a prime example of exactly how and what can happen when a virus mutates? Look up the virus that started out as SIV. You will find it first mutated into two strains; SIVcpz and SIVgon. Then you will find both of those strains mutated into another virus called - HIV. Yep, nothing worry about when it comes to viruses mutating.

"Two types of Human Immunodeficiency Virus (HIV) infect humans: HIV-1 and HIV-2. HIV-1 is the more virulent and easily transmitted, and is the source of the majority of HIV infections throughout the world; HIV-2 is largely confined to west Africa. Both types originated in west and central Africa, jumping from primates to humans. HIV-1 has evolved from a Simian Immunodeficiency Virus (SIVcpz) found in the common chimpanzee subspecies, Pan troglodytes troglodytes, native to southern Cameroon. Kinshasa, in the Democratic Republic of Congo, has the greatest genetic diversity of HIV-1 so far discovered, suggesting that the virus has been there longer than anywhere else. HIV-2 crossed species from a different strain of SIV, found in the Sooty Mangabey, monkeys in Guinea-Bissau."

 

[cuttlefish]

Hey, Charlie. Can you point me to the info where DFO confirmed ISAv in BC in 1998? It did not come out during the Commission hearings and Cohen should know about this.

 

[Charlie]

All one has to do is read Dr Kristy Millers transcript regarding to her study and what she found out about concerning DFO's (and DFO's only) new disease found in the 90's killing farmed Chinook that they named Salmon Leukemia (SL). You will note when she probed into it she found no one even new what SL was, even the ones who did the actual research. That is actually in their testimonies. When Miller started asking questions concerning just exactly what was SL was - is when DFO stopped funding her work.

Read up on ISAv, its symthoms, how it will cause death, and hard ISA is to detect; that is until it has actually caused mass deaths in "farmed" fish. Look at the reasearch for SL, which was only preformed by DFO. They do have both published and unpublished articles on it. Compare the symthoms of SL and ISAv. ( why at it, even BKD). Then ask DFO why "they" stopped the uncompleted research work on a new disease "they" found that was killing "farmed" Chinook salmon they called Salmon Leukemia in 1998. Then ask why DFO doesn't test for ISAv in every Atlantic salmon "they" say died from sympthoms of both SL and BDK - those are all sympthoms of ISAv.

I believe that will answer your questions and one will find Salmon Leukemia (SL) is in fact a mutated form of ISAv. You will also find DFO (and now, including Dr Miller) know it!

 

[Whole in the Water]

Very interesting stuff Charlie. You should forward this info to Alexandra Morton and others and talk to her about it before she goes the the special ISAv session of the Cohen Commission next week!

Here is her link: http://alexandramorton.typepad.com/

 

[Charlie]

Alex is under a "gag order"! Not speaking for Alex (she does that quite well herself) IMHO she has
already found and knows this; however, she and all the others under the gag order, can't discuss it even among themselves! The unpublished 2004 report completed by DFO confirming ISAv found in "wild" salmon tested 2002 has already been sent to Cohen.

However, if you read up on ISAv - you will find there are NO confirmed cases of ISAv in BC (or anywhere else. Might I add, there never will be any "confirmed", that is until it kills off all those "farmed" Atlantic salmon!

 

[GLG]

Thanks Charlie for the info. Sure would like to know all the different types of ISA and there other names.
I suppose I could have asked at this seminar but I missed it today.
Perhaps those that attended will now be able properly ID this virus we are seeing here on the west coast.

Pacific Biological Station 2011 Seminar Series


Speaker: Dr. Knut Falk, Norwegian Veterinary Institute, Oslo, Norway
Subject: An overview of ISA and characteristics of ISA virus
Date: December 5, 2011, 9:00am-10:00am.
Abstract:
Infectious salmon anaemia (ISA) is an emerging disease in the Atlantic salmon (Salmo salar) farming industry, it may cause high mortality with detrimental effects on production, and is listed by the OIE. ISA was first described in 1984 in Norway and since then, disease outbreaks have occurred in Canada, Scotland, the Faroe Islands, USA and Chile. The host range of ISA virus in which disease occurs naturally, is restricted to farmed Atlantic salmon. However, natural infection without disease has been detected in wild brown trout (Salmo trutta), wild Atlantic salmon and wild rainbow trout (Oncorhynchus mykiss).
The causative agent of ISA is the ISA virus, the prototype virus of genus Isavirus in the family Orthomyxoviridae. Accumulated mortality in ISA outbreaks varies from 10% to 95% during a several month period, but intervention by fish health authorities implies that outbreaks normally are not allowed to run to completion. Clinical signs and pathological findings in the terminal stage are severe anaemia with haematocrit below 10%, exophthalmia, pale gills, ascites and haemorrhages in several organs suggesting circulatory failure.
The presentation will give an overview of ISA virus characteristics, and ISA pathology, epidemiology, diagnostics and disease control.
For more information please contact: seminar@pac.dfo-mpo.gc.ca
http://www.pac.dfo-mpo.gc.ca/science/events-evenements/index-eng.htm#Pacific_Biological_Station

 

[cuttlefish]

Thanks, Charlie.
I'll look for that testimony and those SL papers.

 

[agentaqua]

One of the bigger problems with that scenario is that taxpayers (NOT the open net-cage fish farm corporations) would be on the hook to pay - like they did in New Brunswick like 10 years ago. Today, with the price per lb paid for farm-gate Atlantic salmon so low - we would be doing the fish farmers a favour if we paid them insurance costs. It wouldn't get them out of the water and into closed containment...

 

[agentaqua]

Thanks Charlie for the info. Sure would like to know all the different types of ISA and there other names.
I suppose I could have asked at this seminar but I missed it today.
Perhaps those that attended will now be able properly ID this virus we are seeing here on the west coast.

Pacific Biological Station 2011 Seminar Series


Speaker: Dr. Knut Falk, Norwegian Veterinary Institute, Oslo, Norway
Subject: An overview of ISA and characteristics of ISA virus
Date: December 5, 2011, 9:00am-10:00am.
Abstract:
Infectious salmon anaemia (ISA) is an emerging disease in the Atlantic salmon (Salmo salar) farming industry, it may cause high mortality with detrimental effects on production, and is listed by the OIE. ISA was first described in 1984 in Norway and since then, disease outbreaks have occurred in Canada, Scotland, the Faroe Islands, USA and Chile. The host range of ISA virus in which disease occurs naturally, is restricted to farmed Atlantic salmon. However, natural infection without disease has been detected in wild brown trout (Salmo trutta), wild Atlantic salmon and wild rainbow trout (Oncorhynchus mykiss).
The causative agent of ISA is the ISA virus, the prototype virus of genus Isavirus in the family Orthomyxoviridae. Accumulated mortality in ISA outbreaks varies from 10% to 95% during a several month period, but intervention by fish health authorities implies that outbreaks normally are not allowed to run to completion. Clinical signs and pathological findings in the terminal stage are severe anaemia with haematocrit below 10%, exophthalmia, pale gills, ascites and haemorrhages in several organs suggesting circulatory failure.
The presentation will give an overview of ISA virus characteristics, and ISA pathology, epidemiology, diagnostics and disease control.
For more information please contact: seminar@pac.dfo-mpo.gc.ca
http://www.pac.dfo-mpo.gc.ca/science/events-evenements/index-eng.htm#Pacific_Biological_Station

Did anyone attend that lecture?

Another lecture series that may be of interest:

Free Public Lecture
Salmon Farming in BC: Innovation and Evolution

http://web.viu.ca/simmsw/Lecture2011/2. posterRoberts.pdf

Wednesday, January 19, 2011
7:00 - 8:00 p.m.
Vancouver Island University, Building 356, Room 109

 

[agentaqua]

Holmes, that is an interesting twist. Would be sweet justice. Salmon Farming brought to an end by a disease wild Pacific Salmon are immune to....love it.

One of the bigger problems with that scenario is that taxpayers (NOT the open net-cage fish farm corporations) would be on the hook to pay - like they did in New Brunswick like 10 years ago. Today, with the price per lb paid for farm-gate Atlantic salmon so low - we would be doing the fish farmers a favour if we paid them insurance costs. It wouldn't get them out of the water and into closed containment...

 

[Charlie]

holmes, I wouldn't be laughing to much and you might want to check... Canada DID pay them, and that actually did happen! YOUR tax dollars at work!