More Information from Investigation of KHV

[Steve E.]

I recently received a pdf copy of another study into KHV. I have posted the abstract here along with a link that will take one to the complete report. Please note the copywrite statement in bold below that permits distribution with acknowledgement. I have posted only the Abstract as the entire report is very large. The authors listed are the main authors with another page in the document that shows others who have contributed equally. I have also put the url for the complete article in bold.

Steve


Skin mucus of Cyprinus carpio inhibits cyprinid herpesvirus 3 binding to
epidermal cells
Veterinary Research 2011, 42:92 doi:10.1186/1297-9716-42-92
Victor STALIN Raj (s.victor@erasmusmc.nl)
Guillaume Fournier (g.fournier@ulg.ac.be)
Krzysztof Rakus (krzysztof.rakus@fish.edu.pl)
Maygane Ronsmans (mronsmans@ulg.ac.be)
Ping Ouyang (ouyang.ping@live.cn)
Benjamin Michel (benjamichel@yahoo.fr)
Cedric Delforges (cedric.delforge@ulg.ac.be)
Berenice Costes (bcostes@live.be)
Frederic Farnir (f.farnir@ulg.ac.be)
Baptiste Leroy (baptiste.leroy@umons.ac.be)
Ruddy Wattiez (ruddy.wattiez@umons.ac.be)
Charles Melard (C.Melard@ulg.ac.be)
Jan Mast (Jan.Mast@var.fgov.be)
Francois Lieffrig (f.lieffrig@cergroupe.be)
Alain Vanderplasschen (a.vdplasschen@ulg.ac.be)
ISSN 1297-9716
Article type Research
Submission date 2 March 2011
Acceptance date 4 August 2011
Publication date 4 August 2011
Article URL http://www.veterinaryresearch.org/content/42/1/92
This peer-reviewed article was published immediately upon acceptance. It can be downloaded,
printed and distributed freely for any purposes (see copyright notice below).
Articles in Veterinary Research are listed in PubMed and archived at PubMed Central.
For information about publishing your research in Veterinary Research or any BioMed Central
journal, go to
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© 2011 Raj et al. ; licensee BioMed Central Ltd.
This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0),
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited

ABSTRACT
Cyprinid herpesvirus 3 (CyHV-3) is the aetiological agent of a mortal and highly
contagious disease in common and koi carp. The skin is the major portal of entry of
CyHV-3 in carp after immersion in water containing the virus. In the present study, we
used in vivo bioluminescence imaging to investigate the effect of skin mucus removal and
skin epidermis lesion on CyHV-3 entry. Physical treatments inducing removal of the
mucus up to complete erosion of the epidermis were applied on a defined area of carp
skin just before inoculation by immersion in infectious water. CyHV-3 entry in carp was
drastically enhanced on the area of the skin where the mucus was removed with or
without associated epidermal lesion. To investigate whether skin mucus inhibits CyHV-3
binding to epidermal cells, tail fins with an intact mucus layer or without mucus were
inoculated ex vivo. While electron microscopy examination revealed numerous viral
particles bound on the fins inoculated after mucus removal, no particle could be detected
after infection of mucus-covered fins. Finally, anti-CyHV-3 neutralising activity of mucus
extract was tested in vitro. Incubation of CyHV-3 with mucus extract reduced its
infectivity in a dose dependent manner. The present study demonstrates that skin mucus
removal and epidermal lesions enhance CyHV-3 entry in carp. It highlights the role of
fish skin mucus as an innate immune protection against viral epidermal entry.

[Brockp]

Thanks for that Steve;

I found the most interesting observation at the end where they show that the mucus contains a virus inhibiting agent. With our modern industrial methods it won't be hard to separate out this inhibiting agent from mucus.

I am not saying that it will lead to a cure but anything that gives us an insight into families of agents that inhibit this nasty virus is a major step forward. In fact it is how the treatment for HIV started and now, if the patients can tolerate the medicines HIV infection is a controllable disease.

I just hope the research group have the funding in these difficult financial times to carry on their work.

Peter

[Duncan]

its no secret that the cutical has antiviral AND antibacterial agents, but sorry guys i just dont buy this:

The skin is the major portal of entry of
CyHV-3 in carp after immersion in water containing the virus.

if i was the virus i would choose the gill for entry over everthing else every day of the week and twice on sundays why go through such a huge thick barrier like the cutical, epidermis, hyperdermis, and dermis when the distance and thickness at the gill between the pond water and the blood is only one cell thick ? thats UNO cell thick!!

this make no sense to me at all

dunc

[Brockp]

Hi;

I agree with Dunc but the finding of a anti viral agent which is dose dependent (the best test to see if agents work) is interesting.
When you read the whole article they refer two previous publications;

http://www.ncbi.nlm.nih.gov/sites/entre ... 00m,isrctn

http://www.ncbi.nlm.nih.gov/sites/entre ... 00m,isrctn

The second abstract did compare the routes of entry of viruses in carp and did look at the gills as well as other organs. I quote from the abstract of this publication;
“To identify the KHV portal of entry, carp were analyzed by bioluminescence imaging at different times post infection with the FL BAC 136 LUC TK revertant strain. These analyses demonstrated that the skin of the fish covering the fins and also the body is the major portal of entry for KHV in carp. Finally, to further demonstrate the role of the skin as the KHV portal of entry, we constructed an original system, nicknamed "U-tube," to perform percutaneous infection restricted to the posterior part of the fish. All the data obtained in the present study demonstrate that the skin, and not the gills, is the major portal of entry for KHV in carp.”

From my limited knowledge of the science the research looks robust.

So I agree with Dunc that it seems unreasonable for the KHV virus to choose to penetrate through the skin and not the gills or the gut but it wouldn’t be unique. Herpes in man is quiet specific about which tissues in enters through as is polio. Other viruses seems to have a strange affinity for only one tissue, the chicken pox virus stays around in just one small set of nerve cells in the spinal column after recovery and may later go on to give rise to Shingles in just the localised area served by those nerve cells.

Isn’t nature funny. Dunc will now prove I am talking total nonsense bit it is nice to see him back stimulating us to think about difficult issues

Peter

[Duncan]

Hi
Steve is one of the good guys and I’m glad he posted this, but its plain off base and wrong on so many levels
This is a huge list of credits to give it credence but is wrong plain and simple, But i dare say many will take it at face value and quote it without putting the grey matter into gear.

The fact that there are antivirus inhibitors in the mucus is not saying much given the virulence of this disease if that’s the best it has to offer I would not like to see the worst case scenario

But apart from that I dare say that some virus molecules enter through the skin but to say it’s the major point of entry again is so wrong it’s the gill and the gut as there is no effort required for entry and no mucus to overcome

But let’s say the mucus has loads of antiviral inhibitors it’s a moot point if the entry at the gill and gut is so easy for the virus and requires no effort

let’s put it another way it’s like fortifying Buckingham palace against a terror attack by bolstering the front gates with everything you can muster and extra troops but the back door is left wide open??

dunc

[Gazza]

Hi Dunc,

I had read through the post and i was basically going to post ni on similar to you that we lock up all the front of the house but leave the patio doors open lol

I have to say although i can see the virus will try to entire any way it can as it will be as opportunistic as a parasite but like most things in mother nature they are bloody clever so why make it harder than it needs to be. So try to get through the mucus layer then try to burrow through the layers of dermis when it can entire into the gills where there is a very very thin layer and an easy access to the blood stream

This is just my take on it from a Muppet's point of view and i am in no way po poing the great work these guys are doing.

[Steve E.]

I have recently received some information referencing a study of environmental-organic places where the virus may be found in concentrations. Not sure what it all means and I post it for your consideration. It is just one of what I have found is a long string of studies completed or in progress around the world. This URL will take one to the site where the abstract came from: http://www.ncbi.nlm.nih.gov/pubmed/21983526. The work is of course not mine and I post it believing that permission has been granted for public dissemination. My take on these studies is a simple one, this virus is complicated and we just plain do not yet have a handle on the where, when, and how of it at this time. Latency has been found in white blood cells (AKCA Project KHV Study) and now this abstract discusses other residing places for the virus. There are several links at the above URL that will take one to more abstracts that are centered around the aquatic environment. Enjoy for the information.

Also, the AKCA's Project KHV Study with Dr Jin has been extend for an additional year for no increase in the fee for the study. Having met Dr. Jin and her team I remain convinced the funds raised for research amongst the Koi Hobbyists are being well spent.

Steve

Vet Microbiol. 2011 Sep 21. [Epub ahead of print]
Reservoirs of Cyprinid herpesvirus 3 (CyHV-3) DNA in sediments of natural lakes and ponds.
Honjo MN, Minamoto T, Kawabata Z.
Source

Research Institute for Humanity and Nature, 457-4 Motoyama Kamigamo, Kita-ku, Kyoto 603-8047, Japan.
Abstract

Cyprinid herpesvirus 3 (CyHV-3) is a lethal DNA virus that infects common carp and koi. It has caused outbreak of the disease within both aquaculture and natural environmental ecosystems. However, there is not enough understanding of the distribution of CyHV-3 in the natural environments, partly because there is no suitable quantification method. In this study, we tested CyHV-3 extraction methods from sediment and then compared its abundance between sediment and water using real-time PCR. Sediment samples were taken from lake and pond, and total viral DNA was extracted using the viral elution method recommended by the US Environmental Protection Agency (manual method), as well as a commercial DNA extraction kit for soil (commercial kit method) before PCR detection. 7 of 12 (58%) and 5 of 10 (50%) sediment samples showed PCR positive signal for CyHV-3 DNA using the manual method and the commercial kit, respectively, and consistent results were obtained from the samples using the manual method between two independent primer sets. The quantification of CyHV-3 DNA in natural sediment using the manual method and external standard virus revealed that its concentration was 1.2×10(4) to 3.3×10(5) copies DNA/kg. The concentration in sediments was 46-1238 times higher than that in water from the same location, suggesting that sediment could act as a reservoir for CyHV-3 in natural freshwater environments. This is the first report of the existence of CyHV-3 in the sediment of a natural lake or pond.

Copyright © 2011 Elsevier B.V. All rights reserved.

PMID:
21983526
[PubMed - as supplied by publisher]

[Brockp]

Hi Steve;
Interesting post and a worrying bit of work. The methodology is state of the art which makes the findings that viable virus can lie dormant in sediment very worrying. It is unusual for viruses to exist for any length of time outside of their hosts. The reason these findings are worrying is, that if true, it is going to prove impossible to eradicate the virus like we have done with small pox and are doing with polio.
As you say we have a lot to learn about this virulent bug before we can hope to produce effective controls.

The next queston is how long are the viral particles detected in the sediment viable for..... I am sure this followup work is being done.

Peter

[kimr]

Hi

Having just read this I am shocked to read that the virus enters the Koi through a broken mucus barrier, the cases of KHV I have known about ALL pond inhabitants were fit and healthy and in tip top condition. The parasite that appears with KHV is Costia which is a symptom not a cause, and is it not a case that Koi suffering from Costia cause more mucus, which would then mean the barrier is even harder to break through. I also firmly believe that part of the answer to find a cure were distroyed along with all pond mates as is the norm when dealing with KHV.

When children we are given a small dose of polio to build up anti-bodies so we dont get polio, so maybe the answer is to inoculate against instead of preventing, maybe the same will happen with people against HIV. I honestly believe this awful virus will be with us for a very long time, and it is going to take a very long time for anyone to agree what is the best action to take as no-one really agrees or knows what we are truely dealing with. There are so many questions to be answered before a final decission on which coarse is best to take. I think if you asked a Koi keeper who have lost there Koi through KHV about the condition of the Koi before the onset of KHV they will in the majourity say their Koi were fine.

[Steve E.]

I think from what I see is that KHV is becoming spread through the natural progression of a disease adapting. I believe there is latent presence of the virus in more fish that we are aware of to date. I think this is just one more thing we will have to watch for in buying new koi and emphasizes the need for quarantine. As I said the virus is being studied in many places and there will be more to come. What I hope you all take from these posts is a knowledge to allow one to effectively manage the health of your fish from one more point of view. There are folks around who remember when aeromonas was the new killer and we have learned to effectively manage that disease.

Steve