1. No category

– konsekvenser for oppdrett og forvaltning
Høgskolen i Bodø: Kjetil Korsnes
www.hibo.no
Nodavirus
Havforskningsinstituttet: Audun Helge Nerland, Egil Karlsbakk, Sonal Patel
OMJ-98
Universitetet i Bergen: Are Nylund, Trond Isaksen
Nodavirus – betanodavirus – nervous necrosis virus NNV
www.hibo.no
The virus:
• Family Nodaviridae
• 25-40 nm
• ssRNA
•RNA1 (encode RdRp)
•RNA2 (encode protein a)
•RNA3 (subgenomic – B1 og B2)
40
40
• Viral nervous necrosis VNN or
• Viral encephalopathy and
retinopathy VER
OMJ-98
The disease:
Phylogenetic relationship of NNV
AJ245641
SJNVCP2
74
AB046377
1. BFNNV
71 AF160473
AHNNV
EF061913
Europe
1.Barfin flounder
EF061912
81
2. SJNNV
1.Striped jack
98
3. RGNNV
1.Red-spotted grouper
86
AY962682
AJ698094
GmH13/06
GmN10/06
GmSF15/07
GmH14/06
90
GmMR12/0
92
6
GmMR11/06
100 AF445800
100 AY547548
98 AY547549
AY547547
4. TPNNV
GMNNV
Norway
ACNNV
North America
94 AY140796
100 SJNVCP3
99
SJNVCP1
72
1.Tiger puffer
BFNNV
79
www.hibo.no
Major clades:
RGNNV
AB045980
TNNV
1.Turbot nodavirus
Nishizawa et al. 1997, TNV: Johansen et al. 2004
TPNNV
AB056572
SJ-G91
SJNNV
0.1
Phylogenetic tree based on RNA2, ML criteria.
From: Nylund et al. 2008
OMJ-98
5. TNV?
VNN in Norway
www.hibo.no
Farmed fish:
• Turbot
•Bloch et al. 1991
•Johansen et al. 2004
• Atlantic halibut
•Grotmol et al. 1995
•Aspehaug et al. 1999
•Patel et al. 2007
Wild fish:
• Nylund et al. 2008
• Unpublished data
Figure 3. Histological sections of Atlantic cod, Atlantic salmon and turbot
immunolabeled (red colour) with NNV specific antibodies, targeting the
viral capside protein. A) Strong immunolabeling (arrows) in a section of
retina in experimental i.p. challenged Atlantic cod. EP = external plexiform
layer, EL = external nuclear layer, BS = basal layer of visual cells. B)
Section of brain in experimental i.p. challenged Atlantic salmon with
immune positive cells (arrows) scattered in the parenchyma. C) Strong
immunolabeling of a large area (indicated by arrows) in a section of brain
from experimental i.p. challenged turbot. In the immunolabeled area,
vacuolation of cells are visible. D) Immunelabeling of a section of retina
from experimental i.p. challenged turbot. Vacuolated cells are visible within
EL and EP layers. Scale bars = 20 mm.
OMJ-98
• Atlantic cod
Order
Genus
Gadus morhua
Gadiformes
Name
Geographical location
Reference
Wild/farmed
Norway
Nylund et al. (2008)
wild
Norway
Patel et al. (2007)
farmed
Canada
Johnson et al. (2002)
farmed
UK
Starkey et al. (2001)
farmed
Japan
Nishizawa et al. (1997)
unknown
Canada
Gagné et al. (2004)
farmed
Canada
Johnson et al. (2002)
farmed
Atlantic cod
Gadus macrocephalus
Pacific cod
Melanogrammus aeglefinus
Haddock
Pollachius virens
Saith
Norway
Nylund et al. (2008)
wild
Pollachius pollachius
Pollock
Norway
Nylund et al. (2008)
wild
Pleuronectes americanus
Winter flounder
USA
Barker et al. (2002)
wild
Solea solea
Dover sole / common sole
UK
Starkey et al. (2001)
farmed
Hippoglossus hippoglossus
Atlantic halibut
UK
Starkey et al. (2000)
farmed
Hippoglossus hippoglossus
Atlantic halibut
Norway
Grotmol et al. (1995)
farmed
Pleuronectes platessa
Plaice
Norway
Nylund et al. (2008)
wild
Verasper moseri
Barfin flounder
Japan
Nishizawa et al. (1997)
unknown
Perciformes
Scomber scombrus
Mackerel
Norway
Nylund et al. (2008)
wild
Lophiiformes
Lophius piscatorius
Monkfish
Norway
Nylund et al. (2008)
wild
www.hibo.no
Susceptible hosts in BFNNV clade
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Pleuronectiformes
OMJ-98
www.hibo.no
Possible routes of transmission of NNV
NNV as a pathogen in Norwegian fish farming
VNN in farmed Atlantic cod. Photo: Sonal Patel
OMJ-98
www.hibo.no
1. Any susceptibility in
Atlantic salmon?
2. Horizontal transmission to
salmon and cod
3. NNV tissue tropism in adult
cod: organ candidates for
non-lethal biopsy
4. Prophylaxis and control
Experimental i.p. challenged Atlantic salmon
www.hibo.no
• I.p. challenged smolt
• NNV from Atlantic halibut
• Results:
1. Clinical signs
2. Mortality
3. Detection of NNV in brain
Korsnes et al. (2005)
OMJ-98
Brain of i.p. challenged salmon. Nodavirus-like particles present in
membrane bound vacuoles. Scale bar = 0.2 mm. Photo: K. Watanabe.
Cohabitation of NNV i.p.
injected turbot and
healthy cod and salmon:
1. No transmission of NNV
from turbot to salmon
Table 3. Real-time RT-PCR detection of NNV positive tissues in experimental Group
IV in i.p. challenged turbot and cohabitant Atlantic cod at Day 90 and 125. Detection
of NNV, where x = Ct values >35, xx = Ct values 35 – 25, xxx = Ct values <25, n/a =
samples not available. Fish challenged with virus strain AAG/03.
Fish no.
Brain
Retina/eye
Spleen
i.p. challenged D125
1 2 3
cohabitant D90
1 2 3 4 5
xxx
xx xx xx
x
xxx
xx xx
x
xx
xx
cohabitant D125
1 2 3 4 5
x
x
x
x
x
x
x
n/a
x
x
x
www.hibo.no
Experimental horizontal transmission of NNV to salmon and cod
2. Transmisson of NNV
from turbot to cod
NNV isolate applied from
Atlantic halibut
Fig. 1. Scophthalmus maximus . Immunohistochemistry staining of turbot brain at Day 125,
challenged with NNV as i.p. injection. (A) Large area of NNV immunepositive brain cells
(arrows) in turbot from experimental Group IV. Vacuolated cells are visible within the
immuneolabelled area. (B) Immunopositive cells (arrow) in brain from turbot in experimental
Group I. Scale bar = 20 mm.
OMJ-98
Korsnes et al. (in prep)
Tissue tropism of NNV in experimental challenged cod
• I.p. challenged cod
35
30
Day 25
Day 130
15
10
5
oo
d
Bl
e
l
ai
n
Br
Ey
ra
La
te
dn
ey
r
Ki
Li
ve
e
le
en
Sp
ric
l
nt
el
m
Ve
At
riu
rb
Ba
ct
or
al
ills
0
Pe
1. Migration of NNV from i.p.
injection locus
2. Neuroinvasive
3. Tissue tropism similar to
natural infected cod
4. Immunosuppression:
20
G
• Results:
25
Ct value
• NNV from Atlantic halibut
www.hibo.no
Ct tissues Day 25 and 130 post challenge
Tissue
Figure 1 Ct analysis of Day 25 and 130. Observed variation in NNV detection in various
tissues from Day 25 and 130. Brain and eye (retina) tissues show a large difference from Day 25
to 130. For the tissues gill, pectoral fin, brain and barbel, the number of samples consisted of one
positive each at Day 25.
Increased replication of NNV
Head-kidney
Korsnes et al. (in press)
OMJ-98
5. Suggested organ for biopsy:
Prophylaxis and control of NNV – some considerations
www.hibo.no
Transmission modes:
1. Vertical and horizontal
2. Ongoing study of vertical
transmission in cod
Host specificity?
1. Low within BFNNV clade
1. Marine reservoir of NNV
2. Infected brood fish
3. Horizontal transmission
”Classic” horizontal transmisson in species Homo sapiens
OMJ-98
Wild and farmed fish
interactions?
Konklusjoner – hvordan leve med NNV?
1. Screen stamfisk for NNV, viktig ved
inntak av villfisk som stamdyr
2. Hold stamfisk i lukket system for å
hindre horisontal spredning screening av fisk ved kjønnsmodning
3. Yngel bør holdes i lukket system og
vaksineres mot NNV før utsett til åpen
vekstlokalitet
4. Vaksinestrategi: Stamfisk kan
vaksineres for å begrense virus
produksjon ved kjønnsmodning.
Settefisk vaksineres før utsett for å
beskytte mot horisontal smitte på
lokalitet
www.hibo.no
Profylaktisk strategi mot NNV:
OMJ-98
1. Laks bør sees på som
potensiell vert. Unngå
samlokalisering med torsk
eller kveite
2. NNV fra kveite kan overføres
horisontalt til torsk. Unngå
samlokalisering av kveite og
torsk
3. Undersøkelser av NNV smittet
(i.p.) torsk antyder at
hodenyre kan brukes for
screening av stamfisk
4. VNN kan forebygges ved å
blokkere for vertikal og
horisontal spredning av NNV
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