Attempted. Vaccination of Rainbow. Trout Oncorhynchus mykiss. Against Bacterial Kidney Disease. Masahiro Sakai,* Shizuo Atsuta,* and Masanori Kobayashi*.
Nippon Suisan Gakkaishi Attempted
55(12),2105-2109(1989)
Vaccination
of Rainbow
Against
Bacterial
Trout
Oncorhynchus
Kidney
mykiss
Disease
Masahiro Sakai,* Shizuo Atsuta,* and Masanori
Kobayashi*
(ReceivedJune 30, 1989) Immune responses in rainbow trout Oncorhynchusmykiss intraperitoneally injected with formalin killed Renibacteriumsalmoninarumcells (with or without Freund's complete adjuvant), were evaluated by monitoring serum agglutinating antibody and phagocytic activity of kidney leukocytes, and the challenge test. Serum antibody titers (range 8-16) were detected in the vac cinated groups 5 weeks after vaccination. The phagocyticactivitiesin the kidneycells of vaccinated groups increasedafter 15 days and 25 days. However, experimental intraperitoneal challenge(107 or 106 cells/fish) of immunized groups showed no protective effects. Bacterial kidney disease (BKD) is a systemic and clonic infection of salmonid fishes characteriz ed by the presence of granulomatous lesions.1) The etiological agent Resnibacterium salmoninarum is a Gram-positive diplobacillus which is very fastidious in its growth requirements. The disease is considered important because of the serious losses it causes and because of the difficulty in controlling it. Some successful results in chemo therapy with erythromycin,2,3) sulfonamides,4) penicillin,5) clindamycin, kitasamycin and spira mycin6) have been reported. However, it has been proved difficult to eliminate the pathogen from the host because of its intracellular existence in macrophages.7) Vaccination against BKD has been attempted by Paterson et al.8) and McCarthy et al..9) Pater son et al.8) reported a significant increase in serum agglutinating titer and a reduced incidence of BKD in Atlantic salmon Salmo salar. However, because the potency of vaccine was demonstrated by the gross lesions of BKD and the detection of R. salmoninarum antigen with the indirect fluore scent antibody technique (IFAT), the exact efficacy of vaccine was not determined. McCarthy et al.9) also reported that rainbow trout Oncorhynchus mykiss formerly Salmo gairdneri, injected intra
mune
小 林 正 典:北
Fisheries
Sciences,
里 大 学 水 産 学 部).
Kitasato
of
salmoninarum
the
effect
of
lenging
rainbow
trout
vaccinated
bacterins
was
investigated
vaccination
with
the
was
with and
estimated
intraperitoneal
by
chal
of
living
50
g and
injection
organisms.
Materials
and
Methods
Animals Rainbow 10
g were
and
a
trout
of
Iwate
University
a mean
an test,
the
Bacterin
The
with
and
salmon
Oncorhynchus
was
used
for
was
cultured
4•Ž.
washed
adjusted The
to
equal
with
g-fish)
killed fish
by
trout
or were
0.05
ml
adjuvant
Sanriku, Iwate
at
15•Ž
by
adjuvant
were
in
sterile
with
O22-01,Japan(酒
at
same
nm.
cells
MS-222
0.1 of
ml
(for
formalin
bacterin. the
and
520
(Difco).
with
g-fish)
h
emulsifying
(FCA)
with 10
7.2)
washed
anesthetized
(for
48
cells
1.0
killed
and
for
pH
prepared
emulsified
administrated
days
(PBS,
intraperitoneally or
1985, strain
formalin
formalin
were
a dise in
centrifugation
was of
15
of O.D.
complete
injected
from
inactivated
saline
bacterin
Rainbow
50
formalin
volume
12•Ž
The
0.3%
a concentration
Freund's
and
for of
times
adjuvant
an
KDM-210)
buffered
at
kisutch
preparation.
addition
three
phosphate
isolated
bacterin
The
kept
Vaccination
coho
at
were
were Station
pellets.
ased
in
fish
Fish
fish
KU8501
the
The
commercial
Preparation
by
experiment
Experimental
salmoninarum
killed
of
response
respectively.
lwate
daily
weight
immune
Prefecture.
fed
R.
in
challenge by
•@ and
with
used
supplied
- peritoneally with lysed cell preparations of R. salmoninarum were protected against the artificial challenge of living cells, although serum agglutinat ing antibody was not investigated. Thus, the relation between immune response and protection must be investigated in fish vaccinated with R. salmoninarum. In the present study, the im * School of
response
R.
Control volume
井 正 博,厚
of
田 静 男,
2106
Sakai,
FCA.
Vaccinated
maintained 12•Ž
and
tanks
fed
Serum fish
of
each
five
fish
water
The
were
temperature
of
in
obtained
from
weeks
after
microtiter
method.
indirect used
detect
kidney
Five
vessels
of
were
The
serum fish
agglutinating
determined
Serum
titer using
the
Agglutinating
antibody
inarum
were
cinated
by
cination The
vaccinated
and
5,
kidney
phagocytosis
to
the
of
was
and
or
non-vaccinated
25
days
after
were
collected.
leukocytes
assay
method
difference
15
was
Sakai
performed
only
et
estimated
al.11)
by
statistical
by
The
Student
Method
Thirty
after
non-vaccinated itoneal
fish
injection
suspended were
were ed
were with
in
PBS. at
daily necropsied
on
of
the
to
the
higher
60
and
kidney
intraper
bacterial
cells
challenge,
and
days.
the
fish
mortalities All
were
moribund
samples
of
value
is
in days all
was
percent
method
Sixty
cinated
for
by
106
the
12•Ž
vaccine
relative
immunity
from
or
and
were
fish inoculat
after
the
A to
calculat according
RPS
of
indicate
60
Phagocytic
5,
fish
the
kidney or
smears non-vac
non-fluorescent
activities
in kidney
without
the
injected
with
days
after the
cination
were
controled
5,
15
and
and
57.6%
respectively. activities
from
8.5
of to
25
higher
the
10.8%.
leukocytes
and
significantly
cells
40.2%
of 15
the
formalin-killed
were
measured
in
Leukocytes
vaccination,
activities
fish
shown
without
48.6%
phagocytic
FCA
phagocytic
and
10.9%,
hand, with
munized
are
respectively.
were
of
leukocytes cells
28.1%
immunized
25
of
of
days
after
than
im vac
those
of
fish.
Protection
Experiments
slides.
leucocytes
pattern
controled cells died
fish are by
of rainbow
shown 25
days
trout
of
mortalities
challenged in after
of with
Fig.
1.
the
challenge.
vaccinated
with
vaccinated
2.2•~107 All
or bacterial
controled In
that
Renibacterium
Vaccine
Phagocytic (n =5) activity (%)
Singificance
FK*t FK with FCA*2 Control
14.7±3.9 10.9±1.6 9.5±2.1
N.S*3 N.S
15
FK FK with FCA Control
28.1±1.9 40.2+6.6 10.6±2.0
p
