微胞子虫 Microsporidium seriolaeによるブリ類のベこ病の感染動態

誌名誌名 魚病研究

ISSNISSN 0388788X

著者著者

横山, 博綾戸, 大地宮原, 治郎松倉, 一樹高見, 生雄横山, 文彦小川, 和夫

巻/号巻/号 46巻2号

掲載ページ掲載ページ p. 51-58

発行年月発行年月 2011年6月

農林水産省 農林水産技術会議事務局筑波産学連携支援センターTsukuba Business-Academia Cooperation Support Center, Agriculture, Forestry and Fisheries Research CouncilSecretariat

f.(I.41̂i (iJfヲ''e Fish Pathology， 46 (2)， 51-58， 2011. 6 。2011The Japanese Society 01 Fish Pathology

Infection Dynamics of Microsporidium serio/ae (Microspora) Causing the Beko Disease of Serio/a Spp.

Hiroshi Yokoyama l:i¥Oaichi Ayado

1， Jiro Miyahara

2， Kazuki Matsukura

2， Ikuo Takami

2，

Fumihiko Yokoyama3 and Kazuo Ogawa

1

lDepartment of Aquatic Bioscience， Graduate School of Agricultural and Life Sciences，

The University of Tokyo， Tokyo 113-8657， Japan

2Nagasaki Prefecturallnstitute of Fisheries， Nagasaki 851-2213， Japan

3Nagasaki Prefecture Iki District Office， Nagasaki 811-5133， Japan

(Received February 15， 2011)

ABSTRACT -Infection dynamics of Microsporidium seriolae causing the beko disease of yellowtail

Seriola quinqueradiata and amberjack S. dumerili were investigated at fish farms and an experimental

facility. A 4-year survey (2006-2009) showed a sharp increase in prevalence of infection with M

seriolae in June or July soon after stocking net cages with yellowtail and amberjack wild fingerlings

Most cysts blackened and finally disappeared in November in the following year. However， in 2006

when the levels of infection were relatively high， some visible cysts still remained even in autumn of the

following year. Infective periods and effects of fish size on M. seriolae inlection were investigated via

natural exposure of hatchery-reared yellowtail juveniles by translerring to an open sea cage in an

endemic area. Invasion of the parasite into lish occurred in June and July but not after the mid目 August.A comparison 01 M. seriolae infection among lour size classes of yellowtail showed that the smallest lish

(1 9 in mean weight) had the lowest prevalence 01 infection

Key words: Microsporidium seriolae， Seriola quinqueradiata， yellowtail， Seriola dumerili， amberjack，

infection dynamics， beko disease

Beko disease 01 yellowtail Seriola quinqueradiata

is caused by Microsporidium seriolae， which is an obl卜

gate intracellular microsporidian parasite (Egusa， 1982)

Infection with this parasite in the trunk muscle is visible

as elongate white masses which are called “cysts"

Disintegration 01 massive cysts in the trunk muscle

results in the characteristic concave fish body surface

This disease has often been reported in yellowtail and

goldstriped amberjack S. lalandi cultured in open sea

cage from the Kyushu area of Japan (Egusa， 1982;

Yokoyama et al.， 1996; Sano et al.， 1998; Bell et al.，

1999; 2001).

Heavy infection with M. seriolae causes emaciation

and mortality of yellowtail juveniles， whereas lightly or

moderately infected fish usually recover from the dis田

ease (Egusa， 1982). Beko disease has not been con-

sidered as a serious problem by fish farmers. How-

ever， in recent years， commercial fish with even a

minute scar of M. seriolae cysts are often rejected by

consumers in terms 01 hygiene. This public trend has

recalled the beko disease of yellowtail.

The recovering process of lish inlected with M.

Corresponding author E-mail: ayokoh@mail.ecc.u-tokyo.ac.jp

seriolae has not been well documented. Yokoyama et

al. (1996) revealed that disintegration 01 cysts led to

ingestion 01 liberated spores by host's phagocytes，

resulting in lormation of melanomacrophage centers in

the muscle tissues as well as secondary migration of

phagocytized spores into other organs， such as the

heart， intestine， kidney， and gills. Also， Sano et al

(1998) showed that encapsulated cysts in the muscula-

ture were associated with granulation tissue， causing

the scar lormation. These findings indicated that recov-

ery of infected fish was caused by host's inflammatory

responses. However， it is unknown how long it takes

until cysts disappear completely from host fish

Experimental infections of fish with M. seriolae

were attempted by immersion， intramuscular injection

and oral administration of fresh spores but all were

never successful (Yokoyama et al.， unpublished data)

Failures of direct transmission suggest that an obligate

intermediate host is required in the life cycle of M

seriolae. Thus， studies on the host-parasite relation-

ships have depended solely on observation of naturally

infected fish in the open sea cage or hatchery-reared

fish experimentally exposed with the parasite by trans-

ferring to the open sea cage in endemic area. Various

questions still remain to be answered for the beko dis-

52 H. Yokoyama， D. Ayado， J. Miyahara， K. Matsukura， 1. Takami， F. Yokoyama and K. Ogawa

ease， such as the infective period， process of recovery

and a size effect of host fish. In the present study， we

aimed to monitor the seasonal occurrence of M. seriolae as well as to determine several factors responsible for

the infection of the parasite in the open sea

Materials and Methods

Field monitoring at a fish farm Yellowtail and amberjack S. dumerili (O-year and 1目

year) cultured at a private fish farm in Miyazaki Prefec-

ture were examined for M. seriolae between 2006 and

2009. The origin of 2006 and 2007 fingerling cohorts

was Kagoshima Prefecture， whereas 2008 and 2009

cohorts were from Miyazaki Prefecture. After the intro-

duction of fingerlings into sea cages in June or July，

samples of 10-20 fish were randomly collected from

the same lot several times during 6-18 months. Fish

were weighed， measured and filleted， and the number

of cysts in the trunk muscle was counted by naked eyes.

Prevalence and mean intensity of infection was defined

as the number of infected fish/total number of fish exam-

ined， and the number of cysts/the number of infected

fish， respectively. For the 2007 and 2009 cohorts， the

presence of spores was also examined with Uvitex 2B-

stained smear slides of muscle homogenates according

to Yokoyama et al. (1996). Briefly， ca. 1 9 of the mus-

cle tissue block was collected from the dorsal side of

the fil同ts，minced in a glass homogenizer， and passed

through a steel mesh (Sigma; No. 40) with distilled

water (DW). The tissue suspensions were adjusted to

15 mL with DW， centrifuged at 500 x 9 for 10 min.

After the supernatant was discarded， the pellets were

smeared onto a glass slide by a cotton swab. After

fixation with methanol， smear slides were stained with

1 % Uvitex 2B solution dissolved in phosphate butfered

saline (PBS) for 10 min， then counterstained with 0.5%

Evans Blue in PBS for 30 sec. Air-dried stained

smears were examined by fluorescent microscopy (UV

excitation; 330-380 nm). For histopathological obser-

vation， some pieces of muscle tissue infected with M

seriolae cysts were fixed in 10% neutral buffered for-

malin， and embedded in paraffin. Sections cut at 5μm

were stained with H & E and Uvitex 2B， and observed

by fluorescent microscopy (Yokoyama et al.， 1996)

Infection of wild yellowtail fingerlings

Wild yellowtail fingerlings (mojako) were captured

at four different occasions from several locations around

Goto and Tsushima Islands， Nagasaki Prefecture from

May to June 2008. Immediately after catch， fish sam-

ples (n = 88) were iced on board and frozen at -20oC

shortly after landing. The samples were thawed at the

laboratory and visually examined for cysts. Presence

of spores was also investigated by the Uvitex 2B stairト

ing with 25 fish (only large fish from which 1 9 of the

muscle tissue could be collected)， because the remain-

ing 63 fish were not large enough to be tested by the

Uvitex 2B method. Additionally， single and nested

PCR-based detection assays for M. seriolae DNA were

performed according to Bell et al. (1999). Briefly， ca

50 mg sample of musculature was collected from each

fish， homogenized， and the DNA was extracted using

DNA Mini Kit (Qiagen). PCR reactions were carried

out in 20 pL total volume. Each PCR mixture con-

tained 0.1μL of Takara Ex TaqTM HS (5 UIμL)， 2 pL of

10xEx Taq Buffer， 1.6μしofdNTP mixture (2.5 mM

each)，O.5μL (25 pMIμL) of each outer primer (MS-F1:

5¥CACCTGTCTGCAATGCGGG-3' and MS-R1: 5二

GGTTGTTTGGCCGTACGGGG-3') and 3，0μL of

extracted DNA suspension. AII PCR reactions were

performed in iCycler (Bio働 Rad). Denaturation of DNA

(950C for 5 min) was followed by 30 cycles of amplifica-

tion consisting of 30 s for denaturation at 950C， 30 s for

annealing at 430C and 1 min for extension at 720C， and

ended by a 10 min extension at 720C， For nested

PCR， 1 pL of the first round PCR product was used as

template in the second round PCR using the internal

primer sets (MS-F2・5'ーGGTCGCAGCAGGAGCTTTC-

3' and MS【 R2・5'ーCTTCCGGCGTATCTTTAGTC-3')

The same PCR conditions were used as for the first

round PCR. PCR products were analyzed by 1.5%

agarose gel electrophoresis with TAE (40 mM Tris-ace-

tate， 1 mM EDTA) running buffer and visualized with

SYBR Safe DNA gel stain (Invitrogen). Expected sizes

of amplicons for the first and second PCRs were 1，205

bp and 549 bp， respectively

Infection of cultured yellowtail juveniles

After wild yellowtail fingerlings were introduced into

sea cages of eight farms located in Nagasaki Prefecture

in June 2007， fish were sampled (n = 10-20 per each

sampling) at 1， 2， 6， 16 and 25 weeks. Details of col-

Table 1， Inlormation on lish larms 01 cultured yellowtail juveniles in Nagasaki Prelecture

Fish larms A B C D E F G H

Site 01 collection 01 South 01

Danjo South 01

Danjo Danjo Westol South 01 South 01

Goto Goto Goto Goto Goto wild lingerlings

Islands Islands

Islands Islands Islands

Islands Islands Islands

Location 01 sea ×

cages' X X X X Y Z Z

X is located in close proximity to the land， whereas Y and Z are located offshore

week periods between 18 June and 25 August 2009 to

confirm the infective stage of M. seriolae in the marine

environment. After each exposure period， the yellow-

tail were transferred back to the land-based tanks

receiving flow-through of filtered sea water to prevent

their re-exposure to the parasite's infective stage.

After 3 weeks of subsequent rearing (the period deter-

mined in the above experiment)， the fish were examined

for the presence of cysts as described above. The

negative control group was kept in the land-based tank

during the study period. To avoid size effects on the

parasite infection， fish were constrained from growing

by feeding only twice a week.

(3) Effects of fish size on infection: Hatchery-reared

yellowtail juveniles were provided from the Goto Station，

NCSE in February (the first batch of breeding) and June

(the second batch of breeding)， 2010. Fish were kept

in the land-based tank described above until use. Fish

from the first batch were divided into three groups

based on their size; mean body weight and fork length

were 123.8 9 and 20.0 cm for 'Iarge-size'， 45.8 9 and

15.2 cm for ‘medium-size¥and 27.目o9 and 12.7 cm for

'small-size'. Fish from the second batch (mean body

weight and fork length were 1.0 9 and 4目4cm， respec-

tively) were used as an 'extremely smal卜size'group

The four different size classes were hold separately in

columnar net cages which were placed in a 2寸oncircu-

lar tank with flow-through unfiltered sea water. After 1

week of exposure， all fish were transferred to land-based

tanks (500 L tank for the large， medium and smal卜size

groups， and 200 L tank for the extremely smal卜size

group) supplied with seawater filtered with a sand filter

and with 1.0 pm-cartridge filters. After 1 month of rear-

53 Infection dynamics of beko disease

lection sites of wild fingerlings and the location of sea

cages for the eight farms are summarized in Table 1

Location of fish farms designated as A-E was located in

close proximity to the land (ca. 20-80 m)， whereas sea

cages of farms F， G and H were set up offshore (ca

100-350 m from the land). The samples were visually

examined for cysts and for spores in Uvitex 2B-stained

smears by fluorescent microscopy as described above

Exposure of hatchery-reared yellowtail juveniles to

infective water

(1) Determination of days for cyst formation: To

decide the appropriate rearing period of exposed fish in

the next experiment， days required for cyst formation

were investigated. Hatchery-produced yellowtail juve-

niles were obtained from the Goto Station， National

Center for Stock Enhancement (NCSE) for the following

experiments. Experimental fish (n = 150， body length = ca. 5 cm， body weight = ca. 2 g) were stocked in a sea

cage of Nagasaki Prefectural Institute of Fisheries

Sciences (NPIFS)， where persistent infection with M

seriolae has been detected every year. After being

transferred to sea cage on 25 June 2009， five fish were

randomly collected every day for 4 weeks. The sarrト

ples were examined for cysts macroscopically and for

spores microscopically as described above.

(2) Determination of infective period: Hatchery-

reared fish were kept in a land-based tank (200 L) sup-

plied with sand-filtered sea water which was further

passed through a cartridge filter with 1-pm openings.

The inflow was adjusted to ca. 10 Umin. The fish were

divided into eight groups (30-40 fish per one group)，

which were held in a sea cage of NPIFS for various 1-

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Prevalence (Iines) and mean intensity (bars) of Microsporidium serio/ae in the 2006-2009 cohorts of amberjack (closed circle， black bar) and yellowtail (open circle， gray bar) cultured at a fish farm in Miyazaki Prelecture. Vertical lines for preva-lence and mean intensity 01 inlection show the confidence intervals at 95% in reliability and standard deviations， respectively Data 01 mean int巴nsity01 infection in 2007 cohort amberjack is not shown

20

Fig.1.

2006 cohort reached up to 16 cysts/fish in August 2006，

and visually detectable cysts still remained at the end of

the survey. In contrast， 2007 cohort yellowtail and

amberjack had lower prevalence and mean intensity

than those of the 2006 cohort even at the early period

of infection (10% in July for yellowtail). Mean intensity

was as low as 2 cysts/fish over the study period both for

yellowtail and amberjack. Most cysts became brown in

color， progressed to black， and then shrunk to negligible

size (< 1 mm long) by the end of the survey (Fig. 2).

However， spores were detected by Uvitex 28 staining

from more than 90% fish of the 2007 cohort even in the

early period of infection (Fig. 3). For the 2008 cohort，

no infection was observed in yellowtail fingerlings just

after catch (May 2008)， but cysts appeared in more

than 80% fish in June 2008 (Fig. 1). Prevalence

declined substantially over time， and only a few cysts

were found in r¥Jovember 2009. Although monitoring

for the 2009 cohort was ended in November 2009， over-

all levels of infection in yellowtail were relatively

moderate. In all cohorts of fish groups， neither relapse

of prevalence nor young cysts (visible as small and

white cysts) were observed in the following year

Histological observations showed that cysts degen-

erated by host responses over time (Fig. 4). Masses

of spores were encapsulated by host's connective tis-

H. Yokoyama， D. Ayado， J. Miyahara， K. Matsukura， 1. Takami， F. Yokoyama and K. Ogawa

Results

Seasonal infection

Annual and seaso円alpa社ernsin prevalence and

mean intensity of infection were observed in the 4-year

cohorts (Fig. 1). Prevalence in the 2006 cohort of

amberjack was 100% in July and August 2006，

decreased in autumn and winter， but was still 80% in

September 2007. Mean nfection of infection in the

54

ing， 44 fish were collected from each group and exam-

ined for prevalence and intensity of M. seriolae.

11 9 7 5

2007 cohort

，、J

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11 9 7 5

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Prevalence 01 Microsporidium seriolae in the 2007 and 2009 cohorts 01 yellowtail. Closed and open cir-cles represent the prevalence determined by visual observation 01 cysts and microscopic examination for Uvitex 2B-stained spores， respectively.

9 7 5 ，、J 11 9 7 5

Fig.3. Recovery process 01 the beko disease in yellowtail. Arrows show a developing cyst (A: 3 months post-introduction to sea cage， 2009-cohort)， a blackened cyst (8: 6 months post-introduction， 2008-cohort) and degenerative scars (C: 17 months post-introduction，

2007-cohort).

Fig.2.

Inlection dynamics 01 beko disease 55

Fig. 4. Histological sections 01 th巴 trunkmuscle 01 amberjack inlected with Microsporidium serio/ae. H & E & Uvitex 2B stain. A: developing cysts in which Iluorescent mature spores are accumulated centrally. B: lully developed cysts packed with mature spores. C: encapsulated spore masses. D: melanin depositions associated with scar lormation. Bars = 100μm

sue， gradually shrunk in size， finally resulting in the

masses as scars associated with melanin deposition目

Infection of wild yellowtail fingerlings

Neither cysts nor spores were observed in any of

wild yeliowtails caught around Goto or Tsushima islands

(88 fish for visual observation and 25 fish for Uvitex 2B

examination). Using the nested-PCR， none of the 88

fish were positive for M. seriolae DNA in the first round

PCR but six fish were positive (6.8%) after the second

amplification (Table 2). There was no remarkable dif-

ference in the prevalence of infection among the four

fish lots investigated

Infection of cultured yellowtail juveniles

Cysts appeared at 2 weeks post-introduction to sea

cages in seven out of eight farms， and prevalence of

infection increased to more than 90% at 6-25 weeks in

all farms studied (Fig. 5). Distribution of M. seriolae

seemed homogenous， regardless of different origin of

wild fingerlings and different locations of sea cages

Days required for cyst formation

Cysts were first observed in yellowtails 20 days

post-exposure (Fig. 6). The water temperature increased

gradually and over 260C when cysts formed in the

muscles. At that time， mean intensity of infection was

8.3 cysts/infected fish， and the mean length of cyst was

3.8 mm

Infective period

Almost 100% of the fish in the initial four groups

exposed between 18 June and 16 July were found to be

infected with M. seriolae (Fig. 7)ー Startingwith the 5th

group exposed between 16 and 23 July， the prevalence

of infection gradually decreased and was down to zero

in the last group exposed between 18 and 25 August

Mean intensity of infection peaked at 25.8 cysts/infected

fish for the 2nd group， and declined over time. Mean

length of cysts was almost constant at approximately 6

Table 2. Epizootiological survey lor Microsporidium serio/ae in wild lingerlings 01 yellowtail just after catch along the coast 01 Nagasaki Prelecture

Date 01 catch Site 01 collection Sample Mean body Mean body Prevalence 01 inlection examined by

slze length (cm) weight (g) Uvitex 2B Single-PCR Nested-PCR

18 May， 2008 West 01 Goto Islands 35 5.1 7.2 0/9"可 0/35 1/35

20 May， 2008 South East 01 Goto

26 4.7 3.9 0/8 0/26 2/26 Islands

21 May，2008 West 01 Tsushima Island 5 9.5 20.6 0/5 0/5 0/5

6 June. 2008 East 01 Tsushima Island 22 3.9 1.8 0/3 0/22 3/22

在 Number01 positive lish/number 01 examined lish

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28

24

26

H. Yokoyama， D. Ayado， J. Miyahara， K. Matsukura， 1. Takami， F. Yokoyama and K. Ogawa

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Fig. 6. Daily monitoring 01 Microsporidium seriolae inlection

in hatchery-reared yellowtail juveniles stocked in a net

cage 01 Nagasaki Prelectural Institute 01 Fisheries

Sciences. Solid and broken lines represent the preva-

lence 01 inlection (n = 5 lor each sampling) and water

temperature， respectively.

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Prevalence (Iines) and mean intensity (bars) 01 Micro-

sporidium seriolae in juvenile yellowtail stocked in a

net cage 01 Nagasaki Prelectural Institute 01 Fisheries

Sciences. Each lish group was reared lor eight 1-

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Fig. 5. Prevalence 01 Microsporidium seriolae in juvenile yelー

lowtail stocked in sea cages at eight larms in Nagasaki

Prelecture. Upper graph shows data lrom each larm

separately. Middle and lower graphs show the data

pooled by the site 01 lingerlings collection and the loca-

tion area 01 sea cages， respectively (see also Table 1)

25

(6.1-6.6) mm among all fish groups except one (8.2

mm for the 6th group).

ge， medium and small-size groups， while it was signifi・

cantly lower (2.3%) in the extremely small-size group

compared to the others (Table 3). Mean intensity of

infection was significantly lower (士9cysts/infected fish)

Relationship between fish size and infection

Water temperature varied between 22.50C and

25.10C during the experimental period (from 23 June to

26 July， 2010). Prevalence 01 infection was not signilト

cantly different (approximately 70-80%) among the lar-

Comparison 01 prevalence and intensity 01 inlection with Microsporidum seriolae among four dilferent size classes 01 yellow-

tail juveniles (n = 44 lor each class). Different let1ers (a and b) in the columns on the prevalence and intensity 01 inlection

mean signilicant differences among different siz巴 groups(P < 0.05; Ryan's test and Steel-Dwass test lor prevalence and

intensity， respectively). The data 01 mean intensity 01 inlection lor extr巴m巴Iysmal卜sizegroup is not included in the statistical

analysis due to the small sample size (n=1)

Table 3.

Prevalence 01 inlection

(%)

Mean intensity 01 inlec-

tion (mean number 01

cysts/inlected lish)

ヱ9a

12.5b

13.6b

6.0

a

a

a

b

8

2

5

3

41aunuηζ

。口

C

U

7

'

Mean lork length at the

end 01 the experiment

(cm)

22.2

17.3

15.9

7.8

Mean body weight

at the end 01 the

experiment (g)

151.2

65.6

54.4

5.4

Fish size group (initial mean

weight and lork length)

Large (123.8 g， 20.0 cm)

Medium (45.8 g， 15.2 cm)

Small (27.0 g， 12.7 cm)

Extremely small (1.0 g， 4.4 cm)

Inlection dynamics 01 beko disease 57

in the large group compared to the medium (12.5) and

small-size (13.6) groups (Table 3).

Discussion

This is the first study that describes the whole infec-

tion dynamics of the beko disease from the infection

period to the recovery phase in cultured yellowtail.

The reason for annual variation in the prevalence and

mean intensity of M. seriolae among 2006-2009

cohorts is unknown. Because it is unlikely that the

variation is caused by the origin of fingerlings and the

culture site， it may be partly explained by different water

temperature， variable biomass of the unknown interme-

diate host or prevalence of infective stage in the interme-

diate host. Disease occurrence must be subject to

complex interactions between the host and the parasite

under specific environmental influences (Sano et al.，

1998). It is notable that most of fish were found to be

subclinically infected in the early period of infection in

the 2007 cohorts. Most of parasites might have

cleared without developing into visible cysts. The

recovery phase depended on severity of infection in fin・

gerlings during the initial 1-2 months. Fish could

recover from the infection in the second year when the

mean intensity of infection was as low as 5 cysts/fish in

June or July in the first year. No recurrence of the

beko disease was found in the following year， suggest-

ing that a resistance to the disease was established in

surviving fish.

Our observations of wild fingerlings and cultured

juveniles suggest that M. seriolae infection occurred

after introduction of wild fingerlings to sea cages

Although some wild fingerlings were found to be infect-

ed， the infection was extremely light. Considering the

lack of direct transmission of M. serio/ae among fish

hosts， it is unlikely that the infected wild fingerlings have

caused a horizontal transmission of the parasite from

fish to fish within a sea cage. These findings imply

that selection of wild fingerling lots is not effective in pre-

venting the beko disease. The parasite infection

occurred both in coastal and offshore cages， suggesting

that the site selection is not a promising prevention

method at least in the farming area examined in this

study

The results on the infective period indicate that

infectivity of M. seriolae decreased after the end of July.

This seems to correspond with the findings by Sano et

al. (1998)， who observed a low infection level in yellow-

tail juveniles stocked in a sea cage in the end of July.

However， we could not determine a starting point of the

infective period， because preval

The prevalence and mean intensity of infection in

the extremely smal卜sizegroup (mean weight; 1 g) was

the lowest among the fish groups studied. It may be

explained by alteration of feeding habit， although the

route of infection with M. seriolae has not yet been

elucidated. On the other hand， Sano et a/. (1998)

reported that two different fish size classes (mean

weight; 0.3 9 and 22.0 g) were infected with M. seriolae

to a similar extent (64.5% and 95%， respectively). The

two experimental designs are different in exposure peri-

ods; fish in the present study were exposed only for one

week， whereas fish in Sano et al. (1998) were exposed

continuously for 43 days. It is possible that fish in

Sano et al. (1998) became susceptible to M. seriolae

with the growth of fish. Low susceptibility of extremely

small-size fish seems to last only for a short time，

because fish grows rapidly. Thus， it may be difficult to

put the result into practice. As far as the three groups

larger than 27 9 in initial weight， larger fish had a lower

intensity than smaller fish. This suggests that it is

effective to promote growth of fingerlings as large as

possible before the peak period of infection. Generally，

no practical chemical treatment has been available for

microsporidian infections. Effective control may be

achieved with integrated management of a combination

of strategies (Ogawa & Yokoyama， 1998). Sano et a/.

(1998) indicated that sand filtration of rearing water is

effective in removing possible infective agents of M

seriolae， although this method is not applicable for cage

culture. Considering the findings of the infective period

obtained by Sano et al. (1998) and the present study， it

is recommended to use a land-based tank supplied with

filtered seawater until after the peak period of infection

(from mid-June to mid司 July). In a practical point of

view， it seems to be most reasonable to depend on

natural recovery process. However， heavily infected

fish (Iike the 2006 cohort) hardly recover， suggesting

that juveniles with signs of the disease should be dis-

posed as early as possible， e.g. at the time of a vaccina-

tion for viral/bacterial infections usually applied around

July in the first year.

Acknowledgements

We greatly acknowledge the Goto Station， National

Center for Stock Enhancement， Fisheries Research

Agency， for providing yellowtail fingerlings as the exper卜

mental fish， and the fish farmers in Miyazaki and

Nagasaki Prefectures for kind offer of fish samples

We also thank Dr. Daniel Grabner of the University of

Tokyo for his reviewing this manuscript.

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Yokoyama， H.， J-H. Kim， J. Sato， M. Sano and K. Hirano

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微胞子虫 Microsporidiumseriolaeによるブリ類のベ

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f(l，病研究. 46 (2). 51-58 (2011)