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CONTINTS

ABSTRACT

I NTRODUCT ION

THE WESTTRN AUSTRALIAN PEARL CULTURE INDUSTRY

|\4ATERIALS AND MITHODS

1 . F i e l d S t u d i e s

1 . I A rea o f s tudy

1 .2 Wate r gua l i t y pa ramete rs

P A G E

7

B

B

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1 1

1 1

1 2

1 4

1 A

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t f ,

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1 7

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1 .3 Indus t r y reco rds

1 . 4 M a n i p u l a t i v e f i e l - d

r . 4 . t S . r z e , d e p t hexperament

1 .4 .2 Con tag ion expe r imen t1 . 4 . 3 D e n s i t y e x p e r i m e n t

1 .4 .4 Long_ l i ne expe r imen t1 .4 .5 T ranspor t expe r imen t

2 . f nves t i ga t i on o f d i sease

2 .1 pa tho fog i ca l p rocedures

2 .2 Bac te r i o log i ca l me thods

2 .3 V i ro log i ca l p rocedures

2 .4 Bac te r i a l coun ts o f wa te r i n t hecarr ier tanks dur ing transport

Experimental protocol for t ransmi ssionand other laboratory experiments

3 . ' i F ie ld t ransmiss ron expe ramen t

3.2 Laboratory experiments

3 . 2 . 1 B a c t e r i a l i n o c u . l a

3 .2 .2 Exper imen ta l rep roduc t i ono f d i sease

3 .3 Bac te r i a l i nvas ron

3 .4 Inves t i ga t i on o f bac te r i a l -toxinogenes i s

3 .5 Cond i t i on rndex (Cr )

Analyt ical rnethods

- 2 -

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and cleani .ng

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TABLES

P h y s i c a l a n d b i o c h e m i c a l c h a r a c t e r s o f V i b n i o s p ' i s o l a t e df r o m m o r b i d o y s t e r s .

Exper imentaf protocof .

Exper imenta l deslgn of bacter ia f invasion exper iment .

D e s i g n o f b a c t e r i a l t o x i n o E e n e s i s e x p e r i m e n t .

Compar ison of nutr lents and chlorophyl -1 a in WesternAustra l ian mar ine envi ronments. Range oF values are^ i r r a n f a v ' a a a h q . a m n I i n r r

' l o r . a i i o n

v r v v r r ! v r

6, Ini t ia l survlvaf of pear l oysters af ter 12 days suspension.

7. Number of surviv ing pearl oysters af ter a total of I14 dayssuspensaon .

8 . Mor ta l i t y ra te i n re la t i on to t ime and pos i t i on du r ing ca r r i age .

9 . Mean and s tandard dev ia t i on (SD) o f V ibz io sp . , /m l o fsu r face wa te r and the ra t i o o f V i | n io sp . i n wa te r o fca r r l e r t anks to tha t l n seawa te r (TANK:SW) .

10. Morbidl ty/Mortal i ty data for t ransmission experj-ments.

11. Morbidl ty and Mortal l ty data for oysters held at 19oarrcl 29 .

12 . Number and (7 . ) o f oys te rs i n wh ich V ibn io sp . were i so la tedfrom haemolymph and muscle.

13 . E f fec t o f cu l t u re f i l t r a tes on ovs te rs .

14. Mean t SD for Condit ion Index.

- 4 -

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ralJe sraJS^o Jo ^1rT€1.rour abeluecJad elrleTnurnJ uear{

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10, Accumulat ion of large numbers of phagocytes ln theconnect ive t issue septa of the digest ive gland.HematoxyLin and Eosin Staln.

l -1. Accumulat ion of f ibrous l ike phagocytes (A) in theaur i c le . The g ranu la r ce f l s (a r rows) a re "b rown ce f l s "that are normal ly present j -n the aur ic le. H E.

12. Sect ion through mant le t issue showing erosion andphagocyte exudat lon (arrow) of the epi thef ium of the outerfo ld (A ) o f t he man t le . H E .

13 . D iges t i ve g land . Dense i n t ranuc lea r i nc lus lon bod ies(a r rows) l n bas iph i l ce l l s o f d iges t l ve tubu les . H E .

14. Electron micrograph of intranuclear inclusion bodycon ta in ing v l rus - I i ke pa r t i c l es . Ba r i nd i ca tes 1 um.

1 6 .

L 7 .

1 8 .

Ap icomp lexan p ro tozoan in au r i c fe . H E .

Cl l lated protozoa attached to the mid-qut

Residual bodies (arrows) in the cytoplasmce I I s o f d iges t i ve g land tubu fes . H E .

E lec t ron m ic rog raph o f a res idua l body .membrane (a r row) . Ba r i nd i ca tes 1 um.

a n i + ] r a l i r t m t { F

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I NTRODU CT I ON

T h e W e s t e r n A u s t r a l i a n p e a r l i n g i n d u s t r y i s A u s t r a l l a ' s f o u r t h m o s t' i m n o r t . a n t f i s h i n o i n d u s t r y . P e a r l p r o d u c t i o n i s b a s e d o n t h eu t i l i z a t i o n o f n a t u r a l l y g r o w n o y s t e r s t h a t a r e r e m o v e d l r o m

thei r natr . ] ra l beds and then t ranspor ted to lease s i tes wheren a : n I f n n m , : t i o n i q i n d r r . - e . l l - 1 v . r r t i f i c i a l f f e a n s . A n e X c e S S i V ep s o r r t

m n r t : 1 i l \ / . ) f n e a r l o r r c l c r ^ q f l ^ r I l o ' ^ r i n . r r e r r o V a l f r o m t h e n a t u r a lb e d s h a s h a d a s e r i o u s i m p a c t u p o n t h e i n d u s L r y s i n c e 1 9 7 4 .Losses on the lease s i tes can amount to 80% ol the L larvesteds h e l f , a l t h o u g h l o s s e s o f 3 0 - 6 0 % a r e m o r e c o m m o n ' T h e s e r a t e sc o n t r a s t m a r k e d l y w i t h a c c e p t e d " n o r m a l " l o s s e s o f r o - 2 o % . I n

a d d i t i o n t o t h e f o s s e s , a n e q u a l l y s i g n i f l c a n t f a c t o r i s t h a ta f f e c t e d o y s t e r s t h a t d o n o t d i e b u t r e c o v e r , d e v e l o p d e f o r m e dn a c r e a n d a r e u s e l e s s f o r h a l f - p e a r l o r m o t h e r - o f - p e a r lp r o d u c t i o n .

F x c e s s m o r L a l i t y h a s t w o m a j o r e f f e c L s o n t h e i n d u s b r y .F i r s t l y , t h e o b v i o u s e f f e c t s a r e a d i r e c t f i n a n c i a l l o s s t oo p e r a t o r s a s s o c i a t e d v . r - i L h t h e c o s t o I c o l l e c t i n g w i 1 d o y s t e r s+ ^ ^ a F h 6 F L ' i + h - ^ n q a . ] r r c n 1 - l n q l n r o d r r r - l i o n n i n p a r l q n r r e r f . h e

e n i u i n g s e a s o n s . s e c o n d f y , e x c e s s i v e m o r t a l - i t y a l s o F o r c e so p e r a l o r s t o r e m o v e m o r e o y s t e r s t h a n b h e y w o u l d r e q u i r e i fm o r b a l i t y w a s n o l p r e s e n t a n d a s t h e i n d u s t r y i s b a s e d o nn a t u r a l s L o c k s t h e n o r t a l i t y p r o b l e m p l a c e s . i n c r e a s i n g p r e s s u r ef r o m f i s h i n q o n t h e n a L u r a l p o p u l a L i o n .

I h e a L n o I L h e i n v e s t i g a L i o n s r e p o r t e d i n L h i s p a p e r w a s L od e t e r m i n e - " t h r e f a c t o r s a n d p o s s i b l e c a u s e s a s s o c i a t e d w i t hmorta l i ty Most of th is work was done in conjunct ion wi th theI r r ^ a q { r ^ r l - r i r - h k r a c . e x n e n i e n c . i n . t l - h F . r r a ^ | c c | : n d m o c . l -! u ' \ , r u J e ! v r ' y u ' r , - " " i i ' Y

p e r s i s t e n t m o r t a l i t y p r o b l e m . I n c i d e n t a l o u t b r e a k s o f o y s t e rm o r l a l i b y o n o L h e r l e a s e s i t e s m a v h a v e o L h e r d e t e r m i n i n gf a c t o r s i n v o l v e d .

I I THT WESTERN AUSTRAL]AN PEARL CULTURE INDUSTRYF o r m o r e t h a n a c e n t u r y p e a r l a n d p e a r l s h e l l f i s h i n g h a s b e e na v a l u a b l e i n d u s t r y a l o n g t h e N o r t h e r n c o a s L o f W e s t e r nA u s t r a l i a . F o r m o s t o f i t s c o l o u r f u f h i s t o r y , t h e i n d u s t r yrevolved around the col lect ion of shel l or mot f ier -of -pear l ,r a t L r e r t h a n t h e n a t u r a f p e a r l s t h e m s e l v e s ( B a i n , 1 9 8 2 ) . T h en n l - h F r - . \ F . n p a n l c h e l I f r O m L h e s i l v e r O r d ^ l - l - l i n ^ a , - l n 6 . h l

o y s t e r , P i n c t a d a m a x i m a ( J a m e s o n ) , w a s u s e d f o r b u t t o n m a n u -f a r - t r t r e . a n r i a n v n F f r t r ^ l n a e n l q f n r r n r l u , o 1 g a b o n u s Broomel - . ^ ^ r - ^ ' r - ^ m - i ^ h ^ - - n l i n r r r - c n ] - r a . : n d h r r I h e c : n l \ r l a n n ' cu r r s I L o J v r P c o | r , r 9 , o , , u u y u r r L L u ! r J ,

i t h a d a f l e e t o f m o r e t h a n 3 O O s a i l i n g v e s s e l s r , c a l l e dI r r n c e r q f i s l - , i n n f n r . g h g l l .

T h e i n a l l r q l n , / q r r n \ / ; v e d q e r r e n e a l i q . r r r n l i 6 n . l r r ] . i n . r i ^ l 6 n l . 1 t ^ t ^ n T T

o n l r r 1 - . r f i n d f h e d c m : n d f n r ^ n c . a n l q h a l l d u r i n d l o d + n h a ^ l i ^ i h l ^L v I r s 9 r L 9 r u ! c

p r o p o r L i o n s f o - L l o w i n g t h e a d v e n t o f p l a s t i c a s a m a t e r i a l f o r

T n e p u r p o s e o f L h i s p a p e r i s t o s u n L ' n a r i z e t h e p r j n c i p a lf i n d . i n n s o f t h e m o r t a 1 . i + \ , c n ' ' r l i ' a c n a - + c o f w h i c h w i I I b ei r r r v i ' ' 9 r v r

r e p o r L e d o n i n d e L a i l e l s e w h e r e ( P a s s a n d D y b d a h l , i n p r e p )

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Shel l -s are gathered dur ing neap t ide per iodsHookah gear whife bei_ng towed behind luggers.deck, these shel ls are cleaned. of sponges andorganisms with a tomahawk.

Flshing regulat ions designed tos . h c l ' 1 f n n a r r ' l t r r n a ^ ", - - _ r_ rposes a l l ownrn ( 8rr ) measured dorso-ventral lv

by divers usingWhen brought on

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encourage the use of l iveonly shel l greater than 2O3from the shel l vaLve hinge to

use as mother-of-pear lthe growth edge, to be k i l l ,ed forproducts .

Llve shel l are transferred to the larger t ransport ship wheretfrey are sorted into arbi t rary s ize c-tegor les- as proptr t ion_af ly larger nuclei wi l l be inserted into the larger oysterswhen they are later operated upon. Before 1982 there werethree size categories (smal l <1O2 rnm; medium 1O2 rm to 14O nm;and large >14o rnrn). These size categories were adjustedsr ight ly upwards dur ing the 1982 se"Jo.r when the n6w minimumpermissibre size of sher l was increased from r-o2 mm to 1l i mm( 4 . s , ' ) .

Oysters of s imi lar s ize are placed into baskets made ofplast ic covered wire with dlmensi-ons of 61 cm x 23 cm x 2g cm.Each basket has nine part i t ions to house 10 oysters in the

vert ical posi t ion, i .e. hir lge dawn. The baskets are thenstacked underwater in the four carr i -er tanks bui l t into thehold. These tanks have a total capaci ty in excess of 20 ooooysters al though usual" ly no more than fS OOO oysters aretransported per t r ip. Seawater is c i , rculateO inrougfr lnesetanks when the boat is at anchor by pumping in outslde waterand spraying i t into each tank from sl i ts in a pipe around. theupper per imeter which then displaces water t f r rough four bottomdraln holes in the hu1l. When the ship is in mot io.r , " . r" . r i r" ,system on two addit ional holes per tank pipes water to thesurface of each tank.

The oysters are transported from the f ishing grounds to theKuri Bay lease si te, a distance of almost zoo-t<m to tne nortrr(Fig. 1), which takes the ship approximately g7 hours. Duringthe l"onger neap f lshing per iods oi seven or eight days, thetwo transport vessels, , 'Merindah pearl" and ' ,Kur i pelr i , ,a l ternately col lect oysters f rom the luggers and transportthem to .Kuri Bay to reduce the amount oi-time oysters are heldon board to a maximum of four or f lve days.

upon arr ival at the rease si te, the baskets of oysters aboard.. ! l : : :_t :?: :port_ships are quickty unloaded onto i loat i . ,g----wooqen rarts. These raf ts are constructed by lashing t6getherpine poles to form basic uni ts 7 m x 7 m. Uhit f lot ; t io i isprovided by 8-1"o (2oo L) empty metal or foam f i l led rubber- 'drums lashed under the poles. Ten uni ts are jo ined in tworows of f ive to complete a raf t . One hundred-baskets aresuspended about 70 cm apart on the ten paral le l poles oi-eachunit i .e. 1 OOO baskets per raf t . fhe taskets " i . =r"p. .rO"afrom the poles with 5-6 m lengths of j .2 mm black p"fy" i f ry i""""op9. During the year-Iong accl imat izat ion per iod Uefor! tnenucleus operat i ,on, the. oysters are only disturbed _pp"o*i_---mateLy every three months to clean off fouling organisms fromthe llving oysters and remove any dead oysteri fr6m thebaske ts .

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At each sampl ing, the fol lowing parameters wgre mea6ured: alrand wa te r t empera tu re , sa l i n i t y , t u rb id i t y , ch lo rophy l l a (asa measure of amount of phytoplankton), orthophosphatephospha te (Po , -P ) , t o ta l phosphorus (ToT-P) , am lnon ium n i t rogen(NH, -N) , n i t r i t b p lus n i t na te n i t rogqn (No . + No- -N) andtotSr ic jeraahl ni t rogen (TKJN) z o

Temperalures were rneasured with a laboratory rnercury thermo*meter ("c ) ; sal lq i ty checked using speci f ic Aravi ty with ahyd ro rne te r ( s / cm- ) ; and tu rb id i t y exp ressed as a Secch i d i scread ing i n me t res ,

chlorophyf l a sampfes were f i l tered from the sampl ing bott leon si te. AppFoxlmately two 1l t res of water were f i f teredthrough a 47 mm glass f ibre f i l ter (GF/c) under fow vacuumpressure, using a Nal- geng f i l ter uni t mounted on a pfast ic twoLitre graduated cy1lnder, The f l l ter was air dr ied by increas-ing the vacuum, removed with forceps, fo lded into quarters,and stored in mani lJa envelopes. The water volume (to thenearest 10 ml-) f i l tered was noted and aIJ envelopes werestored in air t ight pLast ic bags and frozen unt i l required forana lys i s by the Nu t r i en t Labora to ry .

Water sampfes for nutr ient analysis were decanted intowh i r l packs (Namco) , s to red on we t i ce du r ing f i e ld samp l ing ,and kept f rozen upon return to the Nutr lent Laboratory unt i lanalysis could be carr led out, Detai ls of anal-yt ical methodsand the capabl l i t ies of the techniques used by the Laboratoryto determine nutr ient values (expressed in ug,/1) are givenl -n un r r r l nqs ( L9 /9 ) .

From f i f teen of the above bottom samples taken between May1981 and May 1982, an addlt ional loo ml water sample wasdecanted into smal l bott les and prepared as a cheek forhydrogen sul f ide using the sul f ide determlnat ion methrods inSky r ing e t a ! . ( 1979) . These ana lyses were done by Jud iHansen in CSIRo's Divis ion of Oceanography at Marmion W.A.

On three di f ferent occasions dur ing May and JuIy 1981, theamount of dissol-ved oxygen in the sea water c irculat ing aroundthe oysters in the carr ier tanks aboard the transport shipsand then fater under the rafts where the oysters weresuspended was es t ima ted us lng a D .O. me te r (De f ta Sc ien t i f i c ) .

1 .5 Indus t r y reco rds

Environmentaf data records based on a more systematic sampl ingprogranune were obtained from biologists stat ioned at the KuriBay fease si te. Water qual i ty parameters Lrad been measured ona bi-monthly basis dur ing 1976 and 1977 r , .ear at feast s ixoys te r ra f t l oca t i ons (M. Yasu i , , pe rs . comm. ) , S ince March1978, water samples from the same three stat ions designatedKuri Bay B, S-Place, and Camden B were sampled regular lydur ing one mid-neap t ide per iod of the month, and stat ionsdesignated Kuri Bay C, Hiro Bay and Sampson were measureddur ing the o the r neap pe r iod (F ig . L ) . A t each s ta t i on , wa te r

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The Kur i Bay fease s i te set t lement aLso acts as a weather

s t a t - i o n . R e c o r d s o f a v e r a g e m o n t h l y a l r L e m p e r a t u r e a n d- ^ - + l - \ t r r n : . i n f : l l t - ^ + ^ l q f n r f h p n e n i o d . l a n r r : r r r 1 O ? ? t h r . ) r l r r hL ' r L P L r

Apr i l 1982 were obta ined f rom the Bureau of Meteoro log-y andp l a c e d o n c o m p u t e r f i - l e f o r c o r r e l a t i o n a n d r e g r e s s i o nanalyses wi t l r o ther envi ronmenta l parameters.

I a + i i , ^ f i a l . l a v n a r i m o n t c

o y s t e r s f o r v a r i o u s f i e l d e x p e r i m e n t s w e r e o b t a - i n e d b yi n d p a : r l q p l 1 / I ] - d ' c 1 - r : n c n n r f s h i o s t - o t h e f i S h i n oq L L v r u P u L r y ! r r 9 r L q , 4 J

n n n r r n d q : n d q c J - f i n r r : q i r ^ l c t h c n e r r r r i r e c l n r r m h a r n f n c a r l . 1 \ / q l - . F I . S9 r v u r r u r r l Y s r r

a s L h e y w e r e b n o u g h L o n b o a r d f r o m t h e p e a r l i n g l u g g e r s a n dw e r e b e i n g s o r b e d i n t o t h e t h r e e s i z e c a t e g o r i e s . B a s k e t s o f. \ / q t e r q f o r a o i y p 6 1 t n o a l n a n l - k , a r p t : r r n c q t e i t h e r W i L h v a r l O u su y - u c r v r w L r , ! rr - - i ^ l - . F l r r n n l n r r n a d n l : q F i r - q l - r ' i n q r T r . r ^ r ' i t h e n r r m l - r a n e d n l a q f . i r -r - r r t \ J i r u r y ! L J v r t y !

sheep ear- tag cf ipped onto the s ide of the basket . Theindiv idual oysters in a g iven basket were ass igned numbers onet h r o u g h t e n o n t h e b a s i s o f t h e i r p o s l t i o n w i t h i n t h e b a s k e tf r o r n l e f t t o r i g h t .

U p o n a r r i v a L a t t h e K u r i B a y I e a s e s i t e , t h e s e t a g g e d b a s k e t swere ur i loaded f rom the carr ier tanks and suspended f rom then a f + i n r a n d o m i - p d n ^ - r + L l . h F o t h c r o v s t e r s f r o ml L u p L r s r u ' ( J r r > o r \ , t r g v u r r u r v r J U ! ] J J r

! l - , : ] - n a l 1 6 ^ t - i ^ h l - F i ^ T h a q o a y n a n i m c n f : l n n q i t i 6 n q l ^ r F r e

t a g g e d w i t h c o l o u r e d p l a s t i c s t r i p s t o p r e v e n t d l s t u r b a n c e b ycompany workers, Counts were made over the fo l lowing weeks ont h e n u m b e r o f d e a d o y s t e r s . T h e s u r v i v i n g o y s t e r s a t l h ef in ish of each exper iment then rever ted back in to thecompanyr s cu l ture system.

I . 1 . L S i z e , d e p t h a n d c l e a n i n g e x p e r i m e n t

T h e f i r s t m a j o r f i e l d e x p e r i m e n t w a s d e s i g n e d t o d e t e r m j n e t h ea F f a n f c a n m n n t : l i + \ r . \ f . \ \ , ' q f F y . q n f d i F f e r p n i q . i z e q o f t h e

di f ferent depths at which they were suspended f rorn the raf t a tthe lease s i te , and of the removal- o f fou l ing organisms' Atota l o f , 9oo oysters (9o baskets) were separated in to 18treatments (5 baskets each) consis t ing of three s izes measureddorso-ventra l l -y f rom the shel l va lve h inge to the growthe d g e ( s m a 1 1 < 1 o 2 n [ n ; m e d i u m l o 2 E o 1 4 o r n r n ; a n d l a r g e > 1 4 o r n m ) ;t h r e e d e p t h s ( 2 , 6 a n d 1 0 m ) ; a n d w h e t h e r t h e s h e l l h a d b e e nc l e a n e d o f f o u l i n g o r g a n l s m s a s i s t h e n o r m a l p r o c e d u r e o rl e f t u n c l e a n e d w h e n c o l l e c t e d .

The baskets of oysters were suspended under the nor thwestcorner of the Kur i Bay C raf t on 4 May 1981. The cumulat ivenumber of dead oysters in each t reatment was recorded in thef i n a l o f t e n o b s e r v a t i o n a l c h e c k s o n 2 4 J u l y 1 9 8 1 ( a f t e r 8 id a v s s u s p e n s i o n o n t h e r a f t ) .

o c o n f e . r i o n e x n e r i m e n L

A contagion experiment was also set up on 4 May 1981 on theadjoining northeast corner of the above raf t to see i f thed i s t r i bu t i on o f mor ta l i t y i n a l a rge number o f s im i l a r l yt rea ted oys te rs resu l ted i n a c lumped pa t te rn ( imp- l y ing some

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{ l ^ i r l - i ^ n n r ^ \ , i . i a . l } - . 1 r " A ^ m . i i a m a + a r h l : n L r n n l \ / a F h \ / l a n a

f i o ^ t s - F i f t v f I o a t s v r a r e q n p r - e d I h r c e r e l r . p q : n e r l - a l n n n t h e' l n n r r - l i n r .

r r , i F h f o r r r a d d i t i O n a - r f l O a b S C l r r c F c r ^ e d l - o r r c l h e r a l .a i r h ^ ^ a n r t i n h , , ^ " , , ^ + h a r h . ' h ^ r r ^ ^ a c T o W - L L h S b a n d t h e1 : n n o I i n a l n : n a e a n d a q q a r i a F a . l c f r . \ n d u , : 1 - o r m n r r e m e n l q f h er u r 9 L L

t - , ^ , n r n n u r a n a : r h r n r r a h I r r 6 O m I n r r a i n . \ n . l a y . l - . ' r h ar v P r J v v ! r

f i v e t i m e s t h e w a t e r d e p t h o f 1 : m a t l o w t i d e . T h e s e a r ) c h o rn o n F s . a l s o o f 2 4 - r f i F , l ^ ^ 1 . n . \ l \ / F f h \ / l F - a W e r e a t t a c h e d t o 5 mh c : r r r r r - h a i n \ r h i r - h " a s i n t u r n s h a C k l e d L . \ ^ n R n k ^ a n . h r l r

n n a ^ . . h l . \ . ^ - l i n a c m l F n . r i - h q . \ { l " m r i r h l r r - k n n l r / p l - h r / l a . F

: + l - : n h a r l r , n . l o n t h a t i F F r r f l n a r ^ - - , ^ - r - r - - ] r , . - ,L a u w o y

o e L w e e n f l o d t s , s o t h a t t h e b a s k e t s c o u L d b e s u s p e n d e d a L t h er r q r r : I , . m r i c r r I h ^n r l ha qn , l r - od 1 \ n ' FnF r t .

T o e g u a l i z e s o m e o f t h e d i f f e r e n c e s i n t h e a m o u n t o f t i m e< n a n 1 - ^ h ^ : n d l h 6 l r . a n q n ^ n l - q . - n F h F h a < L a l q . r l . r \ / e ] - a n q f i r n

l h a C ^ n F \ / i I I ^ r a l - I o n r - l i n e . c - c l e r r n e d o n t h e n e x l L o l a s Lr l : r r n f a n l l a c l - i n n r n r l + h F ^ \ / < l - a r a f n n ] - h o K r r r i R r \ / I n n n - l i n e

r ^ ^ F r f + , . . 6 F 6 + : , r a a z r 6 6 i - h e I a c , t d e v n f c o l l e c t i o n j . e . b e f o r e

. l a n r n F r r n o T h a n r < + h a ^ . / a 1 - a r q r r o n a n l r n a ^ n n + h o I n n a - l i n a

: l - r - a n p 1 / i I l a n a l d r r n i n d : q l . \ n ^ \ . / a n n F > f o ' ^ ' h ^ , , F q . \ n + h F , r ^ \ /

f ^ | 1 r r h i R i \ / r 1 I f h n e a q a J - c n f ^ \ / c t a h c h r , . ] h a a h i r i I i : I l r r

: h n : n r l t h e I n a n q n n r . l q h i i m r l - e l r r l w o o a y s .

1 ^ ^ ^ " ^ r e n s w e r e c o l l e c L e d o n 4 a n d 5 J u l y 1 9 8 2 .u r z r v v v y > u

O n b J u l y , 9 O O o y s t e r s ( 3 O O o f e a c h s i z e ) w e r e s u s p e n d e d f r o mI l - ^ . - ^ ^ \ / : r r ^ h ^ + r n n r - r - l i n e a q i m i l e r q n n n \ / q f F r < ' ^ ' e r " a n l : r : e dJ v v v j J r e r J w ! r ! v r u L

. - \ n + h F I n n n - l i n o a f K l r r ' i R : \ , : q u r a l I e < t h a 1 n ^ i n a l

o y s L e r s ( l 0 O o f e a c h s i z e ) o n L h e r a f t t h e r e o n ? J ! . r l y I 9 A 2 .

O n e h u n d r e d o y s t e r s o f e a c h s i z e w e r e t r a n s f e r r e d f r o m e a c hl o n n - l i n a l - ^ f h a r ^ f + ^ h l - ^ " ^ . r . + f \ l + a n / ) A ^ \ r c c , , c n 6 h c i ^ r I\ o r L c r . ] - v o J 5 r u s P L r r

l h i s m o v e m e n t r v a s n e p e a L e d o n 1 2 , S e p t e m b e r ( a f t e r 7 0 d a y ss u s p e n s i o n ) ; a n d L h e l a s t L h i r d o f b o t h l o n g - l r n e s e t s o fn r r q l - a n < u r e n e r n : n q f a n n a r l n n 1 - < n ^ + ^ h a n f : f a n o o r { r r r c

c r r q n e n q i o n ) T h F f i n : l n n r ^ f : l i l - ' / / - . \ r ' - t w a s m a d e a t t h econclus ion of the exper iment on 27 November 1982 (af .Ler aL o L a L o f 1 4 1 d a y s s u s p e n s i o n ) .

1 / . r : , T r : n c , \ ^ n t a v n 6 ' . i n 6 h +

I n o r d e r t o e x a m i n e t h e e f f e c t o f t i m e o f t r a n s p o r t , p o s i t l o ni n f l - F f r A n q n . r f f ^ n k s f i . e . h o t t o m m i d c l l e . ) r t . } n l c v c ' l ) : r d

p o s i L i o n j n w h i c h L h e o y s t e r i s c a r r i e d ( i . e . v e r t i c a l o rh o r i z o n t a l ) o n t t r e i n c i d e n c e o f m o r t a l i t y , t h e f o l l o w i n g- y ^ a n i m F . l q h , a r . a . - ^ n . l r r , , - f a . l , , r n l - L ' . ) | n = n q n a n l - r r o c q o l c i h M = r r

1 9 8 2 . . O n t h e f l r s t d a y o f o y s t e r c o l l e c t i o n i O b a s k e t s e a c hc o n t a i n i n g 1 0 o y s t e r s w e r e i a g g e d t h e n p l a c e d o n t h e b o t t o m o fthe tank. Ten more baskets were marked and p laced on them i d d l e - L e v e l a n d l O m o r e o n L h e L o p l e v e l T h e p r o c e d u r e w a sr e p e a t e d o n t h e 3 r d d a y o f c o l l e c L i o n i n a d i f f e r e n L t a n k ,The q^mc nn.)cedrr re , r ras carr ied out on the second boat on theq A ' n a n F A n f i d p e n r i r ' 6 ^ 6 : | 6 . 1 ^ h 1 - r ^ I l . r l - ' n r _ F q f . . \ r j - h t ' f i n c t . l a \ / O f

c o f l e c t i o n a t l a t e r n e a p t i d e s .

To compare mor ta l i t y be tween oys te rs ca r r i ed i n a ho r i zon ta lpos l t i on to those ca r r i ed i n t he conven t iona l , ve r t i ca l

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eseas TpT1s s^uI

2 . 2 B a c t e r i o l o g i c a l m e t h o d s

S t e r i l e s w a b s v , J e r e u s e d f o r s a m p l i n g t i s s u e s f o r b a c t e r i o l o g y 'T i s s u e c o v e r l n g t h e p e r i c a r d i a l a n d b o d y c a v i t l e s w a s o p e n e dusing ster i l ized inst ruments. Haemolymph was sampled f rom thep e r i - a r d i a l c a v j t y . T h e s u r f a c e o f L h e d - i g e s t i v e g l a n d w a sinc ised wi th a scalpef and then swabbed as far away f rom theg u t a s p o s s i b l e . S w a b s w e r e t h e n t a k e n f n o m t h e i n c j s e d g u t .When muscle was examined a smal l b lock of t issue was removed,s e a r e d i n b o i l i n g w a t e r a n d i n c u b a t e d i n b r o t h .

M e d i a u s e d w e r e n u t r i e n t a g a r ( N A ) , N A + 2 . 5 % s o d i u m c h l o r i d e ,T C B S ( t h i o s u l p h a t e - c j - t r a t e - b i f e s a l t s ) s o l - l d m e d i a a n d N A +2 . 5 % s o d i u m c h l o r i d e b r o t h .

S e l e c t e d c o l o n i e s w e r e c u l t u r e d o n s h e e p b l o o d a g a r a n d N Aw i t h 1 % , 6 % , B % , 1 0 % a n d 1 1 % s o d i u m c h l o r i d e .

Cuf tures were examined for physica l and b iochemical charactersa n d a n t l b i o t i c s e n s i t i v i t v ( T a b 1 e 1 ) .

I n t h e l a b o r a t o r y , m e d i a w e r e i n c u b a t e d a t 3 z o C . I n t h ef i e I d , m e d i a w e r e i n c u b a t e d a t a m b i e n t t e m p e r a t u r e w h i c hvar ied wl th t j - ryre o! day and seasonal ly . Anbient ternperaturesl . angeO r rom Zl - Jr+ \ - .

C r o w b h o f o n e i s o l a L e ( K B g / 8 7 ) w a s e x a m i n e d a t l 8 0 a n d 3 7 o c i nN A + 2 . 5 % N a c l b r o t h . B a c t e r i a l n u m b e r s a s c o l o n y f o r m i - n gu n i t , s p e r m i l l i l i t r e ( c f u l m l ) w e r e c o u n t e d b y s p r e a d i n g aknown volume of NA broth at 6 end 24 hours incubatlon ontoTCBS whi-ch was lncubated at 37"c fo? 24 hours.

2 . 3 V i r o l o g i c a l p r o c e d u r e s

Virus iso lat ion was at tempted f rom 1o normal ar ld 26 d iseasedoysters. Af ter the upper va lve had been removed, haemolymph,h a A n f ^ n d : n n r t i o n o f d j . r e s t j v e o l a n c l w e r e r e m o v e da s e p t l c a l I y . S a m p f e s o b t a i n e d l n t h e f i e l d w e r e f r o z e n i nI i q u - i d n i t r o g e n u n t i l p r o c e s s e d . F r e s h t i s s u e w a s p r o c e s s e df r o m n o r m a l o y s t e r s .

V i r u s i s o l a t i o n w a s p e r f o r m e d b y D r J . C o p l a n d , R e g - i o n a lV e t e r i n a r y L a b o r a t o r y , D e p a r t m e n t o f A g r i c u l t u r e , B e n a l l a ,V i c t o r i a . H e a r t , d i g e s t i v e g l a n d a n d h a e m o l y m p h w e r e u s e d f o ri n o c u l a t i o n o f t i s s u e c u l t u r e s . T i s s u e s w e r e h o m o g e n i z e d .H e a r t t i s s u e w a s d i v i d e d a n d h a l f w a s f i l - t e r e d t h r o u g h a 0 . 4 5m i c r o n m e m b r a n e f i l t e r . T h e r e m a i n i n g h a l f , d i g e s t l v e g l a n da n d h a e m o l y m p h w e r e t r e a t e d w i t h 1 o 0 0 i . u . p e n i c i l l i n a n d l o o oug st reptomycin/mI . Inocula were d i lu ted 1:5 weight , /vo lumew i t h M E M E a r l e ' s B S S w i t h 2 % f o e L a l c a l f s e r u m , p H 7 . 6 .A l i q u o t s ( O . 2 m I ) w e r e i n o c u l a t e d o n t o d u p l - i c a t e R T G - 2 , F H M ,CAE, BF-2, Grunt f in and BB f ish ce l l l ines and j -ncubated at24" . Two fur ther passages were made at 10 days durat lon.

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Gauze-f i l - tered and membrane - f i f tered inocula f rom the two

p o o l s o f d i s e a s e d o y s t e r t i s s u e a n d c o n t r o l t i s s u e w e r e

inoculated in lo the mant le cav i ty , t l - re adductor muscle ( in t ra-

muscular) and in to the haemolymph of normal oysters. Groups

o f 5 o y s t e r s w e r e u s e d f o r e a c h m e t h o d o f i n o c u l a t i o n . F o r t y

f lve oysters were le f t a t Kur i Bay and 45 were t ranspor ted to

Western Austra l ian Mar lne Research Laborator ies and held in

f towlng seawater at 27" , th is being the water temperature at

K u r i B a y a t t h a t t i m e . O y s t e r s w e r e o b s e r v e d f o r 4 0 d a y s .

3 . 2 L a b o r a t o r y e x p e r i m e n t s

At Murdoch Univers i ty oysters were held in 3O x 61 c m or 3b x 91

c r n g l a s s a q u a r i a w i t h a q u a r L z g r a v e l f i l t r a b j o n s y s t e m .

Tanks were housed in envi ronmenta l ly contro l - led rooms. l t ro

food was g iven but prev ious exper ience had shown that oysters

remain v iable for severa l months wi thout food.

F o r e x p e r i m e n t a f s t u d i e s ' g r o u p s o f 5 , 6 o r 1 3 o y s t e r s w e r e

used for test and contro l . A11 oysters used in an exper iment

h a d b e e n t r e a t e d i n t h e s a m e m a n n e r p r i o r t o b h e e x p e r i m e n l '

In rnost cases oysters had been Lre ld fo t ' 1-8 weeks in the

lahoratory before bei r - rg used. Pre l iminary s tudies to tL Iose

l - is ted ln Tabfe 2 are presented in Appendi .x 1.

3 . 2 . L B a c t e r i a l - i n o c u l a

T h e f o l l o w i n g b a c t e r i a w e r e u s e d : V i b n i o . P ' K B 9 / A l , K B 7 / 4 2

and, Pzeudomonaz putze,dacie-nz ' The Vj -br ios had been iso lated

from the haemolymph of s ick oysters at Kur i Bay and were la ter

i d e n t l f i e d v s V . h a z v e y i P . p u t n i d a c i e . n z w a s i s o l a t e d f r o m

tLre haemolymph of d lseased oysters dur ing pre l iminary investr -

g a t i o n s l n t h e l a b o r a t o r y . ( s e e A p p e n d i x 1 ) .

Bacteraa were grown in 1Oo mI a l iquots of nutr ient brot f l +

2 . 5 % N a C I a t 3 7 " . s i x - h o u r c u l t u r e s w e r e c e n t r i f u g e d , w a s h e d

three t imes in s ter i le sa l ine and resuspended in loo ml-

s t e r i l e s a l i n e .

Bacter ia l concentrat ion was determined by a s tandard ser ia l

d i lu t ion tecLln ique us ing NA + 2.5% Nacl as t l re medlum.

Resul ts were expressed as cfu/ml .

3 . 2 . 2 E x p e r i r n e n t a l r e p r o d u c t i o n o f d i s e a s e

Experimentsi sol-ates KBfo r oys te rsoys te rs he ld

1 - 13 were per formed to establ ish whetherg/8I , KB 7/a2 and ?. putne{ac ia,n4 were pathogenic

and to^determiqe whether pathogenic i - ty d i f fered ina t 1 9 " a n d 2 9 " .

The protocol o f Exper iments 1-13 is presented in Table 2.

Pathologlca l and bacter io logica l procedures were per formed as

indicated above unfess the oysters were used for Condi t j -onr n d e x s t u d i e s ( b e i o w ) '

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Lrorsenur IetJe+cPg

oysters used for cI determinat ion were removed from thelrshel1 by cutt ing the adductor and pal l ia l muscle at tachmentson both valves ind transfefr lng the oysters intact body massto a pre-weighed beaker for measurement of wet wej-ght.Oysters were then hornogenised and repl icates weighed intosnal ler pre-weighed beakens to be frozen overnight. FrozenrepJ- icates of oyster homogenate were t l ren freeze-dr ied, beforebeing heat deslccated to constant weight.

The Condit ion Index was calculated as:

To ta l d ry oys te r mea t we igh t x 100 , (Shaw e t a1 . , 1967) .Total wet oyster meat weight

1. Analyt ical methods

The experimenta] factors af fect ing the proport ions (p. . , ) ofoysters which were dead at a speci f ied t ime per lod weFeexamined us ing a l i nea r l og i t mode l , 1og (p - (1 -p . i ) ) . Th i sanalysl s assumes a binomial error distr ibut ion whlch isgeneral ly considered appropr iate in conslder ing the var iat ionsbetween proport ions (Cox 197O). The stat ist ical packageGENSTAT .(efvey e! al . , 1977 ) which uses the nlaxirnum l ikel ihoodnethod of Nelder and Wedderburn (1972) to est imate the l inearparaf iFters of the models, was used to analyse the data. thesigni f icance of factors and their interact ions in the modelsin explaining the var iabi l i ty in the proport lons of deadoystefs was detertnined by f i t t lng two models, one with thefactor and the other without.

Thq di f ference in the l ikel ihood rat io chi-square stat ist icsG') of the two modefs with the appropr iate degrees of f reedom(d . f . ) was then used to de te rm ine the s ign i f i cance o f t hefactor, providing the second model adequately f i t ted t t re rawda ta (B i shop e t a l . 1975 ) . The ra t i o o f t he pa ramete resplmate to i ts standard error can also be compared to the tdistr ibut l -on in exahining the signi f icance of the parameter3in the model . When parametens of models which did not provideadgood f i t needed to be evaluated' a heterogeneity factor '9 ' / a , t . (F inney IETL) , wag app l l ed to the va r iances o f t heparafteters.

The conrplementary 1og-1og l ink wa6 also corrsidered in theanalysls of the experiments. As this method gave very simi l"arregultb to the logi t l ink, only resul ts using the logi t l lnkwi l f , be ! 'eported.

, IV RESULTS

1. Fie1d stu4i.es

L .1 Wate f qua l i t v na r .amp l -pFq-

Lacking publ ished basel ine data on water qual l ty parameters for thefar north Western Austral ian coastal reglon, the search for someenvironmental factor that might contr ibute to mass mortal i ty in pearoysters was largel-y a process of el iminat ion af ter making in i t ia l

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Lack of d issolved oxygen was not impl icater i as a cause ofmorta l i ty as the min imum level o f oxygen detected e l ther in thec a r r i e r t a n k s f u l l o f o y s t e r s d u r i n g t r a n s p o r t o r u n d e r b h er a f t s a t L h e d e p t h b h e b a s k e t s o f o y s L e r s w e r e s u s p e n d e d w a s 5 . 8n ^ n t q n e r m i l l i o n o r a l c r r e l r - n r r e q n o n d i n r r + . \ R A % n f a i r q a t u r a -I J q T L J P L I

t i o n . S a m p l e s i n c l u d e d r e a d i n g s t a k e n u n d e r t h e r a f b s j u s t

before sunr ise when the oxygen contr ibut lon f rom photosynthet ica r F i r | i + \ / k i . l , r l d h F ! p a l h ^ ^ + - ^ t ^ r . \ ^ - + : - i n i m u m . s e s s i l e a d u l tu y v r v u ! v v u ! r \ y e e

L r i \ / , a I \ / e q o e n e r a l I v h ^ \ r F I c r r n ' r e q n i r a f . o r w n a r r r | i n p r n p n t < a n d e d i [ l gY v r r e r q r r y r r u v L ! v v v r v J y ! r

^ \ / c t a F a F n ^ a w : r n n l a - a r . a L n . \ k n 1 1 - n q t r r r l i r r c f n r r r n t o f i r r c d a- . - * J Y S' i n , , , : + a n ^ ^ ^ F r i h i n n I o c e l h = n 1 o n 6 m ^ v ! , ^ a n f S n r n t q e t a l

:_ : : : - ' I

1 9 s e ) .

K r , r i R : \ / ' q i q n l a t e d 1 ^ . a l i ^ n p F F e r ' l - i r r e l r r n r o r ' l t t d a d p n r r i r n n r r a n l S fr r u r f ! u y

d e g r a d a t i o n d u e t o u r b a n a n d i n d u s t r i a l p o l l u t i o n . S l n c e 1 9 7 5P e a r i s P L y L t d h a s f u r t h e r r e d u c e d m a n - m a d e p o l l u t i o n n e a r t h e i rI o , a q e q i 1 - c h v F . v c n r A q f n i r - t i n n t h c r r q e r ' r f q r r n l - h c F i . . i a l - c r r r F n t S

and ensur ing that any rubbish was burned and bur ied ashorerather tLran dumped i -nto the water at the lease (H. Kan-No and H.U e m o t o , p e r s . c o m m . ) .

A n u m b e r o f p o t e n t i a l 1 y t o x i c p h y t o p l a r r k t o n s p e c i e s h a v e b e e nr e c o g n - l s e d i n t h e t r o p - i c a l w a L e r s o f f N o r t h w e s ' u A u s L r a I i a b e l o n g -i n g t o t h e g e n e r a 9 g m n , o d i n i u m , 6 9 n < t d i n i u m , $ o n y a u l o x ,l i , t t o c e n t t u n , a n d i q i c h o d e . t n i u n ( C . H a j l e g r a e f f , p e r s . c o m m . ) .I n L a a n i r , . r u r i t h l - h c r r c n c r . a l . l ' 1 : r . 4 . - t a r i q l - i r . e r r f t r . ^ n i . : l

n a n m r r n i + i p q \ / a n \ r l . : r . . r a y , ' \ r r m h o } ^ a ^ f e n a . ' i n 7 ' a c a n f h r r { -

r - r s r - - r a I I y w i t h o n l y s m a l l p o p u l a t - i o n s o f e a c h . T h e m o s t s t r i k i n gp L l e n o m e n o n i s a n n u a l s u r f a c e b l c o m s o f T n i c h o d e . z m i u m s p . a l o n gt h F - . \ : < l - e q n c ' r - i a l l w d r r r i n . r O . , t ' . \ h F r . a n . i N o v e m b e f w h e n t h e w i n di s s l i o h t I I n l i k e t h a ^ f t a n h e : r r r r r n n r t - a l i l ' v o f n e a r ' l o v q t e r s

a s s c c i a t e d w i b h r e d - t i d e s i n J a p a n , t h e d e c a y o f r t i c h l d e z n i u naf ter b iooms has not been r roted to cause unusual morta l i t leso f p e a r l o y s t e r s o n l e a s e s i n A u s t r a l i a o r w h e r e i t h a s b e e no b s e r v e d o n c o a s t a l f e a s e s i n f n d i a ( C h e l I u m a n d A l a g a r s w a m i ,1 9 7 8 ) .

Given the non-estuar ine locat lons of the cul ture leases and thelack of apparent anomaf ies in the above water qual l ty para-m e L e r s , a t t e n t i o n w a s f o c u s e d o n w a t e r t e m p e r a t u r e . R e c o r d skept by the Japanese staf f a t Kur i Bay showed a water tempera-ture d i f ference dur ing the winter months^between the col lect ionr - r n n r r n d q r n d f h e i r l e a s e s i t e o f a l m o s t 7 " C . T h i q I F n . r er a m ^ a - r t r r n o r l i f f a n o n n o * h r l - t - h r ' . r \ , / < + a F q : n a q h i e n l - F , , 1 + ^ , - l r 1 '- - - ' 1 n gt ranspoft approaclres the maxlmum annual var iat lon of not more+ r r - n l ^ " . - + - h , , h l a n o i n f h a n n o r r ^ . . a : n f N . i i - . \ ' l

' l A A n ) A! o u o , r y P r q e u r r r v , , e v P L r r

t e m p e r a t u r e d i F f e r e n c e w a s c o n f i r m e d b y o u r s a m p f i n g i n J u n e' ] 9 8 2

w h e n t h e s u r f a c e w a L e r t e m p e r a t u r e a L L h e f i s h i n g g r o u n d s" - ^ r a r r i r r : l a t K r r r i R ^ \ l a f e w d a v s I a t e r . a

q r r r . f a . c f p m n F r ^ i r r r c o f 2 5 . 8 " C w a s r e c o r d e d .

I . 2 I n d u s t r y r e c o r d s

A s f a r a s i s k n o w n , n o p e a r l c u f t u i e c o m p a n y o L h e r t h a n P e a r f sPty Ltd has kept systematic records of any envlronment water gual i ty6 ^ F ^ m a + a r F n n i r r n > ' | - c l \ / l - h a . I : n a n c q o h i n ] n a i c : l q r - = f f r l - K r r n i R r r r

been instructed to keep records of monthly water samples takena t t he same ra f t s i t e s ta t i ons s ince 1978 as pa r t o f t he in

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w h l 1 6 a l ] t n r e e o y s r e r s i z e c a t e g o ' f i 5 s e x h i b i L m o r L a l i t y t h a ti s i n v e r s e l y r e l a L e d t o w a t e r t e m p e r a t u r e a t t h e t i m e o fL r a n s p o r t , a h i g h e r p e r c e n L a g e o f t h e s m a - l l e r s - i z e d o y s t e r s d i eo v e r t h e e n t i r e s e a s o n t h a n d o l a r g e r s i z e d o y s L e r s f o r u n k n o w nr e a s o n s .

T h e m . \ r f , a l i l 1 / . l e l - : f o r i : h e \ / e : r 1 q A 1 ( F i . f l r r . a 6 I u r : c t - r r n + h o n

e x a m l n e d f o r s e a s o n a l e f l e c t s b y c o m p a r i n g L h e f . i r s t h a l f , o ft h e i i s h i n g p e r i o d w i t h t h e s e c o n d h a l f u s i n g a n a l y s i s o fc o v a r - i a n c e . A [ t e r t h e e f f e c t s o f o y s t e r s i z e a n d w a l - e r t e m p e r a -ture were taken in to account , no s lgni f icant d i f ference inm o r L a l L t y b e t w e e n o y s t e r s t r a n s p o r t e d a L L h e b e g i n n i n g o f t h ef ish ing season compared wi th those t ranspor ted dur ing thel a t L e r h a L f w a s d e t e c t e d ( F , , , . . . , . = 1 . 5 5 , n . s . ) . S i g n i f i c a n ts e a s o n a f e f f e c L s m i g h t b e e * p 6 d t e d i f t h e m o r b a l i t y r e s u l t e df r o m s o m e o t h e r s t r e s s f u l a n n u a l p h y s i o l o g i c a l p r o c e s s s u c h a ss p a v ' r n i n g a s w a s i m p l i c a t e d i n m a s s m o r t a l i t i e s o f e d i b l eo y s t e r s j n J a p a n ( S i n d e r m a n n , 7 9 7 9 ) .

I i M ^ n i n r r l . a t i r , ' c f i a l d a : r n a n i m e n l - c

1 . 3 . 1 S i z e , d e p t h a n d c f e a n i n g e x p e r i m e n t

T n | - h a I i n c : r . I n a i l - . r n e l r z q . i q o f . ) \ / q f a n m n n f : l i i - r r i h F L ar r v r v y r u e r v J , L r r u r r L

e x p e r i m e n t o f d i f f e r e n L s i z e d o y s t e r s , c l e a n e d o r l e f tu n c - [ e a n e d o f f o u l i n g o r g a n i s m s a n d s u s p e n d e d a t d i f f e r e n t. l F n f h s . n . \ q i . y n i f i n : n f d i [ [ e r e n C e w a s f o r r n d i n t - h a r - t a : n . i n n

f a c L o r , O y s t e r s i z e , h o w e v e r , w a s f o u n d t o b e v e r y s i g n i f i -c a n t ( P < O . O O 1 ) w i t h t h e p e r c e n t a g e m o r t a l i t y d e c r e a s i n gi n v e r s e l y w i t h o y s t e r s i z e ( s m a l - t 7 2 . 7 % ; m e d i u m 6 3 . 6 % ; a n dI a r g e 3 9 . O % ) .

Whi ls t the depth categor ies of 2 m and 5 m were not found tob e s i g n i f - i c a n t l y d i f f e r e n t , t h e d e p t h c a t e g o r y o f 1 0 m w a sl o u n d t o b e s i g n i f i c a n L l y d i l f e r e n t ( P < O , 0 5 ) w i t h l e s sm o r t a l i t y f o u n d a t t h e 1 0 m d e p t h c o m p a r e d w i L h 2 a n d 5 m .T h i s r e s u l t , h o w e v e r , w a s n o L c o n s i s t e n t o v e r a f f r e p l i c a t e sof the exper iment as there was a lso a s igni f icant in teract ionbetween depth and repl icates which is probably due to thep o s i t i o n i n g o f t h e r e p l i c a L e s o n t h e r a f r .

I . 3 . 2 C o n t a g i o n e x p e r i m e n t

The resul ts of the contagion exper lment ind icated that thedisease was not contagious. The mean number of dead perb a s k e t w a s 4 . 9 2 ( w i t h s a m p l e v a r i a n c e o f 3 . 2 O ) f r o m 9 O b a s k e t sof large oysters. The d is t r ibuton of the number of dead perbasket was f i t ted to a b inomial - d is t r ibut ion in order todetect ev idence of non-randomness and resul ted ln a good f i t( P > o . 0 5 ) . A s e c o n d t e s t f o r n o n - r a n d o m n e s s , t h e b i n o m i a li n d e x o f d i s p e r s i o n ( S e b e r , t 9 7 3 ) , w h i c h i s b a s e d o n c o m p a r i n gthe observed var iance est lmate wi th an est imate of thee x p e c t e d v a r i a n c e u n d e r a b i n o m i a I m o d e l , w a s a l s o u s e d . T h i s

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'(9 arqeJ )

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luaurr.redxa dlrsuag t't'T

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sasuPrJen oM'-l aq1 LtasMJaq ACuaJsJJTp al{l r]T paflnsaJ xsal

There was no s igni f icant d i f ference in mcr ta l i tycarr ied in the hor izonta l pos i t ion wLren comparedc a r r i e d v e r t i c a l l Y .

1nt o tho se

2 .

There was no ev idence of excesslve morta l i ty in oysters ln

thei r natura l env i ronment . Morbld i ty and/or morta l i tyoccurred af ter arr iva l a t the lease s i tes ' The cumulat ivem o r t a l i t y a b t i m e s a f t e r a r r i v a l i s p r e s e n t e d i n F i g . 9 . f na o n n r a l F h p r e w a q l i + l - l a . \ r ^ n . r m . r n t ^ l i ] . v w i t h i n t h e f i r S L 7

days and thren a gradual or rapid r ise over the next 3O-4Od a y s . T h e i n c i d e n c e o f n , o r t a l i t y r a t e v a r i e d c o n s j - d e r a b l y f o r

oysters co l lected wrth in and between neap t ides and wasn r . c . a l p q F d r r r i n . r t h e u r i n l - e r m o n l h q ( F i n - 6 ) .

T h e r a t e o f m o r t a l i t y v a r i e d r n v e r s e l y w i L h o y s t e r s i z e ( F l 9 .8 ) . S m a l l o y s t e r s ( < l o z m m ) s i c k e n e d a n d / o r d i e d a t aq r e a t e r r a b e t h a n m e d i u m l I o 2 - I 1 ; o r n r n ) a n d l a r g e o y s t e r s ( > 1 4 5m m . r .

2 . I P a t h o l o q v

Diseased oysters were recognized by gaping, wi thdrawal of them e n i l a . a n d d i n t v q t a i n e d n a c r e b e t w e e n t h e w i t h d r a w n m a n t l ea n d t h e e d g e o f t h e s h e l l . T h e d e g r e e o f t h e s e c h a n g e s w a svar iable. In many cases gapj -ng oysters c losed rapJ-d1y uponst lmulat ion but soon reopened. Olhers c fosed sIow1y and wlrenclosed the valves were not complete ly apposed. Those oystersthat d id not c fose but s t i l l a t tempted c losure and had obvious^ + 6 h i / i i r | = d { - i v i { - 1 ' : r a . t h / l : n t h a ^ J - i n a r ^ r a r . c c l e q . q . F . lv , u r r v

as mor ibund. Dead oysters gaped fu l ly and putref ied rapi -d lybut c tenid j -a l act iv i ty was occasional - Iy seen in these oysters.

Vis ib le fes ions were not apparent on gross examinat ion.However, h is to logica l examinat ion revealed les ions in 74% ofoysters examined. Vaniabl -e s ized focal accumulat ions ofp h a g o c y t i c h a e m o c y t e s w e r e p r e s e n t i n m a n y t i s s u e spar t icu lar ly the connect ive t i ,ssues of the d lgest ive g land( F i q . 1 0 ) a n d m a n t l e . G e n e r a l - i z e d o r f o c a f a c c u m u l a t i o n o f

phagocyt ic haemocytes were commonLy preserr t in the aur1c le( F i g . 1 1 ) a n d v e n t r i c l e . A c o m m o n f i n d i n g w a s e r o s i o n o f t h eepi thel ium of the outer fo fd of the mant fe and the marginalz o n e ( F i g . 1 2 ) a n d i n f i l t r a t i o n o f t h e s u b e p i t h e l l a l t l s s u ew i t h p h a g o c y t i c c e l l s . G r a m n e g a t i v e b a c t e r i a w e r eoccasional - ly seen wi th in phagocytes in a l - l t issues.

Atrophy of g landular epi the l ium of d i ,gest ive g land tubules anddi la t ion of tubules was conmon.

Ul t rast ructura l examinat ion revealed no more than l ightm i c r o s c o p i c f i n d i n g s .

Other h is to logica l f ind inEs that were seen j "n normal anddj -seased oysters but were not refated to d isease are neferredto in Appendix 2.

- 2 8 -

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-6e-

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uoJJ sa?eTosr a^rd 'r"ro11ed pue JeTncJTc 'xaluoc 'qloouls eJaMsaTuoToc aql Je6e sscJ uo '^zt 1e Lrorleqnsua sJnoq ?z 1e ISeN

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Jo xnn aq+ urorJ srequnu "ur".':i";:{:.::;":::A ru: jiH;;

'suoTseaco TTe uo paurelqo +ou aJarvr saJnlTnc aJndpue (pIaTJ eq+ uT auop lou ereu slunoc aleld lcexe) qlmo.:6

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07 Jo rldurfToureeq aql uoJJ uoT+elos1 .6.reu1.rd uo peJnlInc eJaMeIJalpEq sJalsdo Icas tg luoJg 'oI.tJe+s ,611ect6o1o1Ja?ceq seu

qceeg eTrll-og aql 1e peldues sJals^o TeuJou uro.rJ: qdurdlouraeg

T6oroGFes z'z

t-

Bacter ial counts on water in carr ier tanks

The numbers of Vibr io sp. in the water of the carr ier tanks andthe rat io of Vibr io sp. in the tank water to that in seawaterat the 80 Mi l -e Beachr are presented in Tabfe 9. The number ofvibr io sp, in the surface water of the carr ier tanks roseduring transport but the number varied between and withindays. The numbe r of bacter ia was greatest in the waterimmediatefy af ter the carrying baskets had been removed andwas even greater in the sludge remaining at the bottom of thetanks after they had been pumped out.

3. Experimental disease

The term I 'morbid ' . is used to denote those oysters which becameabnormal dur ing the experiments. Morbid is equivalent tot ts ick". The cr i ter ia used to evaluate whether an oyster wasmorb id were as fo l f ows : (a ) s l i gh t gape , no man t le re t rac t i on :(b) mantfe retract ion, no gape; (c) gape but responds tophotost imul i ; (d) gape but responds to v ibrat ion; (e) gape butresponds to direct st lmuf i ; ( f ) gape and mi l-d mant leretract ion; (g) gape and severe mantfe retract ion; (h) edge ofman t le fo lded ; ( i ) no reac t i on to s t imu l i ; ( j ) r eac ts on l ywhen ou t o f wa te r ; ( k ) s l ime masses i n man t le cav i t y ; ( I ) w ide

3 .1 F ie ld t ransmiss ion expe r lmen ts

Morbidi ty/mortal i ty dld not occur in any group of inoculatedoysters held ei ther in Kur l Bay or t ransferred from Kuri Bayto Pe r th .

3,2 Laboratory experiments

The resul ts of laboratory experiments are summarised in Tabfe10. Disease was not produced in al l exper iments. In experi-men ts 1 ,2 ,3 ,9 and 10 tL re re was a g rea te r l eve l o f morb id i t y , /mortal i ty in infecled than in control oysters. This occurredat both 19" and 29". In experiments 4,5,6 and 8 there was11tt le di f ference in the leve1 of morbi di ty, /mortal i ty betweenj-nfected and controf animafs. In experiments 7 and 11 therewas l i t t le dl f ferenqe between lnfected and non-infectedoysters at 19o or 29o but overal l , the morbldi ty, /mortaAl tyof^both j -nfected and controf animals was greater at 19' than

The morbidi ty/mortal l ty data were combined for al l the aboveexperiments (Tabte 1-1). When evaluated using l inear 1ogi f ,analysis, the morbidi ty exhibi ted by control oysters wassigni f icantfy less than oysters infected with lz. hanve-gi(P<o .oo1 ) . I n t he same ana lys i s , s i gn i f i can t l y l ess morb id i t ywas exhibi ted by oysters kept at 29" compared with those helda t 19 " (P<o .oo1 ) . The p ropo r t l on o f mor ta l l t y was s ign i f i -cant ly fess in controL oysters compRred to infected oysters(P<0.q5), and in oysters kept at 29"C compared with those helda t 1 9 " ( P < 0 . o o 1 ) .

- J U -

- Tt -

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qloq ' 1 luaurr.radxe u1 'fyr etqe;, ) TTEJa,ro IC lseq6Tq aq1 pel{

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sE aues aq+ aJaM sdno;6 IIe uloJJ s.ralsdo uT uaas suorsaT aq;

'ds

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E x p e r i m e n t - s 7 a n d E w e r e r e p l i c a t e d i n e x p e r i m e n t t 1 1 ( T a b l e

2 ) , h o w e v e r t L i e s a m p l e s i z e w a s i n c r e a s e d f r o m 5 t o 1 3 o y s t e r sn F n l - r c ^ f m c n t R e q r r ' l 1 - q o f e . n a n i m F n F 1 l r ^ r c r . r : r - n m n a r : h l e r n r i l - , 1P L I v l . ' " " x r " .

f h a 6 r ^ t i a r 6 v h a F i m a n F c L r i l h ^ \ r e F a F q l - r a r , - ' l ^ + r n " L r i h ^r r e r e ^ P L r v j r v u r l r o v ! L r \ j

s i g n i f i c a n t l y h i g h e r C I ' s t h a n i h o s e a t 2 9 " ( p o o l e d c o n t r o la n d t r e a L e c i v a l u e s a t e a c h t e m p e r a t u r e ; P < O . O l ) . N o s i g n i f i - 'cant d i f ference was observed between oysters wi th in eachc e m p e r a L u r e g r o u p , n o w e v e r t h e r e w a s a s i g n - t f l c q n L d i f f e r e n c eb e t w e e n c e n t r o l c y s t e r s a t e a c h t e m p e r a t u r e ( 1 9 " c o n t r o L s

, o " . - ^ n t r . . 1 1 < ' p < n n 1 ) ^ n r i h a l - h ' F F n ! l ^ F : t - F r i . \ \ / q l c n q F t" - " ' " - " F

e a c h t e m p e r a t u r e ( l 9 " t r e a t e d v e r s u s 2 9 " t r e a t e d i P < o . 0 5 ) .

V D ISCUSSI t )N* r . c l a s a n v . h a n . r c f r o m n o r i n a - L s t r u c t u r e o rr l o y u q

F u n c t i o n c f a ] i v i n g o r g a n i s m a n d a s s u c h i s m a n i f e s t i n m a n y, - . , - - - . - l L . - ^ - , , 1 . r - i ^ l ^ ^ . . , , . . - t i . F i ^ - / - ^ . F ; ^ l ^ ^ i ^ - \ n i - ^ - - ^ ' . -v { c t y S d f l u I l d s l l l u I U r p r e u - s 5 \ d c u t u I U g I e ! ; . / . U - L S c d s e l l , d y

b e d u e l o , n f e c L j o u s a g e n t s s u c h a s b a c L e r i a . v i r u s e s ,p r ^ o E o z o a a n d f u n g i . N o n - i n f e c t i o u s a e t i o l o g i e s i n c f u d ec h e m i c a l L o x i n s , o h v s i c a l a g e n b s , a n d h o r r n o n a l , r n e t a b o l i c o rg e n e t i c a b n o r m a l i t i e s . M o s t d i s e a s e s h a v e a m u l t i f a c t o r i a lc a u s a l i t y . O f t e n a m a j o r f a c t o r i s r e s p o n s i b L e b u L i n m o s ti n s t a n c e s d l s e a s e w i l l n o t b e e v i d e n t u n l e s s o t h e r o r e -r l l n n a c i n n f r n . F n n c r ^ + ^ . + h a ^ n ^ . n . i c mu L - P v - f , 1 ' 9

nc:1-h or mr - rn f a l i t rz i s the most over i man i fes ta t lon o fd i s e a s e . A l l p o p u l a t i o n s h a v e a b a s i c o r " n o r m a l " l e v e l o fm r n l - : l i t r r M : q q m ^ n + : l i t i e q : n a n e a : n r l a r l r < a n a v r n a a n = f e r i

f o r m o f n a t u r a l m o r t a l i t y ( S i r r d e r m a n , 1 9 ? 9 ) a n d a r e o f t e nn : r r i c l j n . \ n q F l = r r d a f { e r l a | : n a e n n n n n n + : ^ h ^ f F } - ^ h ^ } \ , ' l - + i. - F - . L r ( ) I l ( ) l L i I e P ( ) P u l - d L l O n

, . . i ^ ^ ^ ^ ^ ^ - - n r - i r - A r r r ^ H . ) r " , F \ / F n l o c a l i z e d m a s s

m o r t a l i t i e s i n r e s t r i c t e d g e o g r a p h i c a r e a s d o o c c u r . f n m o s tc a s e s m a s s m o r L a l i L i e s i n o y s L e r s a r e p r o b a b l y d u e L o i n L e r -a c L i o n s o f e n v i r o n m e n t a ] a n d b i o l o g i c a l f , l c t o r s , L h e I a L b e r i ns o m e i n s L a n c e s b e i n g i n f e c L L o u s . H o L , r e v e r , o n l y a f e w m a s sm o r t a l i t i e s h a v e b e e n d e f - i r r i t e l y a s s o c i a b e d w i t h s p e c i f i cn a f h n . r a n q n r c d ; r t , r r s o r e n v - i r o n m e n t a l f a c t . ' ) n c . ( S n n a r r r r a 1 q 7 1: t -noerman t9 / J ) -

M . ' r l - : 1 i 1 - r r i n n c . a n l n r r - t e r . q i n 1 - h e n n n l - h - 1 s s t o f W e S t e r n

A u s t r a l i a c o u l d b e c l a s s i f i e d a s " f o c a l i z e d m a s s m o r t a l i t i e s " ,1 l w a s r e a d i l y a p p a r e n L L o t - h e i n d u s L r y t h a t t h e r e w a s l i t t l emorta l i ty of oysters rn the i r natura l env i ronment whi ler r n r f : l i l - i e s r r r e - 1 , - n l h ; r n I l n a l n e n : r d F . i a S " n o l ^ m a l " O c c u r f e dA F t e r o v s l . e n s h a c l h e e n t r , : n s f e r r e d L o I e a s e s i t e s . T h e I e v e Io f m o r t a f i t y c o n s i d e r e d t o b e " n o r m a l " b y o p e r a t o r s w i t h i n t h ei n d u s L r y v a r l e d f r o m 1 t o 2 a % b u t t h e c a u s e s , o b h e n t h a nn r . a d : f i ^ n i n l r ' - : l i z e d a r e : q r ^ r c r e r r n k r r r ; y J n .

T n \ / c q t . i r r a l . i o n o f t h F , - ^ - r = 1 i + " ^ h ^ l a l - - ' { a S C a r r j " e d O U Cn r i m a n i l r r r ^ r i f h . n F . n F r ^ t . . ) r w i f h i n t h e i n r ' l | s t . r v - A l . t h e i i m en f t h e i n r r e q l i n a l - i a n l - h i , : . ] n F n : F . . t - i 1 l A v n p r i a n . . i n . t

h F A \ / \ / m n r f e l i t i c s r ^ r h F r . F ^ < F h F n n . \ h l e m n r 1 6 l h g r l e a s e s i t e s h a d:nn : r . cn t - l r r dc r - l i nc r l O \ rq f p r s , r . r c re 1 - r anq .nnn l - cd h r r sh i n l - r ' r t he

f e a s e s i t e i r r v o l v e d i n L a n k s w i t h f l o w t n r o u g h c r r c u l a t i o n ,T h e L l m e s p e n t w i t h i n t a n k s v a r i c d f r o m I . 5 - 4 . 5 d a y sd e p e n d i n g o n b o a t s c h e d u l e s a n d t h e l e n g t h o f n e a p L i d e s .

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Wol f and Sprague (1978) s ta ted tha t t he i r "p ro t i s tan pa ras i te "was not present in normal oysters and that the bodies wereassoc ia ted w i th "bad ly d i s ln teg ra ted " ep i the l i um. A l though wehave seen these bodies in al l oysters they are of ten more: n n a t . a n t i n . 1 \ / c t a n c u ' i + h : t r . ^ ^ h v a f r l i a a c 1 - i r r a a l r n r l = n r l i n

d i g e s t i v e g l a n d s t h a t a r e a u t o l y t i c . T n o u r e x p e r i e n c e m a n yd i q c a q a d . \ 1 7 q J - a r . q h : \ / a e f n n n h r r n l - { - h p r l i d a < f i l ' a n l r n r l a a l l c

presumably f rom reduced food in take, and auto lys is is comrnoni n t l s s u e s f r o m m o r i b u n d o r r e c e n t l y d e a d o y s t e r s . I t i sn ^ c e i h l a F h . a f 1 - h c r r l - r a r i l \ r r l i q i n f e n n : f a r l f i < e , r a r l a e a n i F a r l l - r r r

Wof f and Sprag 'ue (1978) was i n fac t . au to l ysed and no td e g e n e r a t e o r n e c r o L i c .

We bef ieve that the structures referred to as "prot istanparasi tes are not pathogens but normal const i tuGnts of thed i g e s t i v e c e l f s o f P . m a x t m a .

V ib r i os i s o f mar ine f i sh (gnderson and Conroy , 1970) andshe l l f i sh l - a rvae (Tub iash e t a f . , I 97O; B rown , L973 ; B rown andLosee , 1978 : D isa l vo e t a l l l -T578 ; E l s ton and Le ibov i t z , 198o ;Je f f r i es , I g82 ) i s a f iET l - recogn ized d i sease . I n f i sh thedisease is manifested by haemorrf iagic sept lcaemia and,/or anu lce ra t i ve sk in d i sease . The o rgan lsm respons ib l -e i n t herna jo r i t v o f ep i zoo t i cs i s / . angu i l , Lazum. In she l f f i shv ib r l os i - s was f i r s t desc r ibed as "bac i l l - a ry nec ros i s " (Tub iashe t a I , , 1965 ) and i t occu rs i n na tu ra l l y g row ing and ha tche ryrea red l a rvae and j uven l l e cu l tu red oys te rs (E Is ton e t a I . ,1982) The organisms responsible for th is disease tefrd-E6 fat linto three groups; those resembLing V. ongui l lazum, throseresemb l ing V . o lg ino lg t i cuz , and a th i rd g roup o f l a rge l yur: .named Vibr io species.

V ib r i os i s o f adu l t oys te rs i s a l ess we l l desc r ibed d i sease .Tub iash (1971) repo r ted tha t s t ra ins o f / . angu i l l onum wereassociated wlth increased rates of mortal i ty of matureCnazzoz tnea v i zg in i cc bu t he conc luded tha t V )bn io sp . were o fmarginal s igni- f icance as pr imary pathogens in adul t oysters.

Bo th l / . angu i l l azum and l / . a lg ino l y t i cu4 have been repo r ted tobe pathogenic for mature C. gigaz (Paci f ic oyster ) butmortal i ty was direct fy related to water temperature, moltal- i t i -es being greater when oysters were stressed at hj-gh (2o"C)tempera te wa te r t empera tu res (Gr i schkowsky and L i s ton , I g74 ) .

The rel-at ionship between stress and bacter iaf disease ofmar ine an ima fs i s we l -1 recogn ized (Wedermeyer e t aL . , 1976) .Stressors sucfr as temperature, crowding, compeETtI6n fornutr lents, exhaust ion and spawning aI l predispose toinfect lous disease. Temperature is regarded as the mostimportant stressor af fect ing the balance between the organismand i ts environment (Roberts, 1"978). Temperature has ef fectson both the host and on infect ious agents (Kinne, 1980). Eachspecies of marine animal has i ts normal temperature range andat the extremes of the range disease is more f- ikely to occur(Rober t s , I 978 ) , t he suscep t i b j - l i t y t o d i sease be ing re la tedin part to the animals ' defence mechanisms. In oysters lowtemperatures depress phagocyt ic haemocyte numbers and act iv i t \ /

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l ^ f ^ n \ / o f f i c i e n . - r ; T h o n a c r r l f q n l ^ a Y n a r i m n n l - q

n c n ' - r , n m a r l i n r : l . r r l : L e L l r e c o n d i t i o n i n d o x e h . l , d a . l f h a l - o r r c . t e r Sx . . - . , . - , " " "a t 1 9 " w e r e i n " p o o r e r c o n d j . t i o n " t h a n t h o s e a t 2 9 " . T h e yw e r e L h e r e f o r e o b s s i h l \ / ' n o r e s u s r : e n l . i i r I e t o e n v i r o n m e n t a ls t r e s s a n d d i s e a s e a t I o w e r t e m p e r a t u r e s .

T h e m a j o r s o u r c e o f i n f e c t i o n c s f V i b ' z i o s p , i s t h e w a t e rc o l u m n . O u t b r e a k s o f w i b r i o s i s o c c u r a s t h e b a c t e r i a l l o a c i o fL h e w a L e n c o l . u m n . i n c r e a s e s , u s u a l l y i n c o n j u n c l - i o n w l L hi n r - r c a q i n . r r ^ r . a I a . l - c n n . . r F f r r r c . ^ i n . - i . l p n i W i t h - L n C t " e a S e d

plankton counts (Kaneko and Colwel1 , 7973) . The major sourcea f I - ' r n t a n i : f n n n a : n l ^ \ / < l a . a i n F h a e r r e t 6 n , r n d a n i n r r o < 1 - i r r -

t ion is bel ieved to be the water of the t ransporb tanks oncarr ier vessels , I t has been demonstrated that the nr-mbers ofVibr io in the sur face water of the tanks r ise dur ing t ranspor t -at ion. In the loaded tanks only the sur face waters could bes a m p l e d . ] ' l r e w a t e r e n t e r s t h e t o p o f t h e L a n k s a f t e r e n t e r j n g1 - h F ' . / p q q F l l - h r . ) r r . - r h F \ . p n f r r n i q r r q j - e m T h F . n o f . \ n a f h c r e i q m . r F e

water f low as the vessel moves and th is accounts for thev a r i a t i o n i n b a c t e r i a l n u m b e r s d u r i n g t r a n s p o r L a t . i o n . T h er e s u l t s a r e n o t i n d i c a t i v e o f t h e l e v e l - o f b a c t e r l a d e e p i nthe tanks as the sur face water is the most recent ly addedwater . The bacter ia l numbers remain ing in the tank af terunloadlng are probably a more accurate ref lect ion of theb a c t e r j a l r i u m b e r s i n w o t e r s u r r o u n d - i n g o y s t e r s d e e p i n t h etanks and i t ls apparent f rom these values that there has beenmassive increase in bacter ia l numbers dur ino t rarrsoor t .

T h e r e s e r v o i r f o r b a c t e r i a i s m o s t l i k e 1 y t o b e t h e s u r f a c eand gut of the oysters. F i -sh faeces have been shown to be as o u r c e o f l n f e c t i o n a n d a m e d i u m f o r b a c t e r i a l p r o l . i f e r a t i o n(Ruby dhd l rbr in ,1979) , hence the acc l r rnulat ion of excreta andother orEanic mater ia l in lhe tanks would prov ide a f avourabl .eenvi ronment for bacter ia l qror^Jth.

T L F n f h p r ^ m , r i o r < r , r r r c e O F i n f e c t i o n i S I i k c l v t . ] h c i h e w a t e l "s u r r o u n d i n g o y s t e r s o n r a f t s a t t h e l e a s e s i t e s . P r e l i m i n a r y(unpubl - ished) observat ions i r rd icate that the fevel o f bacterraa r o r r n d , r \ / q f e r c i o n r a F l - c i q m r r r - h . r r c a f a ^ t h a n l - h a t o f s e a w a t e r .aura\ r f nnrr r -ha F^f+q p- l a . -ement of ovsters in Lhis contam- in-a t e d e n v l r c n m e n l a f t e r b h e y h a v e b e e n s u b j e c t e d t o a h i g hb a c t e r i a l - l o a d d u r i n g t r a n s p o r t i s I i k e l y t o p o t e l ) t i - a t e t h einfect ion. There is some evidence to suppor t th ls concept inb h a t m o r t a l i t y w a s s h o w n , i n f i e l d e x p e r i m e n t s , t o b e r e l a t e dL o p o s i t i o n o f o y s t e r s o n t h e r a f t a n d w h e t h e r o y s L e r s w e r en l a c a r l n n n : f f c I n n n - l i n A M ^ ' ^ t . l i F \ r h ! : c I . \ $ ' a a f i n

oysters p laced on the outs lde of the raf t and i t was lower ono y s t e r s p l a c e d o n I o n g - l i n e s c o m p a r e d w i t h t h o s e p l a c e don raf ts . Water c i rcu lat ion around oysters on the per ipheryof raf ts and on long- I ines woul-d be greater than in the centren f r ^ e f f q t h p n a l - \ \ / . l i l , , f . i - - r a - ^ f 6 , i - r h , r - b e r s i n t h e s e s i t e s .

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t-d i s e a s e l n m a t u r e C . g i g o . z e x p e r i m e n t a l l y ( C o 1 w e 1 1 a n d S p a r k s ,1 9 6 7 ) a n d i L w a s s u g g e s t e d b h a t t h i s s p e c i e s i s a 1 i k e 1 ysecondary bacter ia l invader in oysters a l ready weakened bys o m e o t h e r d i s e a s e o r e n v i r o n m e n t a l s t r e s s . B r o w n ( 1 9 7 3 )showed that members of the genus Pseudomonas produced d iseasei n C . v i z g ) n i c a l - a r v a e u n d e r t h e s a m e c i r c u m s t a n c e s t h a tV i b n i o s p . p r o d u c e d d i s e a s e .

7 , p u l n e f o c i e n a i n a s n o t p r e v i o u s l y b e e n i m p l i c a t e d a s a c a u s eo f m o l l - u s c a n d i s e a s e . T h i s o r g a n i s m p r o d u c e s d i s e a s e i nm a t u r e ? , m a x i m a e x p e r i m e n t a l l y t h a t i s i n d i s t i n g u l s h a b f e f r o mthat due Lo Vibz io sp. WLrether natura l in fect ion wi th th isqner : i eq or other snecies of Pseudomonads occurs is not knownbut as th is genus is commonly iso lated f rom mar ine substrates,i n f e c t i o n m u s t b e c o n s i d e r e d a p o s s l b i f i t y .

V i SUI'IIVIARY AND CONCLUS ] ONS

( i ) D u r l n g t h e t h r e e y e a r p e r i o d ( J u l y 1 9 8 o - J u n e 1 9 8 3 )o f t h l s i n v e s t i g a t i o n , t h e m a j o r i t y o f p e a r l o y s b e r

. deaths in the cufture industry occurred after t rans-n ^ r + a l - i n n n f . \ \ r < f p r . c l - r \ r + h F I A r d p q + n n n n a n r r f n a m

Lhe f i sh ing g rounds t , o t he i r l ease s i t e a t Ku r - i Bay .Incidental outbreaks of oyster mortal i ty on otherlease s i t es were obse rved i n l ess de ta i l and wereno t necessa r i l v assoc ia ted w i th the fac to rs desc r ibedb e f o w .

( i i ) M o r t a l i t y i n t h e f i e l d w a s r e l a t e d t o t r a n s p o r t .n \ r q { - F r . c F h : l - $ ' a F A c , , l - r i a r . l - p r l ' i . n f h a q a m a n n n r . a r l r r n p q

fo l lowi-ng removal- f rom the natura l beds but werethen not t ranspor ted suf fered no morta l i ty .

( i i l ) E x a m i n a t i o n o f i n d u s t r y r e c o r d s s h o w e d t h a t m o s tm6rf ar i f r r nr - r - r .11-1 gd wi th in seven weeks r :non arn i wala t t h e I e a s e s i t e . T h e i n c i d e n c e o f m o r t a l i - t y v a r i e dw l t h i n a n d b e t w e e n n e a p t i d e f i s h i n g p e r i o d s b u tthe greatest inc idence occurred ln the winter .

( i v ) A m u l t i p l e r e g r e s s i o n a n a l y s l s o f w a t e r q u a l i t y p a r a -meters wi th lndustry records has shown that waterl e m p e r a t u r e i s t h e e n v i r o n m e n t a l f a c t o r m o s t c l o s e l ya s s o c i a t e d w l t h m o r t a f i t y , t h e g r e a t e s t i n c i d e n c eoccurr ing when water temperatures are at a min imum.

(v) A number of f ie ld exper iments were under taken tos t u d y t h e e f f e c t s o f v a r i o u s t r e a t m e n t s o n p e a r ln r r q t e r m c r r t : l i t v - T h e r e s l l l t . s w e r e a s f o l l o w S :

(a ) The pe rcen tage mor ta l i t y was g rea tes t i n t hesmal ler s ized oysters but the reason for th isi s no t known . C lean ing the oys te rs o f f ou l i ngo rgan lsms d id no t resu l t i n a s lgn i f i can t l yd i f f e ren t mor ta l i t y ra te f rom oys te rs l e f tuncleaned. The examinat ion of the ef fect ofvar ious depths at which oysters are malntainedunder raf ts af ter t ransport was inconclusive.

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(v i i i ) I t has been demonstrated that there is a rapid r isein the numbers of marine Vibr ios in the water of- carr ier tanks dur ing transport despi te a f low-throughcirculat j -on of water. The major source of Vibr iosis l - ikely to the gut of oysters and the major substratefor bacter ia in the water is most probably faecesand other organic matter. I t is therefore postulatedthat oysters, weakened by exposure to low annualambient temperatures, come into contact wi th highconcentrat ions of bacter la and that bacter ial invasionoccurs j -n many oysters dur ing transportat ion. Thisis probably potent iated when oysters are concentratedbe low ra f t s a t t he l ease s i t e .

( ix) In summary i t appears that the major predisposlngfac to rs assoc ia ted w i th h iqh mor ta l i t i es i n pea r l sPty Ltd 's operat ion at Kur i Bay are cold watertemperatures, concentrat lon of oysters dur ing trans-portat ion and inadequate water c irculat ion in carr iertanks. A major consequence of th is is infect ionof oysters with marine Vibr ios. I t must not beconcluded however, that these are the on]y infect iousagents involved or that other factors, as yet unldent i -f i ed , a re no t i nvo l ved .

I t is recommended to the industry that t ransportat ionof oysters be minimized dur ing the winter mon.bhs,that the number of oysters carr ied per t r ip be keptto the Lowest that is economical ly feasible and thattank hygiene and water c lrcul-at ion be opt imized.

( x )

VI I ACKNOl^lLEDGEi{ENTSThis study was financed by a grant from the Fishing Ind.ustryResearch Trust Account. The autlrors wish to ackno;Iedoethe extensive techrnical assistance of Mr M. Mannion anIh i s p redecesso r , Mr C . Harwood . The F i she r ies O f f i ce r i nBroome, Mr John Looby, acted abl-y as a l iasion between thepearl cul ture industry and the research team. Whi le excel lentco-operat lon was given by the whole industry, the personnelof Pearfs Pty Ltd shoul_d be accorded special recognit ionfor their many contr ibut ions towards the successful compLet lonof thj-s project. Further thanks for assistance are expressedto staf f of the western Austrar ian Marine Research Laborator ies.especial ly to Mr N. Caput i for preparat ion of the computerprogrammes used in data analysis and Dr D.A. Hancock forhis construct ive cr i t ic ism of th is reDorr.

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''C'f 'raMoC "I'u 'Ja+xeg "-t'c 'pTaTJueg "5'N '^a^TV

sllNlulilu

D i s a l v o , L . H . , B l e c k a ' J . , a n d z e l a ] , R ' ( 1 9 7 8 ) ' V i b n i o- - - " -

i i g " i l l a n u m a n d i a r v a l m o r t a f l t y i n a c a l l f o r n i a c o a s t a l

s h e f l - f i s h h a t c h e r y . A p p l ' € n v i n o n ' h i c z o b i o l ' 3 5 ' 2 1 9 - 2 2 1 '

E l s t o n , R . , E ] 1 i o t , E . L ' , a n d C o l w e l L ' R ' R ' ( L 9 4 2 ) '- - - - - C o n c n i o l i n i n f e c t i o n a n d s u r f a c e c o a t i n g v i b r i o : s h e l - f

f r a g i l i t y , g r o w t h d e p r e s s i o n a n d . m o r t a L i t i e s i n c u f t u r e d

o y . i " " = a n a - c f a * s , C ' z a z z o t t n ^ e a - v . i n g i n ) c a ' 0 z t n e ' a e ' d u 1 i " t

a n d n Q , l L c Q , n a / L l a m e / L c e - n a ' z i o ' t ' F i z h ' D i z ' 5 , 2 6 5 - I a 1 '

E l s t o n , R . , a n d L e i b o v i t z , L ' ( 1 9 8 0 ) ' P a t h o g e n e s i s o f

exper imenta l v ibr ios is ln larva l Amer ican oysters '

C z a z . t o z t n e , a v t n g l n i c a . C a n ' ! ' F l z h ' A q u ' 5 c i ' 3 7 '

9 6 1 - 9 7 A .

F e n g , S . Y . ( 1 9 6 5 ) . H e a r t r a t e a n d I e u c o c y t e - c i r c u l a t i o n i n -" " " ' '

C 7 o ) o o ) t o n o ' v i n g ) n i c . a ( G m e f i n ) . B i o l ' B u L 1 ' r 2 a ' 1 9 8 - 2 1 o '

F i n n e y , D . J . ( 1 9 7 1 ) . P r o b i t A n a f y s i s ' 3 r d E d n ' ( C a m b r i d g e

Univers i tY Press: carnbr idge ) '

c r i s c h k o w s k y , R . S ' , a n d L i s t o n , J ' ( r 9 7 4 ) ' B a c t e r i a l

pathogln ic i ty in t aboratory- induced morta l i ty of the.

P a c i f i c o y s t e r l C z a z ' : " z t z e - o g i g a z T h u n b e r g ) ' P z o c ' / / a L l '

5 h e ) - 1 { i z h . A z z o c . 6 1 , 8 2 - 9 L .

H a n c o c k , D . A . ( 1 9 7 3 ) . K u r i B a y p e a r l s s o m e o f f j - n e s t i n t h e

w o r I d . A u z t . F i z h . 3 2 ( / * ) , l L - ] . 2 .

I m a i , T . ( E d . ) ( 1 9 7 7 ) . A q u a c u l t u r e i n S h a l l o w S e a s ' ( A m e r i n d

P u b ] l s h j - n g C o . P t y L t d : N e w D e l h l ) '

I v a n o v i c i , A . M . ( 1 9 8 0 ) . T L r e a d e n y l a t e e n e r g y c h a n g e i n t h e

estuar lne mol lusc , ?gzaSuz eoQ'n i r lu4 ' Laboratory s tudies

o f r e s p o n s e s t o s a l i n i t y a n d t e m p e r a t u r e ' C o n p ' B i o c h e n '

? h g z i o l . 6 6 A , 4 3 - 5 5 .

J e f f r i e s , v . E . ( 1 9 8 2 ) . T h r e e v i b r i o s t r a i n s p a t h o g e n i c t o" - - - . - i i . " " " o f C z o z z o z t z e a g i g a ' t a n d ' O z t z e ' a e ' d u l i z '

A q u a c u l t u n e 2 9 , 2 O l - 2 2 6 .

K a n e k o , T . , a n d C o l w e l 1 , R . R . ( 1 9 7 3 ) ' E c o - I o g y o f V i b z i a" - ' - p l " " h l e m o 1 9 t i c u a r n c h e s a p e a k e B a y ' ! ' B a c t e ' z i o l ' 71 1 3 ,

24- 32 .

K inne , O . (198o) . D i seases o f mar ine an ima l - s : genera l. ' . ^ - - - ;= ; ; " i = . . 7 r . o i= " . ses o f Mar ine An ima ls ' (Ed ' o ' K inne ) '

vo i . f , pp ' 13 -73 . ( John Wi ley and Sons , L td : ch i ches te r )

L a w r e n c e , D . R . , a n d S c o t t , G . I . ( 1 9 e 2 ) ' T h e d e t e r m i n a t i o n a n d

use o f cond i t i on i -ndex o f oys te rs ' €z tuan iez , 5 , 23 -27 '

L ipovsky , V .P . . , and Chew ' K .K ' ( rg72 ) ' ^Mor ta l i t y o f Pac i f i c.oy -= t " . " (Cnazzoz tnea g igoz ) : t he i n f l uence o f t empera tu re

and enr iched seawatei 6n oyster survival ' ?noc' / ' lat l '

5 h e l L { i z h . A ' t z o c . 6 2 , 7 2 - 4 2 -

-tn-

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T u b i a s h , H . S . ( 1 9 7 4 ) . S i n g l e a n d c o n t i n u o u s e x p o s u r e o f t h ea d u l t A m e r i c a n o y s t e r , C z a z z o z t n e a v i z g ) _ n i - c c , t o m a r i n ev i b r i o s . C a n . t . h i c n o b i o l . 2 A , 5 l - S - 5 1 7 .

T u b i , a s h , H . S . , C h a n l e y , p . E . a n d L e i f s o n , E . ( 1 9 6 5 ) .B a c i l l a r y n e c r . o s i s , a d i - s e a s e o f f a r v a l a n d j u v e n i l eb i v a l v e m o l l _ u s k s . l . E t i o l o g ' y a n d e p i z o o t i o l o g . y . , .B a c t e t i o / . 9 0 , 1 0 g 6 _ 1 0 4 4 .

T u b i a s h , H . S . , C o 1 w e l 1 , R . R . , a n d S a k a z a k i , R . ( t - 9 7 O ) . M a r i n ev i b r i o s a s s o c i a t e d w i t h b a c i l t a r y n e c r o s i s , a d i s e a s e o fl a r v a l a n d j u v e n i l e b i v a l v e m o l l u s k s . ! . b a c t e j l i o l . I o 3 ,2 7 2 - 2 7 3

W e d e m e y e r , c . A . , M e y e r , F . p . , S m i t h , L . ( 1 9 7 6 ) . E n v i r o n m e n t a ls t r e s s a n d f i s h d i s e a s e s . 7 n D i s e a s e s o f F i s h e s . ( E d sS . F . S n i e s z k o a n d H . R . A x e l r o d . ) B o o k 5 , p p 1 - _ 1 9 2 .( T . H . F . p u b l . I n c . : N e p t u n e c i t y , N e w J e r s e y ) .

W o l f , P . H . a n d S p r a g u e , V . ( 1 9 7 8 ) . A n u n i d e n t i f l e d p r o t i s t a r rp a r a s i t e o f t h e p e a n l o y s t e r , ? i n c t a d a m a < i m a . L nt r o p i c a l A u s t r a l i a . t , t n v e . z t e b z . p a t h c , l , 3 I , 2 6 2 _ 2 6 3 .

W r i g h t , L . D . , N e i l s o n , p . , S h o r t , A . D . , a n d G r e e n , M . O .( 1 9 8 2 ) . M o r p h o d y n a m i c s o f a m a c r o t i d a l b e a c h . h a z i n e$.eo lo gy sO , s7 -L2B .

S p a r k s , A . K . , B o s w e L l , J . L . , a n d M a c k i n , J . c . ( 1 9 5 8 ) . S t u d i e son the comparat ive ut i l izat ion of oxyqen bv l iv inq anddead oys te rs . Pzoc . / , / o . t l . She l l l i zh . -A , t zoc . 4 , 62_ rc2 .

S p r a g u e , V . ( 1 9 7 1 ) . D i s e a s e s o f o v s t e r s . A n n u . R e v ./ 4 i c n o b i o 1 . 2 5 , 2 r r - 2 3 e .

T r ipp , M .R . ( f 970 ) , De fense mechan isms o f mo l l usks .l l e t i c u l o e n d o t h e l . S o c . 7 , t 7 3 - I 8 2 .

l1.e-2. t .

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Appendix 2

H i s t o l o g i c a l f i n d i n g s i n n o r m a l o y s L e r s

1 . I n t r a n u c l e a r i n c l u s i o n b o d i e s

R : q . r ^ h i l i n n r a : n n h . \ n h i I i . i n l - r . a n r r . . l a a I . i n C I U S i O n S W e l . ec n m m o n I \ / c F c n i n + h a h a q i n h i l c a l I c . \ f f h F d i r r p c f i r r e n l a n d

t u b u l e s ( F i q . 1 3 ) a n d l e s s c o m m o n l y i n e p i t h e l i a l c e l - [ s o ft h e g u t . T h e i n c l u s i o n s t e n d e d t o b e c e n t r a l l y l o c a t e dw i t h i n n u c l e i a n d w e r e s u r r o u n d e d b y a c l e a r z o n e . T h e ys t a i n e d p u r p l e w i t h p h l o x l n e t a r t r a z i n e , f a i n t p i n k w i t hm e t h y l g r e e n p y r o n i n , s l i g h L l y b a s o p h i I l c w i t h F e u L g e nq f : i n i r l f = c * n e n r f i r r o ^ n . l p a q n o n r l - i r r a T h a

j - n c l u s i o n s m e a s u r e d 2 . 5 - 5 , O u m i n d i a m e t e r . A f f e c t e dn u c f e i w e r e 4 . 3 - 7 . 5 p m i n d i a m e t e r c o m p a r e d t ou n a f f e c t e d n u c l - e l w h i c h m e a s u r e d 2 , 5 - 3 . O u m .

U l t r a s t r u c t u r a l l y t h e i n c L u s i o n s c o n s i s t e d o f a n e l e c t r o n -dense, f ine ly granular matr ix conta ln lng " doughnut-shaped "

s u b u n i t s s u r r o u n d e d b y a n e l e c t r o n - I u c e n t h a l o ( F i g . 1 1 ) .The subunl ts measured 32.6 - 38 nm and the subuni t andh a l o w e r e 5 3 , 3 - 7 0 . 7 n m i n d i a m e t e r . O c c a s i o n a l l ysmal f inc lus ions were seen adjacent to the nuclearm e m b r a n e . T h e s u b - u n i t s r e s e m b l e v i r a l p a r t i c l e s .

7

There were no other chanoes noted wi th in af fected cef ls .

P a r a s i t e s

S e c L i o n s o f m e t a z o a n p a r a s i t e s s u r r o u n d e d b y p h a g o c y t i cc e I I s w e r e s e e n i n t h e c o n n e c t i v e t i s s u e s o f t h e v i s c e r a lmass ln a number of oysters. An apj -complexan protozoanp a r a s i t e w a s c o m m o n l y s e e n i n t h e a u r i c l e ( F i g . 1 5 ) . T h i sp a r a s i t e c o n s i s t e d o f v e r m i f o r m c e f l s 2 . 5 - 3 . O u md . i a m e t e r , e m b e d d e d i n a n e o s i n o p h i l l c m a t r i x 2 5 - 5 5 p md i a m e t e r ( P e r k i n s , p e r s , c o m m . ) . O n e o r s e v e r a 1 n u c l e ii r a n F 6 c 6 h + i - 6 - ^ r - v e r m i f o r m c e l l . T L r e c e l l s c o i l e d

around each other in the mass. Lesions were not seen ina s s o c i a t i o n w i t h t h e s e D a r a s i t e s .

L a r g e c i f i a t e d p r o t o z o a w e r e c o m m o n i n t h e m i d g u t , T h ec l l i a t e s a t t a c h e d a t t h e i r a n t e r i o r e n d t o t h e m i c r o v i l l ia n d c i l i a o f g u t e p i t h e l i a l c e l l s ( F i g . 1 6 ) . T h e ym e a s u r e d 1 0 - 1 1 x 3 0 - 4 1 } l m . T h e s e o r g a n i s m s w e r enot associated wj- th d isease but there was an apparenti n c r e a s e i n t h e i r n u m b e r l n s o m e d i s e a s e d o v s t e r s .

Resldual bodies ln d j .gest ive g land epi the l ium.

In both normal and d iseased oysters round, brownlsh,c l i r r h t l r r r c f y ' a r - l - i I o b o d i e s h a v e b e e n S e e n i n f h c n n i r - : lu r r L u P r u q r

cy top lasm o f t he d iges t i ve ce l I s o f t he d iges t i ve tubu ]es(F ig , 17 ) . These s t ruc tu res were no t apparen t i n t hedarker basiphi l cel ls of the tubules. The round bodieswere present in al l oysters but the number var iedcons ide rab l y w i th in no rma l - and d i seased oys te rs , Thebod ies measured 18 L 1 .2 um, They fa i l ed to s ta in w i th

-LV-

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Tab Ie 1 Phys ica l and b iochern i ca Ir n o r b i d o y s t e r s .

c h a r a c t e r s o f V i b z i o s p . i s o l a t e d f r o m

K B 9 / 8 i - K B 7 1 8 2 ' V. a lg ino lv t i cus

Gram negat iveFermenta t iveurowtn on )Kt1u

r o d

bagar

+

P O O r+

++

ye r .Low

+

+

+

+

+++

++

+++

++

+++

++

poor

++

y" t iot

++

- +

+

++

+

++++

++

P O o r+

+

y e ! r o l r

:

+

l

.

+

M o r i l eGrowth i n 07 . NaCl

Growth on TCBS agarSwarming on f resh agarONPGArg in iueL y s i n e0rn i th ineCi t ra te ( S immons )H ^ SU i e a s bTryp !ophanelndo leAceto inG e l a t i n u t i l i z a t i o n^ ^ i l C - ^ -

5 r u L w 5 c

Manni to ll n o s i t o lS o r b i t o lRhamnoseS u c r o s eM e l i b i o s eArnygdal inArab inoseLacEoseX y l o s eM a l t o s eL r e n a l o s eC e l l o b i o s e

NO.Ox idaseC a t a l a s e

A n t i b i o t i c s e n s i t i v i t y :o /L29ch lo ramphen ico lc a r b i n i c l l l i nl ^ + - ^ ^ . . ^ I : - ^L E L i a u l s r r r r e

t r ime thop r im/ su lpha s oxaz o I ec t r a n t ^ h w . i n

ampic i 11 inc o n h a l n r i r l i n

+

+

+++t

a ) Ident i f ied as Vibnio hanveyi by Dr R. SakazakiN a t i o n a l I n s t i t u t e o f H e a l t h , T o k y o , J a p a n .

b ) S R B C - s h e e p r e d b l o o d c e l l a g a r .

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TABLE 7'. Number of survj-ving pearl- oysters after a total of 144 dayssUspension.

MONTH OF

TRANSFE R

SMALL

cv* KB"

STZE

MEDIUM

CV KB

LARGE

CV KB

TEMP

DfFFERENCE DAYS ONoc LoNGLTNE

JULY

AUGUST

SE PTEMBE R

OC TOBER

1 . 2

2 . 5

1 . 6

L . 1

o-

1 Z

7 0

9 9

- 1 6 -

3 L 2 A

2 7 3 6

100 60

- 1 5 -

2 8 4 1

5 4 5 0

1 0 0 6 3

- 4 1 -

5 0 8 5

8 1 8 0

1 0 0 8 0

b

Pearl oysters t ransferred from the Cape Vi l laret long-I ine

Pearl oysters t ransferred from the Kuri Bay long- l ine

Pearl- oysters pLaced directly on the Kuri Bay raft

- 5 4 -

o!

!

+J

q, lr

{JF

OB

rn 0)

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9. Mean and standard deviat ion (SD) of Vibzio sp./ml ofaurface \^rater and the rat io of Vib+is sp. in water ofcarr ier tanks to that in seawater (TANK:SW) .

Morning

Mean + SD _ _-b' l anK :5W Mean + SD . IANK: 5W

ro"!L'l9

960

3 6 8 +

4 6 +

3 . 4 +

5 9 . 4 +

L 8 2 . 5 +

2 9 . 3 +

8 5 . 0 +

- ̂ - ^ c .r v 5 . u +

t5e.d;6 1 2 3 . 0 +

n o I ?

- 22-9

- L Z J . L

o:,

1 A E O

64.r

L25.4

2 2 I . 2

2 5 8 1 . 8

r . 1

7 9 . 6

7 r . 7

3 0 , 3

9 1 7 . A

L 7 5 7 . 9

Transport vessel A July 1982. FuIl loatl*.

z 3 + 5 . )

5oo ; Le2o ? a r ' l I I

L 725e+ 357

20 r3d+ 6 so4

Transport vessel B July 1982. Half-load.

o . 6

LA .2 r0 .4

b . J + 5 . 2

4 5 3 . e ; 2 3 7 . L

674 + 81

7 6 8 3 + 1 0 0 9

363 + 10?

8 7 . 7 3 2 . O 4 0 8 . 6 + 6 7 . 9

L 9 . 7 5 . r 1 7 2 . 5 + 4 A . 4

zL.g L4.g 5 573.3e! 232.9

l0 020.c i ' + 2 L20.O

Transport vessel B April, 1983. Half-load.

I

I

2

?

4

9 2 t . 7

1 . 4

> 1 2 ) 5 . t

6 q

6 7 . 4

* 3 . 22 1 9 3 " 0 ! 3 0 2 L 9 . 2

1 9 8 . 0 + 4 4 L . 7

I O 1 2 . O - + 1 1 6 8 . 9f-

t9 093 - C',/ + 7 196 L67 .5

b

T

Refers to full or hal-f load of oysters.

Ratio of nunber of bacteria in tankwater to seawater-

Before tanks were loaded with sheu.

Not Done.

After tanks were unloaded at lease.

Botton sludge after tanks PumPed out.

After tanks were Partially loaded with shel1.

- 5 6 -

B

-

7

B

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T A B L E 1 1 . M o r b i d i r y a n d r n o r t a l i t y d a t a f o r o y s t e r s h e l d a t 1 9 0 a n d 2 9 o , a

l n f e c t e d w i t h V i b r i o s p . C o n t r o l

2 g o 1 9 0

M o r b i d i t y 3 2 / 4 0

M o r r a I i r y t 7 / 4 0

1 6 / 3 8 ( 4 ^ 7 % )

4 / 3 8 ( 1 0 . 5 % )

( 80 . o%)

( 4 2 . 5 % )

75 /34 (44 .1%) 4 /33 (12 . r%)

8 /34 (23 .5%) o /33 (o7 " )

c Number a f fec ted /number tes ted

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o."r6.-ldr{ddd-l{JOO{JOOOOOd{rl{lJH{J{r]r$id.|.rd{JrdddlJSCJdOCOO()Colrolrot{tll'loO.tJO]JUr.rlJ{rOc .Qr I | | |.r.l-HHHHHIo]AAAAoooooa t{ar{<lroli|o

b l e 1 3 . E f f e c t o f c u l t u r e f i l t r a t e s o n. ao y s r e r s

D i s e a s e d

4 / s ( 8 0 )

3 / 4 ( 1 5 )

o / 5 ( 0 )

ND

ND

ND

ND

Treated

D i s e a s e d

3 / 5 ( 6 0 )

4 / 4 ( 1 0 0 )

5 / 5 ( 1 o o )

5 / 5 ( 1 0 0 )

5 / 5 ( 1 o o )

4 / 5 ( 8 0 )

4 / 5 ( 8 0 )

3 / s ( 6 0 )

4 / 4(100)

s /5 ( 100 )

5 / s ( 1 0 0 )

5 /5 ( 100 )

2 / 5 (4o )

o / 5 ( o )

Cont ro l

Group

1

2

3

4

5

6

1

Dead

2 / 5 (40 )

3 / 4 ( 7 5 )

o / 5 ( 0 )

ND

ND

ND

ND

Dead

Groups 1-5 inoculat6d wt th V.haneyi .Groups 6 and 7 inoculated wi th P.putrefac iens.

a Number a f fecred /number tes ted (7 . )

ND = no t done

Tab le 14 , Mean- SD fo r Cond i t ion Index

Expe r irnent Temperature

1 g o

Treated

1 1 . 8 1 1 . 2

1 1 . 0 1 0 . 9

n . g ! 2 . 4

^ ^ o

1

.'

1 1

C o n t r o l

L a . J - 2 . 0

9 . 7 1 0 . 8

8 . 2 = t . 3

C o n t r o l

8 . 7 : o . 5

9 . 9 1 . 0 . 8

1 0 . 9 : 1 . 4

T r e a t e d

1d. 511 . 5

1o. o1o. 7

10 . o1o. 7

- 5 0 -

-T9-

(r) pup s uapuic (e) .arerd-s i;;"T:ili:trt::-"i::i lri"T"::l:g i(eg '1rny (€) 'uoidups (Z) 'rBS orIH (I) :slrolIoJ se ere (g

tasul) dpfl Irn) fE sa11s i(pn1g 'txal aqf ur o3 ParJa]a.r sea.rEtu11:ead u1eu: €ulnoqs pIIe:fsnv fsalqfroN ;o urer8erp uoltpf,o'I'1 a.rn81g

a

Figure | . T ime ser ies ofsampled at t .he

industry regords of four waterKur i Bay C Place between l {arch

qual i ty parameters1 9 7 7 a n d A p r i l 1 9 8 2 .

- 6 2 -

- Lv -

'Z86T TTrdv puE LL6I Arenu€I' uaaAlaq.{"g Trnx rbJ (s) €fn?P.radrual rtt a6Ere € puc (v) ITeJuTP: rtlol

to sproce: dTqxuou s r I6btoroalaH fo luaulredac J6 seTres 3urTJ, 'E a:n6TJ

z86t t86t 086r6/6t8/6rLL6L

OU

'r

|uL

o

oo'U

Eo(,L

oA 1978 1979 1 9 8 1

T i m e s e r i e s o f i n d u s t r y r e c o r d s o f p e r c e n t a g ep e a r l o y s t e r s f o r i n d i v i d u a l g o l l e c t i o n t r i p ss e a s o n s b e t w e e n 1 9 7 8 a n d 1 9 8 2 .

mor ta I i t y o ffo r the pear l ing

1977

F i g u r e 4 .

- 6 4 -

'9ZI -

aAZ ) so:J (lI'Z

in) sor gp'a r /^)

pur? l!5I,=/\,) ,tz's aldrrrPs '16.0

!rt T.n f^Z) sot 61.0' 1^) r;oj 6c'[ + /.\

l' .96.0 = I (rlTf.- /a7\ u1s Co.C -

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- f^:) rrrs Co'O -

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Ja^o sllln()19 alTs alll pue (e5r rJn)i ?P sa:nfP:aCtlc:l .rtl?A eas ai)PJlnS

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