魚病研究 Fish Pathology, 46 (3), 87–90, 2011. 9 © 2011 The Japanese Society of Fish Pathology Blood Fluke Infection of Cage 2 to 10 months (average of 6 months) fed with highly fat content fish, mainly chub mackerel Scomber japonicus, R eared Atlantic Bluefin Tuna A tla n tic mackerel S c o mber scombru s , European pil- Thunnus thynnus in chard C lupea pilchartus and round sardinella S a rd in e lla aurita. After this period, tunas are sacrificed in the Wes t Mediterranean floating cages a n d immediately commercialized fresh or frozen. R ocio Ruiz de Ybañez1, José Peñalver2, Among pathological problems reported in reared C arlos Martínez-Carrasco1, Laura del Río1, tuna, a blood fluke Cardicola forsteri (Digenea: 2 1 Aporocotylidae), has been pointed out as a significant Emilio María Dolores , Eduardo Berriatua risk of tuna health1). Initially identified in the Australian 1 and Pilar Muñoz * population of farmed southern BFT Thunnus maccoyii 2), this blood fluke was later reported in Atlantic BFT3–5), 1 Animal Health Department, University of Murcia, being the only one aporocotylid repo rte d s o fa r in th is Murcia 30100, Spain 2 species. Aporocotylids are parasites of marine and Livestock and Fishery Departmen t, Murcia 6) fre s h wate r fis h . Most species are located in the R egional Gov e rn men t, Murcia 30201, heart, bulbus arteriosus, ventral aorta or branchial ves- S p a in sels, although the cephalic or dorsal vessels are not uncommon habitats7). Once established, adult flukes (Recieved November 24, 2010) lay eggs whic h tra v e l to th e g ills where they lodge. The eggs hatch there and break out of the gill as free liv- in g mira c id ia 6). These miracidia infect an intermediate ABSTRACT—Infection of a blood fluke, possibly C ardicola host into which they penetrate to undergo asexual s p . (Digenea: Aporocotylidae), in reared Atlantic b lu e fin reproduction. Bivalves and polychaetes have been tuna Thunnus thynnus was investigated. Parasitological reported to be intermediate hosts for some marin e analyses included visual examination of the heart for the 6,8) aporocotylids . C ercariae emerg e fro m the intermed i- presence of adult fluke and stereomicroscopic and histopa- ate host and actively search for the definitive host, a thological analyses of the gill to assess the presence of fish, penetrate the skin of the host and juvenile flukes lesions caused by parasite eggs. No adult flukes were a tte mpt to reach the circulatory system in whic h they found in the hearts. Some of the gills exhibited small w h ite 6) undergo a migration to the final site where they matu re . to y e llo w fo c i in v o lv in g s in g le fila men ts . B lo o d flu ke eggs Blood flukes are suspected cause of mor ta litie s in a w ere found in gill tissue sections of 29.6% o f s a mpled tuna. number of farmed fish s p e c ie s , fo r e x a mple a mberjack A s lig h t in fla mmatory response was observed around mos t 9) S e rio la d u merili in S p a in and tiger puffer Takifugu of these eggs, while occasionally individual eggs were 10) rubripes in Japan . These mortalities are generally encapsulated by a granulomatous reaction. D espite the considered to be due to the eggs of the blood flukes, absence of remarkable pathological effects in the infected w h ic h b lo c k c a p illa rie s in g ill la mellae and other organs. tuna, blood flukes combined with other agents may cause 11 ) Although Munday and H allegraeff briefly mentioned ‘a major problem. moderately severe, multifo c a l, g ra n u lo matous myocardi- tis’ associated with the presence of eggs (presumably of Key word s : Thunnus thynnus, Atlantic bluefin tuna, C . fo rs te ri) during a mortality of southern BFT in 1996, Ca rd ic o la , blood fluke, Aporocotylidae more intensive histopathological investigations of the heart, gills and other organs of ranched southern BFT have not yet established a clear and consistent link The Atlantic bluefin tuna (BFT) (Thunnus thynnus) between the burden of eggs of C . fo rs te ri a nd mor ta lity is a fish e x p lo ite d b y c o mmercial fishing during centu- (Valdenegro and Now ak, unpublished data). R ecently, ries in the entire Mediterranean area. Due to the Hayward et al.12) associated high number of blood increasing deman d fo r th is fish b y th e s a s h imi-sushi flukes with increased lysozyme and decreased haemo- market, a “capture-based” aquaculture industry has globin levels in southern BFT. According to these been developed in the Mediterranean Sea over the last authors blood flukes might also be associated with the decade. This activity involves the capture of adult indi- onset of elevated mor ta litie s . viduals, during the months of May-June, w hich enter the Cardicola forsteri infection is present in wild Mediterranean Sea for reproduction. After capture, A tla n tic 13) and southern14) B F T a t low leve ls, a n d it is fish are introduced in floating cages where they stay for exacerbated during the rearing cycle in confined conditions10,15). In fa c t, A ik e n et al.16) suggest that the * Corresponding author infection in southern BFT occurs in the farming zone. E-mail: p ila r [email protected] C . fo rs te ri intensities and prevalences are very low in 88R. Ruiz de Ybañez, J. Peñalver, C. Martín e z -C a rra s c o , L . d e l R ío , E . María Dolores, E. Berriatua and P. Muñoz th e wild 14) and at transfer17) while an antibody Table 2. Es timate s fro m the logistic regression models of response against C . fo rs te ri is only initiated after trans- blood fluke infection in captive bluefin tuna in Spain 14) fe r in to th e fa rming zone from th e wild . A s th e 95% C onfidence Variables Odds ratio P value impact of aporocotylids in cage reared tuna must not be intervals neglected, the objective of the present work was to Body weight (Kg) study the prevalence of blood fluke infectio n in B F T 27–40 1.0 0 reared in west Mediterranean, in the Murcian coasts. 150–300 2.61 6.91, 0.99 0.0543 F a rm C artagena Materials and Methods 1 1.0 0 Between November 2006 and December 2008, a 3 2.23 8.18, 0.61 0.2270 to ta l o f 1 0 8 A tla n tic B F T fro m s ix B F T fa rms located in 4 0.59 5.34, 0.06 0.6369 Murcia region (South East Spain) were sampled. Tw o San Pedro fa rms are located in San Pedro del Pinatar (37°49¢ 1 0 .1 6 1.42, 0.02 0 .1 0 1 6 55.46² N , 0 °3 9 ¢42.33² W) w h ile th e re maining farms a re 2 2.37 6.42, 0.87 0.0919 located in “El Gorguel” (Cartagena, 37°34¢31, 692²N, 00°52¢30, 702²W). Tuna weights in the range of 27–40 kg, 150–210 kg and 240–300 kg for 77.4%, a te ly fixe d in 10 % neutral buffered formalin . The rest 1 8 .3 % and 4.3% of the animals, respectively (Table 1). of the gill arch and the heart of each animal w e re c o l- Fish were shot underwater and then hoisted by hook. lected in individual plastic bags, transported to the labo- Gills and hearts were removed on board immediately ratory and stored at –20°C, until examined with a stereo- after tuna were slaughtered. T h e first gill arch w a s microscope (Nikon, SMZ 800) for parasite eggs and obtained from each tuna. For histology, two to four 1 pathological changes. To do this, approximate ly , cm long pieces of the gill arch were cut out and immed i- 120–150 gill filaments per arch were in d iv id u a lly observed under the stereomicroscope, placed in a large P e tri dish with saline solution. H earts were dissected Table 1. Percentage of captive bluefin tuna individuals positive and flushed with water to dislodge any adult flukes and for blood fluke eggs in Spain in 2006–07 flushes were then poured into Petri dishes and simila rly Variables N o . o f fis h % p o s itiv e examined. To carry out the histopathological study, Location the fixed pieces of gill arch (2–4 pieces per tuna) were C artagena 71 31 d e c a lc ifie d in a mix tu re o f 5 % E D T A a n d 3 % H C l fo r 2 4 San Pedro 37 27 h , fo llow e d b y 6 0 min washing in fresh water, prior to F a rm performing routine histological processing as follow s; San Pedro sample s were e mbedded in paraffin blocks, cut and 1 20 5* stained with haemato x y lin a n d e o s in (H & E ).