Use of Parasites As Biological Tags for Separating Stocks of the Starspotted Dogﬁsh Mustelus Manazo in Japan and Taiwan

Blackwell Science, LtdOxford, UK FISFisheries Science0919-92682003 Blackwell Science Asia Pty Ltd 692April 2003 626 Parasites of Mustelus manazo A Yamaguchi et al.

10.1046/j.0919-9268.2002.00626.x Original Article337342BEES SGML FISHERIES SCIENCE 2003; 69: 337–342

Use of parasites as biological tags for separating stocks of the starspotted dogﬁsh Mustelus manazo in Japan and Taiwan

Atsuko YAMAGUCHI,1* Hiroshi YOKOYAMA,2 Kazuo OGAWA2 AND Toru TANIUCHI3

1Faculty of Fisheries, Nagasaki University, Nagasaki, Nagasaki 852-8521, 2Department of Aquatic Bioscience, Graduate School of Agricultural and Life Sciences, University of Tokyo, Bunkyo, Tokyo 113-8657 and 3College of Bioresource Sciences, Nihon University, Fujisawa, Kanagawa 252-8510, Japan

ABSTRACT: The parasite fauna of Mustelus manazo from seven localities in Japan and Taiwan was assessed for the feasibility of using parasites as biological tags for the identification of different host stocks. Examination of 1038 host specimens resulted in 13 parasite species recorded: one nematode, two myxosporeans, two copepods and eight cestodes. Although relatively low overall prevalence of the first five species at different localities showed some significant differences, the samples collected in Tokyo Bay bore the lowest similarity (19.2%). These results support previous studies, which suggested the existence of a unique host stock in Tokyo Bay. Accordingly, combinations of prevalence data may provide useful information on stock identification. Cestodes probably have the greatest potential for use as biological tags. Detailed examination of 25 specimens of M. manazo from Aomori and Tokyo Bay disclosed eight species of intestinal cestodes. A canonical discriminant analysis based on these species separated the two localities clearly.

KEY WORDS: biological tag, elasmobranch, Mustelus manazo, parasite, shark, stock separation.

INTRODUCTION a continuous steep slope with similar environmental conditions. It was considered, therefore, that The starspotted dogfish Mustelus manazo (Chon- mixing of the two populations may occur. drichthyes: Triakidae) is a wide-ranging, temperate Parasites have been used successfully as biolog- and tropical, bottom-dwelling coastal shark, ical tags in numerous studies, including Pacific caught commonly by small trawlers, set net and herring,8 Greenland halibut9 and Australian long lines off Japan and Taiwan. Recent life-history sailfish10 to provide information on fish popula- studies of M. manazo from Japan and Taiwan have tions. However, few studies of elasmobranchs par- demonstrated great variations among localities, asites as biological tags have been conducted. particularly between Aomori and Taiwan popula- Parasites for such use should be chosen using the tions.1–7 In particular, Yamaguchi et al.5,6 found evi- following criteria: infections should be of relatively dence of geographic variations in maximum length long duration (years rather than months), with no and age, growth rate (including embryos), size at reproduction of parasites on or in the host. birth, length and age at sexual maturity, and repro- MacKenzie11 considered that for stock identifica- ductive cycle among populations from Aomori, tion studies, it is appropriate to use several differ- Tokyo Bay, Maizuru, Shimonoseki of Japan and ent groups of parasites, although parasites do not Taiwan. However, no geographic variations in life- necessarily have long life spans in the host. Dr history parameters occurred between Maizuru and Satyu Yamaguti’s list of the parasite fauna of Shimonoseki populations, except for a difference Japanese fishes included one myxosporean, seven in growth curves.5 Although the distance between cestodes and one nematode from M. manazo.12 these two areas is about 600 km, they are linked by However, no further studies of M. manazo parasites have been made, and there is little ecologic information on them. The purpose of the present *Corresponding author: Tel: 81-95-847-1111. study was to survey the parasite fauna of Fax: 81-95-844-3516. Email: [email protected] M. manazo from seven localities in Japan and Received 18 April 2002. Accepted 17 October 2002. Taiwan and to assess the feasibility of using para-

338 FISHERIES SCIENCE A Yamaguchi et al.

Table 1 List of parasites and infection sites in Mustelus manazo Parasite Infection site Nematoda Acanthocheilus rotundatus Stomach Cestoda Calliobothrium hayhowi Intestine Calliobothrium verticillatum Intestine Calliobothrium creeveyae Intestine Calliobothrium tylotocephalum Intestine Echinobothrium sp. Intestine Tripanorhyncha gen. sp. Intestine Phyllobothrium sp. Intestine Orygmatobothrium sp. Intestine Myxozoa Ceratomyxa sp. Gallbladder Chloromixum sp. Gallbladder Copepoda Sphyriidae gen. sp. Fin Perissopus oblongatus Nostril

The measures of the level of injection used in this study was prevalence only. Prevalence is the Fig. 1 Map of the study area, showing sampling number of M. manazo injected divided by the locations. number examined, expressed as a percentage. Prevalence was compared for significance using the Fisher’s exact probability test from contingency tables (SPSS ver. 10.0 J for Windows; SPSS Inc., sites as biological tags for the identification of dif- Chicago, IL, USA). To test for similarity in preva- ferent host stocks. lence among localities, prevalence (%) of five parasite species in each locality was analyzed by cluster analysis using the average linkage method. MATERIALS AND METHODS A canonical discriminant analysis was performed using prevalence data of cestodes to separate Specimens of M. manazo (n = 1038) were exam- stocks (Statistics Analysis HB for Windows ver. 1.0; ined from collections made from 1994 to 1996 Kyoritsu Syuppan Co., Ltd, Tokyo, Japan). from seven localities, Aomori, Choshi, Tokyo Bay, Uwajima, Maizuru, Shimonoseki and Taiwan (Fig. 1). Those from Choshi and Uwajima were RESULTS taken by small bottom trawlers at depths of 100– 150 m and 50–100 m, respectively. Collection data Parasites for the remaining localities are listed in Yamaguchi et al.5,6 The samples from Shimonoseki and Taiwan Parasite species recorded included one nematode were frozen immediately after collection, while (stomach), two myxosporeans (gallbladder), and others were kept on ice during transportation to two copepods (fin or nostril) (Table 1). Because the laboratory. identification of intestinal cestodes was difficult, External parasites from the skin, fins, oral sur- detailed examination of 25 host specimens, con- faces and nostrils were examined and identified. sisting 12 from Tokyo Bay and 13 from Aomori, Stomachs and spiral intestines were separated and were made, resulting in eight cestode species being opened longitudinally and their contents rinsed recognized (Table 1). into beakers, fixed in 10% buffered formalin and settled to remove endoparasitic helminths. Bile from the gallbladder was inspected using a micro- Prevalence of parasites scope (¥400) to determine the presence or absence of Myxozoa. All parasites were identified to the low- The seasonal and ontogenetic variations of preva- est possible taxon. lence were not found in any cases. The five parasite

Parasites of Mustelus manazo FISHERIES SCIENCE 339

Table 2 Prevalence of parasite species in Mustelus manazo from seven localities (excluding Cestoda) Acanthocheilus Sphyriidae gen. Perissopus rotundatus Ceratomvxa sp. Choromyxum sp. sp. oblongatus Prevalence Prevalence Prevalence Prevalence Prevalence Locality (%) n (%) n (%) n (%) n (%) n Aomori 12.6 230 7.6 171 0.6 171 0 194 0 194 Choshi 29.1 79 3.9 51 0 51 0 90 0 90 Tokyo Bay 40.6 266 33.0 254 0 254 3.3 213 6.6 213 Maizuru 31.9 135 13.5 81 1.2 81 1.0 93 0 93 Uwajima 13.5 37 27.7 36 0 36 0 39 0 39 Shimonoseki 16.9 148 14.7 115 8.7 94 0 97 0 97 Taiwan 16.7 186 12.5 16 6.3 16 0 26 0 26 d.f. 66666 c2 73.1 59.3 38.3 14.8 34.9 P 0.000** 0.000** 0.000** 0.022* 0.000**

n, number of fish examined. species listed in Table 2 showed significant differences in prevalence among the localities. Acan- thocheilus rotundatus was common in Tokyo Bay (40.6%), Maizuru (31.9%) and Choshi (29.1%), but uncommon in Aomori (12.6%), Uwajima (13.5%), Taiwan (16.7%) and Shimonoseki (16.9%). There were no significant differences within high or low prevalence locality groups (P > 0.05), whereas significant differences were found between them (P < 0.001). Ceratomyxa sp. was recorded from all localities, the highest prevalence being from Tokyo Bay (33.0%) and the lowest from Aomori and Choshi (7.6% and 3.9%, respectively). Chloromyxum sp. Fig. 2 Percentage similarity of parasite (excluding ces- was recorded from Aomori, Maizuru, Shimonoseki todes) prevalence in seven study areas. and Taiwan, but not from Choshi, Tokyo Bay and Uwajima (all facing the Pacific Ocean). The occurrence of two copepods on M. manazo was limited. Sphyriidae gen. sp. was recorded from Tokyo Bay and Maizuru (prevalence 3.3%, and Bay in January (n = 25) (Table 3), with significant 1.1%, respectively) and Perissopus oblongatus was differences in prevalence between the two locali- also recorded from Tokyo Bay (prevalence 6.6%). ties being found for five species. The prevalence of The greatest similarity in parasite prevalence four Calliobothrium species ranged from 69 to was found between Taiwan and Shimonoseki 100% in Aomori, but, in contrast, ranged from 17 (91.5%), with that between the two former locali- to 58% in Tokyo Bay. Other cestodes, Phylloboth- ties and Aomori also being high (73.8%). The rium sp. (100% in Tokyo Bay vs 23% in Aomori) and similarity between Choshi and Maizuru was com- Tripanorhyncha gen. sp. (58% in Aomori vs 8% in parable (73.7%), with the overall similarity of the Tokyo Bay) were included. five localities (all facing the Japan Sea, except Cho- A canonical discriminant analysis was per- shi) being 54.6%. However, parasite prevalence in formed on two category variables of eight cestode Tokyo Bay showed the least similarity with the species (0: non-parasitic, 1: parasitic) and Canoni- other localities (19.2%) (Fig. 2). cal variable 1 (CV1) plotted (Wilk’s l = 0.245, F = 6.168(8,16) P = 0.001) (Fig. 3). CV1 clearly separated the two localities, the residuals of Phyllobo- Use of cestodes as biological tags thrium sp. contributing most strongly (Table 4). Table 5 demonstrates that the sharks from Tokyo More than one species of cestodes was recorded Bay and Aomori were correctly reclassified in 100% from each host collected from Aomori and Tokyo and 84.6% of cases, respectively.

340 FISHERIES SCIENCE A Yamaguchi et al.

vs P 0.002** 0.000** 0.015* 0.030* 0.000** omori okyo Bay Bay okyo A 1 1.00 11 0.202 1 1 11 0.480 1 T + + +

Fig. 3 Canonical discriminant analysis from two category variables of eight cestode species. Tokyo Bay Tokyo DISCUSSION

+ Several parasite species were found in M. manazo from seven localities indicating their potential use as biological tags. All showed signiﬁcant differ-

++++++ ences in prevalence between at least one pair of localities. In particular, the prevalence of Cerato- + +++++++++ + + + myxa sp. between Tokyo Bay and Choshi showed a signiﬁcant difference, despite the short distance + between those two areas (ª 180 km). Accordingly, mixing of the two M. manazo populations may not occur or be incomplete, at best. The entrance of Tokyo Bay is restricted, being some 6 km wide. Out- side the Bay it is steep and deep; the environmental conditions and animal fauna thus differ within and +++++ +++++++++ + + + + + + + outside the Bay.6 Indeed, the physical factors are one of the reasons for the M. manazo population remaining within the Bay, evidenced by that intestine from Aomori and Tokyo Bay Tokyo and Aomori intestine from population’s unique life-history characteristics, particularly growth,5 embryonic development and omori 6 7 A sex ratio and feeding ecology. This is now sup- ported by the evidence of the difference in parasite prevalence.

++ The ecologic study of M. manazo by Yamaguchi ustelus manazo

M et al. indicated that mixing of Maizuru and Shi- monoseki populations may occur.6 In the present

++++ ++ + study, no signiﬁcant differences in the prevalence of four parasite species were found, but differences ++ ++ in the prevalence of the stomach-infesting Acan-

+++++++++++ +++++++++++ +++ +++++ + + + + thocheilus rotundatus were signiﬁcant. The possi- A1 A2 A3 A4 A5 A6 A7 A8 A9 A10 A11 A12 A13 T1 T2 T3 T4 T5 T6 T7 T8 T9 T10bility T11 T12 d.f. of mixing of these two populations remains high, although it may not be complete. A further consideration is that differences in the prevalence sp. of stomach parasites may simply reﬂect differences 0.01. sp. sp. in food at different localities, and not in separate P < stocks. ccurrence of cestode species in ccurrence

O The clusters of parasite prevalence were similar 7 0.05, ** to those of food types, observed differences in ticillatum

P < prevalence possibly reﬂecting variability of the ygmatobothrium ver tylotocephalum ipanorhyncha gen. sp. * estode species/Host no. estode species/Host alliobothrium hayhowi alliobothrium creeveyae able 3 ecosystem within the study areas, as well as feed- C Or C T Calliobothrium C Calliobothrium Echinobothrium Phyllobothrium Tr Parasites of Mustelus manazo FISHERIES SCIENCE 341

Table 4 Canonical functions and contribution of variables to discriminant analysis Canonical variable Canonical functions

Cestoda F0 P 1Aomori Tokyo Bay Calliobothrium hayhowi 0.153 NS -0.287 -1.88 -2.85 Calliobothrium verticillatum 0.201 NS -0.557 -17.46 -19.34 Calliobothrium creeveyae 1.318 NS -1.294 33.4 29.04 Calliobothrium tylotocephalum 0.2 NS -0.346 12.56 11.4 Echinobothrium sp. 0.079 NS 0.537 52.87 54.68 Tripanorhyncha gen. sp. 0.273 NS -0.391 15.29 13.98 Phyllobothrium sp. 7.483 * 2.203 22.27 29.7 Orygmatobothrium sp. 0.003 NS -0.064 25.65 25.43

* P < 0.05. NS, not signiﬁcant.

Table 5 Reclassiﬁcation of individual specimens to locality based on parasitological discriminant analysis Correct Number of reclassiﬁcation Locality nnpercentage (%) Aomori Tokyo Bay Aomori 13 11 84.6 11 2 Tokyo Bay 12 12 100 0 12

ing habits of M. manazo. Although food and dant in M. manazo in both Aomori and Tokyo Bay, feeding habits differ in M. manazo at different but more so in Aomori than Tokyo Bay. The overall localities,7 the major dietary components were findings for cestodes in the present study indicated crustaceans in each case. The most important prey that the Aomori and Tokyo Bay M. manazo popu- categories were crabs (Aomori, Shimonoseki and lations are separated, supporting the results of Taiwan), mantis shrimps (Tokyo Bay) and shrimps previous ecologic studies.5,6 Similar examination of (Maizuru). Fishes were a minor food category for cestodes elsewhere may provide additional evi- M. manazo at most localities, but were second in dence on stock structures especially between dietary importance at Maizuru. The difference in Shimonoseki and Maizuru, or Tokyo Bay and diets of the Maizuru and Shimonoseki M. manazo Choshi. populations may have been responsible for the difference in nematode prevalence at these two localities. Although the greatest similarity in parasite ACKNOWLEDGMENTS prevalence was found in the Shimonoseki and Taiwan populations, these two populations are We thank Dr Kazuya Nagasawa, National Research clearly separated by morphometric features and Institute of Far Seas Fisheries, Dr Toshiaki vertebral counts.2 The great similarity of parasites Kuramochi, National Science Museum, and Drs may simply reflect similar ecosystems. Janine Caira and Claire Healy, University of Con- The combination of nematode, myxosporean necticut, for their kind help with the identification and copepod prevalence provides useful informa- of parasites. Dr Janine Caira reviewed our first tion on stock identification. However, cestodes manuscript and gave helpful comments on the probably have the greatest potential as biological manuscript. tags for providing information on the stock struc- ture of M. manazo, because of their high prevalence and the occurrence of many species. Four REFERENCES species of Calliobothrium were found in 1. Yamaguchi A, Taniuchi T, Shimizu M. Age and growth of the M. manazo from two study areas and an interest- starspotted dogfish Mustelus manazo from Tokyo Bay, ing pattern of host specificity has been reported by Japan. Fish. Sci. 1996; 62: 919–922. Nasin et al. that many species of Calliobothrium 2. Yamaguchi A. Studies on stock identification of the are known only from species of the family Triak- starspotted dogfish, Mustelus manazo. PhD Thesis, The idae, with many of the parasites being specific to a University of Tokyo, Tokyo. 1997. single triakid.13 Calliobothrium species were abun- 3. Yamaguchi A, Taniuchi T, Shimizu M. Reproductive biology 342 FISHERIES SCIENCE A Yamaguchi et al.

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