Descriptions and ecological notes of free swimming forms of cymothoid isopods (Crustacea: Peracarida) collected in two coastal waters of Japan

CRUSTACEAN RESEARCH, NO. 43: 1 16, 2014 Descriptions and ecological notes of free swimming forms of cymothoid isopods (Crustacea: Peracarida) collected in two coastal waters of Japan Nobuhiro Saito, Takeo Yamauchi, Hiroyuki Ariyama and Osamu Hoshino Abstract. The objective of the study is to clarify the knowledge of free swimming forms of cymothoid isopods, ectoparasites of fishes, collected in two coastal waters of Japan. The sampling was carried out in two ways, light trap samplings at night in Osaka Bay, Pacific coast of central Honshu and direct observations and collections in the daytime in the coastal water of Izu Ohshima Island, Sagami Sea. A total of 326 cymothoids was collected from both localities and they were identified as three morphotypes of mancae, unidentified early juvenile, and juveniles of three genera: Nerocila Leach, 1818, Anilocra Leach, 1818 and Mothocya Hope, 1851. Morphological characters of them were described in detail, and problems in the identification were discussed. The body shapes and the serrations on inner margins of the dactyli of pereopods are the most important characters to divide mancae. In addition, the seasonal occurrence and the swimming behavior of them are also reported. 2009). The early life history including free swimming forms of cymothoid isopods is still poorly known (Brusca, 1978; Jones et al., 2008). Nunomura (1997) described the juveniles of cymothoid isopods are often collected by plankton net survey, but did not mention the reliable collecting data. Jones et al. (2008) attempted to identify free swimming forms (as natatory stage) of cymothoids from Australia by seeking matches of partial 16S mtdna gene sequences, and the sequences revealed that five morphotypes of free swimming cymothoids are similar to described isopods. Through our studies on isopods, many free swimming forms of cymothoids were collected. The present paper deals with description of their morphological characters and a detailed study on seasonal occurrence and swimming behavior have been reported. Introduction Cymothoid isopods (Crustacea: Peracarida) are ectoparasites of marine, brackish and freshwater fishes (Brusca, 1981). Mature females and males infect in the buccal, branchial or abdominal cavity or attach on the body surface of the hosts; on the other hand, mancae, the spawned stage in the brood pouch of adult females, have been spending a planktonic period after released from the adult female to continue for early juvenile, then, late juveniles infect host fishes (e.g. Brusca, 1978). Currently, over 360 species in 43 genera of the family Cymothoidae have been recorded in the world (Bruce & Schotte, 2013), and about 50 species are known from Japan and its adjacent waters (Saito et al., 2000; Yamauchi, Materials and Methods Cymothoid isopods were collected from two sites in Japan (Fig. 1) in two different ways. In Osaka Bay, Pacific coast of central Honshu, light trap samplings were carried out at three stations: St. 1, inclined seawall near the Marine Fisheries Research Center; St. 2, inclined seawall of the Kansai International Airport; and St. 3, vertical seawall of the Kansai International Airport. The samplings were done monthly from May 2000 to April 2002. At the stations, 500 watt underwater lamp (S 1000, Nodak Co. Ltd.) was set at 10 cm under the surface and illuminated for one hour after sunset. Gathering animals including cymothoid isopods were randomly collected by three persons using two types of hand net (23 16

2 N. SAITO ET AL. Fig. 1. Map showing the collecting sites. Site A, Osaka Bay (collection by light-trap sampling at three stations); Site B, Izu-Ohshima Island (direct observation and collection by SCUBA diving, at the northeastern coast called "Aki no hama"). Arrows indicate the collecting points. cm, mesh size 0.3 mm and 30 20 cm, mesh size 2 mm) (Omi et al., 2007). In the coastal water of Izu Ohshima Island, Sagami Sea, free swimming forms of cymothoid isopods were observed by SCUBA diving in the daytime from June 2010 to April 2012. Their swimming behavior was directory observed and parts of them were collected by one person using hand net (16 13 cm, fine mesh size).

FREE-SWIMMING FORMS OF CYMOTHOIDS COLLECTED FROM JAPAN 3 Fig. 2. Seven free-swimming forms of cymothoid isopods, dorsal views. A, manca type 1, BL: 4.8 mm (KMNH IvR 500,687), Izu-Ohshima Island, 26 March, 2012; B, manca type 2, BL: 5.2 mm (KMNH IvR 500,688), Izu- Ohshima Island, 18 June, 2010; C, manca type 3, BL: 5.2 mm (KMNH IvR 500,689), Izu-Ohshima Island, 20 April, 2011; D, early juvenile, BL: 8.1 mm (KMNH IvR 500,690), Osaka Bay, 18 August, 2000; E, aegathoid stage of Nerocila sp., BL: 20.4 mm (KMNH IvR 500,691), Osaka Bay, 16 July, 2000; F, aegathoid stage of Anilocra sp., BL: 18.2 mm (KMNH IvR 500,692), Osaka Bay, 16 July, 2000; G, juvenile of Mothocya sp., BL: 4.2 mm (KMNH IvR 500,693), Osaka Bay, 16 July, 2000.

4 N. SAITO ET AL. Fig. 3. Manca type 1, BL: 4.8 mm (KMNH IvR 500,687), Izu-Ohshima Island, 26 March, 2012. A, cephalon; B, frons with antennule and antenna; C, pleotelson and uropod; D, pereopod 1; E, pereopod 6; F, pleopod 1 (marginal setae of endopod omitted); G, endopod of pleopod 2; H, exopod of pleopod 2; I, uropod. The collected isopods were fixed in 10 % neutralized formalin seawater or frozen after transportation to the laboratory, and then, preserved in 70 % ethanol. Observations of morphological characters were made under a binocular microscope (Olympus X II). Measurements and drawings were made with the aid of a drawing tube attached to a microscope (Olympus BHB Tr). Body lengths (BL) were measured from tip of head to posterior end of pleotelson on dorsal mid line. Other measurements and terminology essentially followed Bruce (1986). The developmental stages of cymothoids were distinguished in the following stages (Brusca, 1978): manca, the spawned stage in brood pouch of adult females, pereopod 7 lacking; juvenile, next stage of manca, pereopod 7 appearing. In addition, elongated juveniles of Nerocila Leach, 1818 and Anilocra Leach, 1818 called aegathoid stages (Bruce, 1987) were distinguished from other elliptical juveniles. For identification to genus, we referred to Bruce (1986; 1987),

FREE-SWIMMING FORMS OF CYMOTHOIDS COLLECTED FROM JAPAN 5 Saito & Hayase (2000), and Jones et al. (2008). The examined specimens were deposited in the Kitakyushu Museum of Natural History and Human History (KMNH IvR), Kitakyushu, Japan. Results Description of the free swimming cymothoid isopods A total of 326 specimens of cymothoids were examined. They consisted of 265 mancae and 61 juveniles. They were identified as three morphotypes of mancae and three genera of juveniles; however, 41 juveniles could not be identified to the genus level because of the absence of distinct characters. Morphological features of each manca and juvenile are as follows: Manca type 1 (Figs. 2A, 3) Material examined. 252 inds. from Osaka Bay and 5 inds. from Izu Ohshima Island. BL: 2.0 to 4.8 mm. Described material: one manca (KMNH IvR 500,687), BL: 4.8 mm, Izu Ohshima Island, 26 March, 2012, coll. O. Hoshino. Description. Body (Fig. 2A) elliptical, pereon and pleon not distinguishable, maximum width about 35 % of body length. Cephalon (Fig. 3A) semicircular, anterior margin produced rostrum. Pleotelson (Fig. 3C) semicircular, about 0.8 times as long as width; posterior margin armed with long marginal plumose setae. Antennule (Fig. 3A, B) composed of 7 articles, not reaching posterior margin of eye. Antenna (Fig. 3A, B) composed of 8 articles, reaching posterior margin of eye. Pereopods 1 to 6 (Fig. 3D E), inner margins of all dactyli armed with serrated teeth. Pleopod 1 (Fig. 3F), medial margin of peduncle with 4 coupling hooks; endopod rectangular; exopod elliptical; both rami fringed by long marginal plumose setae. Pleopod 2 (Fig. 3G H) similar to pleopod 1, endopod lacking appendix masculina. Uropod (Fig. 3I), endopod elliptical, reaching posterior margin of pleotelson; exopod narrower, subequal to endopod; lateral and medial margins of both rami fringed by long marginal plumose setae. Remarks. This morphotype is characterized by elliptical body shape and serration on perepods dactyli. Nunomura & Ikehara (1985) described Lironeca sp. 2 based on a single juvenile (BL: 7.5 mm), collected from floating seaweeds off Ishikawa Prefecture, Japan. The morphological features of Lironeca sp. 2 were in close agreement to those of the manca type 1, but distinguished by the serration on dactylus of pereopod 1 (present in the type 1 vs. absent in Lironeca sp. 2). Manca type 2 (Figs. 2B, 4) Material examined. 4 inds. from Izu Ohshima Island. BL: 2.2 to 5.5 mm. Described material: one manca (KMNH IvR 500,688), BL: 5.2 mm, Izu Ohshima Island, 18 June, 2010, coll. O. Hoshino. Description. Body (Fig. 2B) rectangular, pereon and pleon distinguishable, maximum width about 40 % of body length. Cephalon (Fig. 4A) semicircular, anterior margin produced rostrum but invisible in dorsal view. Pleotelson (Fig. 4C) semicircular, about 0.7 times as long as wide; posterior margin armed with long marginal plumose setae. Antennule (Fig. 4A, B) composed of 7 articles, reaching pereonite 1. Antenna (Fig. 4A, B) composed of 10 articles, reaching pereonite 2. Pereopods 1 to 6 (Fig. 4D, E), inner margins of all dactyli smooth; pereopods 4 to 6 robust. Pleopod 1 (Fig. 4F), medial margin of peduncle with 6 coupling hooks; endopod rectangular; exopod elliptical; both rami bordered by long marginal plumose setae. Pleopod 2 (Fig. 4G) similar to pleopod 1, but medial margin of peduncle with 5 coupling hooks and exopod oval; endopod lacking appendix masculina. Uropod (Fig. 4C), endopod ovoid, reaching posterior margin of pleotelson; exopod narrower, longer than endopod; lateral margin of exopod lacking marginal setae.

6 N. SAITO ET AL. Fig. 4. Manca type 2, BL: 5.2 mm (KMNH IvR 500,688), Izu-Ohshima Island, 18 June, 2010. A, cephalon; B, frons with antennule and antenna; C, pleotelson and uropod; D, pereopod 1; E, pereopod 6; F, pleopod 1; G, pleopod 2. Remarks. This morphotype can be separated from the other morphotype of mancae by the following features: 1) no serration on inner margins of all pereopods dactyli; 2) pereopods 4 to 6 robust; 3) rectangular body shape, and 4) distinguishable pereon and pleon. Manca type 3 (Figs. 2C, 5) Material examined. 2 inds. from Osaka Bay and 2 inds. from Izu Ohshima Island. BL: 2.9 to 5.2 mm. Described material: one manca (KMNH IvR 500,689), BL: 5.2 mm, Izu Ohshima Island, 20 April, 2011, coll. O. Hoshino. Description. Body (Fig. 2C) elliptical, pereon and pleon not distinguishable, maximum width about 30 % of body length. Cephalon (Fig. 5A), anterior margin produced rostrum but invisible in dorsal view. Pleotelson (Fig. 5C) semi ovoid, about 0.9 times as long as width; posterior margin armed with long marginal plumose setae. Antennule (Fig. 5A, B) composed of 7 articles, reaching posterior margin of eye. Antenna (Fig. 5A, B) composed of 9 articles,

FREE-SWIMMING FORMS OF CYMOTHOIDS COLLECTED FROM JAPAN 7 Fig. 5. Manca type 3, BL: 5.2 mm (KMNH IvR 500,689), Izu-Ohshima Island, 20 April, 2011. A, cephalon; B, frons with antennule and antenna; C, pleotelson and uropod; D, pereopod 1; E, pereopod 6; F, pleopod 1; G, pleopod 2. reaching beyond mid point of pereonite 1. Pereopods 1 to 6 (Fig. 5D, E), inner margins of all dactyli armed with serrated teeth. Pleopod 1 (Fig. 5F), medial margin of peduncle with 4 coupling hooks; endopod rectangular; exopod elliptical; both rami bordered by long marginal plumose setae. Pleopod 2 (Fig. 5G) similar to pleopod 1; endopod lacking appendix masculina. Uropod (Fig. 5C), endopod elliptical, reaching posterior margin of pleotelson; exopod narrower, subequal to endopod; lateral and medial margins of both rami

8 N. SAITO ET AL. Fig. 6. Early juvenile, BL: 8.1 mm (KMNH IvR 500,690), Osaka Bay, 18 August, 2000. A, cephalon; B, pleotelson and uropod; C, antennule; D, antenna; E, pereopod 1; F, pereopod 7; G, pleopod 2; H, uropod. bordered by long marginal plumose setae. Remarks. This morphotype can be distinguished easily from the manca types 1 and 2 by the narrower body shape and the serrations on the inner margins of all pereopods dactyli. Inouye (1941) described manca of Mothocya sajori Bruce, 1986 (as Irona melanosticta Schoedte & Meinert, 1884) collected from the Seto Inland Sea, and illustrated it based on a specimen (BL: 3 mm). Its narrower body shape is similar to that of the manca type 3, but this morphotype can be distinguished easily by the serration on dactyli of all pereopods (present in the manca type 3 vs. absent in the manca of M. sajori). Early juvenile (Figs. 2D, 6) Material examined. 39 inds. from Osaka Bay and 2 inds. from Izu Ohshima Island. BL: 4.9 to 12.1 mm. Described material: one juvenile (KMNH IvR 500,690), BL: 8.1 mm, Osaka Bay, 18 August, 2000, coll. H. Omi et al. Description. Body (Fig. 2D) elliptical,

FREE-SWIMMING FORMS OF CYMOTHOIDS COLLECTED FROM JAPAN 9 Fig. 7. Aegathoid stage of Nerocila sp., BL: 20.4 mm (KMNH IvR 500,691), Osaka Bay, 16 July, 2000. A, cephalon; B, frons with antennule and antenna; C, pleotelson and uropod; D, pereopod 1; E, pereopod 7; F, pleopod 1; G, pleopod 2. pereon and pleon not distinguishable, maximum width about 35 % of body length. Cephalon (Fig. 6A) semicircular, anterior margin produced rostrum. Pleotelson (Fig. 6B) semi ovoid, length subequal to width; posterior margin armed with short marginal

10 N. SAITO ET AL. plumose setae. Antennule (Fig. 6C) composed of 8 articles, not reaching posterior margin of eye. Antenna (Fig. 6D) composed of 8 articles, not reaching posterior margin of eye. Pereopods 1 to 3 (Fig. 6E), inner margins of dactyli armed with serrated teeth; those of pereopods 4 to 7 (Fig. 6F) smooth. Pleopod 2 (Fig. 6G), medial margin of peduncle with 4 coupling hooks and 4 marginal plumose setae; exopod elliptical, part of medial margin with short marginal plumose setae; endopod rectangular, naked; endopod with appendix masculina, appendix masculina not reaching posterior margin of endopod. Uropod (Fig. 6H), endopod elliptical, not reaching posterior margin of pleotelson; exopod narrower, subequal to endopod; lateral and medial margins of endopod and medial margin of exopod bordered by marginal plumose setae, lateral margin of exopod lacking. Remarks. Body shape of this juvenile is similar to manca type 1, but they can be distinguished from the other juveniles by the serration on inner margin of dactyli found on pereopods 1 to 3. Aegathoid of Nerocila sp. (Figs. 2E, 7) Material examined. 10 inds. from Osaka Bay. BL: 8.1 to 20.4 mm. Described material: one aegathoid (KMNH IvR 500,691), BL: 20.4 mm, Osaka Bay, 16 July, 2000, coll. H. Omi et al. Description. Body (Fig. 2E) elongated, lateral body margins slightly curved, pereon and pleon not distinguishable, maximum width about 30 % of body length. Cephalon (Fig. 7A) semi ovoid, anterior margin rounded and lacking rostrum; eyes large, posterior margins expanded; posterior margin of cephalon produced strongly and trilobed. Pleotelson (Fig. 7C) triangular; about 0.9 times as long as wide; posterior margin with short marginal plumose setae. Antennule (Fig. 7A, B) composed of 8 articles, extending beyond posterior margin of eye. Antenna (Fig. 7A, B) composed of 9 articles, extending beyond mid point of pereonite 1. Pereopods 1 to 7 (Fig. 7D, E) small sized; propodi with 4 or 5 remarkable robust setae on palms; inner margins of dactyli smooth. Pleopod 1 (Fig. 7F), medial margin of peduncle with 4 coupling hooks and 1 marginal plumose seta; endopod rectangular; exopod triangular; both rami lacking marginal setae. Pleopod 2 (Fig. 7G) similar to pleopod 1; endopod with appendix masculina, appendix masculina reaching posterior margin of endopod. Uropod (Fig. 7C), endopod triangular, reaching posterior margin of pleotelson; exopod narrower, longer than endopod; lateral margin of exopod lacking marginal setae (lateral margin of left uropodal exopod wounded with a notch). Remarks. Morphological characters of this aegathoid well agree with those of the previous descriptions of Nerocila presented by Bruce (1987) and Saito & Hayase (2000): 1) elongated body with slightly curved lateral margins, 2) posterior margin of cephalon produced strongly and trilobed, and 3) setation of propodal palms of pereopods. Aegathoid of Anilocra sp. (Figs. 2F, 8) Material examined. 6 inds. from Osaka Bay. BL: 13.3 to 18.2 mm. Described material: one aegathoid (KMNH IvR 500,692), BL: 18.2 mm, Osaka Bay, 16 July, 2000, coll. H. Omi et al. Description. Body (Fig. 2F) narrower, pereon and pleon not distinguishable, each lateral body margin parallel, maximum width about 20 % of body length. Cephalon (Fig. 8A) trapezoidal, anterior margin produced thick rostrum, posterior margin straight. Eyes large but not expanded on posterior margins. Pleotelson (Fig. 8C) semi ovoid, about 1.2 times as long as width; posterior margin with short marginal plumose setae. Antennule (Fig. 8A, B) composed of 8 articles, not reaching posterior margin of eye. Antenna (Fig. 8A, B) composed of 9 articles, reaching beyond pereonite 1. Pereopods 1 to 7 (Fig. 8D, E), inner margins of all dactyli smooth; pereopod 4 to

FREE-SWIMMING FORMS OF CYMOTHOIDS COLLECTED FROM JAPAN 11 Fig. 8. Aegathoid stage of Anilocra sp., BL: 18.2 mm (KMNH IvR 500,692), Osaka Bay, 16 July, 2000. A, cephalon; B, frons with antennule and antenna; C, pleotelson and uropod; D, pereopod 1; E, pereopod 7; F, pleopod 1; G, pleopod 2. 7 elongate. Pleopod 1 (Fig. 8F), medial margin of peduncle with 4 coupling hooks and 6 marginal plumose setae; exopod triangular, part of the medial margin with short marginal plumose setae; endopod rectangular, naked. Pleopod 2 (Fig. 8G) similar to pleopod 1, but 8 marginal plumose setae on medial margin of peduncle; endopod with appendix masculina not reaching posterior margin of endopod. Uropod (Fig. 8C), endopod elliptical, not reaching posterior margin of pleotelson; exopod narrower, subequal to endopod; lateral margin of exopod lacking marginal setae. Remarks. Elongated body shape and large eyes of this aegathoid is similar to Nerocila sp., but Anilocra sp. can be distinguished by the following characters: 1) narrower body with parallel lateral margins, 2) not expanded posterior margins of eyes, and 3) produced rostrum on anterior margin of cephalon. These morphological characters well agree with those of the previous descriptions of Anilocra presented by Jones et al. (2008).

12 N. SAITO ET AL. Fig. 9. Juvenile of Mothocya sp., BL: 4.2 mm (KMNH IvR 500,693), Osaka Bay, 16 July, 2000. A, cephalon; B, frons with antennule and antenna; C, pleotelson and uropod; D, pereopod 1; E, pereopod 7; F, pleopod 1; G, pleopod 2. Juvenile of Mothocya sp. (Figs. 2G, 9) Material examined. 4 inds. from Osaka Bay. BL: 2.5 to 4.2 mm. Described material: one juvenile (KMNH IvR 500,693), BL: 4.2 mm, Osaka Bay, 16 July, 2000, coll. H. Omi et al. Description. Body (Fig. 2G) elliptical, pereon and pleon distinguishable, maximum width about 45 % of body length. Cephalon (Fig. 9A) semicircular, anterior margin produced rostrum. Pleotelson (Fig. 9C) semicircular, about 0.6 times as long as width; posterior margin lacking marginal setae. Antennule (Fig. 9A, B) stout, composed of 9 articles, reaching posterior margin of eye. Antenna (Fig. 9B) composed of 8 articles, reaching posterior margin of eye. Pereopods 1 to 7 (Fig. 9D, E), inner

FREE-SWIMMING FORMS OF CYMOTHOIDS COLLECTED FROM JAPAN 13 Table 1. Occurrence of free-swimming forms of cymothoid isopods collected by light-trap samplings from Osaka Bay. 2000 2001 2002 M J J A S O N D J F M A M J J A S O N D J F M A St. 1 Manca type 1 2 1 2 19 2 4 42 10 2 28 13 1 Early juvenile 4 1 1 1 1 Aegathoid of Nerocila 1 2 Aegathoid of Anilocra 1 Juvenile of Mothocya 2 1 total individuals 0 0 4 6 1 3 19 2 0 0 0 0 0 4 1 45 11 3 28 13 1 0 0 0 St. 2 Manca type 1 1 1 5 1 1 3 35 1 Early juvenile 2 1 3 2 2 Aegathoid of Nerocila 1 total individuals 0 2 3 0 1 5 1 0 0 0 0 0 1 0 0 6 2 37 1 0 0 0 0 0 St. 3 Manca type 1 1 1 5 3 54 11 2 1 Manca type 3 2 Early juvenile 1 1 1 17 1 Aegathoid of Nerocila 2 1 1 1 1 Aegathoid of Anilocra 1 1 1 1 1 Juvenile of Mothocya 1 total individuals 0 1 2 1 4 3 2 0 1 0 0 0 1 5 3 0 71 14 4 1 0 0 0 0

14 N. SAITO ET AL. 1851 has not been described sufficiently, the characters of the present individuals, stout antennule and extended uropodal exopod, are similar to those of M. parvostis Bruce, 1986 presented by Bruce (1986). Fig. 10. Photograph of the swimming form of cymothoid isopod taken in the northeastern coast of Izu-Ohshima Island, 4 m deep, 40 cm above rocky sea bottom, on 1 February, 2012. Arrow indicates pleopods used for swimming. margins of all dactyli smooth. Pleopod 1 (Fig. 9F), medial margin of peduncle with 4 coupling hooks and 1 marginal plumose seta; endopod rectangular; exopod elliptical; both rami lacking marginal setae. Pleopod 2 (Fig. 9G) similar to pleopod 1; endopod with appendix masculina not reaching posterior margin of endopod. Uropod (Fig. 9C), endopod oval, reaching posterior margin of pleotelson; exopod elliptical, extending beyond posterior margin of pleotelson, about 1.5 times as long as endopod. Remarks. Although morphological features of juvenile of Mothocya Hope, Seasonal occurrence in Osaka Bay Seasonal occurrence of the free swimming cymothoids collected by light trap sampling is provided in Table 1. The cymothoids collected from Osaka Bay consisted of 254 mancae, 20 identified juveniles, and 39 unidentified juveniles. Although this method was not quantitative, there was a tendency of cymothoids to occur seasonally at each station. These cymothoids occurred mainly in summer to early winter (June to December), while they were almost absent in winter to late spring (January to April). Swimming behavior in the coastal water of Izu Ohshima Island Two to eight individuals of cymothoid isopods were found in a day for the fieldwork by SCUBA diving. This observation indicates that the isopods occur commonly in the area. The occurrence was mainly restricted from May to July, while they decreased from August to November. Of these, 11 mancae and 2 early juveniles were collected (Table 2). The mancae were composed of three mor- Table 2. Observed data of free-swimming forms of cymothoid isopods in the coastal water of Izu-Ohshima Island. No. Date Depth (m) Cymothoid BL (mm) 1 1 June 2010 4 Manca, type 1 3.3 2 18 June 2010 5 Manca, type 2 5.2 3 23 June 2010 4 Manca, type 2 4.0 4 18 December 2010 5 Juvenile 7.6 5 3 February 2011 10 Manca, type 3 4.9 6 20 April 2011 2 Manca, type 3 5.2 7 21 December 2011 5 Juvenile 5.6 8 18 January 2012 4 Manca, type 2 (?) 2.2 9 1 February 2012 4 Manca, type 2 (?) 5.5 10 26 May 2012 2 Manca, type 1 4.8 11 27 May 2012 2 Manca, type 1 4.6 12 28 May 2012 15 Manca, type 1 3.5 13 2 April 2012 4 Manca, type 1 4.7

FREE-SWIMMING FORMS OF CYMOTHOIDS COLLECTED FROM JAPAN 15 photypes as above mentioned. Observed live cymothoids usually kept their body horizontal when free swimming (Fig. 10), using their pleopods for swimming and movement was linear. Their swimming speed was very slow; however, they could move quickly for a distance of 50 cm to 1 m in an emergency. They were mainly observed in 2 15 m depth, but sometimes more than 20 m depth. They occurred about 50 cm above the substrates of rocks or gravel, and no individual was observed near sandy bottom regions. Discussion Most planktonic marine invertebrate larvae, including crustaceans, are supplied from reproductive adult populations (e.g. Saito et al., 2012). In the present study, free swimming forms of cymothoid isopods mainly appeared from summer to early winter in Osaka Bay, and occurred mainly from May to July in the coastal water of Izu Ohshima Island. The appearance is probably dependent on recruitments, but we have little knowledge on the reproductive cycle of Japanese cymothoids. The direct observation by SCUBA diving in the coastal water of Izu Ohshima Island, free swimming forms of cymothoids consisting of mancae and early juveniles were confirmed in the daytime. On the other hand, late juveniles (aegathoids of Nerocila sp. and Anilocra sp. and juvenile of Mothocya sp.) occurred at night in Osaka Bay. Jones et al. (2008) also collected late juveniles by light traps; therefore, we could suggest that late juveniles of cymothoids are probably swimming at night and having a strong positive phototaxis. In the present study, the isopods occurred only around rocky or gravel regions. This fact suggests that the isopods may escape from predators, because they can swim fast and hide underneath rocks or gravel. All specimens of mancae and early juveniles in this study could not be identified to the genus level, because they did not have some important taxonomic characters. Furthermore, the relationship between juvenile and adult cymothoids has not been well understood, due to little information about the shape change with growth. We realize that the body shapes and the serration on inner margins of dactyli of pereopods are the most important characters to divide these mancae into three morphotypes. Richardson (1905) reviewed 27 cymothoid species from North America, and several species were shown with their mancae and/or juveniles. Among them, Cymothoa excisa Perty, 1833 has serration on dactyli of pereopods 1 3, and Glossobius impressus (Say, 1818) (as Ceratothoa impressa) and Mothocya nana (Schiaedte & Meinert, 1884) have displayed serration on all pereopods. It should be noted that the serration of G. impressus was found only in the 2nd stage, but not in the 1st and 3rd stages. To clarify the early life history stages of Japanese cymothoids, it is necessary to describe the morphological characters of mancae (including early stages of juvenile) released from the adult females. Furthermore, the aegathoids of Nerocila sp. and Anilocra sp. in this study may include various growth stages. Hence, we suggest that it is necessary to reveal the morphological changes due to growth of cymothoid isopods in the future studies. Acknowledgments We sincerely thank to Mr. Hiroaki Omi and the other staff of the Marine Fisheries Research Center, Research Institute of Environment, Agriculture and Fisheries, Osaka Prefectural Government, for helping the light trap samplings. We are deeply grateful to Dr. B.A. Venmathi Maran (Marine Ecosystem Research Division, Korea Institute of Ocean Science & Technology) for critically reading of the early draft of this manuscript; and to two anonymous reviewers for their useful comments. Literature Cited Bruce, N. L., 1986. Revision of the isopod crustacean genus Mothocya Costa, in Hope, 1851 (Cymothoidae: Flabellifera), parasitic on marine fishes. Journal of Natural History, 20:

16 N. SAITO ET AL. 1089 1192., 1987. Australian species of Nerocila Leach, 1818, and Creniola n. gen. (Isopoda: Cymothoidae), crustacean parasites of marine fishes. Records of the Australian Museum, 39: 355 412., & Schotte, M., 2013. Cymothoidae. In: Schotte, M., Boyko, C.B., Bruce, N.L., Poore, G.C.B., Taiti, S. & Wilson, G.D.F. (eds). World Marine, Freshwater and Terrestrial Isopod Crustaceans Database. Accessed through: World Register of Marine Species at http://www.marinespecies.org/aphia. php?p=taxdetails&id=118274 on 2013 05 16. Brusca, R.C., 1978. Studies on the cymothoid fish symbionts of the eastern Pacific (Isopoda: Cymothoidae). I. Biology of Nerocila californica. Crustaceana, 34: 141 154., 1981. A monograph on the Isopoda Cymothoidae (Crustacea) of the Eastern Pacific. Zoological Journal of the Linnean Society, 73: 117 199. Inouye, M., 1941. On sexuality in Cymothoidae, Isopoda II Irona melanosticta Schoedte & Meinert parasitic in the branchial cavity of the halfbeak, Hyporhamphus sajori (Temminck & Schlegel). Journal of Science of the Hiroshima University, Series B Div. 1, 9: 219 238, 1 pl. Jones, C.M., Miller, T.L., Grutter, A.S., & Cribb, T.H., 2008. Natatory stage cymothoid isopods: description, molecular identification and evolution of attachment. International Journal for Parasitology, 38: 477 491. Nunomura, N., 1997. Isopoda. In: Chihara, M. & Murano, M. (eds.), An Illustrated Guide to Marine Plankton in Japan, Tokai University Press, Tokyo, pp. xv, 1112 1130. (In Japanese), & Ikehara, K., 1985. Some isopod crustaceans collected in the middle coastal area of the Japan Sea. Bulletin of the Toyama Science Museum, (7): 51 69. Omi, H., Ariyama, H., Kusakabe, T., & Tsujimura, H., 2007. Larvae and juvenile fishes collected by light trap sampling at an inclined sea wall of the southern coast in Osaka Bay. Bulletin of the Osaka Prefectural Fisheries Experimental Station, 17: 9 17. (In Japanese) Richardson, H., 1905. A monograph on the isopods of North America. Bulletin of the United States National Museum, 54: i liii, 1 727. Saito, N., & Hayase, Y., 2000. Note on an aegathoid stage of cymothoid isopod, Nerocila phaiopleura Bleeker, 1857 (Crustacea: Isopoda: Cymothoidae) stranded at Miho beach, Suruga Bay, middle of Japan. I. O. P. Diving News, 127: 2 6. (In Japanese with English abstract), Itani, G., & Nunomura, N., 2000. A preliminary check list of isopod crustaceans in Japan. Bulletin of the Toyama Science Museum, (23): 11 107. (In Japanese with English abstract), Itoh, H., & Furota, T., 2012. Occurrence of the planktonic decapod larvae (Crustacea) in tidal flat covering water off Obitsu River estuary, eastern coast of the Tokyo Bay, in summer. Cancer, 21: 13 18. (In Japanese with English abstract) Yamauchi, T., 2009. Deep sea cymothoid isopods (Crustacea: Isopoda: Cymothoidae) of Pacific coast of Northern Honshu, Japan. National Museum of Nature and Science Monographs, 39: 467 481. Addresses: (NS) Suido sha Co. Ltd., Ikuta 8 11 11, Tama ku, Kawasaki, Kanagawa, 214 0038 Japan; (TY) Institute of Natural and Environmental Sciences, University of Hyogo/ Museum of Nature and Human Activities, Hyogo, Yayoigaoka 6, Sanda, Hyogo, 669 1546 Japan; (HA) Marine Fisheries Research Center, Research Institute of Environment, Agriculture and Fisheries, Osaka Prefectural Government, Tanagawa, Misaki, Osaka, 599 0311 Japan; (OH) Diving Services Chap, Okatashinkai 118 2, Ohshima, Tokyo, 100 0102 Japan. E mails: (NS) nsaitoh@suidosha. co.jp; (TY) cymothoid@yahoo.co.jp; (HA) ariyamah@rinku.zaq.ne.jp; (OH) hossy@chap. jp Received: 26 August 2013. Accepted: 21 January 2014.