Journal of the American Control Association, 20(2):134_137,2004 Copyright @ 2OO4by the American Mosquito Control Association, Inc.

A -HOLE MOSQUITO, OCHLEROTATUSBAISASI, FEEDING ON (: OXUDERCINAE) IN THE RYUKYU ISLANDS,

HARUO OKUDO,' TAKAKO TOMA,' HITOSHI SASAKI,3 YUKIKO HIGA,' MAYUMI FUJIKAWA.3 ICHIRO MIYAGI, eNo TAKAO OKAZAWA4

ABSTRACT. Ochlerotatus baisasi was observed and photographed feeding on a mudskipp er ( argentilineatus). Origin of blood meals in the stomachs of female mosquitoes collected at burrows of land and mud lobsters in the forests of Oura, Okinawa, and Komi, Iriomote (the Ryukyu Islands;, was determined by enzyme-linked immunosorbent assay to be from a source.

KEY WORDS Ochlerotatus baisasi, feeding behavior, blood meal source, mudskipper, Periophthalmus ar- genti I ine atus, Ryukyu Islands

INTRODUCTION Ochlerotatus longifurceps is the lst authentic re- cord of a mosquito biting a fish (mudskipper). In the Ryukyu Islands of Japan, adult and im- The use of immunological techniques to identify mature of Ochlerotatus (Geoskusea) baisasi blood meals is a valuable aid in determining the (Knight and Hull) are commonly found in holes blood source (Sasaki 1988). This paper pre- made by land crabs of the family Grapsidae and the sents observations on the feeding behavior of Oc. mud lobster, Thalassina anomala, near mangrove baisasi in mangrove forests in Oura, Okinawa, and forests (Miyagi and Toma 1978, Tanaka et al. 1979, Komi, Iriomote (Ryukyu Islands), and in the labo- Miyagi and Toma 1980, Mogi et al. 1984, Toma ratory as well as the results of enzyme-linked im- and Miyagi 1986, Ng and Kang 1988). However, munosorbent assays (ELISAs) of blood meals in the no information is available on the blood meal stomachs of wild-caught mosquitoes. source and the feeding behavior of this mosquito. According to Belkin (1962),9 of mos- quitoes belonging to the subgenus Geoskusea have MATERIALS AND METHODS been reported from South Pacific Islands and Observation on biting behavior of Oc. baisasi -tonus (Edwards) and kabaensis vis-d-vis mudskipper in the laboratory.' Mosquito (Brug) from Indonesia; baisasi from the Philippines larvae were collected from crab holes in the man- and Ryukyu Islands, Japan; fimbripes (Edwards) grove forest of Oura on September 21, 20O2, by from New Guinea and the Bismarck Archipelago; hand pump and reared to the adult stage in the lab- dalienensis (Taylor) from northern ; becki oratory. A mudskipper (Periophthalmus argentili- (Belkin), perryi (Belkin) and longifurceps (Ed- neatus Yalenciences) also was collected at a wards) from the Solomon Islands; and daggyi pool in the mangrove forest of Oura. The mudskip- (Stone and Bohart) from Vanuatu. All species use per was confined in a cage (2O x 20 x 30 cm) with holes made by land crabs near the coast for breed- about 148 newly emerged female and male mos- ing and resting. Ochlerotatus (Geoskusea) longifur- quitoes. The cage was placed outdoors, and the ceps, which is closely related biologically and mor- feeding activity of the mosquitoes was observed phologically to Oc. baisasr, has been reported to from October 3 to 16, 2002, through a close-up lens bite mudskipper (Periophthalmidae) on Gizo Island, attached to a camera. Engorged mosquitoes were in the Solomon Islands (Stooff and Marks 1965). transferred to another cage of the same size. A tube To our knowledge, although several mosquito spe- (15 X 5 cm diameter) made of black cardboard cies have been reported to feed on reptiles and am- served as a simulated crab hole for resting and ovi- phibians (Reington 1945, Henderson and Senior position. 1961, Hayes 1965, Nalan et al. 1965, DeFoliart Blood meal determinations in the ,stomachs of 1967, Miyagi 1972), feeding on fish is quite rare. wild-caught female mosquitoes by ELISA: Resting female Oc. baisasi engorged with fresh blood and half gravid were collected by sucking tube from the rMiwa Junior High School, 519 Makabe, Itoman, Oki- upper parts of crab holes in the mangrove forests nawa 901-0336, Japan. of Oura and Komi. Twenty-seven engorged speci- 2 Laboratory of Medical Zoology, School of Health Sci- mens were squashed individually onto a filter paper ences, University of the Ryukyus, 2O7 Uehara, Nishihara, (Whatman No. l; Whatman, Tokyo, Japan) imme- Okinawa 903-0215, Japan. diately after capture. The filter papers were kept in 3 Laboratory of Entomology, Rakuno Gakuen Univer- gel sity, Bunkyoudai-midorimachi, Ebetsu, Hokkaido 069- a desiccator with silica after being air dried in 8501, Japan. the field, and the smeared parts were cut off with 4lnternational Student Center, Kanazawa University, scissors and stored individually in a microvial until Kakuma-machi. Kanazawa. Ishikawa 920-1 192. J apan. assaying. The ELISA procedure for determination

134 JuNs 2004 Oc. BArsAstFEEDTNc oN MuosrtppBn 135

Table 1. Feeding habits of Oc. baisasi on a caged mudskipper, P. argentilineatus.t.z

Time feeding took Case no. Feeding location place (h) Remarks I Between eyes - 2200 Penetrated tissue but no bloodfeeding took place 2 On the head - 2200 Bloodfeeding interrupted by host movement, mosquito returned and completed feeding 3 Between eyes - 2200 Flew away from the host immediately after full bloodfeeding 4 On the pectoral fin - 2200 Flew away from the host immediately after full bloodfeeding 5 On the head 1947 Feeding intemrpted by host, no blood taken 6 Near mouth opening 2007 Feeding interrupted by host, no blood taken 7 On the head 2031 Rested on the host for a few minutes after full bloodfeeding accomplished 8 On the back near the t952 Rested on the host for a few minutes after full dorsal fin bloodfeeding accomplished

' Average time per blood meal was 1O min. z Experiments were conducted Octobq 3-16,2OO2. of blood meal source was that of Sasaki (1988), day in the black paper tube provided in the cage. with the use of anti-fish (serum) sheep serum (Cos- Mating swarms were observed to take place above mo Bio Co., Ltd., Tokyo, Japan) and HRP-conju- the opening of the tube at dusk and dawn. Some gated anti-sheep (IgG) rabbit serum (ICN Pharma- female mosquitoes were observed to hover over the ceuticals, Inc., Costa Mesa, CA) as a conjugate. mudskipper and alight on the dorsal part of the fish, around and eventually feeding between the dorsally protruding eyes or the pectoral fin mus- RESULTS AND DISCUSSION culature of the mudskipper. Eight observations of feeding activity are shown Feeding usually lasted about 10 min for a full in Table 1. Male and female Oc. baisasi seem to bloodfeeding. The female appeared to be motion- be nocturnal or crepuscular in activity, resting by less during the feeding, and the fish did not appear

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Fig. 1. Ochlerotatus baisasi feeding on a mudskipper, P. argentilineatus 136 JounN.lr- op rnr Avgnrcal Mosqurro Corvrnol Assocnlox Vor. 20, No. 2

Table 2. Number of Oc. baisasi for which the blood them had fresh blood meals, and others were newly meal was determined by ELISA to be from fish. emerged nulliparous. It can be assumed that Oc. Engorged Engorged baisasi females feed fully on a mudskipper at night. females females Because of the heavy weight of blood, she will land testing testing on the nearest hollow and rest for several hours positive negative until the blood meal in her stomach has been di- Collection for a fish for a fish gested to a certain extent. The half-gravid female Collection site date (2OO2) blood meal blood meal will fly to crab and mud lobster holes for ovioosi- Oura, Okinawa July 6 1 0 tion and to rest until mornins. Oura, Okinawa July 13 I 0 (Oxudercina-e) are a terrestrially Komi, Iriomote August11 6 5 adaptable fish. They are exposed to air for long Komi, Iriomote August 12 2 7 periods of time and rest on plant roots in tidal mud- Komi, Iriomote August 13 0 5 flats and at night (Clayton 1993). Be- Total 10 T7 cause many mudskippers breed in tide pools scat- tered in the forest, they might be the most readily accessible source of a vertebrate blood meal for the crab hole-dwelling mosquitoes. Although the feed- to be disturbed during the process (Fig. 1). Three ing habits of the 9 species of subgenus Geoskusea females (case nos. 2, 5, and 6) interrupted their are not known, except for baisasi and longiforceps, blood feeding because of movement of the host; 1 most of these species could feed readily on a mud- female (case no. 2) subsequently returned to the skipper in the tidal zone. fish for a full feeding. The engorged females laid a batch of 10-40 eggs; most eggs hatched and de- ACKNOWLEDGMENTS veloped to the adult stage. One of us (Okudo) often exposed his hand to We express sincere appreciation to Yukio Naka- about 100 mature Oc. baisasi females in the rearing sone and Takashi Uchino, University of the Ryu- cage, but the females did not attempt to bite his kyus, for providing information and literature on hand. The females did not seem to be attracted to the crab and mud lobster; and to Yuko Nakazato warm-blooded while confined. for useful information on the mudskipper. Our ap- Blood meal sources of Oc. baisasl as determined preciation also goes to Masaya Tatara and Chieko by ELISA are shown in Table 2.Ten (37.07o) blood Matsumoto, Iriomote Wildlife Center, and Mikako meals of 27 tested females were determined to have Tamashiro for their kind collaboration. Thanks are fed on a fish. Two species of the oxudercine gobies, also due to Zenyo Yamazato and Takafumi Tomo- also called mudskippers-P. argentilineatus and P. yose, Nago City Office, for giving permission to modestus-are distributed in the estuaries, intertidal enter the important cultural asset, Oura mangrove flats, and mangrove forests of the Ryukyu Archi- forest. We are also grateful to Yong Hoi-Sen for his pelago (Murdy 1989, Clayton 1993). According to review of the manuscript. our observations, P. argentilineat rs :ue very com- mon in the mangrove forests of Oura, Okinawa, and REFERENCES CITED Komi, Iriomote, and appear to be a suitable host Belkin JN. 1962. The mosquitoes of the South Pacific for Oc. baisasi. (Diptera, Culicidae)Volume 1. Berkeley and Los An- Seventeen (63.0Vo) blood meals of 27 tested fe- geles: Univ. Calif. Press. males did not respond to antiserum produced by Clayton DA. 1993. Mudskippers. In: Ansell AD, Gibson fish. The small volume of blood meal in the stom- RN, Barnes M, eds. 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