Appl. Entomol. Zool. 41 (2): 217–226 (2006) http://odokon.org/ Cold tolerance of the sweet potato weevil, Cylas formicarius (Fabricius) (Coleoptera: Brentidae), from the Southwestern Islands of Japan Ikuo KANDORI,1,* Toshio KIMURA,1 Hisaaki TSUMUKI2 and Tuyosi SUGIMOTO1 1 Entomological Laboratory, Faculty of Agriculture, Kinki University; Nara 631–8505, Japan 2 Research Institute for Bioresources, Okayama University; Kurashiki 710–0046, Japan (Received 19 July 2005; Accepted 7 December 2005) Abstract The cold tolerance of Cylas formicarius from five islands in the southwest of Japan was studied. The thermal thresh- old of the reproductive activity of adults tended to be lower and the supercooling point of the pupae was a little higher for the Okinawa-Honto population than for the others. The cold tolerance of the adult weevils was slightly accelerated by cold acclimation to 15°C for 4 d. Reproductive diapause induced under 10L : 14D increased cold tolerance signifi- cantly. Therefore, the cold tolerance of adult weevils would be accelerated by both cold acclimation at decreasing tem- peratures from autumn to winter and reproductive diapause induced by short daytime hours. When considering the in- creased cold-tolerence of C. formicarius through such mechanisms together with the temperature conditions in winter at the northern edge of its distribution area in the USA, C. formicarius probably has the capability to survive in the coastal zone of the Pacific Ocean in southern and central Japan. Key words: Cylas formicarius; expanded distribution; cold tolerance; cold acclimation; reproductive diapause In Japan, Nawa (1903) first identified this weevil INTRODUCTION and reported its damage in Okinawa Prefecture. The sweet potato weevil, Cylas formicarius Thereafter, the distribution range of this weevil had (Fabricius), is a serious pest of the sweet potato, expanded by transporting infested tubers, and by Ipomoea batatus (Linnaeus), and is found mainly 1940 the pest thoroughly covered the Amami Is- in the tropics and the subtropics throughout Asia, lands. In 1950, all of the southwestern islands lo- the Pacific islands, Africa, the Caribbean, USA, cated at latitudes of lower than 30°N were desig- Venezuela and Guyana (International Institute of nated as being within the infested zone, and have Entomology, 1993). Though C. formicarius prefers since been regulated by the Plant Quarantine Laws I. batatus, more than 30 species of Ipomoea and of Japan. However, recently the weevil has often other genera are recorded as its host plants (Suther- been found in the infestation-free zone at latitudes land, 1986; Austin, 1991). Adults feed on the epi- higher than 30°N, due to the prosperous trade of dermis of vines, petioles, and the external surface agricultural products among the islands (Setokuchi of tubers. Adult females lay individual eggs in et al., 1996; Sugimoto, 2000). In those areas, the vines and tubers. The developing larvae tunnel weevil has been eradicated in every case by the through these parts, causing significant damage emergent control based on the above law, in which (Sherman and Tamashiro, 1954). In response to the a large amount of pesticides was sprayed and host damage, the tuber produces terpene phytoalexins, plants were thoroughly removed (Fujimoto et al., which make the potato inedible even at low levels 2000; Nishioka et al., 2000). For the development of physical damage by the weevils (Akazawa et al., of an effective plant quarantine strategy in Japan, it 1960; Sato et al., 1981, 1982). More than 50% of is essential to clarify how far C. formicarius can the crops in the southwestern islands were dam- potentially disperse toward northern areas. In other aged when grown without any control. words, it is necessary to clarify its cold tolerance. *To whom correspondence should be addressed at: E-mail: [email protected] DOI: 10.1303/aez.2006.217 217 218 I. KANDORI et al. This study examines the cold tolerance of C. formi- carius from five southwestern islands in relation to cold acclimation and diapause, together with threshold temperatures of reproductive activity and supercooling points. MATERIALS AND METHODS Weevils tested. Weevils were collected from southwestern islands of Japan: Amami-Oshima Is. and Okinawa-Honto Is. in 1996, and Nakanoshima Is., Miyakojima Is. and Ishigakijima Is. in 1997 (Fig. 1). After collection, weevils were maintained for successive generations in an isolated insectar- ium at 27.0Ϯ0.5°C (meanϮrange), 70% RH, 14L : 10D (which we call ‘normal conditions’ here- after) at the Faculty of Agriculture, Kinki Univer- Fig. 1. Five islands in the southwest islands of Japan sity, Nara, Japan. They were reared in a plastic where C. formicarius weevils were collected for this study. container (11 cmϫ30 cmϫ10 cm high) with one Muroto is the northernmost area in Japan where C. formicar- ius has invaded hitherto, though eradicated already by emer- sweet potato tuber (variety; Beni-Ogojo, ca. 200 g). gency control. When we used cut tubers, the cut faces of the tu- bers were coated with paraffin to minimize desic- each female and three males were kept together in cation and decay. In the present study, weevils from a plastic container (7 cm wide, 4 cm high), with a the five islands were tested in 1999 and 2000. In all piece of tuber for 5 d at the same temperatures as the experiments, we used immature adults soon during acclimation in order to allow oviposition. after eclosion, which were obtained from damaged After an additional 10 d under normal conditions, tubers by dissection. Fresh immature adults were these tubers were dissected. The presence of larvae visually distinct from mature ones by their pale was considered to be proof of oviposition. color. In this text, weevils from each island are Supercooling point. Supercooling points of the hereafter called by the name of their original is- 3rd-instar larvae, pupae, and male and female land; e.g., weevils from Nakanoshima Is. are re- adults reared under normal conditions were meas- ferred to as Nakanoshima. This study was carried ured in an isolated laboratory at the Research Insti- out with special permission from the Japan Plant tute for Bioresources, Okayama University, Quarantine Office. Kurashiki. Each weevil was fixed on the tip of a Thermal thresholds of reproductive activity. thermocouple with silicon grease and the tempera- Male and female adults from each island were kept ture was decreased at a rate of about 1.0°C/10 min separately in plastic containers (9.5 cm wide, 6 cm in a freezer. The body temperature of weevils was high) with pieces of a tuber under normal condi- recorded with a multi-channel temperature tions for 10 d to promote sexual maturation. Then recorder (Asai et al., 2002). we tested the thermal threshold of 1) copulation Cold tolerance of non-acclimatized adults. and 2) oviposition for adults under the following Immature adult weevils soon after eclosion were conditions. 1) After males and females were sepa- reared in a plastic container (7 cm wide, 4 cm high) rately acclimatized at 10, 11, 13, 15 and 27°C for with a small piece of tuber under normal condi- 2d in a group, each pair was kept in a container tions for 2 wk. Then, males or females were sepa- with a piece of a tuber for 5 d at the same tempera- rately exposed to 0 or Ϫ3°C for 5, 7 or 10 d and the tures as during acclimation. Copulation was con- survivors were counted. Survival was confirmed by firmed by the presence of sperm in the female sper- touching insects with the tip of a calligraphy brush mathecae checked by microscopic examination. 2) just after transferring them to room temperature After males and females were separately acclima- from cool conditions, and again 1 d later. In this tized at 14, 15, 16, 18 and 27°C for 2 d in a group, study, 0 and Ϫ3°C were adopted as exposure tem- Cold Tolerance of C. formicarius 219 peratures, referring to the mean daily minimal tem- nϭ10 in each). Thus, to ensure reproductive dia- peratures in two areas in the USA (see Table 6), pause, weevils were reared with tubers at 24°C, since these two areas are situated around the most 70% RH, 10L : 14D for 2 wk after adult eclosion. northern part of the distribution area of C. formi- Then, after 2 h of acclimation at 15°C, males and carius in the USA. females were separately exposed to 0°C for 5, 7 Cold tolerance of cold-acclimatized adults. To and 10 d. Survivors were counted by the same determine the effect of cold-acclimation on sur- method described above. vival, soon after eclosion the males and females were separately assigned to the following two con- RESULTS ditions. 1) After 2 wk of rearing under normal con- ditions and 2 h of acclimation at 15°C, weevils Thermal thresholds of reproductive activity were exposed to 0°C for 5, 7 and 10 d. 2) After The numbers of females that stored sperm in 10 d of rearing under normal conditions and 4 d of their spermathecae tended to be the largest for Oki- acclimation at 15°C, weevils were exposed to nawa-Honto at 10, 11 and 13°C, however, it was Ϫ3°C for 5, 7 and 10 d. In the second condition, significant only at 11°C (Table 1). The thermal the acclimation time was prolonged to 4 d because threshold of mating behavior, that is, the maximal the acclimation of 2 h seemed less effective as an temperature under which all of the specimens increment for cold tolerance under the first condi- ceased mating behavior, was 10 to 11°C or lower tion (see RESULTS). for all of the populations tested.