Newly Developed Fungal Diet for Artificial Rearing of the Endangered Long-Horned Beetle Callipogon Relictus (Coleoptera: Cerambycidae) Dae-Am YI1,4, Alexander V

Entomological Research 47 (2017) 373–379 RESEARCH PAPER Newly developed fungal diet for artificial rearing of the endangered long-horned beetle Callipogon relictus (Coleoptera: Cerambycidae) Dae-Am YI1,4, Alexander V. KUPRIN2,YongHoonLEE3 and Yeon Jae BAE1 1 Department of Environmental Science and Ecological Engineering, College of Life Sciences and Biotechnology, Korea University, Seoul, South Korea 2 Federal Scientific Center of the East Asia Terrestrial Biodiversity, Russian Academy of Sciences, Vladivostok, Russia 3 Wonju, Gangwon-do, South Korea 4 Research Center of Natural Monument Insects, Yeongwol, Gangwon-do, South Korea Correspondence Abstract Yeon Jae Bae, Department of Environmental Science and Ecological Engineering, College of In this study, we attempted to develop an artificial diet to effectively rear the Life Sciences and Biotechnology, Korea endangered long-horned beetle Callipogon relictus Semenov to facilitate restoration University, Seoul 02841, Korea. efforts for this species. Fungal mycelia of the white-rot fungus Pleurotus florida Email: [email protected] (Eager) were, for the first time, provided as an artificial diet for the larvae of C. relictus. The experiment started on August 11, 2015 with 53 larvae being Received 10 February 2017; provisioned with the fungal diet under laboratory conditions (25°C, 60 % RH, L:D accepted30March2017. 0:24) without diapause. During the first year, 11 adults (4 males and 7 females, doi: 10.1111/1748-5967.12234 20.8 % of the 53 larvae) emerged between September 10 and October 1, 2016. The body length of the largest male was 106.8 mm, and three of the four males exceeded 100 mm in length. These results show that a fungal diet for rearing C. relictus is useful not only for reducing the larval period to approximately 1/6 of the normal larval period in nature, but also for producing large adults. These results support the notion that fungus may provide nutritional benefits during the larval development of C. relictus, and also indicate that this species is fungivorous. Key words: Endangered insect restoration, Fungal mycelium diet, Insect rearing, Larval development, Relict long-horned beetle, White-rot fungus Pleurotus. Introduction in the field is known to be as long as 5 to 6 years (Li et al. 2012). Therefore, reducing the rearing time of the larvae is The endangered long-horned beetle Callipogon relictus considered crucial for effective population restoration. Semenov (Fig. 1), the largest beetle in the Palearctic region, Recently, it has been reported that C. relictus larvae developed is sporadically distributed throughout the Russian Far East, into adults within 6 years in a semi-natural outdoor the northeastern part of China, and the northern part of the environment when provided with sawdust (Li et al. 2012). Korean Peninsula. It is listed in the Red Data Book (Category Two other experiments have shown that the larval growth I) and protected in the Russian Federation and the Republic of stage can be shortened to 42–44 months by terminating Korea. In Korea, it is also designated as a National Natural diapause. The first experiment was performed during the Monument (No. 218) and protected by law. The beetle has period 2009–2013 in Ussuriysk, Russian Federation, by been widely used as an emblematic species in Korea including feeding the larvae naturally decayed Ulmus japonica wood adoption as the symbol of the Entomological Society of under laboratory conditions (25°C, 65 %–75% RH) (Kuprin Korea. et al. 2014). The second experiment was carried out during Special attention has been paid to the restoration of the period 2010–2014 in Yeongwol, Republic of Korea, by C. relictus in Korea and Russia which requires artificial feeding the larvae an artificial diet comprising the sawdust of rearing of the larvae. However, the larval period of C. relictus Quercus mongolica supplemented with additional nutrients © 2017 The Entomological Society of Korea and John Wiley & Sons Australia, Ltd D.-A. Yi et al. stage. It has also been demonstrated that provision of an artificial fungal diet affects adult size and pupal development time in the bark beetle Dendroctonus ponderosae (Bleiker and Six 2007). Given the findings of these studies, it is very tempting to speculate whether C. relictus, which like many other xylophagous beetles inhabit decayed wood, is also fungivorous. In this study, we tested Lyubarsky’s (1953) assumption that fungal mycelia might be an effective diet for the growth of C. relictus larvae. [Correction added on 12 July 2017, after first online publication: “C. relictusfi” has been changed to “C. relictus”.] For this study, we mainly examined the use of fungal mycelia instead of raw sawdust or sawdust containing additional nutrients as a diet for rearing the larvae of C. relictus. Figure 1 A Callipogon relictus female laying eggs on Ulmus japonica infected by the fungus Pleurotus citrinopileatus in Ussuriysk, Russia. Materials and methods under laboratory conditions at constant temperature (20°C, Insects 65 %–75% RH) (Yi 2014). In the former study, the larval growth period was successfully reduced to 42 months, We imported two pairs of live C. relictus collected in North whereas in the latter it was reduced to 44 months (Fig. 2). Korea (Cheonma-san, Pyeonganbuk-do, N. Korea, coll. July – Although there have been numerous previous studies on the 28 August 4, 2015) via China on August 10, 2015. A customs use of artificial diets for long-horned beetles (Harley and clearance to South Korea was approved (#15408195) by the Wilson 1968; Sugiyama 1969; Gardiner 1970; Yamane 1973, Incheon International Airport Regional Office of Animal and 1974; Akutsu et al. 1980; Murakoshi and Aono 1981; Kevin Plant Quarantine Agency (APQA) under the permission of and Robinson 1982; Dubois et al. 2002), little is known about the Ministry of Environment of Korea. utilizing fungal mycelia as a diet for any species of long-horned The adults were divided into pairs, and induced to mate in beetles (Kukor and Martin 1986). In the case of stag beetles two plastic cages (63 cm in length, 44 cm in width, and (Lucanidae) that consume dead or decayed wood during their 23 cm in height) on August 11, 2015. Each of five pieces larval growth period, it is commonly known to insect breeders of decayed wood of Mongolian oak Quercus mongolica – – and collectors that larvae tend to be of a larger size at sites (8 9cmindiameterand12 13 cm in length) to be used for where white-rot fungi are observed on the base of trees. egg-laying were prepared and soaked in water for 2 h. The Most lucanid beetles are well known to be fungivorous wood pieces were then sterilized in a microwave oven for (Kojima 1996; Tanahashi et al. 2009; Tanahashi and Kubota 30 min. After observing several mating activities, the males 2013), and some species of Anobiidae, such as the death- were removed from the cages and isolated. The females were – watch beetle Xestobium rufovillosum are highly associated left in the cages at room temperature (22 25°C) until post- with fungus (Campbell 1941) for the duration of the larval oviposition and death. Commercial sugar gelatins were provided as the adult’s food and were frequently replanished. On September 11, after a month of rearing, the wood pieces were split in order to collect the grown larvae. Fifty-three 1st instar larvae were taken from one cage, but no larvae were found in the other cage. Artificial fungal diet The fungal diet used for rearing larvae was newly separated and cultivated out of mycelia of the white-rot fungus Pleurotus florida (Eager). We used commercial potato dextrose agar powder (PDA: 213400-BD; Difco, Becton Dickinson, Franklin Lakes, NJ, USA) as the medium for sub-culturing Figure 2 Comparison of the larval development period of Callipogon P. florida. PDA agar powder was mixed in a 1000 mL relictus with that observed in previous studies. graduated cylinder, at a ratio of 19.5 mg of powder in 374 Entomological Research 47 (2017) 373–379 © 2017 The Entomological Society of Korea and John Wiley & Sons Australia, Ltd Fungal diet for Callipogon relictus 500 mL water. To aid solubilization, the solution was Table 1 The total period of larval development and body size of male transferred to a 1000 mL conical flask and mixed with a adults fed with fungal diet magnetic stirrer on a heating plate. When completely solubilized, the solution was divided into 20 mL portions, Emerged Laval period Length Weight which were poured into 50 mL round-bottomed glass test No. date (2016) (day) Sex (mm) (g) tubes (27.3 mm in inner diameter), which were capped with 1Sep.10358♂ 101.4 14.59 silicon stoppers. The glass test tubes were then autoclaved at 2Sep.15363♂ 90.2 13.09 2 121°C for 20 min at a vapor pressure of 1.2 kg/cm ,and 3Sep.16364♂ 106.8 16.90 placed in a tilted position until solidified. 4Sep.21369♂ 100.8 15.84 For production of the sawdust medium, we used crushed Mean (±SE) 363.5 99.80 15.105 Mongolian oak wood to prepare sawdust, which was (±2.3) (±3.47) (±0.821) maintained at a 50 %–60 % moisture content. One thousand- gram amounts of the moistened sawdust were placed into polypropylene bottles (1400 mL). The surface of the sawdust Table 2 The total period of larval development and body size of female was then tamped, and a hole of 1 cm diameter was made in the adults fed with fungal diet middle, from the surface to the bottom of the bottle. The bottles were closed with a sterilized cotton cap. The sawdust Emerged Laval period Length Weight No. date (2016) (day) Sex (mm) (g) medium was autoclaved at 121°C for 2 h, at a vapor pressure 2 of 1.2 kg/cm .

Load more