Distinguishing Male and Female Chinese Rose , sinicus, with an Overview of Adoretus Species of Biosecurity Concern Author(s): Grant T. McQuate and Mary Liz Jameson Source: Journal of Science, 11(64):1-18. 2011. Published By: Entomological Society of America DOI: http://dx.doi.org/10.1673/031.011.6401 URL: http://www.bioone.org/doi/full/10.1673/031.011.6401

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Distinguishing male and female Chinese rose beetles, , with an overview of Adoretus species of biosecurity concern Grant T. McQuate1* and Mary Liz Jameson2

1USDA-ARS, U.S. Pacific Basin Agricultural Research Center (PBARC) 2Department of Biological Sciences, Wichita State University, 537 Hubbard Hall, Wichita, Kansas 67260 Abstract The Chinese rose , Adoretus sinicus Burmeister (Coleoptera: : : Adoretini), is a broadly polyphagous scarab beetle that is economically important and causes damage to a wide variety of host plants including agricultural crops and ornamentals in Southeast Asia, China, the Hawaiian Islands and several other Pacific Islands. The species has become established in numerous regions and is of biosecurity concern because importation of this species to other regions poses a threat to agriculture due to its generalist herbivore feeding habits. Field and laboratory research directed towards control of the species is hampered by the lack of characteristics that allow accurate determination of the sexes on live beetles in the field. Here, three recognizable and reliable non-destructive morphological differences between the sexes of A. sinicus are documented: (1) the form of the terminal sternite; (2) the length to width ratio of protarsomere 1, and; 3) the ratio of the combined length of protarsomeres 2-4 to the length of protarsomere 1. Because many Adoretus species are of biosecurity concern, and because tools to identify Adoretus species are lacking, we review the natural history and research on control associated with A. sinicus as well as the genus as a whole.

Keywords: Scarabaeidae, invasive species Correspondence: a* [email protected], b [email protected], *Corresponding author Received: 26 March 2010, Accepted: 21 January 2011 Copyright : This is an open access paper. We use the Creative Commons Attribution 3.0 license that permits unrestricted use, provided that the paper is properly attributed. ISSN: 1536-2442 | Vol. 11, Number 64 Cite this paper as: McQuate GT, Jameson ML. 2011. Distinguishing male and female Chinese rose beetles, Adoretus sinicus, with an overview of Adoretus species of biosecurity concern. Journal of Insect Science 11:64 available online: insectscience.org/11.64 Journal of Insect Science | www.insectscience.org 1 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson

Introduction Islands” (Muir 1917). In areas where it has been introduced, adults can cause heavy In the Hawaiian Islands, the introduction of damage to crops and ornamental plants. non-native and plants has caused Despite much research devoted to control of millions of dollars in crop losses, extinction of the species (including pesticides, parasitoids, native species, destruction of native forests, pathogens, and pheromones) control has been and in excess of $40.8 million in government- largely ineffective (Beardsley 1993). To date, sponsored spending in 2006 for biosecurity A. sinicus remains an abundant pest initiatives (Hawaii Department of Land and throughout the Hawaiian Islands (e.g., Natural Resource 2008). Well over 2,500 Beardsley 1993; Furutani et al. 1995; Siemann species have been introduced to the and Rogers 2003) and is considered a threat to Hawaiian Islands with continued the west coast of the United States (Ritcher establishment rates at an alarming ten to 1948). The species is included on the USDA- twenty species per year (Asquith 1995). One APHIS Regulated Plant Pest List (USDA- non-native species in the Hawaiian Islands is APHIS 2000) and has been detected on floors the Chinese rose beetle, Adoretus sinicus of empty sea cargo containers in storage in Burmeister (Coleoptera: Scarabaeidae: Australia (Stanaway et al. 2001). Fortunately, Rutelinae: Adoretini) (Figures 1-2). The biosecurity protocols have become more species is native to Japan and Taiwan (Mau efficient in recent years, and A. sinicus has not and Kessing 1991), but its distribution now become established in the mainland United also includes China (including Hong Kong), States. Nevertheless, A. sinicus is considered a Indonesia, Cambodia, Singapore, Thailand, species with moderate potential for invasive Vietnam, the Marianas Islands, the Caroline risk in the United States based on factors that Islands and the Hawaiian Islands (CAB include: 1) areas with potentially suitable International 1981). In the Hawaiian Islands, climate exist; 2) entry potential into a new it was first reported on Oahu in 1891 (Riley country based on historical data and and Howard 1893). By 1893, the species was interceptions; 3) potential economic impact if "rapidly becoming a most serious pest” (Riley the species were to become established; and 4) and Howard 1893). Initial identification specific hosts that the species can damage provided the name Adoretus umbrosus L.; (Gregory et al. 2005). Because it has become later, it was referred to as A. tenuimaculatus established in numerous regions, it is clearly a Waterhouse (Pemberton 1964). Nineteen species of concern for biosecurity. years after its noted pest status, the German ruteline systematist Friedrich Ohaus correctly Methods of control are needed for the Chinese identified the beetle as A. sinicus. rose beetle, both to minimize damage caused to agricultural and ornamental crops in Hawaii Adoretus sinicus probably came to Hawaii in as well as to reduce the risk of its introduction the larval stage in soil with imported plants to the U.S. mainland. The lack of reliable, from China, Taiwan, Java, or Timor (Ohaus non-destructive methods for sex determination 1935; Pemberton 1964). By 1898 it was impedes field and laboratory research. established on all major Hawaiian Islands Determination of males and females in most (Koebele 1898), and in 1917 it was referred to species of Adoretus is problematic. Unlike as “one of the worst garden pests of the many genera of leaf chafers (Rutelinae:

Journal of Insect Science | www.insectscience.org 2 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson Scarabaeidae), there is amazingly little is caused by interveinal feeding that creates a external sexual dimorphism. Within the lace-like appearance and greatly reduces Rutelinae, standard sexual determination can photosynthetic capabilities of the plant be made based on a variety of characters, (Furutani et al. 1995). The adult Chinese rose including differences in the form of the beetle has been reported to feed on over 250 protarsal claws, protarsi, metatibial apex, species and approximately 56 families of metatibia, metacoxa, metafemur, plants (Habeck 1964) or on over 500 plant metatrochanter, and abdominal sternites. species (Hession et al. 1994). Host plants However, as stated by Baraud (1985), most include many economically important plants Adoretus species lack noticeable sexual such as broccoli (Brassica oleracea var. dimorphism. Currently, determination of italica Plenck), cabbage (Brassica oleracea sexes in adult A. sinicus requires killing and var. capitata L.), cacao (Theobroma cacao dissection of specimens (e.g., Hession et al. L.), Chinese broccoli (Brassica oleracea L. 1994). Our research sought more readily var. alboglabra), Chinese cabbage (Brassica accessible, external morphological characters rapa L. subsp. chinensis [L.] Hanelt), chiso that would not necessitate destruction of live (Perilla frutescens [L.] Britton), corn (Zea specimens. In support of further research to mays L.), cotton (Gossypium barbadense L.), improve control of the Chinese rose beetle, an cucumber (Cucumis sativus L.), eggplant assessment of characteristics that can be used (Solanum melongena L.), ginger (Zingiber for non-destructive differentiation of the sexes officinale Roscoe), grape (Vitis labrusca is provided. Additionally, because background Bailey), green beans (Phaseolus vulgaris L.), literature on Chinese rose beetle is scattered jack fruit (Artocarpus heterophyllus Lam.), among a number of publications, many of okra (Hibiscus esculentus L.), peanuts which are not readily available, a review of (Arachis hypogaea L.), Oriental persimmon the basic biology of this species and past (Diospyros kaki Thunb.), raspberry (Rubus efforts of control is also included. The niveus Thunb.), roses (Rosa spp.), salak palm assessment provided here of characteristics (Salacca zalacca Gaerther), soybean (Glycine useful for differentiation of sexes in the max L.), star fruit (Averrhoa carambola L.), Chinese rose beetle may also be helpful in sex strawberry (Fragaria chiloensis [L.] Duch.), determination of other species of Adoretus, sweet potato (Ipomoea batatus [L.]), taro including known pests that are also of (Colocasia esculenta [L.] Schott) and tea biosecurity concern because of the potential (Camellia sinensis L.) (Mau and Kessing for invasion through the movement of potted 1991; Arita et al. 1993; Zee et al. 2003). plants across geographic regions. In recognition of the potential agricultural and Adults emerge at dusk and peak feeding and biosecurity threat of Adoretus species, we also mating occurs about 30 minutes after sunset review closely related and economically (Tsutsumi et al. 1993). As is common with important Adoretus species. many other nocturnal species of scarab beetles, adults are readily monitored using Biology of the Chinese Rose Beetle light traps. Sex ratio in the field is approximately 1:1, although about 27.9% of The Chinese rose beetle is a generalist foliage adults collected at light traps are males feeder and is primarily active at night (Ebesu (Habeck 1964). During the day, adults are 2003). Damage to ornamental plants and crops found under leaves, loose bark, or are

Journal of Insect Science | www.insectscience.org 3 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson shallowly buried in the soil (Williams 1931). beetle (Figures 1, 2) is oblong oval, reddish- Adults preferentially feed on leaves and plant brown and covered with creamy, dense, scale- species that are relatively high in non- like setae, giving the beetle an overall grayish structural carbohydrates (Arita et al. 1993) appearance. Body length varies from ~10-12 and prefer leaves with feeding or other types mm in length. Species identification for A. of damage (Pemberton 1959). Plants produce sinicus is based on form of the male genitalia increased levels of ethylene gas both after (Figure 10). Field-collected adults have been ethephon treatment and after leaves are found to live for as long as eight weeks in the damaged by insect feeding. It appears that the laboratory (Habeck 1964). high levels of ethylene gas emission “are involved in the formation of a plant volatile(s) The characteristic interveinal defoliation that serves as an attractant and feeding pattern of the Chinese rose beetle is caused by stimulus for the Chinese rose beetle” (Arita et the beetle’s unusual mouthparts. The labrum al. 1988). Such an attractant, however, has not is produced ventrally at the middle and forms yet been identified. a tooth-like process. This process completely separates the mandibles and maxillae into two Developmental rate of A. sinicus from egg to independent chewing apparati that are adult varies greatly depending on temperature incapable of meeting in the middle. When and quality of larval substrate. Development feeding on a leaf surface, the beetle uses only from egg to adult averages about 15 weeks in one side of its mouth at a time, thus producing the laboratory (Habeck 1964) or 6-7 weeks in paired holes in leaves and often leaving a the field (Mau and Kessing 1991). Oviposition narrow strip of leaf intact in the middle begins seven days after pairing of mature (Arrow 1917). males and females (Habeck 1964). Females oviposit in the soil about 4 cm below the Because adults cause heavy damage to crops surface. Based on 6 field-collected females, and ornamental plants, research has been females lay 22-89 eggs each (average = 54). devoted to control of the species. Despite Eggs are oval (~1.5 x 1.0 mm) and white numerous attempts using many methods, (Habeck 1963). The egg stage may last control of the Chinese rose beetle has proved approximately 12-16 days at 24.0°C or 7-13 to be problematical. Tests to develop effective days at 28.6°C (Habeck 1964). The larval control techniques have included the use of stage includes three instars. Duration of the parasitoids, pathogens, pesticides, food lures, instars averages 19.6-22.8, 14.5-16.8, and and aggregation pheromones. 34.3-44.4 days, respectively (Habeck 1964). Larvae are c-shaped, white grubs that live in Parasitic Hymenoptera from the families rich soil, leaf litter, decaying vegetation, or Scoliidae (Campsomeris marginella modesta compost, where they feed on dead plant Smith) and Tiphiidae (Tiphia segregata tissues (Mau and Kessing 1991). They Crawford) were introduced to Hawaii from apparently do not attack living vegetable the Philippines to control the introduced tissues (Mau and Kessing 1991; Williams scarab beetles A. sinicus and Anomala 1931). The pupa, which is approximately 6.0- orientalis (Waterhouse) (Scarabaeidae: 12.0 mm, is covered with dense, short setae. Rutelinae: Anomalini) (Muir 1917, 1919). The pupal stage lasts 11-17 days, averaging Over 14,000 eggs, pupae, and adults of C. about 14 days (Habeck 1964). The adult marginella modesta were imported to Hawaii

Journal of Insect Science | www.insectscience.org 4 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson between 1915-1917. In the short term, there pesticides. However, tests on adult A. sinicus were immediate declines in the target scarabs. using the nematodes Steinernema Prior to importation of the wasp, over 3,500 carpocapsae (Weiser) and Heterorhabditis sp. Anomala grubs were found in 1/20th of an MB7 (Maui isolate) were ineffective (Hara et acre; only four Anomala grubs were found in al. 1989), and tests on larvae were also the same area in 1919 (Muir 1919). Muir ineffective (Tsutsumi et al. 1993). (1919) hypothesized that biocontrol success would lead to “extinction” of A. orientalis in Broad spectrum organophosphate pesticides, the region, and that C. marginella would like carbaryl, have been used for Chinese rose maintain itself on grubs of the Chinese rose beetle control, as they have been used for beetle. In the long term, however, control by control of Japanese beetles and other scarab these parasitoids has been only slightly beetles. Because of the potential for effective (Pemberton 1964). In another significant non-target effects of attempt, importation of 24,000 Adoretus grubs organophosphate insecticide use, development from the Philippines that were parasitized of effective reduced risk insecticides is with Tiphia lucida Crawford also failed needed. An azadirachtin-based pesticide has (Pemberton 1964). been tested and found to be promising (Arita Tsutsumi et al. 1995), and imidicloprid-based Treatments using the entomopathogenic fungi systemic pesticides have also been found to Metarhizium and Beauveria have been help in control of Chinese rose beetle. At conducted on the Chinese rose beetle. Success present, though, pesticide registrations are of using the green muscardine fungus, limited for Chinese rose beetle control in Metarhizium anisopliae (Metschnikov), Hawaii, especially considering the wide range against A. sinicus are conflicting. Koebele of crops that can be attacked by A. sinicus. (1897) and Williams (1931) reported some success against the Chinese rose beetle, Food-type lures, as have been developed for particularly during the wet season. Three the Japanese beetle (Popillia japonica strains of M. anisopliae (M346, M2151, and Newman) (Scarabaeidae: Rutelinae: M2162) were tested on larvae of A. sinicus Anomalini) (e.g., Ladd and McGovern 1980), with success (Tsutsumi et al. 1993), but could be developed for Adoretus species. effectiveness needs to be replicated due to the Alternatively, it may be possible to identify high mortality in both controls and attractive herbivory-induced plant volatiles experimental treatments. Consistent infection such as have been found for other scarab in scarabs may be limited to Beauveria beetle species (Loughrin et al. 1995, Reinecke brongniartii (Saccardo) Petch or large-spored et al. 2002). This possibility is encouraged by varieties of M. anisopliae var. majus (Metch.) the observation that “host plants that were Sorokin, and research efforts should be previously fed upon by the beetle were more directed towards these fungal strains and desirable as a food source than untouched species (Jackson and Klein 2006). plants of the same species” (Pemberton 1959, Arita et al. 1988). Patterns of feeding Entomopathogenic nematodes in the families preference could be used for management and Steinernematidae and Heterorhabditidae could control of A. sinicus. For example, A. sinicus be effective control agents for scarab beetle prefers recently matured leaves in the larvae or adults and could minimize the use of uppermost part of the plant, whereas A.

Journal of Insect Science | www.insectscience.org 5 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson versutus prefers younger leaves in the lower (GTM, unpublished). Further semiochemical part of the plant (Tsutsumi et al. 1993). research is clearly needed with A. sinicus.

Pheromones can be used to disrupt behavior, Materials and Methods reduce populations, and monitor for outbreaks of scarab pests (Jackson and Klein 2006). . Beetles were collected at night, in the Preliminary observations indicated that vicinity of Hilo, HI, from four different host pheromones may be involved in the attraction plant species (copperleaf, Acalypha of A. sinicus and A. versutus to mating sites wilkesiana Muell.-Arg. [Euphorbiaceae]; rose, (Tsutsumi et al. 1993), with the probable Rosa sp. [Rosaceae]; pilau maile, Paederia existence of an A. sinicus pheromone sex foetida L. [Rubiaceae]; and wax apple, attractant (Hession et al. 1994). A synthetic Syzygium samarangense (Blume) Merrill and sex attractant, produced by females and Perry [Myrtaceae]) from Jan.-Sept., 2009. primarily attractive to males (“Japonilure”), Collected beetles were held outdoors in an was successfully developed for the Japanese environment that included castor bean beetle (Tumlinson et al. 1977 1979, Doolittle (Ricinus communis L.); [Euphorbiaceae] et al. 1980) and joint presentation of plants on which they could feed for at least Japonilure and food-type lure provided a lure two weeks before beetles were evaluated. The “of outstanding attractancy” (Ladd et al. holding time was selected in order that there 1981). A sex attractant has also been would be an opportunity for wearing of the discovered for Anomala orientalis (Zhang et external teeth of the protibia. The external al. 1994) and shows promise for population teeth of the protibia tend to be more acute in management by mating disruption (e.g., the male (less acute in the female), but Wenninger and Averill 2006). Promising acuteness is subject to wear that rounds the attractants have been successfully developed apices of the teeth. Following the two week for at least one other Adoretus species. The holding time, beetles were killed by freezing synthetic unsaturated terpene alcohols in a So-Low ultra low temperature freezer nerolidol (3,7,11-tri-methyl-1,6,10- (So-Low Environmental Equipment, www.so- dodecatrien-3-ol) and geraniol ([E]3,7,- low.com) maintained at -70o C, and then dimethyl-2,6-octodien-1-ol) were shown to be external morphological characters of potential highly attractive to the Australian A. use in sex determination were evaluated. tessulatus Burmeister (Donaldson 1986). Both chemicals are found widely in the essential The following measurements were made on oils of flowers and plants. These results each individual: total body length (tip of the provide encouragement that comparable food- head to the tip of the closed elytra), maximum based and sex attractant lures could also be body width (maximum width of the elytra, developed for the Chinese rose beetle. typically at about the middle of the elytra), However, initial tests of synthetic lures length of protarsomere 1 (from the top of the developed for Japanese beetles and other condyle to the apex of the protarsomere; see scarab beetles (including geraniol) failed to Figures 3, 4), maximum width of identify effective attractants for A. sinicus and protarsomere 1, and the combined length of A. versutus (Tsutsumi et al. 1993), and protarsomeres 2 – 4 (from the base of preliminary trials of nerolidol also failed to protarsomere 2 to the apex of protarsomere 4; demonstrate attractiveness for A. sinicus see Figures 3, 4). All protarsomere

Journal of Insect Science | www.insectscience.org 6 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson measurements were made using the ventral (mean ± SE: length : 11.65 ± 0.067 mm; : side of the right front leg. For each specimen, 10.71 ± 0.047 mm; F = 130.91, df = 1, 198, p external teeth of the protibia (Figures 3, 4) < 0.0001; width : 5.39 ± 0.014 mm; : 4.97 were assessed as being acute or rounded ± 0.024 mm; F = 228.17, df = 1, 198, p < (Figures 3, 4) and whether the apex of the 0.0001). However, obvious overlap between terminal sternite was quadrate or rounded the sexes make these measurements unreliable posteriorly (Figures 5, 6). Each specimen was for sex determination (Figure 7). Protarsomere then dissected for definitive sex identification 1 was significantly longer in females (: 0.54 based on the presence or absence of the male ± 0.00070 mm; : 0.37 ± 0.00071 mm; F = genitalia. Presence or absence of eggs was 29166.35, df = 1, 198, p < 0.0001; Figures 3, also noted and, if present, maximum length 4). However, without a comparative and width were recorded for two randomly characteristic, this feature is not useful in selected eggs. One hundred specimens of each determining sex. Width of protarsomere 1 (: sex were evaluated for the above 0.199 ± 0.00027 mm; : 0.195 ± 0.00050 morphological characters. Observations on the mm) and the combined length of protarsomere form of the terminal sternite were made with a 2-4 (: 0.61 ± 0.00 mm; : 0.61 ± 0.00 mm) Leica Wild M3Z dissecting microscope (Leica varied little between sexes. However, both the Microsystems, Inc., www.leica- ratio of protarsomere 1 length: width (: 2.69 microsystems.com) that, in conjunction with a ± 0.0045; : 1.88 ± 0.0059; F = 11971.93, df measurement scale etched in a glass slide, was = 1, 198, p < 0.0001) and the ratio of the also used to measure body length and body combined length of protarsomeres 2-4 to the width. Tarsomeres were measured with a length of protarsomere 1 (: 1.14 ± 0.0015; Leica Wild M8 microscope with a 1.6X : 1.67 ± 0.0033; F = 21027.18, df = 1, 198, p objective adapter and a calibrated ocular < 0.0001) differed significantly by sex and micrometer. Digital images of specimens and there was complete separation (no overlap) for structures were captured using the Auto- males and females (Figure 8). The form of the Montage imaging system by Syncroscopy apex of the terminal sternite provided clear (Synoptics Inc., www.synoptics.co.uk). separation of the sexes. The terminal sternite Images were edited in Adobe Photoshop CS2 is always rounded posteriorly in females (Adobe Systems Inc., www.adobe.com) (Figure 6) and always quadrate in males (background removed, contrast manipulated). (Figure 5) based on our observations of 100 males and 100 females. Although the apices Statistical Analysis. of the male protibial teeth can become more Significance of differences by sex in body rounded through wear over time, the more length and width, protarsomere 1 length, acute (males) versus the less acute (females) protarsomere width, protarsomere length : distinction was reliable in over 90% of the width ratio, and protarsomeres 2 – 4: cases for all three protibial teeth, with the protarsomere 1 ratio were tested by ANOVA apical tooth in males being the most (JMP 2007). consistently acute of the three teeth (Figures 3, 4, 9). Results

On average, females were significantly larger (based on body length and width) than males

Journal of Insect Science | www.insectscience.org 7 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson Discussion relative proportions of protarsomere 1 is also a reliable character for sex determination. In A. Three reliable characteristics that allow non- sinicus males, the length averages less than 2 destructive determination of males and times the width. In females, the length females of A. sinicus were found: 1) the form averages greater than 2.5 times the width. of the terminal sternite (Figures 5, 6); 2) the Protarsomere characteristics must all be length to width ratio of protarsomere 1 viewed ventrally; examination in dorsal view (Figures 3, 4, 8), and; 3) the ratio of the may lead to incorrect assessment of the length combined length of protarsomeres 2-4: length of tarsomeres. Protarsomere characteristics of protarsomere 1 (Figures 3, 4, 8). These may be useful in sex determination of some characteristics are most easily discerned in the other species of Adoretus, but are not broadly laboratory with chilled specimens and with applicable within the entire genus. the aid of a microscope. It is also possible to Characteristics of the protarsomeres have discern sex differences in the field with a hand some applicability to some other scarabaeids lens. For field application, it is easier to see (e.g., Japanese beetle, Popillia japonica, the leg characters than the terminal sternite USDA-APHIS 2004) but must be assessed on character. a species by species basis.

In A. sinicus males, the apex of the terminal Less reliable methods for separating the sexes sternite is quadrate (Figure 5), whereas the involved relative characters that required apex of the female terminal sternite is rounded comparison between known males and posteriorly (Figure 6). Cream-colored setae at females as well as characteristics that were the apex of the sternites may obscure this subject to abrasion and wearing. One of the region slightly, but the characteristics are still characteristics was the difference in the form discernable. This distinction was valid in all of the protibial teeth (acute in the males specimens examined. This characteristic is versus rounded in the females). Even useful in many groups of Scarabaeidae, and it restricting our observations to adults “aged” at is broadly applicable within the genus least two weeks, the protibial teeth of males Adoretus and other members of the tribe were more acute than those of females in over Adoretini. 90% of specimens examined. However, A. sinicus adults use the protibia for digging in The relative dimensions of the protarsomeres the soil, and the teeth are subject to wear. (Figures 3, 4, 8) in ventral view are also Thus, this is an unreliable characteristic for reliable for separating the sexes. This sex determination and does not provide 100% difference can be observed by comparing the accuracy. combined length of protarsomeres 2 to 4 to the length of protarsomere 1, or by comparing For the sexing that we have done in the the relative length and width of protarsomere laboratory, we typically look first at the 1. In A. sinicus males, the combined length of relative dimensions of protarsomere 1, and protarsomeres 2 - 4 is over 1.5 times the then confirm sexing based on this character length of protarsomere 1 (Figures 3, 8), with subsequent observations of the terminal whereas in females, the combined length of sternite and the protibial teeth. As one protarsomeres 2 to 4 is less than 1.2 times the develops familiarity with the differences, one length of protarsomere 1 (Figures 4, 8). The can recognize the differences in the relative

Journal of Insect Science | www.insectscience.org 8 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson dimensions of protarsomere 1 without having whereas in females the fifth protarsomere is to make any actual measurements. It’s helpful gracile and only slightly developed to have more than one characteristic to use to internomedially. This character was not reduce chance of incorrect decision-making useful, however, in separating males and based on assessment of only one character. females of A. sinicus. Other characters that may be useful in sex determination of other Other characters useful for determination of Adoretus species include the form of the the sexes in other Rutelinae are, for example, pygidium (short and oblique in the female; the form of the abdomen (males with sternites longer and more convex in the male), size of concave or planar; females with sternites the eyes (larger in the male than in the weakly convex), but this is not useful in the female), the length of the antennal club Adoretus species that we examined. In (longer in the male than in the female), and addition, characters that are useful in other form of the pro- and mesotarsal claws Adoretus species are not reliable for use in A. (unequally split in the male; equally split in sinicus. Those characters include: (1) the the female). These additional characteristics presence (female) or absence (male) of a should not be applied generally to all species longitudinal furrow on the disc of the of Adoretus. However, they may provide pygidium; (2) the sternites appearing slightly some utility for research on other species. convex (females) versus slightly concave (males); and (3) the 5th protarsomere similar in The genus Adoretus includes approximately width to other protarsomeres and armed with 460 species (Krajcik 2007), and its , less developed internomedial teeth or lacking biology, and evolutionary history are poorly internomedial teeth (female) versus the 5th known. Adoretus sinicus is not the only protarsomere being generally wider and better Adoretus species of biosecurity concern. The developed than other protarsomeres and genus Adoretus includes several pests and armed with well developed internomedial invasive species in addition to the Chinese teeth (males) (e.g., Figures 12, 13). rose beetle (Table 1): A. bicolor Brenske, A. caliginosus Burmeister, A compressus This latter characteristic is apparent in A. (Weber), A. hirsutus Ohaus, A. ranunculus tenuimaculatus. Adoretus sinicus was Burmeister, A. tenuimaculatus (Figure 11), originally referred to as A. tenuimaculatus in and A. versutus Harold. Species in the genus Hawaii. Some literature and databases still are distributed in the Palearctic, Afrotropical, confuse these species. The comparisons Indomalaysian, Australasian, and Oceanic between these species are useful to illustrate biogeographic regions. Based on limited differences (and similarities) between biological information, adults are herbivore Adoretus species. Comparisons are more generalists and are known to feed on a wide readily made between these two species variety of plants. Larvae are associated with because A. tenuimaculatus is one of the most roots and, depending on the species, feed on commonly encountered Adoretus in living or non-living plant tissues. Eggs and collections, thus it is a good species for larvae may be easily transported with comparison due to accessibility of specimens. cultivated plants in soil or roots (Ohaus 1935) In males of A. tenuimaculatus, the fifth which, combined with the generally broad protarsomere is slightly thickened and armed host ranges, are the basis for the biosecurity with an internomedial tooth (Figure 13), concern for this group in relation to movement

Journal of Insect Science | www.insectscience.org 9 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson of potted plants across geographic regions. identified to species. In comparison, all four This biosecurity concern makes improved morphotypes within the genus Maladera taxonomic knowledge of the group essential. (Coleoptera: Scarabaeidae: Melolonthinae; Adult species in the genus are externally another scarab genus that includes pests) were similar (about 10.0 mm in length, brownish identified to species. Clearly, additional with cream-colored scales) making systematics research is needed within the identification of Adoretini problematic. Other genus Adoretus, a group that includes many than scattered and older regional works (e.g, economically important species. Péringuey 1902: Arrow 1917; Baraud 1985), there are few resources for identification of The reliable, non-destructive means of A. species. Form of the male genitalia is sinicus sex determination reported here will probably the best method of identification be instrumental in research aimed at (e.g., A. sinicus versus A. tenuimaculatus developing improved integrated pest [Figures 10, 11]), and this necessitates management systems for A. sinicus as well as dissection of the male genitalia. Molecular for other Adoretus species. Non-destructive tools, however, are being developed to aid in means of sex determination of live specimens the identification of adults and larvae. DNA in will facilitate progress in research that seeks larval-adult species associations within scarab to develop sex-dependent detection, beetle communities from Nepal have been monitoring and control methods that make use examined (Ahrens et al. 2007). Based on of pheromones, mating, or reproductive about 1600 base pairs of mitochondrial COX1 parameters. Such tools are of critical and RRNL and 700 base pairs of nuclear 28S importance for managing existing pest rRNA, larval specimens could be associated populations as well as for countering new with 19 identified, known adult species invasions of Adoretus species that are likely to between 86.1% and 92.7% of the time occur given the magnitude of transport of (Ahrens et al. 2007). Nine morphotypes in the agricultural goods across geographic regions. sample were members of the genus Adoretus, but only two morphotypes (22%) could be Table 1. Adoretus species of biosecurity concern

Journal of Insect Science | www.insectscience.org 10 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson Acknowledgements treated plants. Journal of Economic Entomology 81(5): 1373-1376. We thank Charmaine D. Sylva and Janice T. Nagata (USDA-ARS-PBARC) for night-time Arita LH, Furutani SC, Fukuda MT, field collections of Chinese rose beetles and Nakayama TR. 1993. Feeding response of the Janice T. Nagata and M. Roberta St. Chinese rose beetle (Coleoptera: Ambrogio for permitting night-time beetle Scarabaeidae) to non-structural carbohydrate collection on their properties. We also thank in plants. Journal of Economic Entomology Charmaine D. Sylva for patience and 86: 1416-1419. perseverance in making observations and measurements of the 200 specimens Arita Tsutsumi LH, Furutani SC, Yoshimura examined. Additionally, we thank Brett C. ER, Sakai DT. 1995. Feeding responses of the Ratcliffe, Michael Reding, Lorna Arita- Chinese rose beetle, Adoretus sinicus Tsutsumi and an anonymous reviewer for Burmeister, to azadarachtin treated host constructive comments on earlier drafts of this plants. Journal of Hawaiian and Pacific manuscript. This article reports the results of Agriculture 6: 49-54. research only. Mention of a proprietary product does not constitute an endorsement or Arrow GJ. 1917. The Fauna of British India a recommendation by the USDA for its use. including Ceylon and Burma. Coleoptera USDA is an equal opportunity provider and Lamellicornia Part II. (Rutelinae, employer. Desmonychinae, and Euchirinae). Taylor and Francis. References Asquith A. 1995. Alien species and the Ahmed MA, Kumar ST, Dharmaraju E. 1978. extinction crisis of Hawaii's invertebrates. Cockchafer beetle, Adoretus bicolor Brenske Endangered Species UPDATE 12: 1-8. damaging grape vine. Indian Journal of Available online, Entomology 39(4): 389-390. http://www.hear.org/articles/asquith1995

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Figures 1-2. Habitus of Adoretus sinicus: 1) Male. 2) Female. Inset shows close-up of dorsal setae. High quality figures are available online.

Figures 3-4. Sexual characteristics associated with the protarsus of Adoretus sinicus: 3) Male. 4) Female. High quality figures are available online. Journal of Insect Science | www.insectscience.org 16 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson

Figures 5-6. Sexual characteristics associated with the terminal sternite (ventral view) of Adoretus sinicus: 5) Male. 6) Female. High quality figures are available online.

Figure 7. Comparative length and width of male and female adult Chinese rose beetles. Lower and upper boundaries of the boxes are 25th and 75th percentile values. Whiskers below and above the boxes are 10th and 90th percentile values. Black line is median value. Red line is average value. Outlier values are indicated by points outside the whiskers. High quality figures are available online. Journal of Insect Science | www.insectscience.org 17 Journal of Insect Science: Vol. 11 | Article 64 McQuate and Jameson

Figure 8. Comparative ratios of lengths of protarsomeres 2 - 4 Figure 9. Percentages of observations of the form of the tips of the combined divided by protarsomere 1 and protarsomere 1 length three external teeth of the protibia noted to be acute versus divided by width for male and female adult Chinese rose beetles. rounded for male and female adult Chinese rose beetles (based on Lower and upper boundaries of the boxes are 25th and 75th observations of 100 males and 100 females). High quality figures are percentile values. Whiskers below and above the boxes are 10th and available online. 90th percentile values. Black line is median value. Red line is average value. Outlier values are indicated by points outside the whiskers. High quality figures are available online.

Figures 10-13. Characteristic differences between A. sinicus and A. tenuimaculatus. 10) Male parameres of A. sinicus. 11) Male parameres of A. tenuimaculatus. 12-13) Protarsus, dorsal view: 12) A. sinicus, male (with acute internomedial tooth on fifth protarsomere). 13) A. tenuimaculatus, male (with rounded internomedial tooth on fifth protarsomere). High quality figures are available online. Journal of Insect Science | www.insectscience.org 18