Phylogenetic relationships of the subfamily Melolonthinae (Coleoptera, Scarabaeidae) Mª MILAGRO COCA-ABIA Insect Syst.Evol. Coca-Abia, M.: Phylogenetic relationships of the subfamily Melolonthinae (Coleoptera, Scarabaeidae). Insect Syst. Evol. 38: 447-472. Copenhagen, November, 2007. ISSN 1399-560X A cladistic analysis of the subfamily Melolonthinae was undertaken using 48 taxa and 47 external morphology and male and female genitalia characters. Serica brunna and Pachydema hirticollis were chosen as outgroups. The aim of the present study is to assess supraspecific relationships and to test the monophyly of Melolonthinae and included tribes, genera and sub- genera. A general parsimony heuristic search was used to find the most parsimonious trees. Successive character weighting was used to assess the internal consistency of the data. The robustness of the clades was assessed with Bootstrap and Bayesian phylogenetic analysis. Melolonthinae is found to be a non-monophyletic group, whereas the monophyly of Holotrichia, Trichesthes, Phyllophaga and its subgenera, is here questioned. Mª M. Coca-Abia, Centro de Investigaciones y Tecnologías Agroalimentarias (CITA). Unidad de Sanidad Vegetal. Apartado de Correos 727. 50080 Zaragoza, Spain. ([email protected]) Phone: 00 34 976 716323. Fax: 00 34 976 716335. Introduction Thus, in this point, Lacroix (1989, 1993, 2000) According to some of the current checklists classification is going to be followed. This author (Evans, 2003), the subfamily Melolonthinae in- includes seven tribes in Melolonthinae based on cludes around 800 genera and 12,000 species only morphological features shared by all of them. It is in the New World. This huge biodiversity, togeth- not possible to confirm that these characters are er with the fact that some species are crop pests, synapomorphies until a phylogenetic analysis be has led to attempt to classify this group (Lacroix, done, but this grouping (Lacroix 1989, 1993, 1989, 1993, 2000; Evans, 2003; Baraud, 1992). 2000) was established on the basis of morpholog- Other authors (Scholtz, 1990; Smith et al. 2006) ical criteria, which is the most consistent in a con- have carried out high-level scarabaeoid classifica- text where there has been no previous attempt to tion on base to phylogenetic structure. But, in spite reconstruct the phylogeny of this subfamily and of these efforts to classify the family Scarabaeo- there is no objective reference to propose a tribal idea based on the phylogenetic structure, the classification (Smith et al. 2006). monophyly of Melolonthinae and the internal rela- According to Lacroix (1989, 1993, 2000), Mel- tionships of the tribes included have not been olonthinae includes the tribes Diplotaxini, Enari- established reliably (Smith et al. 2006). ini, Leucopholini, Melolonthini, Pegylini, Rhizo- Because high-level phylogenetic analyses do not trogini and Schizonychini. However, the high tax- throw any light on the monophyly of Melolonth- onomic rank of Melolonthinae (subfamily or tribe) inae or on the phylogenetic relationships of its has long been in dispute. Also, some genera such tribes (Browne & Scholtz, 1998; Scholtz, 1990; as Holotrichia Hope, 1837 (Bunalski 1995; Itoh Smith et al. 2006) and other classifications are 1995, 2002) and Phyllophaga (Morón 1986), have arbitrary (Lacroix, 1989, 1993, 2000; Evans, 2003; been taxonomically well studied, but rarely sub- Baraud, 1992); any classification followed to start jected to a thorough (strict) cladistic analysis. a phylogenetic analysis will be reprehensible. The Melolonthini and Rhizotrogini are the most © Insect Systematics & Evolution (Group 2) 448 Coca-Abia, M. INSECT SYST. EVOL. 38:4 (2007) diverse tribes of Melolonthinae and are distributed The specimens are deposited at the Museo all over the world. They include many endemic Nacional de Ciencias Naturales, Madrid (Spain) genera at local level, such as Hypotrichia LeConte, (MNCN); National Museum of Natural History 1862 from Florida; Dinacoma Casey, 1889, Plec- (Smithsonian Institution), Washington D.C (USA) trodes Horn, 1867 and Parathyce Hardy, 1974 (USNM); American Museum of Natural History from western United States and Thyce LeConte, of New York (USA) (AMNY); Natural History 1856 and Hypothyce Howden, 1968 from southern Museum London (England) (BMNH); Linnean United States and northern México. But other gen- Society of London (England) (LSL); Humbold- era are distributed worldwide, such as Polyphylla Universität zu Berlin, Museum für Naturkunde Harris, 1841 from the Nearctic and Palaearctic Re- (Germany) (HUMN); Hungarian Natural History gions, Phyllophaga (s. lato) Harris, 1827, Tri- Museum Budapest (Hungary) (HNHM); Zoologi- chesthes Erichson, 1847 (Coca-Abia 2002) from cal Museum, University of Copenhagen (Denmark) the Nearctic and Neotropical Regions and Melo- (UZM); Agro-Food Institute Ottawa (Canada) lontha Fabricius, 1775 and Rhizotrogus Berthold, (AFI); Instituto Nacional de la Biodiversidad 1827 from the Paearctic Region. (Costa Rica) (INBIO) and Howden Collection LeConte (1856) and Bates (1888) provided ini- (HC). tial taxonomic treatments of Melolonthidae from Characters of the external morphology and male the United States and Central America, respective- and female genitalia of each species were used for ly. Other authors focused on Phyllophaga (Horn the analysis. For the study, mouthparts (mentum, 1887; Smith 1889; Luginbill & Painter 1953; maxillae, mandibles and labrum) were removed, Morón 1986; Coca-Abia et al 1993) and Poly- immersed in distilled water and studied under a phylla (Young 1988; Wailly 1993, 1997a,b, 1998, dissecting microscope and scanning electron 2000, 2001) from the United States, Mexico and microscope (SEM). Europe or from more restricted geographic areas A similar procedure was used to study the such as, Arizona (Sanderson 1958), Texas (Rein- endophallus and female genitalia but here the hard 1939, 1946, 1950), Florida (Woodruff & structures were also rinsed with hot 5% KOH solu- Beck 1989) or Nebraska (Ratcliffe 1991). None of tion, dehydrated through a progressive ethanol se- these papers analyzed the phylogenetic structure ries (70%, 90%, 95% and 100%), and finally of Melolonthinae nor did they intent to assess the mounted in Euparal ® for examination under dis- monophyly of included tribes, genera and subgen- secting microscope. The aedeagus and genital seg- era. ments were also examined immersed in distilled The aim of the present study is to assess su- water, and some of the aedeagus were studied praspecific relationships, identify monophyletic under the SEM. groups, and to test the monophyly of Melo- SEM services were provided by the AMRAY lonthinae and included tribes, genera and subgen- 1810 Scanning Electron Microscopy of the United era. States Department of Agriculture (USDA) at the A cladistic analysis of Melolonthinae could be- National Museum of Natural History (Smithso- come a basis for later studies but the phylogeny of nian Institution). the lowest taxonomic ranks is likely to change For phylogenetic analysis, a parsimony general with the addition of new species (e.g., from Phyl- heuristic search was run in PAUP 4.0 (Swofford lophaga (s. lato) and Holotrichia, especially if 2003) to find the most parsimonious trees. The these imply the addition of new characters states to maximum number of trees saved was set to 1000; the analysis. searches were run with the simple addition se- quence and, the tree bisection reconnection Material and methods branch-swapping algorithm with the Mulpars option on. Successive character weighting based Table 1 lists the species studied for each genus, on Consistence Index (CI) (Farris 1969; Carpenter and representative percentage (%) of the total 1988) - base weight 1000 - was used to assess the number of species, as well as the total number of internal consistency of the data. species included in the genus and the data source The robustness of the clades was assessed by (in parentheses). The specimens studied can be using the bootstrap method (Felsenstein 1985) obtained from the author on request. with a heuristic research of 100 replicates and INSECT SYST. EVOL. 38:4 (2007) Phylogeny of Melolonthinae 449 Table 1. Species studied for the phylogeny of the Melolonthinae. The total number of species included in the genus, the number of species studied and their percentage(%) of the total number of species for each genus is indicated. Genus (Subgenus) Species studied Total species (references) % total Diplotaxis D. connata Schaeffer, 1905 317 (Evans, 2003) 1,89 D. ingenua Fall, 1909 D. marginicollis Fall, 1909 D. muricata Schaeffer, 1907 D. sordida (Say, 1825) D. tristis Kirby, 1837 Pachrodema P. abnormis Moser, 1918 9 (Evans, 2003) 11,1 Apicencya A. calva (Waterhouse, 1882) 9 (Lacroix, 1993) 22,2 A. mucronata (Klug, 1833) Bisencya B. gutticollis (Waterhouse, 1882) 8 (Lacroix, 1993) 25 B. invulnerata (Waterhouse, 1882) Cherbezatina C. depressiuscula (Waterhouse, 1882) 32 (Lacroix, 1993) 6,25 C. latifrons (Waterhouse, 1882) Enaria E. marginata Waterhouse, 1882 37 (Lacroix, 1993) 2,70 Enthora E. polita Waterhouse, 1878 8 (Lacroix, 1993) 12,5 Eulephida E. mashona Arrow, 1902 22 (Lacroix, 2000) 4,54 Dinacoma D. caseyi Blaisdell, 1930 2 (Evans, 2003) 100 D. marginata (Casey, 1887) Hypothyce H. mixta Howden, 1968 2 (Evans, 2003) 100 H. osburni (Cartwright, 1967) Hypotrichia H. spissipes LeConte, 1861 1 (Evans, 2003) 100 Melolontha M. melolontha (L., 1758) 41 (Dalla Torre, 1912) 2,44 Parathyce P. bidentata (Fall, 1932) 6 (Evans, 2003) 100