A peer-reviewed open-access journal ZooKeys 147: 229-260 (2011) doi: 10.3897/zookeys. 147.187 | #7Z,00Ke y S www.zookeys.o rg Launched to accelerate biodiversity research Phylogeny of minute carabid beetles and their relatives based upon DNA sequence data (Coleoptera, Carabidae, Trechitae) David R. Maddison!', Karen A. Ober? | Department of Entomology, University of Arizona, Tucson, AZ 85721, USA; Current address: Department of Zoology, 3029 Cordley Hall, Oregon State University, Corvallis, OR 97331, USA 2. Department of Ento- mology, University of Arizona, Tucson, AZ 85721, USA; Current Address: Department of Biology, College of the Holy Cross, Worcester, MA 01610, USA Corresponding author: David R. Maddison (david.maddison@science.oregonstate.edu) Academic editor: 7 Erwin | Received 30 July 2011 | Accepted 25 August 2011 | Published 16 November 2011 Citation: Maddison DR, Ober KA (2011) Phylogeny of minute carabid beetles and their relatives based upon DNA sequence data (Coleoptera, Carabidae, Trechitae). In: Erwin T (Ed) Proceedings of a symposium honoring the careers of Ross and Joyce Bell and their contributions to scientific work. Burlington, Vermont, 12-15 June 2010. ZooKeys 147: 229-260. doi: 10.3897/zookeys.147.187 1 Abstract The phylogeny of ground beetles of supertribe Trechitae is inferred using DNA sequences of genes that code for 28S ribosomal RNA, 18S ribosomal RNA, and wingless. Within the outgroups, austral psydrines are inferred to be monophyletic, and separate from the three genera of true Psydrina (Psydrus, Nomius, Laccocenus); the austral psydrines are formally removed from Psydrini and are treated herein as their own tribe, Moriomorphini Sloane. All three genes place Gehringia with Psydrina. Trechitae is inferred to be monophyletic, and sister to Patrobini. Within trechites, evidence is presented that Zasmanitachoides is not a tachyine, but is instead a mem- ber of Trechini. Perileptus is a member of subtribe Trechodina. Against Erwin’s hypothesis of anillines as a polyphyletic lineage derived from the tachyine genus Paratachys, the anillines sampled are monophyletic, and not related to Paratachys. Zolini, Pogonini, Tachyina, and Xystosomina are all monophyletic, with the latter two being sister groups. The relationships of the subtribe Bembidiina were studied in greater detail. Phrypeus is only distantly related to Bembidion, and there is no evidence from sequence data that it belongs within Bembidiina. Three groups that have been recently considered to be outside of the large genus Bembidion are shown to be derived members of Bembidion, related to subgroups: Ci/enus is related to the Ocydromus complex of Bembidion, Zecillenus is related to the New Zealand subgenus Zeplataphus, and Hydrium is close to subgenus Metallina. The relationships among major lineages of Trechitae are not, however, resolved with these data. Copyright David R. Maddison, Karen A. Ober. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 230 David R. Maddison & Karen A. Ober / ZooKeys 147: 229-260 (2011) Keywords ground beetles, DNA, molecular phylogeny, Bembidiini, Trechinae, Carabidae, Trechitae Introduction The supertribe Trechitae comprises over 5,300 described species (Lorenz 2005) of ground beetles. Although this is comparable to the number of mammal species (Wil- son and Reeder 2005), trechites are much more poorly known. Trechites are diverse on all continents except Antarctica. Most adults of this group are relatively small (less than 10 mm in length), and include the smallest known carabids, about 0.7 mm in length (Erwin 1973; Jeannel 1963). Division of this group into suprageneric taxa var- ies among authors, with most North American authors favoring four tribes: Trechini and Bembidiini, with over 2,500 species each, and the smaller groups Pogonini and Zolini, with about 85 and 55 species respectively (Lorenz 2005). Trechini includes many troglobitic species, and is most speciose in temperate areas. Bembidiini is world- wide, with many species living along bodies of water; it includes the smallest adults. This tribe includes the largest carabid genus, Bembidion, with over 1,200 recognized species (Lorenz 2005). Most pogonines are halobiontic; the majority live in the Old World. The Zolini is a strictly south-temperate lineage, except for the monotypic genus Sinozolus from China (Deuve 1997). A brief review of the diversity within each tribe is given in Grebennikov and Maddison (2005). The basic structure of the phylogeny of trechites is not well known. The only explicit, modern analyses have been based upon limited characters of adult and larval structure (Grebennikov 2008; Grebennikov and Maddison 2005; Roig-Jufent and Cicchino 2001); these have inferred a few aspects of the phylogeny (Fig. 1). We present here the first detailed examination of relationships within Trechitae based upon DNA sequences, using portions of genes for small (18S) and large (28S) subunits of ribosomal RNA, as well as the nuclear protein-coding gene wingless. This paper has been over a decade in gestation, and some results have already been reported in other publications. For example, the discovery from the DNA sequence data reported herein of Tasmanitachoides place in Trechini rather than in Tachyina was the inspiration for Grebennikov’s search for Tasmanitachoides larvae, which as he recently reported (Grebennikov 2008) confirms its placement in Trechini. Methods Taxa examined. The 14 outgroup species included are listed in Table 1. Morpho- logical data and previously collected 18S rDNA data suggests that the sister group of Trechitae is likely Patrobini (Arndt 1993; Deuve 1993; Maddison et al. 1999a; Miiller 1975; Zamotajlov 2002), and we include three species of this near outgroup. More Phylogeny of Trechitae... 231 Trechodina, including q Trechina Trechina a Tasmanitachoides : Trechodina, including Trechina Tasmanitachoides Xystosomina Xystosomina Tachyina Tachyina Pogonini Pogonini Zolini Zolini Anillina Anillina Phrypeus Phrypeus A Sinechostictus B Sinechostictus : Asaphidion : Asaphidion Grebennikov (2008) =| | Bembidion Grebennikov (2008) +; Bembidion larval characters larval characters analysis 1: all characters unordered analysis 2: characters unordered & ordered Xystosomina Tachyina Bembidiina Roig-Junent + Cicchino (2001) adult characters Figure |. Phylogenies of Trechitae from morphological studies A Strict consensus tree of most parsimo- nious trees from larval data, with all characters treated as unordered, from Grebennikov (2008); this is the tree presented in Grebennikov (2008:Fig. 3) B Strict consensus tree of most parsimonious trees from larval data, with some characters treated as ordered, as specified by Grebennikov (2008); this tree is not presented in that paper, but was inferred from the described conditions C “Best fit” tree presented by Roig-Jufent and Cicchino (2001) based upon adult morphological data. distant relatives are less clear. We include representatives of carabid groups that are of a similar grade as trechites, that is, they are members of Carabidae Conjunctae but are not members of Harpalinae or Brachininae. These include all three genera of Psydrini (s.str.), as well as six genera of Moriomorphini. ‘The latter is the group referred to as “austral psydrines” in Maddison et al. (1999a), and includes all traditional psydrines except for Psydrini in the strict sense. In addition, as Gehringiini has been proposed to be a psydrine relative (Erwin 1985), or potentially within Trechitae (Erwin 1984), we 2352 David R. Maddison & Karen A. Ober / ZooKeys 147: 229-260 (2011) Table |. Outgroup taxon sampling. Four-digit numbers in entries are D.R. Maddison voucher numbers; further information on the specimens is given in the Appendix; where two numbers are listed, the sequence was formed by combining data from both specimens. Other entries are GenBank numbers of previously published sequences from Maddison et al. (1999a, 1999b), Ober (2002), and Ober and Maddison (2008). 18S wingless Pterostichini Pterostichus melanarius Mliger AF002779 AF398707 AF398623 Moriomorphini Amblytelus curtus (Fabricius) AF012484 AF398566 Mecyclothorax vulcanus (Blackburn) AF012482 AF398648 AF398601 Melisodera picipennis Westwood AF012481 AF398640 AF398602 Meonis sp. AF398722 AF398692 AF398603 Sitaphe parallelipennis Baehr 0669 0669 Tropopterus sp. AF012483 2200 Psydrini — Laccocenus ambiguus Sloane AF012486 AF398675 AF398596 Psydrus piceus LeConte AF002784 AF398684 L627, Nomius pygmaeus (Dejean) 0893 AF438100 AF437971 Gehringiini Gehringia olympica Darlington AF012512 AF398702 AF398591 Patrobin —— Diplous californicus (Motschulsky) AF002785 AF398699 AF398587 Patrobus longicornis (Say) AF002786 AF398700 AF398613 Penetretus temporalis Bedel 0631,1710 0631 include one member of Gehringiini, Gehringia olympica Darlington. We also include one representative of Harpalinae, Pterostichus, as the most-distant outgroup. Within Trechitae, 64 species in 40 genera are sampled, with all tribes represented, and an emphasis on subtribe Bembidiina (Table 2). The classification used here is modified version of Lorenz's (2005), with ranks similar to those typically used in North America (e.g., Lindroth 1963). The sequences obtained for Trechus came from two dif- ferent specimens from Montana; one of these is Zrechus oregonensis Hatch, the other, a female, cannot be identified with certainty to species, but belongs to the 7’ chalybeus species group, to which 7’ oregonensis also belongs. In analyses combining different genes, sequences from these two individuals were combined into a chimeric taxon. Locality information for the specimens newly sequenced in this paper is given in the Appendix. Voucher specimens are deposited in the David Maddison voucher col- lection in the Oregon State Arthropod Collection at Oregon State University. DNA sequencing. Methods for obtaining DNA sequences, including extraction methods and cycling reactions, are described in Maddison et al. (2008). Primers used are listed in Table 3; see Maddison et al. (2008) for information about original source of primer sequences. In brief, we obtained ca. 2000 bases of sequence data of 18S ribo- somal DNA (18S rDNA or 18S), about 1000 bases in the D1 through D3 domains of Phylogeny of Trechitae... 233 Table 2. Taxon sampling of trechites. Four-digit numbers in entries are D.R. Maddison voucher num- bers; further information on the specimens is given in the Appendix; where two numbers are listed, the sequence was formed by combining data from both specimens. Other entries are GenBank numbers of previously published sequences from Maddison et al. (1999a, 1999b), Ober (2002), and Ober and Mad- dison (2008). iss [288 | wingless Trechini: Trechodina Cnides sp. 1808 0691 Pachydesus sp. 0678 AF438112 AF437978 Perileptus areolatus (Creutzer) 1707 0824 1707 Perileptus sloanei Moore 0767 Thalassophilus longicornis (Sturm) 0823 0823 Trechodes bipartitus (MacLeay) 0705 0705 0705 Trechodes jeanneli jeanneli Mateu 0606 Trechosiella sp. 1709 0723 1709 Tasmanitachoides fitzroyi (Darlington) 1575 0762 Trechini: Trechina Trechus chalybeus species group AF002793 Trechus oregonensis Hatch AF398673 0587 Omalodera limbata Blanchard 0571 0571 0571 Homaloderodes germaini Jeannel 1066 Kenodactylus audouini (Guérin-Méneville) 0670 0670 Paratrechus sp. 1076 Trechinotus flavocinctus Jeannel 0575 0575 0575 Zolini Merizodus angusticollis Solier AF012487 0453 0453 Oopterus sp. AF012488 0387 0387 Oopterus helmsi (Sharp) AF002787 0354 0354 Sloaneana tasmaniae (Sloane) AF002788 0339 0339 Pogonini Diplochaetus planatus (G.H. Horn) AF002789 AF438060 AF437938 Pogonus (Pogonus) chalceus (Marsham) 1711 0679 0679 Thalassotrechus barbarae (G.H.Horn) 0703 0530 Bembidiini: Tachyina Lymnastis sp. 0988 0988 0988 Micratopus sp. 0605 0605 Paratachys vorax (LeConte) 0410 0410 Elaphropus obesulus LeConte 0411 0411 0411 Elaphropus cf. nigrolimbatus Peringuey 0761 Elaphropus sp. 3 0713 0713 Pericompsus laetulus LeConte AF002790 0429 0429 Polyderis rufotestacea (Hayward) 0717, 0718 0718 Tachys vittiger LeConte 0760 Tachys corax LeConte 0604 0604 Tachyta nana inornata (Say) 0573 AF438141 AF438002 234 David R. Maddison & Karen A. Ober / ZooKeys 147: 229-260 (2011) iss [288 | winless Bembidiini: Xystosomina Erwiniana hilaris (Bates) AF012489 0409 0409 Erwiniana crassa (Erwin) 0989 0989 Mioptachys flavicauda Say 0684 0684 0684 Philipis bicolor Baehr 0592 0592 Bembidiini: Anillina Anillinus (langdoni group) sp. 0690 0690 Serranillus sp. 1084 1084 Typhlocharis armata Coiffait 0572,1718 0572 0572 Nesamblyops sp. 0696 Bembidiini: Bembidiina Asaphidion alaskanum Wickham 0585 0585 Asaphidion championi Andrewes 0574 Asaphidion curtum (Heyden) AF002792 0267 0267 Amerizus (Amerizus) sp. 0576 0576 Ocys harpaloides (Audinet-Serville) 0569 0569 Phrypeus rickseckeri Hayward 0776 0692 Sinechostichus solarii (G. Miiller) 0603 0603 Bembidion (Antiperyphanes) sp. nz. chilense 0714 0714 Solier Bembidion (Hoquedela) cf. csikii Jedlicka 0916 0916 Bembidion (Cillenus) laterale (Samouelle) 0602 0602 Bembidion (Notaphus) insulatum (LeConte) 0444 0444 Bembidion (Bracteon) balli Lindroth EF648613 EF648838 EF649474 Bembidion (Odontium) coxendix Say EF648618 EF648837 EF649481 Bembidion (Metallina) dyschirinum LeConte 0896 0896 Bembidion (Pseudoperyphus) integrum Casey | EF648659 EF649056 EF649609 Bembidion (Bracteon) levettei carrianum Casey EF648620 EF648842 EF649480 Bembidion (Hydrium) levigatum Say 0763 0763 Bembidion (Ocydromus) mexicanum Dejean | AF012490 0260 0262 Bembidion (Phyla) obtusum Audinet-Serville 0895 0895 Bembidion (Melomalus) planatum (LeConte) 0601 0601 Bembidion (Bembidion) quadrimaculatum 0676 0676 0676 dubitans (LeConte) Bembidion (Zeplataphus) tairuense Bates 0607 0607 0607 Bembidion (Zecillenus) sp. 0595 0595 0595 28S ribosomal DNA (28S rDNA, or 28S) and about 450 bases of the nuclear protein- coding gene wingless (wg). Amplified products were cleaned, quantified, and sequenced at the University of Arizona's Genomic and Technology Core Facility using either a 3730 or 3730 XL Applied Biosystems automatic sequencer. Assembly of multiple chromatograms for each gene fragment and initial base calls were made with Sequencher (Gene Codes Corporation) or using Phred (Green and Ewing 2002) and Phrap (Green 1999) as orchestrated by Mesquite’s Chromaseq pack- Phylogeny of Trechitae... 235 Table 3. Primers used for DNA amplification and sequencing. Dir: direction of primer, either forward (F) or reverse (R). Syn: primer synonym. Kind: primer used for original PCR amplification and sequenc- ing (A) or primer used only for sequencing (S). Original references for primer sequences are given in Mad- dison et al. (2008). Primer pairs used in earlier PCR reactions for wingless were 5wg-3weg, SwgB-3weg2, and B5wg1-B3weg2; more recent, and reliable, reactions used the pairs wg550F-wgABRz or wg578F-wgABR. Gene Sequence GGGAGGAAAAGAAACTAAC LS998R_ {D3 |R |A |GCATAGTTCACCATCTTTC 18S TATGCTTGTCTCAAAGATTAA CACCYACGGAAACCTTGTTACGACTT SS398F ie IBS ) CCTGAGAAACGGCTACCACATC SS1054F F S ATCAAGAACGAAAGT SS1090R i, [IR 4S GAGTCTCGTTCGTTATCGGA GTCCTGTECCATTATICCAT |e ess ATGCGTCAGGARTGYAARTGYCAYGGYATGTC R |A CACTTNACYTCRCARCACCARTG Fa TGCACNGTGAARACYTGCTGGATG lweAbR | ~=|R |A |YTCGCAGCACCARTGGAA P5wgl | JF JA GARTGYAAGTGTCAYGGYATGTCTGG GARTGYAARTCYCAYGGYATGTCTGG = ACBTGYTGGATGCGNCTKCC 3we2 CTCGCARCACCARTGGAATGTRCA Pima | Ik a ACTCGCARCACCAGTGGAATGTRCA awe | [R_ |A _ |ACTCGCARCACCARTGGAATGTRCA WE age (Maddison and Maddison 2009a; Maddison and Maddison 2009b), with subse- quent modifications by Chromaseq and manual inspection. Multiple peaks at a single position in both reads were coded using IUPAC ambiguity codes. Newly obtained sequences have been deposited in GenBank with accession num- bers GU556024 through GU556153. Alignment and resulting matrices. The two ribosomal genes, 18S and 28S, were aligned using ClustalW 1.8.3, with a gap opening cost of 10, gap extension of 0.1, then adjusted by eye; areas of uncertain alignment were excluded. The amino acid translation of the wingless gene was aligned using Clustal W version 1.83 (Chenna et al. 2003) using gap opening cost of 5, gap extension cost 0.2, and a Gonnet series matrix. The central region of the wingless alignment evidently had a rich his- tory of insertion and deletions; the alignment of this region was adjusted by eye in Mac- Clade (Maddison and Maddison 2005). An alignment of nucleotides was then created, with the nucleotides forced to match the amino acid alignment using MacClade. There were two wingless matrices produced, one with the alignment-ambiguous region included (“all nucleotides”), and another with that region excluded (“well-aligned nucleotides”). Phylogenetic inference. Each of the four matrices (28S, 18S, and the two wingless matrices) were subjected to parsimony, Bayesian, and maximum likelihood analyses. 236 David R. Maddison & Karen A. Ober / ZooKeys 147: 229-260 (2011) Most-parsimonious trees were sought using PAUP* (Swofford 2002). For each search, 2000 replicates were conducted, each beginning with a starting tree formed by the random addition sequence option, with subsequent TBR branch rearrangement. The number of most parsimonious trees (MPTs) ranged from 15 to 502 across the four matrices, and for each matrix the MPTs were found in at least 460 of the 2000 replicates. For parsimony bootstrap analyses in PAUP*, 1000 bootstrap replicates were con- ducted, each of which used a heuristic search with five replicates, each beginning with a starting tree formed by the random addition sequence option, with TBR branch rearrangement, with each replicate saving no more than 25 trees. Models of nucleotide evolution chosen with the aid of ModelTest (Posada 2005), with the aid of PAUP* (Swofford 2002). For 18S and 28S genes, the model chosen by the Akaike Information Criterion (AIC) was a General Time Reversible rate ma- trix with a proportion of sites being invariant and the remainder following a gamma distribution (the GTR+I+I° model). For the wingless gene, the GTR+I+I° model was chosen for the region without extensive insertions and deletions, but for the indel-rich region a GIR+I° model was preferred. When codon positions were allowed separate models, GTR+I+I was preferred for first positions, GTR+I for second positions, and HKY85+I+I for third positions. Bayesian analyses were conducted using MrBayes (Huelsenbeck and Ronquist 2005). Two runs of four chains each were run for between 8 million and 30 million generations, with trees sampled every 1,000 generations. Runs were terminated once the average standard deviation of split frequencies went below 0.01 (Huelsenbeck and Ronquist 2005). For each analysis, the trees in a burn-in period were excluded, and the majority-rule consensus tree of remaining trees was calculated by PAUP to determine Bayesian Posterior Probabilities (BPP) of clades. The burn-in period was at least 25% of the total length of the run (as only the remaining 75% were used to calculate the average standard deviation of split frequencies used as a convergence diagnostic), and extended until the likelihood scores and all parameter values reached a stable plateau, as judged by visualization tools in Tracer (Rambaut and Drummond 2004). The burn- in period ranged from 3 million generations to 25 million generations. The number of trees sampled for each analysis varied from 10,000 to 30,000. Likelihood analyses of nucleotide data were conducted using RAxML version 7.0.4 (Stamatakis 2006). For each matrix, 1000 search replicates were conducted to find the maximum likelihood trees. 2000 non-parametric bootstrap replicates were used to calculate bootstrap values for groups of interest. Several of the analyses of 18S rDNA yielded trees with the trechine Cvides, whose terminal branch was extremely long, well outside of Trechitae. As morphological data indicates definitively that Cvides is a trechite, some analyses were performed that forced it to reside within Trechitae. Two full suites of constrained analyses were conducted, one with Trechitae constrained to be monophyletic, and other with Trechini constrained to be monophyletic. For the latter, the position of 7asmanitachoides was not constrained in likelihood and parsimony analyses, allowing it to move anywhere on the tree. All trees are shown rooted within the outgroup, arbitrarily next to Pterostichus. Phylogeny of Trechitae... OS7 Grebennikov’s (2008) larval morphological data were reanalyzed using TNT ver- sion 1.1 (Goloboff et al. 2008). Most parsimonious trees were found using the follow- ing commands: rseed[, hold 1000, xmult: hits 100 ratchet 5 norss nocss, xmult. This caused TNT to do multiple searches, each beginning with a tree with taxa added in random order, with up to 1000 trees held in memory, with each search using five cycles of ratcheting; enough searches were done until the best trees found were found 100 times. 332 equally parsimonious trees of length 140 were found. Results of phylogenetic analysis Phylogenetic trees inferred from individual genes are shown in Figs 2-4, and from the merged matrix in Fig. 5. Support values for various hypotheses are shown in Tables 4 and 5. Summaries of supported phylogenetic hypotheses are presented in Figs 6—7. Support values for various groups in analyses of 18S constrained to keep Trechitae or Trechini monophyletic are shown in Table 6. Results for the reanalysis of Grebennikov’s (2008) larval data are presented in Fig. 1B, and described in more detail in the text, below. Discussion We here discuss in turn the evidence available for various relationships within trechites. As significant results have been found within the outgroups sampled, we will discuss these first. Outgroup structure: Monophyly of Moriomorphini Among the carabids currently considered to belong to Psydrini (sens. /at.), only three genera belong to Psydrina in the strict sense: Psydrus (North America), Nomius (Hol- arctic and Africa), and Laccocenus (Australia). The remaining genera are arrayed in multiple subtribes (Baehr 1998; Moore 1963), and are restricted to the Southern Hemisphere, primarily in temperate areas, except for members of the genus Mecyclo- thorax, which occur as far north as Hawai’i (Liebherr 2006; 2008; 2009). Sequences of 18S rDNA indicate that all psydrines other than Psydrus, Nomius, and Laccocenus belong to a clade, termed the “austral psydrines” by Maddison et al. (1999a). Three apomorphies of adult structure (Baehr 1999) also indicate monophyly of the austral psydrines. In addition, 18S rDNA indicated that austral psydrines were not closely related to psydrines in the strict sense (Maddison, et al. 1999a). Our data indicate that Psydrina are not closely related to austral psydrines. While a strong test of this hypothesis with 28S and wingless would require more extensive sampling of non-trechites than we have done, all three genes we studied suggest that Psydrini in the classical sense, containing Psydrus, Nomius, Laccocenus, and the austral 238 David R. Maddison & Karen A. Ober / ZooKeys 147: 229-260 (2011) - Pterostichus r Me onts 4 ee elisodera aa 2 Sitaphe LJ Mecyclothorax —- Amblytelus Tropopterus Laccocenus Psydrus Nomius Diplous — Patrobus Penetretus Phrypeus Sinechostichus os Bembidion dyschirinum - Bembidion levigatum Bembidion insulatum — Bembidion quadrimaculatum ™— Bembidion sp. nr. chilense -- Bembidion tairuense Bembidion (Zecillenus) a - Bembidion cf. csikii Bembidion obtusum - g Bembidion laterale — Bembidion mexicanum Bembidion coxendix - Bembidion levettei Bembidion integrum Bembidion planatum Bembidion balli Ocys | | Asaphidion alaskanum Gehringia 28S rDNA Mm BPP mam BPP | BPP === BPP Asaphidion championi BPP Asaphidion curtum ed Soe loaneana zz . Oopterus sp. | Merizodus - Oopterus helmsi Nesamblyops a ee ocnats nillinus = Serranillus Diplochaetus a — Pogonus Thalassotrechus Trechinotus = | -— Kenodactylus = Homaloderodes Omalodera Paratrechus Trechus Pachydesus Thalassophilus _————— Perileptus areolatus = Perileptus sloanei Trechosiella - Trechodes bipartitus — Trechodes jeanneli Philipis = Mioptachys a -Erwiniana crassa - Erwiniana hilaris Lymnastis Micratopus Paratachys | Pericompsus Polyderis Tachys corax a Tachys vittiger Tachyta Elaphropus obesulus 0.1 a F Elaphropus cf. nigrolimbatus : Elaphropus sp. 3 Tasmanitachoides 90, MLBoot 90, MLBoot 90, MLBoot 75, MLBoot 50 90, parsBoot 90 75, parsBoot 50 50, in MPTs 50 or in MLTs, in MPTs Cnides Figure 2. Majority-rule consensus tree of trees sampled in Bayesian analysis, with branch lengths propor- tional to average branch lengths across trees that contain that branch, for 28S rDNA data. Branch lengths were reconstructed by MrBayes; scale bar units are substitutions per site. Thickness and shade of branches indicate support for that clade, based upon estimated Bayesian Posterior Probability percentages (BPP), Maximum Likelihood bootstrap values (MLBoot), and parsimony bootstrap values (parsBoot). Phylogeny of Trechitae... 239 - Pterostichus Cnides | - Meonis (0028200 Fr Mecyclothorax | '—— Tropopterus Melisodera — Sitaphe - Gehringia - Laccocenus - Nomius ——— _ Psydrus ——— Diplous - Patrobus Penetretus Omalodera L__| a - Trechinotus L an Trechus | — Pachydesus - Perileptus areolatus | -—— — Tasmanitachoides Trechodes bipartitus - Trechosiella - Serranillus (|__ Diplochaetus - Pogonus Typhlocharis | i. — Asaphidion curtum | - Bembidion mexicanum - Bembidion tairuense Bs — Bembidion quadrimaculatum 18S rDNA — Bembidion (Zecillenus) P Bembicion-Dall MME BPP 90, MLBoot 90, parsBoot 90 | Bembidion levettei r— Bembidion coxendix mmm BPP 90, MLBoot 75, parsBoot 50 ——~ Bembidion integrum mm BPP 90, MLBoot 50, in MPTs * Elaphropus obesulus Pcie a ii m= BPP 75, MLBoot 50 or in MLTs, in MPTs - Pericompsus " BPP 50 Lymnastis ——- Erwiniana hilaris - Mioptachys - Sloaneana Ss —— verizodus = Oopterus sp. ——— Oopterus helmsi 0.1 Figure 3. Majority-rule consensus tree of trees sampled in Bayesian analysis, with branch lengths pro- portional to average branch lengths across trees that contain that branch, for 18S rDNA data. See caption of Fig. 2 for additional details. psydrines, is not monophyletic (Figs 2-5). Combined with evidence provided by the more extensive 18S rDNA taxon sampling of Maddison et al. (1999a), and the mor- phological data (Baehr 1999), these two groups should be in separate taxa. We there- fore remove austral psydrines from Psydrini, and place them in their own tribe. The valid name for this tribe is Moriomorphini Sloane (1890: 646). Our results indicate strong support for monophyly of Moriomorphini from both 18S rDNA and 28S rDNA (Table 4). Most analyses of the wingless gene suggest in- stead that the moriomorphines form a grade, although parsimony analysis of the well- aligned nucleotides does support monophyly of the group. More extensive sampling of wingless sequences of non-trechites is needed to examine this further. 240 Pterostichus - Sitaphe - Melisodera > Amblytelus | /=—— Tropopterus _ ' - Mecyclothorax | —— Meonis BPP - Psydrus BPP ill L Laccocenus = Gehringia BPP BPP BPP - Diplous Patrobus - Penetretus Anillinus - Typhlocharis r Trechus - Omalodera rt - Kenodactylus : Trechinotus - Thalassophilus Pachydesus - Philipis ———— Mioptachys - Erwiniana crassa Erwiniana hilaris — Lymnastis Elaphropus obesulus - Elaphropus sp. 3 Paratachys Tachyta Micratopus 0.1 — Pericompsus Polyderis Tachys corax - Sloaneana Manzagus ~~, _Copterus sp. : - Oopterus helmsi — Phrypeus —— Diplochaetus ee Pogonus Thalassotrechus - Amerizus Ocys l Sinechostichus | »—— Asaphidion alaskanum '— Asaphidion curtum - Bembidion obtusum a r Bembidion insulatum __ - Bembidion quadrimaculatum p=; Bembidion sp. nr. chilense | (ggg Bembidion tairuense — Bembidion (Zecillenus) -- Bembidion cf. csikii | gay. Bembidion dyschirinum - Bembidion levigatum - Bembidion planatum ™ —— Bembidion laterale -— Bembidion mexicanum - Bembidion levettei 1— Bembidion balli — - Bembidion coxendix —— Bembidion integrum = — Trechosiella David R. Maddison & Karen A. Ober / ZooKeys 147: 229-260 (2011) wingless 90, MLBoot 90, MLBoot 90, MLBoot 75, MLBoot 50 90, parsBoot 90 75, parsBoot 50 50, in MPTs 50 or in MLTs, in MPTs — Perileptus areolatus — Trechodes bipartitus Figure 4. Majority-rule consensus tree of trees sampled in Bayesian analysis, with branch lengths propor- tional to average branch lengths across trees that contain that branch, for the complete wingless data. See caption of Fig. 2 for additional details. Outgroup structure: relationship of Gehringiini Gehringiini are a small group (five known species; Baehr et al. 2009) of minute car- abids of uncertain relationships. Morphological data places them variously as sister group of Paussinae (R.T. Bell at the 1983 Entomological Society of America meetings; Beutel 1992), with Trachypachidae (Kryzhanovskij 1976; Lindroth 1969), among the basal carabids (Bell 1967; Darlington 1933; Jeannel 1941), or a member of Trechitae or a relative of Psydrini (Bell 1967; Darlington 1933; Erwin 1984; 1985; Hammond Phylogeny of Trechitae... 241 Pterostichus Meonis Melisodera Sitaphe Tropopterus Amblytelus Mecyclothorax EACeOCe nus enringla Psydrini+ Gehringi Psydrus syd Gehringia Nomius 08 pipes Patrobini atrobus Int eH Penetretus Nesamblyops UD Typhlocharis i: 008 | 559. — fastiiene Anillina rolled 4 lomaloderodes OD a omaledss j enodactylus : O06 Fechinoths Trechina Oo Paratrechus Trechus Cnides Pachydesus 2 Thalassophilus Trechodina Perileptus areolatus ‘ : Dae 606 Porlleptus sean (with Cnides and anD Tasmanitachoides Tasmanitachoides) ’ Trechosiella CD a Trechodes bipartitus Trechodes jeanneli Diplochaetus