Arachnologische Mitteilungen 54

Arboricaria barcoding 25 2007) and initial phylogenetic analysis using the neighbor joining and parsimony algorithms implemented in PHYLIP 3.5c (Felsenstein 1989). Some particularly short or redun- dant (identical) barcodes were removed from the analysis, to minimize the computational effort. The resulting dataset contained barcodes for 144 specimens of 12 Micaria species, including representatives from a wide range of morphologi- cally defined species groups. The final phylogenetic analysis was performed using phylogeny.fr (Dereeper et al. 2008), with twelve different workflows, using sequence alignment by MUSCLE version 3.8.31 (Edgar 2004) or ClustalW 2.1, cu- ration using Gblocks 0.91b (Castresana 2000) or by removing positions with gaps, and phylogenetic inference using the Maximum Likelihood approach implemented in PhyML 3.1 (Guindon & Gascuel 2003), the Neighbor Joining method of BioNJ 3.66 (Gascuel 1997) or the Maximum Parsimony algorithm of TNT 1.1 (Goloboff et al. 2008), using default settings. Bootstrap support was estimated for each of the tree building methods in combination with MUSCLE alignment and gap removal. The nucleotide substitution model for the maximum-likelihood analysis was the very general default Generalised Time-Reversible (GTR) model, with Gamma shape parameter 0.725. Phylogenetic trees were visualized and explored in iTOL v3 (Letunic & Bork 2016). All the conclusions discussed below are independent of the exact choice of sequences, alignment method and tree inference algorithm. No attempt was made to optimize the parameters of any of the methods or to optimize the alignments by manual editing. Also, the choice of tree building methods was dic- tated by a desire to cover a wide range of conceptually di- verse methods (including the neighbour-joining approach, which is not strictly a phylogenetic inference method), rather than trying to use a few theoretically preferred inference ap- proaches. Such an intentionally diversified strategy would be suboptimal in the context of a comprehensive phylogenetic analysis, where maximal resolution and careful assessment of the support of each node in the tree is the aim. It is, however, a suitable approach in the present analysis, which has a more focused ambition, namely to test if any of the methods tried would allow us to reject Platnick’s hypothesis that Arboricaria is nested within a paraphyletic Micaria sensu stricto. Results and discussion The two different alignment methods resulted in identical alignments, and results were independent of the treatment of gaps in the alignments. Overall, relationships among the Micaria species were very similar in all three tree building approaches. A summary of the preferred majority-rule con- sensus tree resulting from the phylogenetic analysis is shown in Fig. 1 (the full trees for all methods are included in the electronic supplementary files, including sequence accession numbers, branch lengths and bootstrap support information). While this tree, based exclusively on mitochondrial COI data for a limited sample of species, should not be considered as a strongly supported and reliable phylogeny of Micaria in ge- neral, it allows a clear answer to Platnick’s questions: while Micaria sensu lato is a consistently recovered monophyletic group, Arboricaria subopaca , as the representative member of Arboricaria (i.e., Wunderlich’s subopaca group), is never reco- vered as sister to the remaining Micaria species, and Mica- ria sensu stricto would be paraphyletic. More specifically, in all analyses that provided sufficient phylogenetic resolution A. subopaca was found to be more closely related to, e.g., M. aenea, M. longipes , M. alpina and the species of the pulica- ria species group than to the members of the dives or scenica groups. More diverse sequence data would be required to re- solve the exact relationships: bootstrap support for the exact placement of A. subopaca is low, and different methods place it closer to either M. aenea (as suggested already by Wunderlich (1980)) or to M. alpina/longipes , and the entire clade contai- ning these four species is nested within the pulicaria group in some of the analyses. The pulicaria group according to Wun- derlich (1980) includes the type species of Micaria ( M. ful- gens ). Obviously, no conclusion is possible regarding the mo- nophyly of Arboricaria , as only one species is represented in the analysis, but this monophyly has not been contentious in earlier discussions of the status of the genus (Platnick 2001, Wunderlich 2017) and is irrelevant for the question at hand. Confidence in the phylogenetic results is provided not only by the stability of these findings towards the choice of analytical methodology, but also by the fact that all indivi- dual species represented by more than one specimen are ro- bustly monophyletic (with bootstrap support between 66 and 100 %). A single exception is the closely related species pair Fig. 1: Preferred phylogeny of barcoded Micaria species, based on a majority-rule consensus of analyses in phylogeny.fr, using three different phyloge- netic inference algorithms (BioNJ, PhyML and TNT). The bootstrap support for each clade in each of the analyses is indicated above the branches (–: clade not recovered in this analysis, +: clade not consistently resolved in this analysis). A set of gnaphosid species from 14 genera was used as outgroup to root the tree. The number of sequences (n) included in the analysis is indicated for each species.