DOI: 10.25338/b8490t
title: Delimitation of tribes in the subfamily Leptanillinae (Hymenoptera: Formicidae), with a description of the male of Protanilla lini Terayama, 2009

description: The subfamily Leptanillinae Emery, 1910 (Hymenoptera, Formicidae) is a
 clade of cryptic subterranean ants, which is restricted to the tropics and
 warm temperate regions of the Old World. Due to acquisition bias against
 the minute and hypogaeic workers, most known leptanilline specimens are
 male, with four genera described solely from males. The sexes have been
 associated in only two out of 68 described species, meaning that redundant
 naming of taxa is likely. Herein the phylogeny of the Leptanillinae,
 sampled with emphasis on largely undescribed male material, is inferred
 from ultra-conserved elements (UCEs) using maximum-likelihood inference.
 This method associates the male of Protanilla lini Terayama, 2009 with
 corresponding workers collected on Okinawa-jima, Japan, allowing the first
 published description of male ants belonging to the Anomalomyrmini Taylor,
 1990, one of the two established tribes within the Leptanillinae. The
 first male-based diagnosis of these tribes is provided, along with a
 dichotomous key to both all described male-based species within the
 Leptanillinae and male morphospecies sequenced in this study. With
 genome-scale data enabling the association of separately collected sexes
 and phylogenomic inference contextualizing morphological observations, the
 parallel taxonomy that afflicts this enigmatic group of ants can begin to
 be resolved.
description: DNA was extracted non-destructively from 39 leptanilline
 specimens, plus <em>Martialis heureka </em>as an outgroup,
 using a DNeasy Blood & Tissue Kit (Qiagen Inc., Valencia, CA)
 according to manufacturer instructions. Genomic concentrations were
 quantified for each sample with a Qubit 2.0 fluorometer (Life Technologies
 Inc., Carlsbad, CA). Phylogenomic data were generated using the UCE probe
 set <i>hym-v2 </i>(Branstetter <i>et al.</i>
 2017), with libraries being prepared and target loci enriched using the
 protocol of Branstetter <i>et al.</i> (2017). Enrichment
 success and size-adjusted DNA concentrations of pools were assessed using
 the SYBR FAST qPCR kit (Kapa Biosystems, Wilmington, MA) and all pools
 were combined into an equimolar final pool. The contents of this final
 pool were sequenced on an Illumina HiSeq 2500 at the High Throughput
 Genomics Facility, University of Utah, Salt Lake City, UT or an Illumina
 HiSeq 4000 at Novogene, Sacramento, CA. The FASTQ
 output was demultiplexed and cleansed of adapter contamination and
 low-quality reads using <i>illumiprocessor</i> (Faircloth
 2013) in the PHYLUCE package. Raw reads were assembled with
 <i>trinity </i>v. 2013-02-25 (Grabherr <em>et al.
 </em>2011) or with SPAdes v. 3.12.0 (Bankevich <em>et al.
 </em>2013). All PHYLUCE commands hereinafter are cited from
 Faircloth (2016). Species-specific contig assemblies were obtained with
 the ant-specific <i>hym-v2</i> probe set (Branstetter
 <i>et al.</i> 2017) using
 <i>phyluce_assembly_match_contigs_to_probes.py
 </i>(min_coverage=80), with min_identity=90 to minimize the
 influence of possible contamination; and a list of UCE loci shared across
 all taxa was generated using
 <i>phyluce_assembly_get_match_counts.py</i>, and<i>
 </i>separate FASTA files for each locus were created using these
 outputs. Sequences were aligned separately by locus using MAFFT L-INS-i
 (Katoh & Toh 2008), rather than the default version of MAFFT
 implemented in <i>phyluce</i>, implemented with the command
 <i>phyluce_assembly_seqcap_align.py</i>. These sequences were
 then trimmed with Gblocks (Castresana
 2000) as implemented by the wrapper script
 <i>phyluce_assembly_get_gblocks_trimmed_alignment_from_untrimmed.py</i> (settings: b1=0.5, b2=0.5, b3=12, b4=7). Alignment statistics for the output FASTA files were calculated with <i>phyluce_align_get_align_summary_data.py</i>.<i> </i>Finally, a dataset that was 90% complete with respect to taxon coverage per locus was generated using the script <i>phyluce_align_get_only_loci_with_min_taxa.py</i>, consisting of 580 loci. The final alignment was then concatenated and converted to PHYLIP format with <i>phyluce_align_format_nexus_files_for_raxml</i> and was 368,656 bp in length, with 19.07% missing data, 135,832 parsimony-informative sites, and mean locus length being 634 bp.