The Systematics and Biogeography of the Mite Harvestman Family

The systematics and biogeography of the mite harvestman family Sironidae (Arachnida : Opiliones : Cyphophthalmi) with the description of five new species Author(s): Gonzalo Giribet, Ligia R. Benavides and Izaskun Merino-Sáinz Source: Invertebrate Systematics, 31(4):456-491. Published By: CSIRO Publishing URL: http://www.bioone.org/doi/full/10.1071/IS16086

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The systematics and biogeography of the mite harvestman family Sironidae (Arachnida : Opiliones : Cyphophthalmi) with the description of ﬁve new species

Gonzalo Giribet A, Ligia R. Benavides A,C and Izaskun Merino-Sáinz B

AMuseum of Comparative Zoology & Department of Organismic and Evolutionary Biology, Harvard University, 26 Oxford Street, Cambridge, MA 02138, USA. BDepartamento de Biología de Organismos y Sistemas, Universidad de Oviedo, Calle Catedrático Rodrigo Uría s/n, 33071 Oviedo, Asturias, Spain. CCorresponding author. Email: [email protected]

Abstract. Sironidae, the first described family of Cyphophthalmi, is among the least understood phylogenetically. After examining recent collections across their distribution range, we provide the first comprehensive treatment of Sironidae by including molecular data from most of the known species, and all genera except for the monotypic Odontosiro Juberthie, 1961. We also revisit the male genitalic morphology for most genera by using confocal laser scanning microscopy and provide descriptions of five new species belonging to Iberosiro de Bivort & Giribet, 2004 (monotypic until now), Paramiopsalis Juberthie, 1962 and Siro Latreille, 1802. While the monophyly of Sironidae remains poorly supported using traditional Sanger-based markers, with the Mediterranean Parasiro Hansen & Sørensen, 1904 and the Japanese Suzukielus Juberthie, 1970b sometimes branching basally with respect to the other sironids, the remaining genera form a well- supported Laurentian/Laurasian clade. This group divides into a Western European/North American clade of Siro and the remaining genera, Iberosiro, Paramiopsalis and Cyphophthalmus Joseph, 1868. Iberosiro and Paramiopsalis form a well- supported clade from the NW corner of the Iberian Peninsula, while Cyphophthalmus is widespread in the Balkan region and Eastern Mediterranean. Finally, the following new taxa are described: Iberosiro rosae Giribet, Merino-Sáinz & Benavides, sp. nov., Paramiopsalis anadonae Giribet, Merino-Sáinz & Benavides, sp. nov., Paramiopsalis ramblae Benavides & Giribet, sp. nov., Siro ligiae Giribet, sp. nov., and Siro richarti Benavides & Giribet, sp. nov.

Additional keywords: diversity, Iberian Peninsula, Iberosiro, Paramiopsalis, Siro.

Received 19 December 2016, accepted 20 February 2017, published online 7 July 2017

Introduction exception of a morphological cladistic analysis (de Bivort and The Opiliones suborder Cyphophthalmi has received ample Giribet 2004). More recently, Dreszer et al.(2015) published attention from phylogenetic and biogeographic points of view an updated phylogenetic analysis of the family, but focused (Juberthie and Massoud 1976; Shear 1980; Boyer et al. 2007; mostly on the genus Cyphophthalmus and on previously Giribet et al. 2012). From its six currently recognised families, described taxa, and the outgroup sampling did not allow for Sironidae is restricted to the former Laurentian/Laurasian terranes testing of the monophyly of Sironidae. A study by Giribet et al. (Fig. 1), while the ﬁve remaining families have a Gondwanan (2012), which combined morphological and molecular data, was origin (Giribet and Prieto 2003; Sharma and Giribet 2009; Clouse ambiguous with respect to the monophyly of Sironidae, but found and Giribet 2010; Benavides and Giribet 2013; Giribet et al. four main clades: Parasiro Hansen & Sørensen, 1904 (with three 2016) (but see discussion about Marwe coarctata Shear, 1985 described species; endemic to the Western Mediterranean); below, a putative sironid found in Kenya). Recent work on sironid Suzukielus Juberthie, 1970b (monotypic; Honshu, Japan); Siro phylogeny has focussed on particular clades – the genera Siro (with 12 described extant species plus two amber fossils; Latreille, 1802 (Giribet and Shear 2010), Cyphophthalmus North America and Europe); and the clade formed by Joseph, 1868 (Boyer et al. 2005; Karaman 2009; Murienne Paramiopsalis (two described species; NW Iberian Peninsula) et al. 2010; Dreszer et al. 2015) and Paramiopsalis Juberthie, and Cyphophthalmus (33 species; Balkan region to Asia Minor). 1962 (Murienne and Giribet 2009) – but few studies have Siro, Paramiopsalis and Cyphophthalmus constituted the core addressed the global phylogeny of the family (Giribet and of Sironidae, sharing many of the characters typically assigned to Boyer 2002; Boyer et al. 2007; Giribet et al. 2012), with the the family and forming a clade in virtually all analyses. However,

Journal compilation CSIRO 2017 www.publish.csiro.au/journals/is Systematics and biogeography of Sironidae Invertebrate Systematics 457

(A)

(B)(C) (D)

(E) (F)(G)

(H)

Fig. 1. Geographical distribution of the sironid taxa included in our molecular study. (A) Distribution of the family worldwide. B–H, Detailed maps showing the generic sampling. (B) Cyphophthalmus depicted in Cyan; (C) Iberosiro represented in red (I. rosae, sp. nov. represented by a triangle); (D) Paramiopsalis in orange (P. anadonae, sp. nov. represented by a triangle; P. ramblae, sp. nov. by a square); (E) Parasiro in green; (F) Suzukielus in magenta; (G) European Siro and (H) North American Siro both in blue (S. ligiae, sp. nov. represented by a triangle; S. richarti, sp. nov. by a square). 458 Invertebrate Systematics G. Giribet et al. the membership of Parasiro and Suzukielus in the main sironid sputter-coated with gold using an EMS 300T D Dual Head clade is poorly supported, if at all, in Sanger-based molecular Sputter Coater (Electron Microscope Sciences, Hatfield, PA). analyses and in morphological analyses (e.g. Giribet and Boyer Images were taken using a Zeiss Supra 55VP Field Emission 2002; de Bivort and Giribet 2004; Boyer et al. 2007; Giribet Scanning Electron Microscope at the Harvard Center of et al. 2012). Iberosiro de Bivort & Giribet, 2004, a monotypic Nanoscale Systems (CNS). genus described from Portugal (de Bivort and Giribet 2004), was found to form a clade with the sympatric genus Paramiopsalis Auto-fluorescence imaging (de Bivort and Giribet 2004; Dreszer et al. 2015). We used an LSM 880 with Airyscan (Carl Zeiss, Jena, Here we provide a comprehensive phylogeny of the family Germany) system from the Harvard Center for Biological fi Sironidae based on ve genetic markers, including molecular Imaging (HCBI) to image the spermatopositor of the new data from multiple individuals of the genus Iberosiro, for which species. Spermatopositors were dissected from the ventral side we describe a second species from Asturias, Spain. This new of the opisthosoma and placed on slides containing Rapiclear species lives in a rather unorthodox habitat for Cyphophthalmi, (SunJin Lab Co., Hsinchu City, Taiwan). The master pinhole was and occurs in sympatry with a species of Paramiopsalis, set to 1–1.25 Airy unit (AU), and the 20Â Plan Apochromat long which we describe here as well. In addition, we name a second working distance objective was used. Laser wavelengths of 481 Paramiopsalis from Asturias, to bring the genus to four species and 565 nm were used to detect the auto-fluorescence of the (Juberthie 1962; Rambla and Fontarnau 1984; Murienne and chitin. A series of images (100–270) were taken in the z direction Giribet 2009), and two new species of Siro from North America, to generate a Z-stack. Zen 2 blue edition modular image- bringing the number of living Siro species to fourteen (Ewing processing software (Carl Zeiss, Jena, Germany) was used 1923; Hoffman 1963; Shear 1980; Giribet and Shear 2010). to assemble, edit and generate a 3D reconstruction of the Species descriptions are accompanied with detailed scanning spermatopositor for each species. All videos were deposited in electron micrographs and with novel imaging of their genitalia the MCZ database MCZbase (http://mczbase.mcz.harvard.edu) with confocal laser microscopy, as in previous recent studies associated with the specimen records. (Murienne and Giribet 2009; Dreszer et al. 2015). Molecules Methods Laboratory protocols Taxon sampling Foreach specimen, DNAwas extracted and isolated from up to All specimens used for this study were preserved in ~96% ethanol two legs using the Qiagen DNeasy tissue kit (Valencia, CA, USA) and stored at –20 Cor–80 C. Fig. 1 shows the sampling localities following the manufacturer’s guidelines; the remainder of the for our sironid samples. The majority of specimens were acquired specimen was kept as a voucher and deposited in the Museum of over a series of field trips, but a few were sent by colleagues. The Comparative Zoology (Harvard University), and is accessible list of specimens with corresponding MCZ voucher numbers, through MCZbase. Genes or gene fragments targeted for PCR GenBank accession numbers and collection details is given in amplification and sequencing included the nuclear ribosomal Table 1. genes 18S rRNA (18S hereafter, ~1800 bp) and 28S rRNA The ingroup is represented by 65 sironid specimens. (28S hereafter, fragment of ~2700 bp); two mitochondrial The outgroups include five species belonging to the ribosomal genes, 12S rRNA (12S hereafter, ~400 bp) and 16S Cyphophthalmi families Neogoveidae, Pettalidae, Stylocellidae rRNA (16S hereafter, ~550 bp) and the mitochondrial protein- and Troglosironidae. encoding gene cytochrome c oxidase subunit I (COI; 771 bp). Morphology PCR was used to amplify the genes mentioned above for a total of ~5.4 Kb per specimen. For detailed methods, amplification and Digital light microscopy analysis sequencing primer sequences, fragment lengths and annealing temperatures used for PCR amplification and sequencing see our Using a JVC-KY-F75U digital camera mounted on a Leica earlier work (e.g. Giribet and Shear 2010; Giribet et al. 2012). MZ 12.5 stereomicroscope with the Plan 0.5Â objective, we took photographs of the habitus of male holotype and a female Phylogenetic analyses paratype for each species in dorsal, lateral and ventral view. A series of 5–15 images were taken at different focal planes We conducted analyses under parsimony and model-based with the software package Auto-Montage Pro ver. 5.02.0096 approaches to investigate Sironidae relationships under a broad (Synoptics group, Cambridge, UK). set of assumptions.

Scanning electron microscopy analysis Parsimony When available, we took scanning electron microscopy Parsimony analyses were conducted under the dynamic (SEM) images of a male and a female for each species. For homology framework with the software POY v. 5.1.1 SEM procedures, specimens were cleansed by ultrasound for (Wheeler et al. 2015), with genes analysed individually and 1–5 min. Following ultrasound cleansing, left side appendages with all gene partitions combined; the results presented are (chelicera, pedipalp and legs) were dissected from the body and based on the combined analyses, and the individual genes air-dried on a Petri dish for a few minutes. Once dried, all parts were only analysed to test for possible biases or sequence were mounted on an SEM stub with carbon bi-adhesive tape and contamination. Tree searches were executed using a timed Systematics and biogeography of Sironidae Invertebrate Systematics 459

Table 1. Specimen data with genes sequenced and GenBank accession numbers New sequences appear in bold

Species Catalogue number Region 18S rRNA 28S rRNA 12S rRNA 16S rRNA COI Family Sironidae Cyphophthalmus duricorius MCZ-135009.2 Slovenia KJ857509 KJ857512 – KJ857515 KJ857527 Cyphophthalmus sp. DNA100497ii KY352065 KY352095 ––– C. ere MCZ-135018 Serbia AY639462 DQ825593 – AY639527 AY639557 C. gjorgjevici MCZ-135017 Macedonia AY639464 DQ825587 – AY639529 AY639559 C. gordani MCZ-135014 Montenegro AY639467 DQ825592 – AY639532 – C. hlavaci MCZ-135052 Croatia –––KJ857544 – C. markoi MCZ-135016 Macedonia –––AY639534 AY639561 C. martensi MCZ-135013 Montenegro AY639471 DQ825589 – AY639536 AY639563 C. minutus MCZ-135012 Montenegro AY639473 DQ825591 – AY639537 AY639565 C. ognjenovici MCZ-135027 Bosnia and Herzegovina AY639475 DQ825594 ––AY639567 C. rumijae MCZ-135011 Montenegro AY639477 DQ825588 – AY639539 AY639569 C. solentiensis MCZ-129787 Croatia KJ857518 KJ857522 – KJ857532 KJ857528 C. solentiensis MCZ-135079 Croatia KJ857519 KJ857523 – KJ857533 KJ857529 Cyphophthalmus sp. MCZ-135031 Bulgaria – AY918873 ––– Cyphophthalmus sp. MCZ-135032 Bulgaria – KY352096 ––– C. teyrovskyi MCZ-135025 Montenegro AY639482 DQ513118 – AY639544 AY639571 C. trebinjanum MCZ-135026 Bosnia and Herzegovina AY639483 DQ513119 ––AY639572 C. zetae MCZ-135022 Montenegro AY639485 AY639515 – AY639546 AY639574 Iberosiro rosae, sp. nov. MCZ-135072 Asturias, Spain KY352066 KJ857524 KY352124 KJ857534 KJ857530 I. rosae, sp. nov. MCZ-135073.2 Asturias, Spain KY352067 KY352097 KY352125 KY352146 KY352166 I. rosae, sp. nov. MCZ-135073.3 Asturias, Spain KY352068 KY352098 KY352126 KY352147 KY352167 I. rosae, sp. nov. MCZ-135073.4 Asturias, Spain KY352069 KY352099 KY352127 KY352148 KY352168 Paramiopsalis anadonae, sp. nov. MCZ-135070 Asturias, Spain KY352076 JF934991 KY352128 JF935024 JF786390 P. anadonae, sp. nov. MCZ-135073 Asturias, Spain KY352075 KY352100 KY352129 KY352149 KY352169 P. anadonae, sp. nov. MCZ-135075 Asturias, Spain KY352072 KY352101 KY352130 KY352150 KY352170 P. anadonae, sp. nov. MCZ-135076 Asturias, Spain KY352073 KY352102 KY352131 KY352151 KY352171 P. eduardoi MCZ-135034 Galicia, Spain EU638284 EU638287 EU638281 EU638288 P. eduardoi MCZ-135035 Galicia, Spain KY352074 KY352103 KY352132 KY352152 KY352172 P. ramblae, sp. nov. MCZ-133850 Asturias, Spain KY352077 KY352104 KY352133 KY352153 KY352173 P. ramulosus MCZ-132365 Portugal KY352070 KY352105 KY352134 KY352154 KY352174 P. ramulosus MCZ-132368 Portugal KY352071 KY352106 KY352135 KY352155 KY352175 P. ramulosus MCZ-132370 Portugal JF934956 JF934990 KY352136 JF935023 – P. ramulosus MCZ-135006 Galicia, Spain AY639489 DQ513121 – AY639550 DQ825641 Parasiro coiffaiti MCZ-132372 Catalonia, Spain AY918872 DQ513122 – AY918877 DQ825642 P. coiffaiti MCZ-135033 Catalonia, Spain EU638283 EU638286 – KY352156 – P. minor MCZ-132374 Sardinia, Italy JF934958 JF934992 – JF935025 JF786391 P. minor MCZ-132376 Sardinia, Italy KY352078 KY352107 KY352137 KY352157 KY352176 Siro acaroides MCZ-132381 California, USA KY352080 KY352108 KY352138 KY352158 KY352177 S. acaroides MCZ-130002 Oregon, USA KY352079 KY352109 KY352139 KY352159 KY352178 S. acaroides MCZ-132379 Oregon, USA DQ513143 DQ513130 ––DQ513114 S. acaroides MCZ-132380 California, USA DQ513144 DQ513131 ––– S. acaroides MCZ-134454 Oregon, USA AY639490 DQ513128 – AY639551 DQ825643 S. acaroides MCZ-134495 Oregon, USA KY352081 KY352110 KY352140 KY352160 KY352179 S. boyerae MCZ-92901.1 Washington, USA DQ513139 DQ513125 ––DQ513112 S. boyerae MCZ-92901.2 Washington, USA KY352082 KY352112 ––– S. boyerae MCZ-132390 Washington, USA KY352083 KY352111 KY352141 – KY352180 S. calaveras MCZ-92889 California, USA KY352084 KY352113 KY352142 KY352161 KY352181 S. carpaticus MCZ-135071 Poland KJ857536 KJ857539 – KJ857545 KJ857542 S. clousi MCZ-130003 Oregon, USA KJ857537 KJ857540 ––KJ857543 S. clousi MCZ-130003.2 Oregon, USA KY352094 KY352114 ––– S. crassus MCZ-68552 Slovenia KY352085 KY352115 ––– S. exilis MCZ-134551 Maryland, USA AY639491 DQ825585 ––AY639579 S. kamiakensis MCZ-132388 Idaho, USA KJ857538.1 KJ857541.1 ––– S. kamiakensis MCZ-132389 Idaho, USA JF934959 JF934993 ––– S. kamiakensis MCZ-132386 Idaho, USA KY352087 KY352116 ––KY352182 S. kamiakensis MCZ-134453 Washington, USA KY352088 KY352117 ––– S. ligiae, sp. nov. MCZ-130001.1 Oregon, USA DQ513141 DQ513127 ––KY352183 (continued next page ) 460 Invertebrate Systematics G. Giribet et al.

Table 1. (continued )

Species Catalogue number Region 18S rRNA 28S rRNA 12S rRNA 16S rRNA COI S. ligiae, sp. nov. MCZ-130001.2 Oregon, USA KY352086 KY352118 ––KY352184 S. richarti, sp. nov. MCZ-134488 Idaho, USA KY352089 KY352119 ––– S. rubens MCZ-132391 France AY428818 DQ825584 ––DQ513111 S. shasta MCZ-130004 California, USA KJ857521 KJ857525–6 – KJ857535 KJ857531 S. shasta MCZ-130004.2 California, USA KY352090 KY352120 – KY352162 KY352185 S. valleorum MCZ-135008 Italy AY639492 DQ513123 – AY639552 AY639580 Suzukielus sauteri MCZ-132256 Japan DQ513138 DQ513116 – DQ518086 DQ513108 S. sauteri MCZ-132260 Japan KY352091 KY352121 KY352143 KY352163 KY352186 S. sauteri MCZ-132261 Japan KY352092 KY352122 KY352144 KY352164 KY352187 S. sauteri MCZ-132262 Japan KY352093 KY352123 KY352145 KY352165 KY352188 S. sauteri MCZ-132263 Japan DQ825541 DQ825583 DQ825615 DQ825640

Outgroups Chileogovea oedipus MCZ-134709 DQ133721 DQ133733 – DQ518055 DQ133745 Fangensis spelaeus MCZ-134248 DQ133712 DQ825554 – GQ488195 AY639583 Metasiro americanus MCZ-133799 DQ825542 DQ825595 – DQ825616 JF786394 Ogovea cameroonensis MCZ-132315 JF934960 JF934994 – JF935026 JF786392 Troglosiro aelleni MCZ-134764 AY639497 DQ518044 – AY639555 AY639584

Table 2. Length of the individual gene partitions MAFFT alignment trimmed datasets using Gblocks v. 0.91b (Castresana 2000; before and after treatment with GBlocks (GB) Talavera and Castresana 2007), even though only 5.6% of the data were discarded. Gblocks parameters were defined as MAFFT MAFFT+GB follows: minimum number of sequences for a conserved fl 12S rRNA 353 310 position (50%), minimum number of sequences for a anking 16S rRNA 510 439 position (50%), maximum number of contiguous non- 18S rRNA 1764 1763 conserved positions (8), minimum length of a block (5, instead 28S rRNA 2180 2087 of the default value of 10), since this value is preferable for COI 657 657 rRNA-like alignments that contain many small but very well- conserved blocks (Gblocks online documentation), and allowed gap positions (with half). Gene partitions were concatenated search function that combines multiple cycles of tree building with the software SequenceMatrix v. 1.8 (Vaidya et al. 2011), with TBR branch swapping, parsimony ratchet and tree fusing. and PartitionFinder v. 1.0.1 (Lanfear et al. 2012) was used to Timed searches of two hours with four processors were executed choose the partition schemes and models of sequence evolution for the individual genes and the combined dataset. Nodal stability of the concatenated dataset. was measured with a sensitivity analysis that considers different Maximum likelihood analyses were conducted in RAxML- values of indel/transversion ratio, transversion/transition ratio HCP2 v. 8.2.6 (Stamatakis 2014) as implemented in the and indel opening and extension. For this study, we analysed CIPRES science gateway portal (Miller et al. 2010). Nodal six different parameter sets as in previous studies (Giribet et al. support values were calculated with 1000 replicates of the 2012, 2016). A modified Wheeler incongruence length difference fast bootstrap (BS) algorithm using the GTRGAMMA model (WILD) metric (Wheeler 1995; Sharma et al. 2011) was calculated (Stamatakis et al. 2008). to choose among the different hypotheses resulting from the A Bayesian inference analysis of the combined data was sensitivity analysis; the optimal parameter set being the one executed with the online version of MrBayes v. 3.2.6 that minimises overall incongruence among the different (Ronquist et al. 2012) as implemented in the CIPRES science partitions. The resulting WILD values are shown in Table 2. gateway portal (Miller et al. 2010). The analysis consisted of Nodal support for the optimal tree was estimated via 200 two independent runs of 10 million generations each, sampled jackknife pseudoreplicates using auto_sequence_partition as every 1000 generations. A 50% majority-rule consensus tree was discussed in an earlier work (Giribet et al. 2010). estimated after discarding a burn-in of 10% of the sampled trees. Convergence of the independent runs was confirmed with Tracer v. 1.6 (Rambaut et al. 2014). Posterior probability (PP) values Maximum likelihood and Bayesian inference analyses >0.95 were plotted on the tree. For the analyses under maximum likelihood (ML) and Bayesian inference (BI) we generated multiple sequence Specimen repositories alignments for each individual gene partition with the online MCZ, Museum of Comparative Zoology, Harvard University, version of MAFFT v. 7 (Katoh and Standley 2013). With the Cambridge, Massachusetts, USA; ZUPV, Departamento de aim of comparing the results with those of the direct optimization Zoología y Biología Celular, Universidad del País Vasco, approach, we used the complete datasets, but also evaluated Bilbo, Spain. Systematics and biogeography of Sironidae Invertebrate Systematics 461

Results and discussion direct optimization analyses S. crassus groups with the American species (Fig. 2). Our molecular matrix consists of 70 terminals, 65 sironid Within the North American clade, two main groups appear in specimens and representatives of five outgroup species. most analyses, one constituted by Siro species from California Lengths of the MAFFT alignments for each individual gene, and Oregon (S. calaveras Giribet & Shear, 2010, S. shasta Giribet before and after Gblocks treatment, are shown in Table 3. The & Shear, 2010 and S. acaroides Ewing, 1923), and a second clade total length of the concatenated alignment was 5464 bp before comprising species from Idaho, Oregon and Washington removal of non-conserved positions with Gblocks and 5256 bp (S. richarti, sp. nov., S. kamiakensis Newell, 1943, S. boyerae after treatment. Fig. 2 summarises the parsimony results of our Giribet & Shear, 2010 and S. ligiae, sp. nov.), plus S. exilis POY analysis, Fig. 3 the results of the analysis of ML with Hoffman, 1963, from Maryland. This latter clade is interesting as RAxML, and the BI analysis with MrBayes is presented in Fig. 4. both S. richarti and S. kamiakensis males have a unique Phylogenetic analysis of the assembled sironid dataset is bisegmented tarsus IV (Fig. 28G, H), a character otherwise congruent with prior studies in identifying three main clades – only found in some pettalids and in Suzukielus (Giribet and Parasiro, Suzukielus and the remaining Sironidae – these Boyer 2002). However, this character is not an apomorphy appearing under all analyses, data matrices and parameter sets for S. richarti and S. kamiakensis, as the latter is closest to (except for parameter set 3211). Monophyly of Sironidae as S. boyerae + S. ligiae, sp. nov.+ S. exilis, which present an currently circumscribed is found only in the trimmed ML undivided tarsus IV in the males. As shown in pettalids, the analysis (figure not shown), albeit with 40% bootstrap support divided tarsus IV is more labile than previously thought (Boyer (BS). Sironidae, including Suzukielus but excluding Parasiro,is and Giribet 2007). The widespread species S. acaroides shows found in the BI analysis of the complete dataset (posterior deep genetic structure, suggesting the possibility of additional probability, PP = 0.99; Fig. 4), trimmed BI analysis (PP = 0.97, species within this group. figure not shown) and untrimmed ML dataset (BS = 85%; Fig. 3). Iberosiro constitutes the sister group of Paramiopsalis in all The parsimony analyses of direct optimization with POY analyses and methods (Figs 2–4), as suggested also by the fusion place both Parasiro and Suzukielus within the outgroups of the second and third walking leg coxae in these two genera (Fig. 2). The placement of Parasiro and/or Suzukielus outside (e.g. Figs 6D, E, 10E, 17C) – the second coxae are free in all of the core Sironidae is a recurrent theme of prior analyses using other sironids (e.g. Figs 21E, F, 26E, F) – a condition otherwise morphology – these two genera differ in fundamental characters only found in some neogoveids and in all stylocellids. This from most other sironids – and molecular data (Boyer et al. clade is also supported by the presence of a pronounced 2007;Giribetet al. 2012). Nevertheless, a sironid clade ventral process in the palp trochanter (also found in some including Parasiro, Suzukielus and Siro is strongly supported pettalids and in Odontosiro) (Juberthie 1970a; Giribet and in a transcriptome-based phylogenomic analysis of Opiliones Boyer 2002; Giribet et al. 2012). Iberosiro and Paramiopsalis (Fernández et al. 2017). Therefore, the instability and low show a nearly sympatric distribution, with species from both support of either alternative placement of Parasiro and genera occurring under the same rocks in at least one locality Suzukielus and disagreement among analytical methods with (I. rosae, sp. nov. and P. anadonae, sp. nov.). Despite an attempt respect to these taxa is not discussed further in the context of to visit the type locality of Iberosiro in Portugal, we were not the present work. able to obtain fresh specimens of I. dystilos de Bivort & Giribet, Within the core sironid clade, the Western European/North 2004, but morphological analyses have also placed Iberosiro American Siro is often sister group of a clade of the European as sister group of Paramiopsalis (de Bivort and Giribet 2004; genera Cyphophthalmus, Iberosiro and Paramiopsalis. Giribet et al. 2012). However, monophyly of Siro is supported only in some BI The genus Paramiopsalis comprises at least four species (two analyses, as S. clousi Giribet & Shear, 2010 appears as sister newly described here) difficult to distinguish morphologically. group of the Iberosiro–Paramiopsalis–Cyphophtlalmus clade While some species seem to have a broad geographic range in most POY analyses and in the untrimmed ML data analysis. (e.g. P. ramulosus Juberthie, 1962), others are single locality The remaining Siro species generally divide into a European endemics with different species found closely geographically clade – S. valleorum Chemini, 1990, S. crassus Novak & (i.e. P. ramblae, sp. nov. and P. anadonae, sp. nov.), although Giribet, 2010, S. rubens Latreille, 1802 and S. carpaticus their habitat preference differs, as P. ramblae was found in dense Rafalski, 1956 – and an American clade, although in some forest while P. anadonae was found under a rock in grassland,

Table 3. Tree lengths for different data partitions (TOT, all molecular partitions) analysed and incongruence length difference

(WILD) values between datasets The optimal parameter set (the one that minimises the WILD value) is indicated in bold

Parameter 18S 28S 12S 16S COI TOT WILD 111 162 1289 674 2250 3766 8282 0.017024873 121 229 1948 1067 3642 5725 12 801 0.01484259 211 173 1537 776 2560 3831 9040 0.018030973 221 247 2417 1252 4170 5811 14 134 0.016768077 3211 234 2077 1128 3794 5796 13 214 0.014000303 3221 332 2727 1421 4732 7669 17 206 0.01888876 462 Invertebrate Systematics G. Giribet et al.

Chileogovea oedipus MCZ-134709 Suzukielus sauteri MCZ-132263 Suzukielus sauteri MCZ-132262 Suzukielus sauteri MCZ-132260 Suzukielus sauteri MCZ-132256 Suzukielus sauteri MCZ-132261 Parasiro coiffaiti MCZ-132372 100 Parasiro coiffaiti MCZ-135033 Parasiro minor MCZ-132374 Parasiro minor MCZ-132376 Fangensis spelaeus MCZ-134248 Troglosiro aelleni MCZ-134764 Metasiro americanus MCZ-133799 Ogovea cameroonensis MCZ-132315 Siro valleorum MCZ-135008 Siro carpaticus MCZ-135071 Siro rubens MCZ-132391 Siro crassus MCZ-68552 96 Siro calaveras MCZ-130005 Siro shasta MCZ-130004 Siro shasta MCZ-130004.2 Siro acaroides DNA101621 Siro acaroides MCZ-132380 Siro acaroides MCZ-130002 Siro acaroides MCZ-134454 Siro acaroides MCZ-132379 Siro acaroides MCZ-134495 Siro richarti MCZ-134488 Siro kamiakensis MCZ-132388 97 Siro kamiakensis MCZ-134453 Siro kamiakensis MCZ-132386 Siro kamiakensis MCZ-132389 Siro boyerae MCZ-132390 Siro boyerae MCZ-92901.1 Siro boyerae MCZ-92901.2 Siro exilis MCZ-134551 Siro ligiae sp. nov. MCZ-130001 Siro ligiae sp. nov. MCZ-130001.2 Siro clousi MCZ-130003 Siro clousi MCZ-130003.2 Iberosiro rosae sp. nov. MCZ-135073.3 Iberosiro rosae sp. nov. MCZ-135073.2 Iberosiro rosae sp. nov. MCZ-135072 93 Iberosiro rosae sp. nov. MCZ-135073.4 Paramiopsalis ramblae sp. nov. MCZ-133850 Paramiopsalis ramulosus MCZ-132370 100 Paramiopsalis ramulosus MCZ-135006 Paramiopsalis ramulosus MCZ-132365 Paramiopsalis ramulosus MCZ-132368 Paramiopsalis eduardoi MCZ-135034 Paramiopsalis eduardoi MCZ-135035 96 Paramiopsalis anadonae sp. nov. MCZ-135070 Paramiopsalis anadonae sp. nov. MCZ-135073 Paramiopsalis anadonae sp. nov. MCZ-135075 Paramiopsalis anadonae sp. nov. MCZ-135076 Cyphophthalmus sp. MCZ-135031 Cyphophthalmus gjorgjevici MCZ-135017 Cyphophthalmus solentiensis MCZ-129787 Cyphophthalmus solentiensis MCZ-135079 Cyphophthalmus duricorius MCZ-135009.2 98 Cyphophthalmus rumijae MCZ-135011 Cyphophthalmus ere MCZ-135018 Cyphophthalmus eratoae DNA100497ii Cyphophthalmus markoi MCZ-135016 Cyphophthalmus hlavaci DNA102099 Cyphophthalmus zetae MCZ-135022 Cyphophthalmus martensi MCZ-135013 Cyphophthalmus sp. MCZ-135032 Cyphophthalmus gordani MCZ-135014 Cyphophthalmus teyrovskyi MCZ-135025 Cyphophthalmus trebinjanum MCZ-135026 Cyphophthalmus minutus MCZ-135012 Cyphophthalmus ognjenovici MCZ-135027

200.0

Fig. 2. Phylogenetic relationships of Sironidae based on the parsimony direct optimization analysis of all molecular data under the parameter set 3211. Sironid genera colours as in Fig. 1. Navajo rugs denote monophyly (black) or non-monophyly (white) of a given node under the parameter set speciﬁed in the legend. Numbers above nodes indicate jack-knife support values. A colour version of this ﬁgure is available from the journal online. Systematics and biogeography of Sironidae Invertebrate Systematics 463

Chileogovea oedipus MCZ-134709 Fangensis spelaeus MCZ-134248 Parasiro minor MCZ-132376 * Parasiro minor MCZ-132374 Parasiro coiffaiti MCZ-135033 Parasiro coiffaiti MCZ-132372 Troglosiro aelleni MCZ-134764 * Metasiro savannahensis MCZ-133799 Ogovea cameroonensis MCZ-132315 Suzukielus sauteri MCZ-132263 * Suzukielus sauteri MCZ-132262 Suzukielus sauteri MCZ-132261 Suzukielus sauteri MCZ-132260 Suzukielus sauteri MCZ-132256 * Siro rubens MCZ-132391 Siro carpaticus MCZ-135071 Siro valleorum MCZ-135008 Siro crassus MCZ-68552 Siro calaveras MCZ-92889 * Siro shasta MCZ-130004.1 Siro shasta MCZ-130004.2 Siro acaroides MCZ-132380 Siro acaroides MCZ-132381 * Siro acaroides MCZ-134454 Siro acaroides MCZ-130002 Siro acaroides MCZ-134495 Siro acaroides MCZ-132379 Siro richarti sp. nov. MCZ-134488 Siro kamiakensis MCZ-132389 * Siro kamiakensis MCZ-132388 Siro kamiakensis MCZ-132386 * Siro kamiakensis MCZ-134453 Siro exilis MCZ-134551 * Siro ligiae sp. nov. MCZ-130001.2 Siro ligiae sp. nov. MCZ-130001.1 Siro boyerae MCZ-92901.2 Siro boyerae MCZ-92901.1 Siro boyerae MCZ-132390 Siro clousi MCZ-130003.2 Siro clousi MCZ-130003.1 Iberosiro rosae sp. nov. MCZ-135072.1 * Iberosiro rosae sp. nov. MCZ-135073.4 Iberosiro rosae sp. nov. MCZ-135073.3 Iberosiro rosae sp. nov. MCZ-135073.2 * Paramiopsalis ramulosus MCZ-132370 Paramiopsalis ramulosus MCZ-135006 Paramiopsalis ramulosus MCZ-132365 * Paramiopsalis ramulosus MCZ-132368 Paramiopsalis ramblae sp. nov. MCZ-133850 Paramiopsalis eduardoi MCZ-135034 Paramiopsalis eduardoi MCZ-135035 * Paramiopsalis anadonae sp. nov. MCZ-135070 Paramiopsalis anadonae sp. nov. MCZ-135073 Paramiopsalis anadonae sp. nov. MCZ-135076 Paramiopsalis anadonae sp. nov. MCZ-135075 Cyphophthalmus sp. MCZ-135031 Cyphophthalmus gjorgjevici MCZ-135017 Cyphophthalmus solentiensis MCZ-135079 Cyphophthalmus solentiensis MCZ-129787 Cyphophthalmus rumijae MCZ-135011 * Cyphophthalmus duricorius MCZ-135009 2 Cyphophthalmus ere MCZ-135018 Cyphophthalmus hlavaci MCZ-135052 Cyphophthalmus markoi MCZ-135016 Cyphophthalmus zetae MCZ-135022 Cyphophthalmus eratoae DNA100497ii Cyphophthalmus sp. MCZ-135032 Cyphophthalmus martensi MCZ-135013 Cyphophthalmus gordani MCZ-135014 Cyphophthalmus teyrovskyi MCZ-135025 Cyphophthalmus trebinjanum MCZ-135026 Cyphophthalmus minutus MCZ-135012 Cyphophthalmus ognjenovici MCZ-135027

0.4

Fig. 3. Maximum-likelihood tree (lnL = –41665.28540) resulting from the analysis of the combined molecular dataset aligned with MAFFT and analysed with RAxML under GTRGAMMA. Genera colours as in Fig. 1. Black asterisks on nodes denote bootstrap values between 95% and 100%. A colour version of this ﬁgure is available from the journal online. 464 Invertebrate Systematics G. Giribet et al.

Chileogovea oedipus MCZ-134709 Fangensis spelaeus MCZ-134248 Parasiro coiffaiti MCZ-132372 * Parasiro coiffaiti MCZ-135033 Parasiro minor MCZ-132374 Parasiro minor MCZ-132376 Troglosiro aelleni MCZ-134764 Metasiro savannahensis MCZ-133799 Ogovea cameroonensis MCZ-132315 Suzukielus sauteri MCZ-132263 * Suzukielus sauteri MCZ-132262 Suzukielus sauteri MCZ-132256 Suzukielus sauteri MCZ-132260 Suzukielus sauteri MCZ-132261 Siro clousi MCZ-130003.1 Siro clousi MCZ-130003.2 Siro carpaticus MCZ-135071 Siro rubens MCZ-132391 Siro crassus MCZ-68552 Siro valleorum MCZ-135008 Siro calaveras MCZ-92889 * Siro shasta MCZ-130004.1 Siro shasta MCZ-130004.2 * * Siro acaroides MCZ-132381 Siro acaroides MCZ-132380 * Siro acaroides MCZ-130002 Siro acaroides MCZ-134454 Siro acaroides MCZ-132379 Siro acaroides MCZ-134495 Siro richarti sp. nov. MCZ-134488 Siro kamiakensis MCZ-132386 * * Siro kamiakensis MCZ-134453 Siro kamiakensis MCZ-132388 * Siro kamiakensis MCZ-132389 Siro exilis MCZ-134551 * Siro ligiae sp. nov. MCZ-130001.1 * Siro ligiae sp. nov. MCZ-130001.2 * Siro boyerae MCZ-92901.2 * Siro boyerae MCZ-92901.1 Siro boyerae MCZ-132390 Iberosiro rosae sp. nov. MCZ-135073.1 * Iberosiro rosae sp. nov. MCZ-135073.4 Iberosiro rosae sp. nov. MCZ-135073.2 Iberosiro rosae sp. nov. MCZ-135073.3 * Paramiopsalis ramulosus MCZ-132370 Paramiopsalis ramulosus MCZ-135006 Paramiopsalis ramulosus MCZ-132365 * Paramiopsalis ramulosus MCZ-132368 Paramiopsalis ramblae sp. nov. MCZ-133850 Paramiopsalis eduardoi MCZ-135034 Paramiopsalis eduardoi MCZ-135035 Paramiopsalis anadonae sp. nov. MCZ-135070 * Paramiopsalis anadonae sp. nov. MCZ-135073 Paramiopsalis anadonae sp. nov. MCZ-135075 Paramiopsalis anadonae sp. nov. MCZ-135076 Cyphophthalmus gjorgjevici MCZ-135017 Cyphophthalmus sp. MCZ-135031 Cyphophthalmus solentiensis MCZ-129787 Cyphophthalmus solentiensis MCZ-135079 Cyphophthalmus duricorius MCZ-135009.2 * Cyphophthalmus rumijae MCZ-135011 Cyphophthalmus ere MCZ-135018 Cyphophthalmus eratoae DNA100497ii Cyphophthalmus zetae MCZ-135022 Cyphophthalmus hlavaci MCZ-135052 Cyphophthalmus markoi MCZ-135016 Cyphophthalmus martensi MCZ-135013 Cyphophthalmus sp. MCZ-135032 Cyphophthalmus gordani MCZ-135014 Cyphophthalmus teyrovskyi MCZ-135025 Cyphophthalmus trebinjanum MCZ-135026 Cyphophthalmus minutus MCZ-135012 Cyphophthalmus ognjenovici MCZ-135027

0.05

Fig.4. Phylogenetic hypothesisof sironidrelationships basedon the Bayesianinferenceanalysis of thecombined genes. Clade colours as in Fig. 1 Black asterisks indicate posterior probabilities between 0.95 and 1.00. A colour version of this ﬁgure is available from the journal online. Systematics and biogeography of Sironidae Invertebrate Systematics 465

along with I. rosae, sp. nov. Among the few characters that we (A) were able to match to the broad genetic diversity of this genus is the adenostyle and whether it is plumose or not, with P. ramulosus and P. eduardoi Murienne & Giribet, 2009 constituting the two extremes (Rambla and Fontarnau 1984; Murienne and Giribet 2009). The clade of NW Iberian species including Iberosiro and Paramiopsalis is sister group to the Balkan/Eastern Mediterranean Cyphophthalmus, as indicated in most earlier studies of sironid phylogeny (e.g. Murienne and Giribet 2009; (B) Giribet et al. 2012; Dreszer et al. 2015). Relationships within Cyphophthalmus mostly reﬂect earlier work on the group (Boyer et al. 2005; Karaman 2009; Murienne et al. 2010; Dreszer et al. 2015), and since no additional data are provided, it is not discussed further here. We were not able to add to this study molecular data of the monotypic genus Odontosiro Juberthie, 1961 due to lack of specimens for molecular work, but it has been suggested that it (C) is the sister group of Parasiro, based on morphological characters (de Bivort and Giribet 2004; Giribet et al. 2012). This is congruent with its geographical location on the NW Iberian Peninsula (Rambla and Fontarnau 1986), an area with two other sironid genera. Another potential sironid from a cave in Kenya, Marwe coarctata Shear, 1985, found to form a clade with Iberosiro and Paramiopsalis, again based on morphological characters (de Bivort and Giribet 2004; Giribet et al. 2012), was likewise not available for molecular study; however, including this taxon in Fig. 5. Iberosiro rosae, sp. nov. Holotype male (MCZ 135072) in (A) dorsal; future analyses would be important to test the supposed Laurasian (B) lateral; and (C) ventral view. Scale bars = 0.5 mm. distribution of the family. Future directed ﬁeldwork should target these two interesting but rare and understudied species: Marwe is Paratypes. 1 < and 1 , (MCZ 141134), mounted for SEM), Spain: known from three adult and two juvenile specimens from a single Asturias: Finca Experimental el Carbayal, Municipio de Illano, R. Rosa leg., cave (Shear 1985) and Odontosiro, despite having been reported 9.vii.2007, pitfall. Additional material. 3 juveniles (MCZ 135073; ex MCZ DNA106394) from a few localities (Juberthie 1961; Rambla and Fontarnau used for DNA sequencing; same collecting data as holotype. 1984, 1986), has never been found by the authors despite multiple attempts at searching near the type locality and in many of the Diagnosis other known localities for the species, and no museum specimens Iberosiro with a small male gonostome, shorter than the suture seem to be available. along coxae IV, as opposed to I. distylos, which has a larger gonostome with respect to the endites of coxae IV. Systematics Description Family SIRONIDAE Simon Male Slender sironid, total length of holotype 1.63 mm; width at Genus Iberosiro de Bivort & Giribet widest point 0.52 mm; length/width ratio 3.13; width across ozophores 0.49 mm. Body pale yellow (in ethanol; Fig. 5), Type species: Iberosiro dystilos de Bivort & Giribet, 2004. Species included: Iberosiro dystilos de Bivort & Giribet, 2004; Iberosiro with tuberculate-microgranulate morphology (sensu Murphree rosae Giribet, Merino-Sáinz & Benavides, sp. nov. 1988), except for the distal portions of sternites VI and VII, which are smooth (Figs 6A, 7A). Opisthosomal mid-dorsal Iberosiro rosae Giribet, Merino-Sáinz & Benavides, sp. nov. longitudinal sulcus absent. Eyes absent. Ozophores conical, (Figs 5–8) type II sensu Juberthie (1970a), facing laterally, but elevated from the margin of the scutum (Fig. 5B); with uniform and non- http://zoobank.org/NomenclaturalActs/86532F3A-8567-4F93-8E3E- directional granular ornamentation all over its surface and with a 53AE7BA3287F subterminal opening sensu Novak and Giribet (2006) (Fig. 7E). Material examined Ventral prosomal complex with coxa I free and coxa II fused to Holotype. < (MCZ135072;exMCZDNA106393;DNAextracted from coxa III (Fig. 6D); coxae I, II and IV meet along the midline one leg), Spain: Asturias: Finca Experimental el Carbayal (Giribet station (Fig. 6D); coxal pores present between coxae III and IV (Fig. 6D, 869: 43.3451, –6.8953; 898 m elevation), Municipio de Illano, A. Anadón, F); endites of coxa IV running adjacent to the midline suture for G. Giribet, I. Merino-Sáinz, R. Rosa leg., 16.v.2011, direct collecting, under a a length longer than the gonostome (Fig. 6D, F). Gonostome large boulder. semicircular/subtriangular with horn-like anterior apophyses on 466 Invertebrate Systematics G. Giribet et al.

(A) (B)

(C)

(D)

(E)

(F)

(G)

Fig. 6. Iberosiro rosae, sp. nov. Male and female (MCZ 141134). (A) Male ventral view; (B) female ventral view; (C) female dorsal view; (D) male thoracic complex; (E) female thoracic complex; (F) detail of male thoracic complex; (G) detail of female thoracic complex. Scale bars: A–E = 100 mm; F, G =20mm. Systematics and biogeography of Sironidae Invertebrate Systematics 467

(A)(B)

(E)

Fig. 7. Iberosiro rosae, sp. nov. Male and female (MCZ 141134). (A) Male anal region; (B) female anal region; (C) male anal glands; (D) male spiracle; (E) male ozophore. Scale bars: A, B =20mm; C–E =10mm. the endites of the fourth coxae (Fig. 6D, F). Sternal opisthosomal segment; without dorsal or ventral processes (Fig. 8A); second glands absent. Spiracles circular sensu Giribet and Boyer (2002) article not attenuated, sub-cylindrical for mostof its length, except (Fig. 7D). Sternites 8 and 9 medially fused. Male anal plate with for the fixed finger; with two small teeth on the distal end of a medial longitudinal non-ornamented carina. Two exocrine the fixed finger; mobile finger with uniform dentition (Fig. 8B). glands on tergite IX (Fig. 7C). Chelicerae measurements of male paratype (in mm length/ Chelicerae smooth along most of their surface, only with width): proximal segment 0.54/0.11; second segment 0.50/ sparse ornamentation on the ventral portion of the proximal 0.07; movable finger 0.15/0.09. Trochanters, femurs, patellae 468 Invertebrate Systematics G. Giribet et al.

(A) (B)

(D)

(E)

(F)

(I)

(J)

Fig. 8. Iberosiro rosae, sp. nov. Male and female (MCZ 141134). (A) Male chelicera; (B) detail of male chelicera; (C) male tarsus I showing the dorsal solenidia; (D) detail of dorsal solenidia in male tarsus I; (E) male tarsus II; (F) male tarsus IV; (G) dorsal adenostyle; (H) detail of adenostyle tip; (I) ventral adenostyle; (J) ovipositor. Scale bars: A, H = 100 mm; B–D, F, G =10mm; E =20mm. Systematics and biogeography of Sironidae Invertebrate Systematics 469 and tibiae of legs I, II and III ornamented. Leg I metatarsus and Remarks tarsus smooth (Fig. 8C); tarsus I with dorsal solenidia (Willemart The holotype of Iberosiro rosae, sp. nov. is poorly preserveddue to and Giribet 2010) (Fig. 8C, D); without a solea. Metatarsus of the DNA extraction procedure, and the spermatopositor was not legs IV ornamented and tarsus IV smooth (Fig. 8B). Tarsus IV found in the vial. However, the presence of a double adenostyle on entire (not divided) and with a double adenostyle, the dorsal, the male tarsus IV confirms that this species belongs to the genus larger, located towards the centre of the tarsus, opening at the Iberosiro. Iberosiro rosae is described based on a few specimens – tip (Fig. 8F H), and a second smaller adenostyle-like structure collectedbypitfallanddirectsearchingunderlargeboulders,where located ventrally and opposite to thedorsal adenostyle (Fig. 8F, I). it was found together with specimens of Paramiopsalis anadonae Dorsal adenostyle 0.46 mm long, 0.11 mm wide (Fig. 8E). under the same rocks. Ventral adenostyle 0.13 mm long, 0.07 mm wide bearing six terminal setae (Fig. 8I; one broken at the base). Pedipalps and remaining leg segments not imaged. Spermatopositor not Etymology available for study. The species is named after Rocío Rosa, a local biologist who collected the first specimens of this species and who accompanied Female us in the field when looking for additional specimens for this work. Total length of female (in mm) 1.40; width at widest point 0.61; length/width ratio 2.29 (Fig. 6C); width across ozophores Genus Paramiopsalis Juberthie 0.56 mm (Fig. 6C). Ventral prosomal complex with coxa I and II meeting along the midline (Fig. 6E, G); gonostome sub- Type species: Paramiopsalis ramulosus Juberthie, 1962. pentagonal with an elevated posterior margin (Fig. 6E, G); Species included: Paramiopsalis ramulosus Juberthie, 1962; P. eduardoi coxal pores present between coxae III and IV (Fig. 6G). Body Murienne & Giribet, 2009; P. anadonae Giribet, Merino-Sáinz & with granular surface except on sternite VII, which is smooth Benavides, sp. nov.; P. ramblae Benavides & Giribet, sp. nov. (Fig. 7B). Ovipositor 0.71 mm long and 0.08 mm wide (Fig. 8J). Paramiopsalis anadonae Giribet, Merino-Sáinz & Distribution Benavides, sp. nov. This species is only known from the type locality, an experimental (Figs 9–15) prairie with scattered boulders without trees or leaf litter, in an http://zoobank.org/NomenclaturalActs/B1CF4729-75F4-4C3F-828C- ecosystem known in Spain as ‘brezal-tojal’, or an area for 0A5EC0D88DC0 increasing yield of pasture for cattle (Fig. 1C, red triangle). Paramiopsalis sp. MCZ IZ-135070: Dreszer et al., 2015:3,5,6.

(A) (D)

(B) (E)

(F)

(C)

Fig. 9. Paramiopsalis anadonae, sp. nov. (A–C) Holotype male (MCZ 135070) in (A) dorsal; (B) lateral; and (C) ventral view; (D–F) Paratype female (MCZ 135070) in (D) dorsal; (E) lateral; and (F) ventral view. Scale bars = 0.5 mm. 470 Invertebrate Systematics G. Giribet et al.

(A)(B)

(E) (F)

(G)

(H)

Fig. 10. Paramiopsalis anadonae, sp. nov. Male and female (MCZ 135074). (A) Male ventral view; (B) female ventral view; (C) male lateral view; (D) female lateral view; (E) male thoracic complex; (F) female thoracic region; (G) detail of male thoracic complex; (H) detail of female thoracic complex. Scale bars: A–F = 100 mm; G, H =20mm. Systematics and biogeography of Sironidae Invertebrate Systematics 471

(A) (B)

(C)

(E)

(D)

Fig. 11. Paramiopsalis anadonae, sp. nov. Male and female (MCZ 135074). (A) Male anal region; (B) female anal region; (C) male spiracle; (D) male ozophore. Scale bars: A =50mm; B =40mm; C =10mm; D =20mm.

Material examined Concejo de Allende, A. Anadón, G. Giribet, I. Merino-Sáinz leg., 18.v.2011, litter sifting. Holotype. < (MCZ 135070; ex DNA104624), Spain: Asturias: Illano Additional material. Spain: Asturias: Municipio de Illano: 12 specimens (43.3451 , –6.8953 ; 556 m elevation), C. Prieto leg., 15.v.2009 (ex ZUPV- incollectionofUniversidaddeOviedo(GiribetLocality871:43.3195,–6.8712; 4190.29TPH 72684 98759). 576 m elevation), La Montaña, A. Anadón, G. Giribet, I. Merino-Sáinz leg., Paratypes.1, (MCZ 135070; ex DNA104624), Spain: Asturias: Illano 16.v.2011, litter sifting; 12 specimens in collection of Universidad de Oviedo, (43.3451, –6.8953; 556 m elevation) same data as holotype; 1 ,, 2 juveniles Road to Berguño, 8 km S of Cangas del Narcea (Giribet Locality 878: 43.1205 , (MCZ 135070; ex DNA104624; DNA extracted from 1 juvenile), C. Prieto –6.5723 ; 540 m elevation), A. Anadón, G. Giribet, I. Merino-Sáinz leg., leg., 15.v.2009 (ex ZUPV-4190.29TPH 72684 98759); 1 <,1,, 1 juvenile 18.v.2011, litter sifting; 1 , (MCZ 135076; ex MCZ DNA106400; used for (MCZ 135073; ex MCZ DNA106394), Finca Experimental el Carbayal DNA extraction), Monte de San Pedro de Corias, Cangas del Narcea (Giribet (Giribet station 869: 43.3451, –6.8953; 898 m elevation), Municipio de Locality 879: 43.2005 , –6.5311 ; 356 m elevation), A. Anadón, G. Giribet, Illano, A. Anadón, G. Giribet, I. Merino-Sáinz, R. Rosa leg., 16.v.2011, I. Merino-Sáinz leg., 18.v.2011, litter sifting. direct collecting, under a large boulder; 6 <,8,, 2 juveniles (MCZ 135074; ex DNA106395, 1 <,1, mounted for SEM, 1 < for genitalia), La Montaña (Giribet Locality 871: 43.3195, –6.8712; 576 m elevation), Municipio de Diagnosis Illano, A. Anadón, G. Giribet, I. Merino-Sáinz leg., 16.v.2011, litter sifting; < , < Paramiopsalis with a fringed adenostyle – as opposed to the 2 ,10 , 2 juveniles (MCZ 135075; ex DNA106399; 1 used for DNA – – extraction), Road to Berguño, 8 km S of Cangas del Narcea (Giribet Locality plumose one of P. ramulosus and a broad anal gland region 878: 43.1205, –6.5723; 540 m elevation), A. Anadón, G. Giribet, I. Merino- again, differing from the smaller one present in P. ramulosus. Sáinz leg., 18.v.2011, litter sifting; 1 < (MCZ 135077;ex MCZ DNA106401), It resembles P. eduardoi and P. ramblae in these two characters, near Linares (Giribet station 880: 43.2328, –6.5693; 468 m elevation), but has a male leg tarsus IV more elongated than in these species 472 Invertebrate Systematics G. Giribet et al.

(A) (B)

(C)

(D) (E)

Fig. 12. Paramiopsalis anadonae,sp. nov.Male (MCZ135074).(A) Malechelicera; (B) detailof malechelicera; (C) male palp;(D) detailof ﬁrst segmentof male palp; (E) male palp tip. Scale bars: A, C = 100 mm; B, D =20mm; E =10mm.

(0.35 mm long and 0.11 mm wide; L/W = 3.18). With a truncated granular ornamentation throughout its surface and with a posterior region, differing from the other species in this group. subterminal opening (Fig. 11E). Ventral prosomal complex with coxa I free and coxa II fused to coxa III (Fig. 10A, E); Description coxae I, II and IV meet along the midline (Fig. 10E); coxal pores between coxae III and IV (Fig. 10E, G); coxa IV endites running Male adjacent to the midline suture for a length longer than the Total length of male holotype 2.00 mm; width at widest gonostome (Fig. 10E); gonostome semicircular/subtriangular point 0.83 mm; length/width ratio 2.40; width across with horn-like anterior apophyses on the endites of coxa IV ozophores 0.79 mm. Body light brown and legs light orange (Fig. 6E, G). Sternal glands absent. Spiracles circular (in ethanol; Fig. 9A–C); body with tuberculate-microgranulate (Fig. 11E). Sternites 8 and 9 medially fused. Anal plate with a morphology (Fig. 10A, C). Opisthosomal mid-dorsal longitudinal medial longitudinal non-ornamented carina (Fig. 11A). Exocrine sulcus absent. Eyes absent. Ozophores conical, type II sensu glands with multiple openings along a distinctive broad area of Juberthie (1970a), facing laterally but elevated from the margin tergite IX (Fig. 11C). Distal end of the opisthosoma truncated of the scutum (Fig. 10C), with uniform and non-directional (Figs 10A, 11A).

Fig. 13. Paramiopsalis anadonae, sp. nov. Male (MCZ 135074). (A) Leg I; (B) claw of leg I; (C) leg II; (D) detail of solenidia on dorsal side of leg II; (E) claw of leg II; (F) leg III; (G) claw of leg III; (H) leg IV; (I) claw of leg IV. Scale bars: A, C, F, H = 100 mm; B, D, E, G, I =10mm. Systematics and biogeography of Sironidae Invertebrate Systematics 473

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(I) (H) 474 Invertebrate Systematics G. Giribet et al.

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Fig. 14. Paramiopsalis anadonae, sp. nov. Male and female (MCZ 135074). (A) Tarsus of male leg IV; (B) adenostyle; (C) detail of adenostyle tip; (D) tarsus of female leg IV; (E) detail of dorsal side female tarsus IV showing solenidia. Scale bars: A =40mm; B =10mm; C =5mm; D =50mm; E =20mm.

Chelicerae robust without dorsal or ventral processes on the bases (Fig. 15A). Distal part of terminal lobe sub-triangular/ proximal segment (Fig. 12A). Proximal segment ornamented semicircular. Pars apicalis with enlarged, hooked, mobile throughout its surface and distal segment mostly smooth, with digits, almost as tall as the terminal lobe. Ventral plate sparse ornamentation on dorsal side (Fig. 12A). Chelicerae (Fig. 15B) with three ventral microtrichia ~0.23 mm long and measurements of male paratype (in mm length/width): with wide bases; the central microtrichia in a more basal position proximal segment 0.54/0.14; second segment 0.50/0.12. than the lateral ones. Movable and fixed cheliceral fingers with uniform dentition (Fig. 12A, B). Movable finger 0.17 mm long. Pedipalp with scale-like cuticle; dorsal and ventral sides of palp trochanter Female with thorn-like apophyses (Fig. 12C, D). Palp trochanter with Total length of female paratype (in mm) 2.31; width at widest large denticles both in the ventral and dorsal bulging parts. Palp point 0.88; length/width ratio 2.63 (Fig. 9D–F); width across distal end without modifications (Fig. 12E). Leg segments ozophores 0.81 mm (Fig. 9D). Ventral prosomal complex with densely ornamented with exception of metatarsi I and II and coxae II and III meeting along the midline (Fig. 10F); gonostome tarsi of all legs, which have a smooth surface (Fig. 13A, C, F, H). sub-pentagonal with an elevated posterior margin (Fig. 10F, H). Tarsus of leg I with dorsal solenidia and without a ventral solea Anal region without modifications (Fig. 13B). Tarsi of legs III and (Fig. 13A); tarsus of leg II with dorsal solenidia (Fig. 13D); IV with abundant dorsal solenidia (Fig. 14D, E). Ovipositor not tarsus IV entire; claws of all legs smooth (without teeth, pegs or studied. any other modifications; Fig. 13B, E, G, I); adenostyle located one-third from the proximal end of tarsus (Figs 13H, 14A), fringed but not plumose, with a subterminal pore (Fig. 14A–C). Distribution Table 4 shows appendage measurements in mm. This species is known from multiple locations along the valleys of Spermatopositor (Fig. 15) 0.31 mm long, 0.13 mm wide. the rivers Navia and Narcea, which for some distance run along Dorsal plate with 4/4 long microtrichia (~0.15 mm) with wide parallel valleys at ~25 km apart (Fig. 1D, orange triangles). Systematics and biogeography of Sironidae Invertebrate Systematics 475

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Fig. 15. Paramiopsalis anadonae, sp. nov. Spermatopositor. (A) Dorsal view; (B) ventral view. Scale bars = 0.05 mm.

Table 4. Palp and leg measurements (length/width, mm) in Paramiopsalis anadonae, sp. nov. Measurements refer to male paratype mounted for SEM

Trochanter Femur Patella Tibia Metatarsus Tarsus L W L/W L W L/W L W L/W L W L/W L W L/W L W L/W Total Palp 0.14 0.07 2.00 0.27 0.06 4.50 0.18 0.06 3.00 0.20 0.06 3.33 0.21 0.05 4.20 1.00 Leg I 0.12 0.10 1.20 0.45 0.12 3.75 0.20 0.12 1.67 0.26 0.13 2.00 0.20 0.09 2.22 0.41 0.12 3.42 1.64 Leg II 0.35 0.11 3.18 0.17 0.11 1.55 0.24 0.11 2.18 0.13 0.10 1.30 0.36 0.13 2.77 1.25 leg III 0.12 0.10 1.20 0.31 0.11 2.82 0.15 0.11 1.36 0.22 0.11 2.00 0.13 0.08 1.63 0.29 0.09 3.22 1.22 Leg IV 0.20 0.10 2.00 0.35 0.10 3.50 0.20 0.11 1.82 0.23 0.11 2.09 0.15 0.09 1.67 0.35 0.11 3.18 1.48

Remarks Paramiopsalis ramblae Benavides & Giribet, sp. nov. Paramiopsalis anadonae is very close morphologically to (Figs 16–19) P. eduardoi and P. ramblae. It is found along the valleys of http://zoobank.org/NomenclaturalActs/B98D9A17-EBE4-46EF-A1A4- two parallel rivers in Asturias, on both sides but north of 82766E6C58D1 Muniellos, which, interestingly, hosts a different species in Paramiopsalis sp.: Merino Sáinz & Anadón, 2009: 557, 558. this genus, P. ramblae. While this may suggest possible micro-endemism in the area, P. anadonae is sister group to the Material examined Galician species P. eduardoi and not to P. ramblae, therefore Holotype. < (MCZ 133850; ex DNA106396) Spain: Asturias: Reserva probably indicating ancient speciation. Integral de Muniellos (Giribet Locality 874: 43.0359, –6.6942; 824 m elevation), A. Anadón, G. Giribet, I. Merino-Sáinz leg., 17.v.2011, litter sifting. Paratypes. 1 < (MCZ 133852; ex DNA106398, mounted for SEM) Etymology Spain: Asturias: Reserva Integral de Muniellos (Giribet Locality 877: 43.0341, –6.6872), A. Anadón, G. Giribet, I. Merino-Sáinz leg., The species is named after Araceli Anadón, mentor of IMS for her 17.v.2011, under log; 3 juveniles (MCZ 133850; ex DNA106396; 1 help with logistics and ﬁeldwork in Asturias. specimen used for DNA extraction), same locality as the holotype; 1 476 Invertebrate Systematics G. Giribet et al.

(A) broken but distinguishable); coxal pores present between coxae III and IV (Fig. 17C); sternal opisthosomal glands absent. Spiracles circular (Fig. 17H). Sternites 8 and 9 medially fused and tergite IX free (Fig. 17E); anal plate with a longitudinal middle carina (Fig. 17E). Exocrine glands with multiple openings along a distinctive broad elevated area of tergite IX (Fig. 17F); tergite IX depressed forming a bi-lobed posterior end (Fig. 16A, C). (B) Chelicerae with proximal segment ornamented and distal segment smooth (Fig. 18A); proximal segment without prominent dorsal or ventral processes; chelicerae measurements of male paratype (in mm length/width): proximal segment 0.46/ 0.13; second segment 0.55/0.12. Pedipalps smooth throughout their surface. Pedipalp trochanter armed with ventral and dorsal thorn-like denticles (Fig. 18B, C). Legs relatively robust (Fig. 18D, H, K, M). All leg segments ornamented with exception of metatarsi IandIIandtarsi ofall legs,whichhavea smoothcuticle(Fig.18E,I, (C) L, N). Tarsus of leg I with dorsal solenidia (Fig. 18F) and without a solea. Tarsus IV entire, robust (Fig. 18M, N); adenostyle located approximately one-third from the proximal end of tarsus (Fig. 18N), fringed but not plumose, with a subterminal opening on its retrolateral side (Fig. 18P). Appendage measurements (in mm) shown in Table 5. Spermatopositor (Fig. 19) 0.25 mm long, 0.10 mm wide. Dorsal plate with 4/4 dorsal long microtrichia (~0.15 mm) with widened bases (only the lateral ones seen in Fig. 19A). Distal margin of terminal lobe sub-triangular. Pars apicalis with Fig. 16. Paramiopsalis ramblae, sp. nov. (A–C) Holotype male (MCZ short, hooked, mobile digits, not surpassing the terminal lobe. 133850). (A) Dorsal; (B) lateral; and (C) ventral view. Scale bars = 0.5 mm. Ventral plate (Fig. 15B) with three microtrichia (~0.13 mm), with very wide bases; the central microtrichia in a more basal position juvenile (MCZ 133851; ex DNA106397), Reserva Integral de Muniellos than the lateral microtrichia. (Giribet Locality 876: 43.0376, –6.6957), A. Anadón, G. Giribet, I. Merino- Sáinz leg., 17.v.2011, litter sifting. Female Diagnosis Unknown. Paramiopsalis with a fringed adenostyle – as opposed to the plumose one of P. ramulosus – and a broad anal gland region – Distribution again, differing from the smaller one present in P. ramulosus. It resembles P. eduardoi and P. anadonae, sp. nov. in these two This species is only known from a few sites in the Reserva Integral characters, but differs from P. anadonae in the shape of the male de Muniellos or Bosque de Muniellos (Fig. 1D, orange square), tarsus IV, which is less elongated and wider (0.33 mm long and a temperate rainforest dominated by oak and designated a 0.14 mm wide; L/W = 2.36). With a characteristic bi-lobed biosphere reserve by UNESCO in 2000, and since 2003 part posterior end. of the larger Natural Park of Fuentes del Narcea, Degaña e Ibias (see a description of the reserve in Merino Sainz and Anadón Description 2008). A female, probably of this species, was reported by Merino Male Sáinz and Anadón (2009), but was not examined here. Robust sironid. Total length of male holotype 1.56 mm; width at widest point 0.69 mm; length/width ratio 2.26; width across Remarks ozophores 0.63 mm. Body brown and legs light brown in ethanol Paramiopsalis ramblae appears as an early lineage that is sister – (Fig. 16A C); entire body with tuberculate-microgranulate group to a clade formed by P. eduardoi from Galicia and cuticle (Fig. 17A, B). Opisthosomal mid-dorsal longitudinal P. anadonae from Asturias (Figs 3, 4), or as the sister group to sulcus absent (Fig. 16A). Eyes absent. Ozophores of type II all other Paramiopsalis species (Fig. 2). It is not sister group to its (Juberthie 1970a), entirely ornamented and with a subterminal closest geographic species, P. anadonae. opening (Fig. 17H). Ventral prosomal complex with coxa I free and coxa II fused to coxa III (Fig. 17A, C); coxae I, II and IV meet along the midline (Fig. 17C); coxa IV endites running Etymology adjacent to the midline suture for a length longer than the This species is named after the late Maria Rambla, who pioneered gonostome (Fig. 17C); gonostome semicircular/subtriangular Cyphophthalmi work in the Iberian Peninsula, and who inspired with anterior apophyses on the endites of coxa IV (Fig. 17C, D; generations of Opiliones workers, including G.G. Systematics and biogeography of Sironidae Invertebrate Systematics 477

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Fig. 17. Paramiopsalis ramblae, sp. nov. Male (MCZ 133852). (A) Lateral view; (B) ventral view; (C) thoracic complex; (D) gonostome; (E) anal region; (F) detail of anal region; (G) spiracle; (H) ozophore. Scale bars: A–C = 100 mm; D, E =20mm; F–H =10mm. 478 Invertebrate Systematics G. Giribet et al.

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Fig. 18. Paramiopsalis ramblae, sp. nov. Male (MCZ 133852). (A) Chelicera; (B) palp; (C) palp ﬁrst segment; (D) leg I; (E) tarsus I; (F) dorsal solenidia in tarsus I; (G) claw I; (H) leg II; (I) tarsus II; (J) claw II; (K) leg III; (L) tarsus III; (M) leg IV; (N) tarsus IV; (O) claw IV; (P) adenostyle. Scale bars: A–D, H,K,M = 100 mm; C, E, I, L, N =20mm; F, G, J, O, P =10mm. Systematics and biogeography of Sironidae Invertebrate Systematics 479

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Fig. 19. Paramiopsalis ramblae, sp. nov. Spermatopositor. (A) Dorsal view; (B) ventral view. Scale bars = 0.05 mm.

Table 5. Palp and leg measurements (length/width, mm) in Paramiopsalis ramblae, sp. nov. Measurements refer to male paratype mounted for SEM

Trochanter Femur Patella Tibia Metatarsus Tarsus L W L/W L W L/W L W L/W L W L/W L W L/W L W L/W Total Palp 0.13 0.70 0.19 0.26 0.07 3.71 0.18 0.06 3.00 0.19 0.06 3.17 0.19 0.05 3.80 0.95 Leg I 0.08 0.09 0.89 0.36 0.11 3.27 0.18 0.10 1.80 0.23 0.09 2.56 0.14 0.08 1.75 0.32 0.11 2.91 1.31 Leg II 0.10 0.09 1.11 0.33 0.10 3.30 0.16 0.11 1.45 0.23 0.12 1.92 0.14 0.09 1.56 0.33 0.12 2.75 1.29 leg III 0.31 0.11 2.82 0.16 0.11 1.45 0.19 0.12 1.58 0.13 0.09 1.44 0.29 0.10 2.90 1.08 Leg IV 0.17 0.09 1.89 0.34 0.10 3.40 0.19 0.12 1.58 0.22 0.12 1.83 0.12 0.10 1.20 0.33 0.14 2.36 1.37

Genus Siro Latreille Material examined Type species: Siro rubens Latreille, 1804. Holotype. < (MCZ 130001; ex. MCZ DNA101617) USA: Oregon: Species included: Siro rubens Latreille, 1804; S. acaroides (Ewing, 1923); Clatsop: Ecola State Park (Giribet Locality 383: 45.9157 , –123.9645 ;62m S. boyerae Giribet & Shear, 2010; S. calaveras Giribet & Shear, 2010; elevation), S. L. Boyer, R. M. Clouse, G. Giribet leg., 20.vi.2005 S. carpaticus Rafalski, 1956; S. clousi Giribet & Shear, 2010; S. crassus Paratypes. 3 <,3, (MCZ 130001; ex. MCZ DNA101617; 1 < used for Novak & Giribet, 2006; S. exilis Hoffman, 1963; S. kamiakensis (Newell, DNA extraction and genitalia, 1 < and 1 , mounted for SEM), same data as 1943); Siro ligiae Giribet, sp. nov.; Siro richarti Benavides & Giribet, holotype. sp. nov.; S. shasta Giribet & Shear, 2010; S. sonoma Shear, 1980; S. valleorum Chemini, 1990. Diagnosis Small Siro with an entire tarsus IV, similar to S. acaroides and Siro ligiae Giribet, sp. nov. S. boyerae, from which it is distinguished by its slenderer body, with a length/width ratio of 2.34 in S. ligiae, sp. nov., versus 1.84 – (Figs 20 24) in S. boyerae and 1.5 in S. acaroides. The number of dorsal http://zoobank.org/NomenclaturalActs/FA7BF30D-25CA-44DC-B436- microtrichiae is also larger in S. ligiae (8+8) than in S. boyerae 68A56412332E (5+5). It is also distinguished from S. acaroides in the spiracles, Siro boyerae Giribet & Shear, 2010, partim. which are open in S. acaroides, but circular in S. ligiae and 480 Invertebrate Systematics G. Giribet et al.

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Fig. 20. Siro ligiae, sp. nov. (A–C) Holotype male (MCZ 130001). (A) Dorsal view; (B) lateral view; and (C) ventral view; (D–F) Paratype female (MCZ 130001). (D) Dorsal view; (E) lateral view; and (F) ventral view. Scale bars = 0.5 mm.

S. boyerae. For differences with all other North American Siro, chelicerae measurements of male paratype (in mm length/ see Giribet and Shear (2010). width): proximal segment 0.54/0.16; distal segment 0.68/0.17; proximal segment sparsely ornamented and distal segment fi fi Description smooth (Fig. 23A); movable and xed cheliceral ngers with uniform dentition (Fig. 23B). Pedipalps smooth; ventral side of Male palp trochanter without apophyses (Fig. 23C). Legs relatively Total length of male holotype 1.87 mm; width at widest point slender (Fig. 23D–G); all leg segments densely ornamented with 0.80 mm; length/width ratio 2.34; width across ozophores exception of tarsi and metatarsi I and II and tarsi III and IV, which 0.70 mm. Body light brown and legs light orange (in ethanol; have a smooth cuticle with a scale-like surface (Fig. 23D–G); Fig. 20A–C); body with tuberculate-microgranulate surface tarsus of leg I with dorsal solenidia and without a solea; tarsus (Fig. 21A–C). Opisthosomal mid-dorsal longitudinal sulcus of leg II with solenidia; tarsus of leg IV entire (Fig. 23G–H); absent. Eyes absent. Ozophores of type II sensu Juberthie adenostyle located two-fifths from the proximal end of the tarsus (1970a), with spiral ornamentation throughout their surface (Fig. 23I), lamelliform and with a subterminal opening (Fig. 23J). (Fig. 22F). Ventral prosomal complex with coxae I and II free Appendage measurements (in mm) are shown in Table 6. and coxae III and IV fused (Fig. 21A, E); coxae I, II and IV Spermatopositor (Fig. 24) short, 0.25 mm long, 0.11 mm meet along the midline (Fig. 21E); coxa IV endites running wide, smooth, typical of sironids; with two moveable fingers adjacent to the midline suture for a length longer than the recurved outward, ending as hooks, about the same length gonostome (Fig. 21E); gonostome semicircular, with anterior as the membranous ventral lobe; microtrichial formula 4, 6, apophyses on the endites of coxa IV (Fig. 21G, broken but 8+8 (n = 1). distinguishable); coxal pores between coxae III and IV (Fig. 21E); sternal opisthosomal glands absent. Spiracles circular (Fig. 22E); corona analis formed by the fusion of Female sternites 8 and 9 and tergite IX (Fig. 22A); anal plate with a Total length of female paratype 1.73 mm; width at widest point longitudinal middle carina lacking ornamentation and three 0.76 mm; length/width ratio 2.27; width across ozophores exocrine glands on tergite VIII facing ventrally (Fig. 22A, C, D). 0.67 mm (Fig. 20D–F); ventral prosomal complex with coxae Chelicerae robust; with a ventral process towards the base I, II and III meeting along the midline (Fig. 21F); gonostome of the proximal segment but no dorsal process (Fig. 23A); sub-pentagonal with an elevated posterior margin (Fig. 21H). Systematics and biogeography of Sironidae Invertebrate Systematics 481

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Fig. 21. Siro ligiae, sp. nov. Male and female (MCZ 130001). (A) Male ventral view; (B) female ventral view; (C) male lateral view; (D) female lateral view; (E) male thoracic complex; (F) female thoracic complex; (G) detail of male thoracic complex; (H) detail of female thoracic complex. Scale bars: A–D = 200 mm; E, F = 100 mm; G, H =20mm. 482 Invertebrate Systematics G. Giribet et al.

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Fig. 22. Siro ligiae, sp. nov. Male and female (MCZ 130001). (A) Male anal region; (B) female anal region; (C) detail of male anal plate; (D) detail of male anal region showing anal glands; (E) male spiracle; (F) male ozophore. Scale bars: A, B =40mm; C = 50; D =5mm; E =10mm; F =20mm. Systematics and biogeography of Sironidae Invertebrate Systematics 483

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Fig. 23. Siro ligiae, sp. nov. Male and female (MCZ 130001). (A) Male chelicerae; (B) detail male palp; (C) male palp; (D) male leg I; (E) male leg II; (F) male leg III; (G) male leg IV; (H) claw of male leg IV; (I) male tarsus IV; (J) adenostyle; (K) female tarsus IV. Scale bars: A, C–G = 100 mm; B =20mm; H–K =10mm. 484 Invertebrate Systematics G. Giribet et al.

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Fig. 24. Siro ligiae, sp. nov. Spermatopositor. (A) Dorsal view; (B) ventral view. Scale bars = 0.05 mm.

Table 6. Palp and leg measurements (length/width, mm) in Siro ligiae, sp. nov. Measurements refer to male paratype mounted for SEM

Trochanter Femur Patella Tibia Metatarsus Tarsus L W L/W L W L/W L W L/W L W L/W L W L/W L W L/W Total Palp Leg I 0.13 0.12 1.08 0.45 0.13 3.46 0.21 0.13 1.62 0.29 0.12 2.42 0.19 0.11 1.73 0.40 0.13 3.08 1.67 Leg II 0.13 0.13 1.00 0.40 0.12 3.33 0.16 0.13 1.23 0.27 0.14 1.93 0.17 0.11 1.55 0.39 0.13 3.00 1.52 leg III 0.16 0.10 1.60 0.26 0.12 2.17 0.18 0.13 1.38 0.21 0.15 1.40 0.16 0.09 1.78 0.32 0.09 3.56 1.29 Leg IV 0.40 0.13 3.08 0.24 0.15 1.60 0.26 0.15 1.73 0.18 0.11 1.64 0.39 0.15 2.60 1.47

Anal region without conspicuous modiﬁcations (Fig. 22B). analyses S. ligiae appears instead as the sister group of the eastern Tarsus of leg IV cylindrical, smooth (Fig. 23K). Ovipositor North American S. exilis (Fig. 2). not studied.

Etymology Distribution The species is named after Ligia R. Benavides for her dedication This species is known from its type locality only (Fig. 1H, blue to Cyphophthalmi. This species was recognised before this paper triangle). was put together and the species author wished to retain the name that has been used since realising that this was a cryptic species Remarks originally considered S. boyerae Giribet & Shear, 2010. Siro ligiae was originally thought to be conspeciﬁc with S. boyerae (Giribet and Shear 2010), from which it is nearly Siro richarti Benavides & Giribet, sp. nov. indistinguishable morphologically, but has subsequently been – found to be quite distinct molecularly (unpubl. data), as illustrated (Figs 25 29) here with the deep divergence shown between the representatives http://zoobank.org/NomenclaturalActs/A3E4A36B-0C21-49F5-A79B- of these putative sister species (Figs 3, 4), although in some 6287F77DF3C4 Systematics and biogeography of Sironidae Invertebrate Systematics 485

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(B) (E)

Fig. 25. Siro richarti, sp. nov. (A–C) Holotype male (MCZ 134870). (A) Dorsal view; (B) lateral view; and (C) ventral view; (D–F) Paratype female (MCZ 134870). (D) Dorsal view; (E) lateral view; and (F) ventral view. Scale bars = 0.5 mm.

Material examined ozophores 0.63 mm. Body light orange and legs light chestnut – Holotype. < (MCZ 134487; ex MCZ DNA105684) USA: Nez Perce (in ethanol; Fig. 25A C); entire body with tuberculate- – National Forest: Idaho: FS Rd 311 via Crooked River Rd, 13 mi [~21 km] S microgranulate surface (Fig. 26A C). Opisthosomal mid-dorsal of SR 14 (45.6853, –115.5427; 1460 m elevation), C. Richart, longitudinal sulcus absent (Fig. 25A). Eyes absent. Ozophores S. Derkarabetian, J. Underwood leg., 9.vii.2008 (C. Richart code CHR2733). of type II sensu Juberthie (1970a), with spiral ornamentation Paratypes. 1 <,2, (MCZ 134487; ex MCZ DNA105684) (C. Richart throughout their surface (Fig. 27E). Ventral prosomal complex code CHR2733), same collecting data as holotype; 1 < (MCZ 134488; ex with coxae I and II free (Fig. 26A–C); coxae I, II and IV meet along MCZ DNA105692, used for DNA extraction and genitalia), USA: Nez Perce the midline; coxa IV endites run adjacent to the midline suture for National Forest: Idaho: FS Rd 311 via Crooked River Rd, 13 mi S of SR 14 a length much longer than the gonostome (Fig. 26E); gonostome – (45.6853 , 115.5427 ; 1460 m elevation), C. Richart, S. Derkarabetian, semicircular with anterior apophyses on the endites of coxa IV J. Underwood leg., 9.vii.2008 (C. Richart code CHR2732), Picea forest, in the anterior part of gonostome (Fig. 26E, G); coxal pores stream-side woody debris. between coxae III and IV (Fig. 26E). Sternal opisthosomal glands Diagnosis absent. Spiracles circular (Fig. 27D). Corona analis formed by the Siro with a divided male tarsus IV, as in S. kamiakensis, but with a fusion of sternites 8 and 9 and tergite IX (Fig. 27A); posterior end smaller male gonostome and larger coxa IV in S. richarti, sp. nov., of the opisthosoma truncated and deeply concave; anal plate which overgrows coxa III and touches the endites of coxa II, semicircular, with two bi-concave depressions (Fig. 27A); three unlike in S. kamiakensis. It also differs from S. kamiakensis in the anal glands on a distinctive smooth area of tergite IX and facing male anal region, where the distal area of the opisthosoma posteriorly. in S. richarti is truncated with a bi-concave smooth anal plate, Chelicerae robust (Fig. 28A). Chelicerae measurements (in while in S. kamiakensis the posterior is unmodified, and the anal mm): proximal segment broken, length measurements not plate has a broad longitudinal carina and conspicuous lateral provided, 0.19 wide; distal segment 0.70 long, 0.17 wide; ornamentation. The orientation of the three anal gland openings proximal segment with sparse ornamentation on the ventral fi also differs between these two species, facing posteriorly in side; distal segment smooth; movable and xed cheliceral fi S. richarti and ventrally in S. kamiakensis. ngers with uniform dentition (Fig. 28A). Pedipalps smooth; ventral side of palp trochanter with thorn-like cuticular spines. – Description Legs slender (Fig. 28D G) with all leg segments densely ornamented with exception of tarsi and metatarsi I and II and Male tarsi of legs III and IV, which have a smooth surface; leg I without Total length of male holotype 1.66 mm; width at widest a solea. Claws of all legs smooth, without teeth or lateral pegs point 0.68 mm; length/width ratio 2.44; width across (i.e. Fig. 28E). Tarsus of leg IV bipartite (Fig. 28G, H); adenostyle 486 Invertebrate Systematics G. Giribet et al.

(A)(B)

(C)(D)

(E)(F)

(G)(H)

Fig. 26. Siro richarti, sp. nov. Male and female (MCZ 134487). (A) Male ventral view; (B) female ventral view; (C) male lateral view; (D) female lateral view; (E) male thoracic complex; (F) female thoracic complex; (G) detail of male thoracic complex; (H) detail of female thoracic complex. Scale bars: A–D = 200 mm; E, F = 100 mm; G, H =20mm. Systematics and biogeography of Sironidae Invertebrate Systematics 487

(A)(B)

(E)

Fig. 27. Siro richarti, sp. nov. Male and female (MCZ 134487). (A) Male anal region; (B) female anal region; (C) male spiracle; (D) male ozophore. Scale bars: A =40mm; B =50mm; C =10mm; D =20mm. lamelliform located two-thirds from the proximal edge of the first (Fig. 26F); gonostome sub-pentagonal with an elevated (most proximal) tarsus segment (Fig. 28H); with a subterminal posterior margin (Fig. 26H). Anal region without conspicuous opening (Fig. 28J). Appendage measurements (in mm) shown in modifications (Fig. 27B). Tarsus of leg IV undivided, cylindrical, Table 7. smooth (Fig. 28K). Spermatopositor (Fig. 29) short: 0.20 mm long, 0.11 mm wide; Ovipositor not studied. smooth, typical of sironids; with two moveable fingers curved outwards, ending as hooks much longer than the membranous Distribution ventral lobe; microtrichial formula 4, 6, 5+5 (n = 1). This species is known from its type locality only (Fig. 1H, blue square). Female Total length of female paratype (in mm): 1.81; width at widest Remarks point: 0.71; length/width ratio: 2.54; width across ozophores As in S. kamiakensis (Newell, 1943), S. richarti has a divided 0.66 mm (Fig. 25D, F); coxae I and II meet along the midline male tarsus IV; however, these two species form a grade at the 488 Invertebrate Systematics G. Giribet et al.

(A) (G)

(B)

(H)

(E)

(J)

(F)

(K)

Fig. 28. Siro richarti, sp. nov. Maleand female(MCZ 134487).(A) Male chelicera; (B) male palp; (C) male palp trochanter; (D) male leg I; (E) male leg II; (F) male leg III; (G) male leg IV; (H) male claw leg IV; (I) male metatarsus and tarsus IV; (J) adenostyle; (K) female tarsus IV. Scale bars: A, B, D–G = 100 mm; C =20mm; H, K =50mm; I, J =10mm. Systematics and biogeography of Sironidae Invertebrate Systematics 489

Table 7. Palp and leg measurements (length/width, mm) in Siro richarti, sp. nov. Measurements refer to male paratype mounted for SEM

Trochanter Femur Patella Tibia Metatarsus Tarsus L W L/W L W L/W L W L/W L W L/W L W L/W L W L/W Total Palp 0.21 0.07 3.00 0.38 0.07 5.43 0.25 0.05 5.00 0.27 0.07 3.86 0.24 0.07 3.43 1.35 Leg I 0.15 0.12 1.25 0.51 0.12 4.25 0.24 0.13 1.85 0.36 0.12 3.00 0.22 0.09 2.44 0.45 0.13 3.46 1.93 Leg II 0.16 0.12 1.33 0.47 0.11 4.27 0.21 0.13 1.62 0.29 0.14 2.07 0.19 0.09 2.11 0.44 0.11 4.00 1.76 leg III 0.33 0.12 2.75 0.24 0.12 2.00 0.25 0.13 1.92 0.17 0.08 2.13 0.34 0.09 3.78 1.33 leg IV 0.25 0.13 1.92 0.42 0.13 3.23 0.25 0.13 1.92 0.29 0.13 2.23 0.18 0.10 1.80 0.38 0.15 2.53 1.77

(A)(B)

Fig. 29. Siro richarti, sp. nov. Spermatopositor. (A) Dorsal view; (B) ventral view. Scale bars = 0.05 mm.

base of the clade including S. exilis, S. boyerae and S. ligiae, specimens of a new species. Assistance from the Harvard Center of Nanoscale suggesting that the divided male tarsus IV may have appeared Systems (CNS) and the Harvard Center for Biological Imaging (HCBI) was twice in this clade or been lost in the common ancestor of the instrumental for this study. Erin McIntyre assisted with DNA sequencing; geographically widespread clade including S. exilis plus the Sebastian Gliem and Douglas Richardson (HCBI) provided support with the cryptic species S. boyeare and S. ligiae. confocal microscope. We would like to thank Associate Editor Mark Harvey and two anonymous reviewers for their comments that improved this manuscript. This work was supported by internal funds from the MCZ and Etymology by NSFgrants(nos0236871and1144417)toGG. Thisworkwasconductedin The species is named after Casey Richart, for providing the part at the Center for Nanoscale Systems (CNS, Harvard), a member of the specimens for study and in recognition of his contributions to National Nanotechnology Coordinated Infrastructure Network (NNCI), the study of US leaf litter Opiliones. which is supported by the National Science Foundation under NSF award no. 1541959. Acknowledgements References Several colleagues and students have helped to collect sironids for more than a decade, and they all are acknowledged here. Special thanks to Araceli Anadón Benavides, L. R., and Giribet, G. (2013). A revision of selected clades of and Rocío Rosa for their assistance with ﬁeldwork in Asturias that resulted in Neotropical mite harvestmen (Arachnida, Opiliones, Cyphophthalmi, three of the species described here, and to Carlos Prieto, who made us aware of Neogoveidae) with the description of eight new species. Bulletin of the the new material being collected in Asturias. To Sarah Boyer and Ron Clouse Museum of Comparative Zoology 161,1–44. doi:10.3099/0027-4100- for their ﬁeld company in the NW USA and to Casey Richart for providing 161.1.1 490 Invertebrate Systematics G. Giribet et al.

Boyer, S. L., and Giribet, G. (2007). A new model Gondwanan taxon: A molecular phylogeny of the temperate Gondwanan family Pettalidae systematics and biogeography of the harvestman family Pettalidae (Arachnida, Opiliones, Cyphophthalmi) with biogeographic and (Arachnida, Opiliones, Cyphophthalmi), with a taxonomic revision of taxonomic implications. Zoological Journal of the Linnean Society genera from Australia and New Zealand. Cladistics 23, 337–361. 178, 523–545. doi:10.1111/zoj.12419 doi:10.1111/j.1096-0031.2007.00149.x Hoffman, R. L. (1963). A new phalangid of the genus Siro from eastern United Boyer, S. L., Karaman, I., and Giribet, G. (2005). The genus Cyphophthalmus States, and taxonomic notes on other American sironids (Arach., (Arachnida, Opiliones, Cyphophthalmi) in Europe: a phylogenetic Opiliones). Senckenbergiana Biologica 44, 129–139. approach to Balkan Peninsula biogeography. Molecular Phylogenetics Juberthie, C. (1961). Étude des opilions cyphophthalmes (arachnides) and Evolution 36, 554–567. doi:10.1016/j.ympev.2005.04.004 du Portugal: description d’Odontosiro lusitanicus g. n., sp. n. Bulletin Boyer, S. L., Clouse, R. M., Benavides, L. R., Sharma, P., Schwendinger, P. J., du Muséum National d’Histoire Naturelle, 2e série 33, 512–519. Karunarathna, I., and Giribet, G. (2007). Biogeography of the world: Juberthie, C. (1962). Étude des opilions cyphophthalmes Stylocellinae du a case study from cyphophthalmid Opiliones, a globally distributed group Portugal. Description de Paramiopsalis ramulosus gen. n., sp. n. Bulletin of arachnids. Journal of Biogeography 34, 2070–2085. doi:10.1111/ du Muséum National d’Histoire Naturelle, 2e série 34, 267–275. j.1365-2699.2007.01755.x Juberthie, C. (1970a). Les genres d’opilions Sironinae (Cyphophthalmes). Castresana, J. (2000). Selection of conserved blocks from multiple alignments Bulletin du Muséum National d’Histoire Naturelle, 2e série 41, for their use in phylogenetic analysis. Molecular Biology and Evolution 1371–1390. 17, 540–552. doi:10.1093/oxfordjournals.molbev.a026334 Juberthie, C. (1970b). Sur Suzukielus sauteri (Roewer, 1916) opilion Clouse, R. M., and Giribet, G. (2010). When Thailand was an island – the cyphophthalme du Japon. Revue d’Écologie et de Biologie du Sol 7, phylogeny and biogeography of mite harvestmen (Opiliones, 563–569. Cyphophthalmi, Stylocellidae) in Southeast Asia. Journal of Juberthie, C., and Massoud, Z. (1976). Biogéographie, taxonomie et Biogeography 37, 1114–1130. doi:10.1111/j.1365-2699.2010.02274.x morphologie ultrastructurale des opilions cyphophthalmes. Revue de Bivort, B. L., and Giribet, G. (2004). A new genus of cyphophthalmid d’Écologie et de Biologie du Sol 13, 219–231. from the Iberian Peninsula with a phylogenetic analysis of the Sironidae Karaman, I. M. (2009). The taxonomical status and diversity of Balkan (Arachnida: Opiliones: Cyphophthalmi) and a SEM database of sironids (Opiliones, Cyphophthalmi) with descriptions of twelve new external morphology. Invertebrate Systematics 18,7–52. doi:10.1071/ species. Zoological Journal of the Linnean Society 156, 260–318. IS03029 doi:10.1111/j.1096-3642.2009.00446.x Dreszer, T. B., Rađa, T., and Giribet, G. (2015). Cyphophthalmus solentiensis Katoh, K., and Standley, D. M. (2013). MAFFT multiple sequence alignment sp. nov. (Cyphophthalmi, Sironidae) a new endogean mite harvestman software version 7: improvements in performance and usability. species from Croatia, with an application of confocal laser microscopy Molecular Biology and Evolution 30, 772–780. doi:10.1093/molbev/ to illustrate genitalia in Opiliones. Breviora 543,1–15. doi:10.3099/ mst010 MCZ18.1 Lanfear, R., Calcott, B., Ho, S. Y., and Guindon, S. (2012). PartitionFinder: Ewing, H. E. (1923). Holosiro acaroides, new genus and species, – the only combined selection of partitioning schemes and substitution models for new world representative of the mite-like phalangids of the suborder phylogenetic analyses. Molecular Biology and Evolution 29, 1695–1701. Cyphophthalmi. Annals of the Entomological Society of America 16, doi:10.1093/molbev/mss020 387–391. doi:10.1093/aesa/16.4.387 Merino Sainz, I., and Anadón, A. (2008). La fauna de Opiliones (Arachnida) Fernández, R., Sharma, P. P., Tourinho, A. L., and Giribet, G. (2017). The de la Reserva Integral Natural de Muniellos (Asturias) y del noroeste de la Opiliones Tree of Life: shedding light on harvestmen relationships through Península Ibérica. Boletin de la SEA 43, 199–210. transcriptomics. Proceedings of the Royal B: Biological Sciences 284, Merino Sáinz, I., and Anadón, A. (2009). Primera cita del género 20162340. doi:10.1098/rspb.2016.2340 Paramiopsalis Juberthie, 1962 (Arachnida: Opiliones, Sironidae) para Giribet, G., and Boyer, S. L. (2002). A cladistic analysis of the Asturias (España). Boletin de la SEA 45, 556–558. cyphophthalmid genera (Opiliones, Cyphophthalmi). The Journal of Miller, M. A., Pfeiffer, W., and Schwartz, T. (2010). Creating the CIPRES Arachnology 30, 110–128. doi:10.1636/0161-8202(2002)030[0110: science gateway for inference of large phylogenetic trees. In ‘Proceedings ACAOTC]2.0.CO;2 of the Gateway Computing Environments Workshop (GCE), New Giribet, G., and Prieto, C. E. (2003). A new Afrotropical Ogovea (Opiliones, Orleans’. pp. 1–8. Cyphophthalmi) from Cameroon, with a discussion on the taxonomic Murienne, J., and Giribet, G. (2009). The Iberian Peninsula: ancient history of characters in the family Ogoveidae. Zootaxa 329,1–18. a hot spot of mite harvestmen (Arachnida: Opiliones: Cyphophthalmi: Giribet, G., and Shear, W. A. (2010). The genus Siro Latreille, 1796 Sironidae) diversity. Zoological Journal of the Linnean Society 156, (Opiliones, Cyphophthalmi, Sironidae), in North America with a 785–800. doi:10.1111/j.1096-3642.2008.00512.x phylogenetic analysis based on molecular data and the description of Murienne, J., Karaman, I., and Giribet, G. (2010). Explosive evolution of an four new species. Bulletin of the Museum of Comparative Zoology 160, ancient group of Cyphophthalmi (Arachnida: Opiliones) in the Balkan 1–33. doi:10.3099/0027-4100-160.1.1 Peninsula. Journal of Biogeography 37,90–102. doi:10.1111/j.1365- Giribet, G., Vogt, L., Pérez González, A., Sharma, P., and Kury, A. B. (2010). 2699.2009.02180.x A multilocus approach to harvestman (Arachnida: Opiliones) phylogeny Murphree, C. S. (1988). Morphology of the dorsal integument of ten opilionid with emphasis on biogeography and the systematics of Laniatores. species(Arachnida,Opiliones). The Journal of Arachnology 16, 237–252. Cladistics 26, 408–437. doi:10.1111/j.1096-0031.2009.00296.x Novak, T., and Giribet, G. (2006). A new species of Cyphophthalmi Giribet, G., Sharma, P. P., Benavides, L. R., Boyer, S. L., Clouse, R. M., de (Arachnida, Opiliones, Sironidae) from Eastern Slovenia. Zootaxa Bivort, B. L., Dimitrov, D., Kawauchi, G. Y., Murienne, J. Y., and 1330,27–42. Schwendinger, P. J. (2012). Evolutionary and biogeographical history Rambaut, A., Suchard, M. A., Xie, D., and Drummond, A. J. (2014). Tracer of an ancient and global group of arachnids (Arachnida: Opiliones: v1.6. Available at http://beast.bio.ed.ac.uk/Tracer [veriﬁed 7 April 2017]. Cyphophthalmi) with a new taxonomic arrangement. Biological Rambla, M., and Fontarnau, R. (1984). Les Opilions Cyphophthalmes Journal of the Linnean Society. Linnean Society of London 105, (Arachnida) de la faune ibérique: I. Sur Paramiopsalis ramulosus 92–130. doi:10.1111/j.1095-8312.2011.01774.x Juberthie, 1962. Revue Arachnologique 5, 145–152. Giribet, G., Boyer, S. L., Baker, C., Fernández, R., Sharma, P. P., de Bivort, Rambla, M., and Fontarnau, R. (1986). Les Opilions Cyphophthalmes B. L., Daniels, S. R., Harvey, M. S., and Griswold, C. E. (2016). (Arachnida) de la faune ibérique: III. Sur Odontosiro lusitanicus Systematics and biogeography of Sironidae Invertebrate Systematics 491

Juberthie, 1961. Mémoirs de la Société Royale Belge d’Entomologie 33, Stamatakis, A., Hoover, P., and Rougemont, J. (2008). A rapid bootstrap 171–178. algorithm for the RAxML Web servers. Systematic Biology 57, 758–771. Ronquist, F., Teslenko, M., van der Mark, P., Ayres, D. L., Darling, A., doi:10.1080/10635150802429642 Höhna, S., Larget, B., Liu, L., Suchard, M. A., and Huelsenbeck, J. P. Talavera, G., and Castresana, J. (2007). Improvement of phylogenies after (2012). MrBayes 3.2: efﬁcient Bayesian phylogenetic inference and removing divergent and ambiguously aligned blocks from protein model choice across a large model space. Systematic Biology 61, sequence alignments. Systematic Biology 56, 564–577. doi:10.1080/ 539–542. doi:10.1093/sysbio/sys029 10635150701472164 Sharma, P. P., and Giribet, G. (2009). The family Troglosironidae Vaidya, G., Lohman, D. J., and Meier, R. (2011). SequenceMatrix: (Opiliones: Cyphophthalmi) of New Caledonia. In ‘Zoologia concatenation software for the fast assembly of multi-gene datasets Neocaledonica 7. Biodiversity Studies in New Caledonia’. (Ed. with character set and codon information. Cladistics 27, 171–180. P. Grandcolas.) Mémoires du Muséum national d’Histoire naturelle doi:10.1111/j.1096-0031.2010.00329.x 198,83–123. Wheeler, W. C. (1995). Sequence alignment, parameter sensitivity, and the Sharma, P. P., Vahtera, V., Kawauchi, G. Y., and Giribet, G. (2011). phylogenetic analysis of molecular data. Systematic Biology 44, 321–331.

Running WILD: the case for exploring mixed parameter sets in doi:10.1093/sysbio/44.3.321 sensitivity analysis. Cladistics 27, 538–549. doi:10.1111/j.1096-0031. Wheeler, W. C., Lucaroni, N., Hong, L., Crowley, L. M., and Varón, A. 2010.00345.x (2015). POY version 5: phylogenetic analysis using dynamic homologies Shear, W. A. (1980). A review of the Cyphophthalmi of the United States and under multiple optimality criteria. Cladistics 31, 189–196. doi:10.1111/ Mexico, with a proposed reclassiﬁcation of the suborder (Arachnida, cla.12083 Opiliones). American Museum Novitates 2705,1–34. Willemart, R. H., and Giribet, G. (2010). A scanning electron microscopic Shear, W. A. (1985). Marwe coarctata, a remarkable new cyphophthalmid survey of the cuticle in Cyphophthalmi (Arachnida, Opiliones) with the from a limestone cave in Kenya (Arachnida, Opiliones). American description of novel sensory and glandular structures. Zoomorphology Museum Novitates 2830,1–6. 129, 175–183. doi:10.1007/s00435-010-0110-z Stamatakis, A. (2014). RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30, 1312–1313. doi:10.1093/bioinformatics/btu033 Handling editor: Mark Harvey