Finding Padaeus bovillus (Hemiptera: Pentatomidae): A Phylogenetic Placement and the Description of Its Sister Species

© 2021 Academia Sinica, Taiwan Open Access Finding Padaeus bovillus (Hemiptera: Pentatomidae): A Phylogenetic Placement and the Description of Its Sister Species Filipe Michels Bianchi1,* , Verônica Krein1, David Rider2, and Jocelia Grazia1 1Laboratório de Entomologia Sistemática, Departamento de Zoologia, Instituto de Biociências, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brazil. *Correspondence: E-mail: [email protected] (Bianchi) E-mail: [email protected] (Krein); [email protected] (Grazia) 2Entomology Department, School of Natural Resource Sciences, North Dakota State University, Fargo, ND, USA. E-mail: [email protected] (Rider) Received 28 November 2020 / Accepted 14 January 2021 / Published 22 March 2021 Communicated by Y. Miles Zhang The pentatomids (Hemiptera: Heteroptera) are the third most speciose family within the Heteroptera or the true bugs. The family occurs worldwide and comprises around five thousand valid species within 950 genera. Padaeus Stål belongs to a complex of other genera of Carpocorini Mulsant and Rey related to Euschistus Dallas. These genera present similarities in color, size, and shape, and share common features. However, among its four congeneric species, Padaeus bovillus Distant has been highlighted as an outlier by the posterior margins of the bucculae evanescent, while its congeneric species present posterior margins of the bucculae lobed. Thus, herein we redescribe P. bovillus and present a hypothesis regarding its phylogenetic placement within the Carpocorini. Furthermore, a new species similar to P. bovillus is described. Four molecular markers (COI, CytB, 16S, and 28S) plus 86 morphological characters were used to infer the phylogeny under Maximum likelihood and Bayesian inference. For the descriptions, we measured 16 morphometric parameters and dissected the genitalic structures. We also include illustrations of the habitus, internal and external genitalic structures, and provide distribution maps. Mitripus seclusus sp. n. Bianchi, Krein, Rider, and Grazia is recovered as the sister species to Mitripus bovillus comb. n., and both within Mitripus Rolston. Among other shared characters, species of Mitripus have the femora unarmed, they have a macula near the apex of the radial vein, and the mandibular plates tapering apically. Mitripus bovillus comb. n. and Mitripus seclusus sp. n. have the posterior margin of the pygophore projecting as a spine, a unique pattern within the genus. According to our results, Mitripus including M. bovillus comb. n. and Mitripus seclusus sp. n. now includes five species. Key words: Carpocorini, Pentatominae, Phylogenetics, Stink bug, Neotropics. Citation: Bianchi FM, Krein V, Rider D, Grazia J. 2021. Finding Padaeus bovillus (Hemiptera: Pentatomidae): A phylogenetic placement and the description of its sister species. Zool Stud 60:11. doi:10.6620/ZS.2021.60-11. BACKGROUND Within Heteroptera (Insecta: Hemiptera), Pentatomidae is one of the three largest families (Schuh and Weirauch 2020). Pentatomids are known as stink bugs, and the family contains about 950 genera containing almost 5,000 valid species (Rider et al. 2018). Although well-supported monophyly of Pentatomidae has been inferred using distinct sources of data and analytical approaches, the phylogenetic relationships within the family are still unclear (e.g., Henry 1997; Li et al. 2005; Grazia et al. 2008; Wu et al. 2016). For the subordinate taxa within Pentatomidae, phylogenetic hypotheses are scarce (Grazia et al. 2008). The taxonomy and classification of Pentatomidae have mostly been based on morphological similarities. Thus, most tribes and groups of genera have never been studied under a phylogenetic framework, and Zoological Studies 60:11 (2021) doi:10.6620/ZS.2021.60-11 1 © 2021 Academia Sinica, Taiwan the relationships within the family lack phylogenetic hypotheses (Rider et al. 2018). Euschistus Dallas is currently placed in Carpocorini Mulsant and Rey, and it is one of the most speciose genera within the Pentatomidae, containing 67 species (Bianchi et al. 2017a), all of which occur exclusively in the New World (Rolston 1974a). The delimitation of the genus is imprecise, resulting in a considerable progressive accretion of species since its proposition, and also a subsequent creation of new genera for species formerly placed within Euschistus (e.g., Ladeaschistus Rolston, Adustonotus Bianchi) (Rolston 1974a; Bianchi et al. 2017a). Moreover, during the taxonomic history of the Pentatomidae, many genera have been hypothesized to be related to Euschistus (Euschistus group, hereafter), mainly due to morphological similarities and distribution. Barão et al. (2020) recovered the monophyly of the Euschistus group containing 22 genera. Padaeus Stål is one of the genera that Rolston (1974a) speculated might be related to Euschistus; and its placement of Padaeus within Euschistus group was hypothesized by Barão et al. (2020). Currently, Padaeus includes P. bovillus Distant, P. teapensis (Distant), P. trivittatus Stål, P. verrucifer Stål, and the type species P. viduus (Vollenhoven). A diagnostic character of Padaeus is the posterior margins of bucculae are lobed and prolonged to the base of the head (Stål 1862; Rolston 1974a; Rolston and McDonald 1984). However, this feature is not common to all species of Padaeus. Padaeus bovillus has the posterior margins of the bucculae evanescent, similar to many other species of Euschistus group. This condition has put the current placement of P. bovillus in question (e.g., Rolston 1974a; Rolston and McDonald 1984). Even in the original description, Distant (1900) stated that P. bovillus differs from its congeneric species and highlighted its resemblance to Sibaria armata (Dallas). From an in-depth investigation of the morphology of specimens identified as P. bovillus, we discovered that some specimens differed slightly in general somatic characters, and greatly in genitalic structures. Thus, we provide a phylogenetic hypothesis for the systematic placement of P. bovillus and the similar species found by us, redescribe P. bovillus and update the knowledge of this species, and describe the new species similar to P. bovillus. MATERIALS AND METHODS Phylogenetic analyses Bianchi et al. (2017b) provided a backbone for the phylogenetic relationships among genera related to Sibaria and Ladeaschistus. Thus, we used the morphological and molecular matrices of Bianchi et al. (2017b) to serve as the base for our phylogenetic analyses. This matrix contained 32 taxa scored for 85 morphological characters and 2327 bp from the mitochondrial markers cytochrome c oxidase subunit I (COI), cytochrome b (Cytb) and ribosomal 16S, and 28S. Since our focus was to accommodate P. bovillus and a new species, the matrix was reduced to 21 terminal taxa, and P. viduus (the type species of Padaeus), the recently described Sibaria amazonica Krein, Rider & Grazia (see DISCUSSION), P. bovillus and the new species (Table 1) were added. The selection of terminal taxa prioritized availability of molecular data, although we kept the generic sampling. We also proposed an additional morphological character (i.e., character 11: Head, proportion of eyes related to head width: (0) less than 0.43; (1) greater than 0.5), and a new state to character 68 (i.e., Pygophore, ventral rim at middle, form: (5) pointed). Both states for each character were re-evaluated and the scores of the whole matrix were double-checked (Table S1). The specimens used to score the morphological matrix were identified according to the literature (Table 1). The specimens of S. amazonica, P. viduus, P. bovillus and the new species were preserved on pins; they were collected many decades ago (see material examined). The genomic DNA extractions followed Bianchi et al. (2017b) protocol using the DNeasy Blood and Tissue kit (Qiagen, Valencia, CA, USA). The results, however, were of low quality and quantity. Therefore, only the morphological partition was scored for S. amazonica, P. viduus, P. bovillus and the new species. The other four molecular partitions were scored as missing data for these species. Morphological data were coded in Mesquite 3.61 (Maddison and Maddison 2019), and the matrix was exported as a NEXUS file for phylogenetic analyses. Unobserved states were scored with ‘?’ and inapplicable states with ‘–’. All characters were treated as nonadditive (Table 2). Morphological characters analyzed using probabilistic methods were treated under the Mk evolutionary model (Lewis 2001). The alignments of individual molecular markers, evolutionary models for each partition, maximumlikelihood routine (ML) including bootstrap (BS), and other parameters not stated here followed Bianchi et al. (2017b). Bayesian inference (BI) using the concatenated matrix was performed in the multithreading version of the program MrBayes 3.2.0 (Ronquist and Huelsenbeck 2003), setting nst = 1 rates = equal for morphological partition and nst = 6 rates = invgamma for each molecular marker, for 2.5 million generations (nruns = 2 page 2 of 13Zoological Studies 60:11 (2021) © 2021 Academia Sinica, Taiwan nchains = 4) with trees sampled every 1000 generations. Tracer v.1.6.0 (Rambaut et al. 2014) was used to inspect the convergence with the stationary distribution of the chains. The first 20% of the generations were discarded as “burn-in”, and then the chains were combined. The combined ESS values for each parameter were higher than 200. The posterior probability (PP) was estimated for the remaining generations. For both ML and BI values, the nodes presenting PP < 0.50 and BS < 50 were collapsed. Phylogenetic trees were visualized and edited using FigTree v1.4.0 (Rambaut et al. 2014) (http://tree.bio.ed.ac.uk/software/figtree/). Glyphepomis spinosa Campos & Grazia was used to root the trees in both ML and BI analyses based on Bianchi et al. (2017b) and Barão et al. (2020). Taxonomy All the specimens were observed and evaluated using a light stereomicroscope. The measurements of the following 15 morphometric parameters were taken under a light stereomicroscope: total body length (BL) (measured from the apex of head to the apex of the abdomen, excluding the hemelytral membranes), maximum abdominal width (AW), medial length of head (HL) (disc of the head parallel to observer), maximum width of head (HW) (including eyes), length of head anterior to the eyes (LE), interocular distance (ID), length of antennomeres I (I), II (II), III (III), IV (IV), V (V), medial pronotal length (PL), maximum pronotal width (PW), medial scutellar length (SL), and maximum scutellar width (SW). Measurements (mean ± standard deviation) were given in millimeters. The entire abdomen was removed from each female to access the internal genitalia. For males, only the pygophore was removed. Each female abdomen and male pygophore was then cleaned in aqueous supersaturated KOH solution and boiled for about 10 minutes. Female internal genitalic structures were stained with Congo Red. The terminology of genitalic structures follows Baker (1931), Dupuis (1970), and Schaefer (1977), and Tsai et al. (2011) exclusively for parameres. Kment and Vilímová (2010) were followed for terminology concerning the external scent efferent system of the metathoracic scent glands. Macrophotographs of dorsal, ventral and lateral habitus, Table 1. Taxon sampling for the phylogenetic analysis of Padaeus bovillus (Distant) and related carpocorines, including Genbank accession number for respective DNA marker. Molecular markers not sequenced marked with “-”. Identification literature brings the main reference used to identify specimens used in the morphological analysis Species COI Cyt b 16S 28S Identification literature Adustonotus grandis (Rolston) KU892549 KU853795 KU853775 KU853759 Bianchi et al. 2017b Adustonotus hansi (Grazia) KU892550 KU853796 - KU853760 Bianchi et al. 2017b Adustonotus saramagoi (Bianchi, Cioato and Grazia) KU892552 KU853798 KU853778 - Bianchi et al. 2017b Adustontous paranticus (Grazia) KU892551 KU853797 KU853777 KU853761 Bianchi et al. 2017b Agroecus scabricornis (Herrich-Schäffer) KU892539 KU853783 KU853764 - Rider and Rolston 1987 Caonabo pseudoscylax (Bergroth) KU892540 KU853784 KU853765 KU853749 Rolston 1974b Diceraeus furcatus (Fabricius) U892541 KU853785 KU853766 KU853750 Barão et al. 2020 Dichelops (D.) leucostigmus (Dallas) U892542 KU853786 KU853767 KU853751 Barão et al. 2020 Euschistus (E.) crenator (Fabricius) - KU853787 KU853768 KU853752 Rolston 1974a Euschistus (E.) heros (Fabricius) KU892543 KU853788 KU853769 KU853753 Rolston 1974a Euschistus (E.) taurulus Berg KU892545 KU853789 KU853770 KU853754 Hickmann et al. 2019 Euschistus (L.) circumfusus Berg - KU853790 KU853771 KU853755 Weiler et al. 2016 Euschistus (L.) cornutus (Dallas) U892546 KU853791 KU853772 KU853756 Weiler et al. 2016 Euschistus (L.) triangulator (Herrich-Schäffer) - KU853792 KU853773 KU853757 Weiler et al. 2016 Glyphepomis spinosa Campos and Grazia KU892553 KU853799 - - Bianchi et al. 2016 Ladeaschistus bilobus (Stål) KU892554 KU853800 KU853779 KU853762 Rolston 1973 Ladeaschistus borgesi Bianchi, Cioato and Grazia KU892555 - KU853780 Cioato et al. 2015 Mitripus acutus Dallas KU892547 KU853793 KU853774 KU853758 Bianchi et al. 2017b Mitripus convergens (Herrich-Schäffer) KU892548 KU853794 - - Bianchi et al. 2017b Mitripus seclusus sp. n. Bianchi, Krein, Rider and Grazia ---- Mitripus bovillus (Distant) comb. n. - - - - Distant 1900 Padaeus viduus (Vollenhoven) - - - - Vollenhoven 1868 Sibaria amazonica Krein, Rider and Grazia - - - - Krein et al. 2020 Sibaria armata (Dallas) KU892556 KU853801 KU853781 KU853763 Krein et al. 2020 Sibaria englemani Rolston KU892557 - KU853782 - Krein et al. 2020 page 3 of 13Zoological Studies 60:11 (2021) © 2021 Academia Sinica, Taiwan and also genitalic structures of the both sexes were taken in sequential focus using a Nikon AZ100M and digitally stacked in the NIS Elements software, available in the Zoology Department of Universidade Federal do Rio Grande do Sul. The specimens used for this study were borrowed from: DARC: David A. Rider Collection, Department of Entomology, North Dakota State University, Fargo, North Dakota; UFRG: Coleção do Departamento de Zoologia, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil; Joseph E. Eger personal collection (JEEC) USNM: National Museum of Natural History, Smithsonian Institution, Washington D.C., USA; and NHMUK: The Natural History Museum, London, United Kingdom. RESULTS TAXONOMY (The taxonomies in this section are based on the phylogenetic analyses. See below) Pentatomidae Leach, 1815 Pentatominae Leach, 1815 Genus Mitripus Rolston, 1978 Mitripus seclusus sp. n. Bianchi, Krein, Rider and Grazia (Figs. 1–5, Table 3) urn:lsid:zoobank.org:act:ECBED1B6-EC60-440D-A82689322D5F6674 Type Material: Holotype: ECUADOR: 1 male, Paramba, Ecuador / Coll. I. R. Sc. N. B, Equateur: Paramba, Ecuador (UFRG); 1 female, Paramba, Ecuador / Collection Rosenberg / NMNH. Etymology: The name is allusive to the seclusion among the authors while describing this species (L. seclusus: shut up, separated, recluse, seclude). Most of the intellectual work was made during the pandemic of COVID-19, with the authors respecting proper social distancing. Diagnosis: Mitripus seclusus sp. n. may be distinguished from M. convergens and M. legionarius by each humeral angle developed laterally as a stout spine; from M. acutus by the clypeus being slightly longer than the mandibular plates, and the anterolateral margins of the pronotum entirely smooth; and from M. Table 2. Morphological partition based on Bianchi et al. (2017b). Character state matrix for the phylogenetic analysis of Mitripus bovillus (Distant) and related carpocorines. Taxa in bold are not present in Bianchi et al. (2017b). -. inapplicable data; ?. missing data Adustonotus grandis 100110000011111121312000100-11111101010010101000000001110001011101020001001-111111?-11 Adustonotus hansi 100010100001011121001001100-11111000110010121000000002010201211000-40001001-1011111-11 Adustonotus saramagoi 100110000101011121001001100-11112110010010121000000001010201211001040001001-1310111211 Adustontous paranticus 100010000001111121000101100-11111101110010101001000000010201011100-40001001-131011?-11 Agroecus scabricornis 120110000010121122101010100-0010011000-1100000???001000???11210101000111101-1110100?10 Caonabo pseudoscylax 1011101111001210-2101000000-1101311001002100101?110000021000010110?20200-0001031111010 Diceraeus furcatus 112010100001110100112100100-00001100110001101110010110011000110121010001101-1011110-11 Dichelops (D.) leucostigmus 112010011110111102101000100-0101111010-0111000???11100010200110121130001101-112111??01 Euschistus (E.) crenator 221010100101100102101110100-0100210010-020100110100000000001211110-3010110011200110-00 Euschistus (E.) heros 221010110101110102112010100-0100311110-020020110110201010000010110-1010111011211110-01 Euschistus (E.) taurulus 021010101011100122102010100-0100110000-021120010000201020100000110-2010110011211110-00 Euschistus (L.) circumfusus 0210000010011001122001001010010?011011001001111010010101001100011103011111011221111211 Euschistus (L.) cornutus 011010111001010122302011100-0011011001100101110010020103001121011101011111011221111211 Euschistus (L.) triangulator 22111011100110012210201011100011011001100102111011020103001120011102011110011221111211 Glyphepomis spinosa 2110111101010100-2102000000-1001011100-011000000000101030000110110?100110000100011-?00 Ladeaschistus bilobus 1210000010111011?2001100100-11102100110120121002000001011200010001021001001-10100?0-11 Ladeaschistus borgesi 020010000011111122100101101011102000110120021002000001011200010001031001001-11110?0?11 Mitripus acutus 220100000001101122101100101011101001110020001001000102011201001100-30001001-1211100-11 Mitripus bovillus comb.n. 1200000000111010?2111100101011102100110020001001010100011200001001150001001?1301110?01 Mitripus convergens 000010100011101121000100101011101001010020121001000002011200010100-30001001-111111?-11 Mitripus seclusus sp. n. 1200000000111010?210110010101110210011002000100101010?????000010011500?1001-1301110-01 Padaeus viduus 201000010010011102101000101000113010111010100001010010020000111110?00101100?1221101201 Sibaria amazonica 020110100111121112101110101011102100110110001000010000011200000101030001001-111111-?01 Sibaria armata 020100000111131112101100101011101000110110021000010000011200010101040001001-111111-?01 Sibaria englemani 020000000111131112101110101011100000110120001000010000011200010101030001001-111111-?01 page 4 of 13Zoological Studies 60:11 (2021) © 2021 Academia Sinica, Taiwan bovillus by the humeral angles not depressed anteriorly, and by the shape of the parameres (Fig. 3). Description: Coloration: Dorsal surface ochraceous with black punctures, giving an overall dark brown matte aspect; antennae ochraceous ventrally and basally, brownish dorsally. Scutellum darker basally than apically. Connexivum brownish, middle third ochraceous. Ventral surface yellowish ochraceous; head and abdomen impunctate, thorax with brownish-yellow punctures. Legs ochraceous with reddish-brown spots on femora and tibiae; each femur with brownish ring at apex; apex of each tarsomere brownish. Head: Clypeus slightly longer than mandibular plates, both rounded apically. Head tapering to apex, lateral margins of mandibular plates sinuous, concave near eyes. Ocelli red or yellow. Antennomere Fig. 1. Bayesian Inference consensus tree based on the analysis of four molecular markers and morphological characters from 25 species of carpocorines. Numbers close to nodes are Bayesian posterior probability / Maximum-Likelihood bootstrap support, respectively. Only nodal support above PP = 0.5 or BS = 50 were collapsed (“*” indicates uninformed value); A–B, Padaeus viduus (Vollenhoven); C–D, Mitripus bovillus comb. n. (Distant): A: capsula seminalis; B: pygophore; C: capsula seminalis; D: pygophore (numbers near to the genitalic structures indicate character and state); red dashed line delimitates Mitripus clade. page 5 of 13Zoological Studies 60:11 (2021) © 2021 Academia Sinica, Taiwan Fig. 2. Habitus of Mitripus seclusus and Mitripus bovillus. A–C, Mitripus seclusus sp. n.: A: dorsal; B: ventral; C: lateral; D–G, Mitripus bovillus comb. n. (Distant): D: dorsal; E: ventral; F: labels; G: lateral. Scale bars = 1 mm. Table 3. Measurements: mean (± standard deviation) given in millimeters of morphometric parameters of Mitripus bovillus comb. n. (Distant) and Mitripus seclusus sp. n. Mitripus bovillus comb. n. Mitripus seclusus sp. n. Male (n = 3) Female (n = 3) Male (n = 1) Female (n = 1) BL 10.93 (0.35) 11.60 (0.66) 10.00 9.90 AW 5.53 (0.15) 6.07 (0.31) 5.00 5.40 HL 2.00 (0.00) 2.13 (0.06) 1.80 1.60 HW 2.30 (0.10) 2.40 (0.10) 2.10 2.10 LE 1.10 (0.10) 1.13 (0.06) 1.00 0.80 ID 0.97 (0.06) 1.03 (0.15) 1.00 1.00 I 0.60 (0.00) 0.67 (0.06) 0.50 0.50 II 1.17 (0.06) 1.20 (0.00) 1.00 - III 1.33 (0.06) 1.47 (0.06) 1.20 - IV 1.90 (0.00) 2.25 (0.07) - - V 1.95 (0.07) 2.10 - - PL 2.43 (0.06) 2.70 (0.10) 2.30 2.20 PW 8.50 (0.36) 8.87 (0.45) 7.60 7.60 SL 3.80 (0.20) 4.00 (0.35) 3.60 3.70 SW 3.57 (0.06) 3.90 (0.26) 3.30 3.60 BL: total body length; AW: abdominal width; HL: head length; HW: head maximum width; LE: length of head before eyes; ID: interocular distance; I; II; III; IV; V: antennomere length I, II, III, IV, V respectively; PL: pronotal length; PW: pronotal maximum width; SL: scutellar length; SW: scutellar maximum width. page 6 of 13Zoological Studies 60:11 (2021) © 2021 Academia Sinica, Taiwan Fig. 3. Male genitalic structures of Mitripus seclusus sp. n. and Mitripus bovillus comb. n. (Distant). A–J, Mitripus seclusus: A–C: pygophore: A: dorsal view; B: posterior view; C: ventral view; D–G: left paramere: D: dorsal view; E: ventral view; F: lateral view; G: mesial view; H–J: phallus: H: ventral view; I: lateral view; J: dorsal view; K–T, Mitripus bovillus: K–M: pygophore: K: dorsal view; L: posterior view; M: ventral view; N–Q: left paramere: N: dorsal view; O: ventral view; P: lateral view; P: mesial view; R–T: phallus: R: ventral view; S: lateral view; T: dorsal view. App: apical process of paramere, bpp: basal process of paramere, dr: dorsal rim, spdr: superior process of dorsal rim, tr: transverse ridge, vr: ventral rim. Scale bars = 1 mm. page 7 of 13Zoological Studies 60:11 (2021) © 2021 Academia Sinica, Taiwan proportions: I < II < III (IV and V lacking). Anterior margins of bucculae truncate with sharp projections, posterior margins evanescent. Gena flat. Rostrum reaching anterior margins of metacoxae. Thorax: Anterolateral margin of pronotum smooth on anterior half, slightly sinuous; anterolateral angles developed as small yellowish-ochraceous spines, projected laterally. Each humeral angle produced laterally as a black stout spine. Pronotal cicatrices brownish, with an ochraceous spot posterior to each mesial angle. Basal angles of scutellum with small foveae. Apex of radial vein with a small ivory dot at endocorium. Membrane of hemelytra fumose, with subparallel veins. Evaporatoria each extending halfway from ostiole to metapleural lateral margin; surface impunctate, presenting gyrification near ostioles; lateral fold present; peritreme spout-like. Abdomen: Each posterolateral angle of connexiva developed as a tiny black spine. Posterolateral angles of urosternite VII slightly projected as a spine. Spiracles concolorous with abdominal disc. Male genitalia: Pygophore (Fig. 3A–C): In dorsal view, trapezoidal; genital cup not well exposed; each posterolateral angle of pygophore projected as a small triangle, rounded apically; median projection of dorsal rim short. Dorsal rim interrupted by diagonal depressions flanking median projection of dorsal rim; lateral margins slightly sinuous, discontinuous near median projection of dorsal rim; superior process of dorsal rim exposed, rectangular; ventral rim concave with a triangular projection at middle. In posterior view, genial cup opening ellipsoid; dorsal rim concave, smooth; superior process of dorsal rim long, bladelike; transverse ridge concave medially, U-shaped; inferior layer of ventral rim medially straight, tumescent laterally. In ventral view, ventral rim concave with a triangular projection at middle; posterolateral angles projected posteriorly, slightly bifid. Tenth segment: posterior margin trapezoidal with setae; disc smooth; tubercles on basal third slightly developed. Parameres (Fig. 3D–G): In dorsal and ventral view: each with basal apodeme smaller than crown; stem slightly smaller than crown; crown bending outward nearly at a 45° angle. Basal process of paramere narrow, rounded, with long setae; apical process of paramere enlarged basally, tapering towards truncate apex. In mesial and lateral views: apical process of paramere stout, triangular, with scale-like structures on its outer surface. Phallus (Fig. 3H–J): Phallotheca tubular, slightly constricted basally; ventral basal processes of phallotheca quadrangular; dorsal processes of phallotheca hook-like, bent ventrally in about 45°, short, not surpassing expanded conjunctiva; vesica process spout-like; conjunctiva lacking processes; ductus seminis distalis short, not exposed out of phallotheca. Female genitalia: In posteroventral view (Fig. 4A), gonocoxites VIII with shallow punctures, concolorous with ochraceous disc; surface flat with a slightly depressed area near posterolateral margin; mesial margins straight, overlapping, with a brown spot apically; apex squared. Laterotergites VIII with black margins; apex spine-like. Gonocoxites IX trapezoid, about three times wider than long, anterior and lateral margins straight, posterior margin concave. Laterotergites IX rounded apically, mesial margins forming a right angle, lateral margins convex, slightly surpassing tergite VIII; segment X rectangular. Distribution: Mitripus seclusus sp. n. is known only from Ecuador (Fig. 5). Pentatomidae Leach, 1815 Pentatominae Leach, 1815 Genus Mitripus Rolston, 1978 Mitripus bovillus (Distant) comb. n. (Figs. 1–5, Table 3) urn:lsid:zoobank.org:act:6910E7CD-BB6F-4222-A9E2F2E155BE3C59 Padaeus bovillus Distant, 1900: 689, 690 (original description); Kirkaldy, 1909: 69 (catalog); Rolston, 1976: 7 (revision). Material examined: Type Material: Holotype female: COSTA RICA: Tuis, Cartago, Terralba 650 m., A. Pittier, BRIT. MUS. (NHMUK) TYPE HEM/050. COSTA RICA: 1m#, Costa Rica, Prov. Heredia, F. La Selva, 3km S Pto. Viejo, 10°26'N 84°01'W / 25.III.1987, H. A. Hespenheide / D. A. Rider Collection (DARC); 1f#, Costa Rica, Pr. Heredia, Puerto Viejo, Finca La Selva / R J. Marquis, coll. No. I, 11.IX.1986 / Piper aricianum (?) / NMNH; 1f#, Collection SchildBurgdorf, Costa Rica, San Carlos / Padaeus bovillus Distant / NMNH ; 1m# / Costa Rica: Cartago Prov., Mon. Nac. Guayabo, 22.XII.1994 / M. J. Tauber, C. A. Tauber, P. J. Tauber Collectors / UC Berkeley EMEC 1240641. PANAMA: 1m#, 1f#, Panama: Bocas Del Toro Pr. 3km. n. Continental Div. on Fortuna Hwy 925 m., 13.VII.1996, A. R. Gillogly / EGER / Sibaria n. sp. Det. J. E. Eger, 1997 (UFRG). Diagnosis: Mitripus bovillus may be distinguished from M. convergens and M. legionarius by the humeral angles which are developed laterally as stout spines; from Mitripus acutus by the clypeus slightly longer than mandibular plates, anterolateral margins of pronotum entirely smooth; and from Mitripus seclusus by the humeral angles depressed anteriorly and the shape of parameres (Fig. 3). Redescription: Coloration: Dorsal surface ochraceous with black punctures, giving an overall dark page 8 of 13Zoological Studies 60:11 (2021) © 2021 Academia Sinica, Taiwan brown matte aspect; head and thorax with ochraceous punctures; antennae ochraceous, antennomeres I–II with irregular dark brown spots, antennomeres III–IV dark on apical 3/4, and antennomere V dark brown on apical half. Scutellum dark brown on base. Connexivum blackish, middle third ochraceous. Ventral surface yellowish-ochraceous; abdomen impunctate. Legs ochraceous with brown spots on femora and tibiae; apex of each tarsomere brownish. Head: Clypeus slightly longer than mandibular plates, rounded apically. Head tapering to apex, lateral margins of mandibular plates sinuous, concave near eyes. Ocelli red or yellow. Antennomere proportions: I < II < III < IV = V. Anterior margins of bucculae truncated, each with sharp projection, posterior margins evanescent. Rostrum reaching metacoxae. Thorax: Anterolateral margins of pronotum smooth on anterior half, anterolateral angles developed as small yellowish-ochraceous spines, projected laterally. Each humeral angle produced laterally as a black, stout spine, somewhat depressed anteriorly. Pronotal cicatrices brownish, with ochraceous spot posterior to mesial angles. Basal angles of scutellum with small fovea. Apex of radial vein with a small ivory dot at endocorium. Hemelytral membrane fumose, with veins subparallel. Evaporatoria each extending halfway from ostiole to metapleural lateral margin, and present on posterior margin of mesopleuron; surface impunctate, with gyrification near ostiole; lateral fold present; peritreme spout-like. Abdomen: Posterolateral angles of connexiva developed as tiny black spines. Posterolateral angles of urosternite VII slightly projected as spines. Spiracles concolorous with abdominal disc. Male genitalia: Pygophore (Fig. 3K–M): in dorsal view, pygophore trapezoidal; genital cup not well exposed; posterolateral angles of pygophore developed as rounded projections; median projection of dorsal rim short. Dorsal rim interrupted by diagonal depressions flanking median projection of dorsal rim, lateral Fig. 4. Female genitalic structures. A: Mitripus seclusus sp. n.; B–C Mitripus bovillus comb. n. (Distant). A–B: posteroventral view of female terminal abdominal segments; C: internal genitalic structures. cs: capsula seminalis, mw: median wall, pi: pars intermedialis, rs: ring sclerites, tvi: thickening of vaginal intima, va: vesicular area. Scale bars: 1 mm. page 9 of 13Zoological Studies 60:11 (2021)