New species of the endemic Neotropical caddisfly genus Contulma from the Andes of Ecuador (Trichoptera: Anomalopsychidae)

The genus Contulma Flint (Trichoptera: Anomalopsychidae) is composed mostly of regionally endemic species occurring above 2,000 m, with a few more widespread species and some that are found at lower elevations. Adults of three new species of Contulma are described and illustrated from the Andes of Ecuador, Contulma lina, new species, Contulma quito, new species, and Contulma sangay, new species. These species are similar to previously described species from the region, including C. paluguillensis, C. nevada, and C. lancelolata. New provincial records are provided for C. bacula, C. cataracta, and C. echinata. Contulma duffi Oláh, 2016 is considered a junior, subjective synonym of C. penai, Holzenthal & Flint, 1995. Also, we provide an identification key to males of the 30 Contulma species now known.


INTRODUCTION
Neotropical Trichoptera currently includes more than 3,200 described species representing 155 genera and 25 families (Holzenthal & Calor, 2017) occurring in Mexico, the Caribbean, Central, and South America. Remarkably, 115 genera, or ca.75% of the total, are endemic to the region, making the fauna the second most diverse in the world for endemic genera after the Australasian (De Moor & Ivanov, 2008). Moreover, several Neotropical genera are highly endemic regionally at the species level such as Amphoropsyche Holzenthal (Holzenthal, 1985;Holzenthal & Rázuri-Gonzales, 2011), Atananolica Mosely (Holzenthal, 1988;Henriques-Oliveira & Santos, 2014), and Contulma Flint (Holzenthal & Robertson, 2006), the latter the main subject of this paper. Nevertheless, the Neotropical caddisfly fauna is incompletely known, mainly because there are regions in the Neotropics where the aquatic ecosystems are far from being well studied (e.g., Ríos-Touma et al., 2017). There is also a lack of regional researchers studying the order, especially in the Andean countries of Venezuela, Colombia, Ecuador, Peru, and Bolivia, which undoubtedly harbor hundreds Table 1 Described species in the genus Contulma with geographic distribution and known life stages.

Contulma sana Jardim & Nessimian (2011) Brazil male
Contulma sancta Holzenthal & Flint (1995) Costa Rica male Contulma sangay, n. sp. Ecuador Contulma spinosa Holzenthal & Flint (1995) Colombia, Ecuador male, female, larva Contulma talamanca Holzenthal & Flint (1995) Costa Rica male, female Contulma tapanti Holzenthal & Flint (1995) Costa Rica male, female Contulma tica Holzenthal & Flint (1995) Costa Rica male Contulma tijuca Holzenthal & Flint (1995) Brazil male, female, probable larva Contulma tripui Holzenthal & Robertson (2006) Brazil male Holzenthal & Flint (1995) Costa Rica male, female, larva of species. For example, co-authors Ríos-Touma is the first Ecuadorian to describe new species of caddisflies and Rázuri-Gonzales only the third Peruvian to do so. A comprehensive revision of the endemic Neotropical genus Contulma was completed by Holzenthal & Flint (1995) and included 21 species, 18 described as new. Since then, seven new species have been described, including one we synonymize here (Table 1). Species in the genus are known from Costa Rica, the Andes of Colombia to Chile, and in the mountains of southeastern Brazil (Holzenthal & Calor, 2017). This genus seems to display a high degree of local endemism among its species (Holzenthal & Robertson, 2006), and they are rarely collected using standard light trap techniques. Hand netting during the day, especially at high elevations, or the use of Malaise traps is generally more effective (Holzenthal & Ríos-Touma, 2012). The infrequency of collection does not mean the species are rare in nature, but most have been described from only 1-5 individuals. Perhaps this is a reflection of minimal collecting effort or low temperatures at high elevations that reduce adult flying activity. The habitats of these species are small waterfalls, seeps, and small streams in lush forested mountainous areas as well as high elevation páramo streams above the tree line in the Andes (Holzenthal & Flint, 1995).

Contulma valverdei
Of the 30 species now known in the genus, including three new species described here, 19 occur in the tropical Andean countries (Ecuador, Colombia, Peru, Bolivia) and all occur above 2,000 m, except for two of these species also found in lower elevations (Holzenthal & Flint, 1995;Holzenthal & Robertson, 2006). Eight species occur in Ecuador, five endemic and three are also present in Colombia. The three new species described here are from localities were no caddisfly collecting occurred previously.

MATERIALS AND METHODS
We used the methods described by Blahnik & Holzenthal (2004) to prepare adult specimens for taxonomic study. Genitalia were cleared in 85% lactic acid heated to 125 • C for 20 min (Blahnik, Holzenthal & Prather, 2007). An Olympus BX41 compound microscope outfitted with a drawing tube was used to observe specimens. Genital structures were drawn with pencil on paper and final illustrations were rendered in Adobe Illustrator. Morphological terminology follows that of Holzenthal & Flint (1995). Descriptions of species and generation of the identification key were accomplished using the software packages DELTA and INTKEY (Dallwitz, 1980;Dallwitz, Paine & Zurcher, 1999).
Types of the new species are deposited in the collections of the Museo Ecuatoriano de Ciencias Naturales, Quito, Ecuador (MECN) and the University of Minnesota Insect Collection, St. Paul, Minnesota, USA (UMSP). All specimens examined in this study were affixed with a barcode label containing a unique nine digit numeric code starting with the prefix UMSP. These codes are provided here for holotypes only. All associated specimen data are stored in the UMSP database. This study was performed under the Environmental Ministry of Ecuador study permits 36-2010-IC-FLO/FAU-DPA-MA and 005-15-IC-FAU-FLO-DNB/MA.
The electronic version of this article in Portable Document Format (PDF) will represent a published work according to the International Commission on Zoological Nomenclature (ICZN), and hence the new names contained in the electronic version are effectively published under that Code from the electronic edition alone. This published work and the nomenclatural acts it contains have been registered in ZooBank, the online registration system for the ICZN. The ZooBank LSIDs (Life Science Identifiers) can be resolved and the associated information viewed through any standard web browser by appending the LSID to the prefix http://zoobank.org/. The LSID for this publication is: urn:lsid:zoobank.org:pub:54BC56DC-5CC1-4DA0-82E4-AF69599C2F5D. The online version of this work is archived and available from the following digital repositories: PeerJ, PubMed Central and CLOCKSS.

Species descriptions
Contulma lina, new species, Holzenthal, Ríos-Touma, Rázuri-Gonzales LSID urn:lsid:zoobank.org:act:310DDC54-0008-4535-A385-BD71BB630F21 Figs. 1A-1E, 2 Diagnosis: Contulma lina, n. sp., as well as C. quito, n. sp., and the recently described C. paluguillensis Holzenthal & Ríos-Touma, 2012, in addition to C. echinata, C. nevada, and C. papallacta, all described by Holzenthal & Flint (1995), and all from either Ecuador or Colombia, share several features. All have some degree of development of a dorsolateral process on segment IX in the male genitalia and a development of setae or a setose process on the mesal face of the same segment below the dorsolateral process. These three species formed a clade within the cranifer-group of Holzenthal & Flint (1995) to which C. lina, n. sp., C. quito, n. sp., and C. paluguillensis also belong. In some species, one or the other of these characters may be more or less developed, but the combination of characters shared by these species indicate that they may have a common origin. The phallic structures seem to be unique to each species and attest to their distinctiveness. All are known from only a handful of specimens, most from only the type and a few paratypes, and the species each occur from only one or a few high altitude localities spread across a vast expanse of the northern Andes. We predict that additional collecting will discover yet more new species in this radiation of Andean Trichoptera biodiversity. Contulma lina is distinguished from the above mentioned species, all illustrated by Holzenthal & Flint (1995) and Holzenthal & Ríos-Touma (2012), by the following combination of characters: mesal surface of segment IX bearing a sinuous band of setae from below dorsolateral process and continuing to near sternum IX; mesal process of sternum IX with prominent posteromesal, heavily sclerotized, spatulate projection, without excavation; and phallus without spine-like setae, but instead with the apex bearing a lightly sclerotized scale-like structure, shallowly excavate at apex. Description: Male: Forewing length 5.5 mm (n = 1). Forewing color gray brown, immaculate, vestiture rubbed (specimen was netted during the day in light rain and mist). Male genitalia: Segment IX very short dorsally, narrow, deeply excavate mesally; in lateral view, IX quadrate, extended anterolaterally; posteriorly with short, broad, dorsolateral, spatulate process; posterior margin of IX produced medially to form broad, prominent, quadrate, heavily setose, paired lateral lobes; lobes widely separated ventrally; mesal face of segment IX bearing sinuous band of setae from below spatulate process continuing to near sternum IX; sternum IX with prominent posteromesal, heavily sclerotized, spatulate projection. Inferior appendages short, subtriangular, apices broadly rounded and bearing apical setae; inferior appendages apparently fused to base of IXth sternal projection, together forming highly complex structure as in Figs. 1A, 1C. Processes of subphallic membranes present, membranous, mound-like, setose. Segment X entirely membranous, apex entire, extending beyond apices of dorsolateral processes. Phallus complex; phallobase tubular, elongate, slender, sclerotized; phallicata very lightly sclerotized, dorsally with paired, very lightly sclerotized, semi-membranous lobes; apicoventral phallic membranes with paired, broad, lightly sclerotized, dorsolateral plates, membranes inflated anteroventrally, apically with lightly sclerotized scale-like structure, shallowly excavate at apex; phallotremal sclerite present, tubular, curved. Female: Forewing length 6.5 mm (n = 1). Color and vestiture as in male. Vaginal apparatus in ventral view elongate, hourglass shaped, narrowest in middle; base widely emarginate, subtriangular; apex trident shaped with wide, paired, sclerotized, slightly sinuate midlateral processes, their apices truncate; single medial process elongate, narrow, slerotized, about same length as midlateral processes; medial membranes highly convoluted, as approximated in Fig. 2  Diagnosis: This new species is very similar to C. nevada of the group of species mentioned above (C. echinata, C. lina, C. nevada, C. paluguillensis, and C. papallacta). Contulma quito, n. sp., and C. nevada have similar short dorsolateral processes on segment IX, a small setal patch or setal bearing process below the dorsolateral process, emarginate sternal projection of sternum IX, and short spines in the phallus. The major differences between C. quito and C. nevada are the broader excavation of the IXth sternal projection and the many more spine-like setae in the phallus of C. quito.
Description: Male: Forewing length 5.5 mm (n = 3). Forewing color brown, with small patch of cream colored hairs at apex of subcosta and arculus, vestiture intact. Male genitalia: Segment IX very short dorsally, narrow, deeply excavate mesally; in lateral view, IX quadrate, slightly extended anterolaterally; posteriorly with very short, narrow, dorsolateral, spatulate process; posterior margin of IX produced medially to form broad, prominent, broadly rounded, heavily setose, paired lateral lobes; lobes widely separated ventrally; mesal face of segment IX with short, sclerotized, setose process below spatulate process. (Variation: in male paratype UMSP000148995, the spatulate processes and the short setose processes below them are each fused on both sides of the specimen to form a thin, crecentic shelf (Fig. 3F). In male paratype UMSP000148996, the spatulate processes and the setose processes below them are variously slightly longer or shorter on either the right or left sides than they are in the holotype). Sternum IX with prominent posteromesal, heavily sclerotized, strongly emarginate projection. Inferior appendages short, crescentric, apices rounded and bearing apical setae; inferior appendages apparently fused to base of IXth sternal projection, together forming highly complex structure as in Figs. 3A, 3C. Processes of subphallic membranes present, membranous, mound-like, setose. Segment X entirely membranous, apex divided, with lightly sclerotized lateral flanges. Phallus complex; phallobase tubular, elongate, slender, sclerotized; phallicata very lightly sclerotized, dorsally with paired, very lightly sclerotized, semi-membranous lobes; apicoventral phallic membranes with paired, broad, lightly sclerotized, dorsolateral plates forming broadly rounded trough, apicoventrally with paired patches of numerous (ca. 50) very short spine-like setae; phallotremal sclerite very lightly sclerotized, difficult to discern.
Female: Unknown.  Diagnosis: This new species is similar to C. lanceolata, Holzenthal & Flint, 1995, also from Ecuador. Both share a similar shape of segment IX, which is strongly extended anterolaterally and with the posterior portion excavated medially in both species. In addition, both have an elongate dorsolateral setose lanceolate process on segment IX, but in C. sangay the process is divided apically into a pair of narrow, terete lobes. Most distinctively, the new species lacks the ventrolateral process of segment IX seen in C. lanceolata (Holzenthal & Flint, 1995, fig. 70).
Female: Forewing length 4.5 mm (n = 1). Color as in male, but vestiture rubbed. Vaginal apparatus in ventral view short, oval, widest in middle; base rounded, urn-shaped; apex trident shaped with narrow, paired, lightly sclerotized, indistinct midlateral processes, their apices acute; single medial process elongate, narrow, sclerotized, longer than midlateral processes; medial membranes highly convoluted, as approximated in Fig. 5 Holzenthal & Flint, 1995:11 Holzenthal & Flint, 1995:12 Holzenthal & Flint, 1995:18  Contulma duffi, described from Antioquia, Colombia, fits clearly within the variation we have seen in the species C. penai, which also occurs in Antioquia, Colombia, and Ecuador, where it is rather common compared to other species in the genus. There are no features illustrated or diagnosed in the original description that distinguish the species from C. penai. Most of the characters discussed by Oláh (2016) are those common to the genus as a whole. The membranous structure of tergum X is identical in both species. The dorsolateral processes of segment IX are illustrated as slightly curved in C. duffi compared to those illustrated for C. penai, but this slight difference is variable, and the inferior appendages are identical in the two species. In Oláh's diagnosis, the species is said to be most similar to C. bacula, but it shares little in common with that species. The inferior appendages, the posteromesal process of sternum IX, and the phallus are completely different between the 2 species.  13(12). Projection of sternum IX subtriangular in ventral view, with broad, angulate mesal excavation (Fig. 3C)  The species is known only from the male holotype and a male paratype. In the holotype, there are no dorsolateral processes (HR 3A), but in the paratype, a rudimentary process occurs (HR 3G). However, it shares all other diagnostic features of this group, including the posterior, setose extension of segment IX, the broad, shelf-like structure of sternum IX, including the flat, tooth-like, apical setae, and the complex, membranous lobes of the phallus. It appears artificially in couplet 26, separated from related species because it lacks the strongly downcurved, apically rugose dorsolateral processes indicated in couplet 2.

Key to males of Contulma
The collection of additional specimens may prove that the absence of the dorsolateral process is an aberration; if so, the specimens would lead to C. fluminensis in the key presented above.

DISCUSSION
Contulma species appear to be highly regionally endemic (Holzenthal & Flint, 1995;Holzenthal & Robertson, 2006). We observed this pattern in our collections in Ecuador. Patterns of high endemicity in macroinvertebrate benthic larvae are well known in high altitude glacial streams; these habitats are highly threatened by climate change (Jacobsen et al., 2012). On the other hand, we added records to the Ecuadorian fauna of species that were previously known only from Colombia, demonstrating that some species may have spread across the Andes. Despite this distribution pattern, aquatic insect species, especially mayflies, have shown genetic isolation in the various ranges of the Ecuadorian Andes, even with relatively short distances between populations and with recent volcanic eruption history (Finn, Encalada & Hampel, 2016). No similar study has yet been performed with any Andean caddisfly species, but we expect a similar pattern of isolation among populations. This isolation could increase by the intense land use changes and water pollution occurring throughout the inter-Andean valleys (Ríos-Touma, Acosta & Prat, 2014) potentially preventing dispersal among mountain populations and also local extinctions.