Description of the late- or last-instar larva of Ischyomius Chevrolat, with comments on the family placement of the genus (Coleoptera: Tenebrionoidea: Pythidae)

Study of recently collected larvae of two species of Ischyomius Chevrolat, 1871 from Napo and Pichincha, Ecuador, tentatively identified as I. denticollis Champion, 1916 and I. singularis Chevrolat, 1871 confirm the classification of the genus Ischyomius within the family Pythidae. Salient larval characters which solidify this placement are: 5 stemmata arranged into an anterior column of 3 and posterior column of 2, parabasal ridges along anterior margins of abdominal tergites, prominent urogomphi with inner urogomphal teeth and various dorsal tubercles, and transverse row of dentiform asperities along anterior margin of sternite 9. Important features of the larva are illustrated with photographs. A key to larvae of world genera of Pythidae is provided as well as natural history information on Ischyomius. Key-Words. Immatures; Neotropical; Classification; Zingiberales; Ecuador; Venezuela.


INTRODUCTION
The Neotropical genus Ischyomius Chevrolat, 1878 currently contains eight species, encompassing a geographical range from Costa Rica in the north, to Bolivia in the south (Pollock, 1998(Pollock, , 2007(Pollock, , 2009. As with many other relatively obscure taxa of Tenebrionoidea, Ischyomius has had an unstable position within the classification of the tenebrionoid Coleoptera; it was placed in various families at different times, including Tenebrionidae, Melandryidae, Tetratomidae, and Trictenotomidae (Pollock, 2010). Placement of Ischyomius in Pythidae was first suggested by Lawrence (1982), who offered no detailed supporting evidence for this placement. In their treatment of beetle classification, Lawrence & Newton (1995) placed Ischyomius as Pythidae, incertae sedis. As mentioned by Pollock & Lawrence (1995: 467), "an additional three genera, Ischyomius, Trimitomerus and Osphyoplesius, should be placed there [Pythidae] tentatively, pending more detailed morphological studies and/or the discovery of the immature stages". This statement was repeated by Pollock (1998), as Ischyomius historically has been somewhat enigmatic taxonomically.
In a revision of the genus, Pollock (1998) provided evidence from various adult structural features that Ischyomius is closely related to genera already recognized as Pythidae. It was stated that discovery of the immature stages of a species of Ischyomius would be of great value, and would either corroborate or refute this adult-based family placement. Subsequent to Pollock (1998), larvae of Ischyomius became available and though not formally described, Pollock (2010) mentioned that features of the head, legs, and urogomphal plate were very similar to larvae of Pytho Latreille and supported placement of Ischyomius in Pythidae. More recently, important larval material of Ischyomius (with adults of I. denticollis Champion, 1916 andI. singularis Chevrolat, 1878) was collected by the second author in Napo and Pichincha Provinces, Ecuador.
Despite the lack of a publication of an actual larval description, Ischyomius has been placed in Pythidae in some recent publications (e.g., Lawrence et al., 2011;Pollock, 2018). The description of the mature larva of Ischyomius (either I. denticollis or I. singularis) removes any remaining doubt as to the family placement of the genus. The purpose of the present paper is to describe and illustrate the larva of Ischyomius, provide an updated key to larvae of world genera of Pythidae (originally in Pollock & Lawrence, 1995), and comment on classification of the family.

MATERIAL AND METHODS
This study was based on 14 medium-large larvae and 13 adults of Ischyomius (I. singularis and I. denticollis) collected in association with these larvae. Details are provided below in "Material examined". One larva was macerated in cold KOH before mouthparts were dissected from it. Photographs were taken of specimens in 70% isopropanol with a Zeiss Discovery V20 stereoscope, acquired with ZEN 2.3 Lite and stacked using CombineZP (Alan Hadley).
Though no larvae were reared to adult or DNA matched to absolutely confirm the association presumed in this paper, we are confident that the larvae described herein are those of Ischyomius because of multiple co-occurrences with adults. At one site ("km 36, Rte 20, Pichincha, Ecuador"), adults of both I. denticollis and I. singularis were collected in addition to larvae representing two species (especially notable in their different urogomphi). The relative size of both adults and larvae, I. denticollis being larger (Pollock, 1998), led the authors to think that the larger of the two larvae is likely I. denticollis and the smaller I. singularis. However, a species not represented by adults cannot yet be ruled out in either case. Two of the vials from the Pichincha site contain adults of only I. denticollis and only the larger of the two species of larvae. These collections were made at a slightly higher elevation and approximately 0.5 km away from the other locality where a mixture of both species was collected. The separation of the two sites further suggests an association of the large larval type from the uphill sites with the adult of I. denticollis. In the case of the Napo site, only the smaller species of larvae was collected along with I. singularis adults, an additional reason to suppose that our larval association is valid. This will be confirmed or refuted when/if larvae are reared to adults or matched by DNA profiles.
Adults of one other Ischyomius species, I. bicolor Champion, are known from Ecuador. The Museum of Zoology -Invertebrates Section (QCAZ-I) of the Pontifical Catholic University of Ecuador, to which the Ecuador material will be returned, contains specimens of I. bicolor from the Yasuni National Park, ca. 200 m elevation. A search is planned for its yet undescribed larvae.
Terminology used in the description follows that of Lawrence (1991); the format of the description follows (as closely as possible) that of the larva of Anaplopus tuberculatus Blackburn (Pollock & Lawrence, 1995) for ease of comparison. Description: Body ( Fig. 1) elongate, parallel-sided, slightly dorsoventrally flattened, light yellow, very slightly darker on head; labrum, antennae, tergite 9, apices of tubercles and asperities dark red-brown; setae slender, elongate, sparsely positioned over body surface, with serially repeated pattern on abdominal segments 1-8.

Natural history and collection notes
Pollock (2018) provided a summary of information on the natural history of Ischyomius adults and larvae, based in large part upon work done by the second author. These insects seem to be associated with a restricted group of plant species within the monocots (e.g., Musa, Strelitzia, and Heliconia), specifically within Order Zingiberales. Label data from adult specimens also suggest a possible association with palms (Arecaceae). Additional information is presented below, giving greater detail on the collection and habitat of Ischyomius in Ecuador.
In addition to furnishing the specimens on which the genus description is based, the collections in Ecuador provided valuable information on the host plants for Ischyomius. These beetles seem to be restricted to the dead and fermenting petioles of several families within the monocot order Zingiberales, especially the genera Musa L. and Heliconia L. (Pollock, 2018). The second author has since found an adult of I. denticollis in the axils of a Renealmia plant (also in Zingiberales) where the leaf was dead and the stem fermenting, leading to speculation that eggs might be deposited there also.
Ischyomius larvae are not subcortical in dead trees as in most other Pythidae but are found inside fermenting, fleshy petioles or central leaf veins of Zingiberales. Adults of at least two Ischyomius species have been found hiding between the folded halves of dead Zingiberales leaves, usually next to the large, still-moist, central vein of recently dead leaves. Leaves that are shaded much of the day seem to be preferred by the beetles over more Figures 14-16. Ischyomius sp. larva: (14-15) urogomphal plate (dorsolateral, dorsal); (16) urogomphal plate and segment X (ventral). Scale bars = 0.5 mm. Figures 12-13. Ischyomius sp. larva: (12) anterior face of hind leg; (13) thorax and first abdominal segment, ventral. Scale bars = 0.25 and 0.5 respectively. exposed leaves. The second author's method of collecting adult Ischyomius and other Coleoptera from the same microhabitat has evolved from simply opening dead Zingiberales leaves to look for beetles to now placing the still-attached leaves into a deep insect net before cutting the stem or central leaf vein, thereby reducing the number of escapees. Leaves are shaken inside the net before being opened and examined. Adult beetles that were frequently found sharing the same microhabitat as Ischyomius include the silvanid genera Euplatamus Sharp, Silvanops Grouvelle, and Telephanus Erichson, erotylids from tribes Langurini and Toramini, and endomychids from the genera Anidrytus Gerstaecker, Archipines Strohecker, Corynomalus Chevrolat, and Epopterus Chevrolat. Occasional members of the families Carabidae, Staphylinidae, Hydrophilidae, Tenebrionidae, Chrysomelidae, Cerambycidae, and Curculionidae and miscellaneous others were also found. Ischyomius larvae were sought by carefully slitting soft, fermenting petioles with a knife and opening them by hand. Adult cillaeine Nitidulidae have been found as well inside the dead petioles.

DISCUSSION
As mentioned above, the larvae of Ischyomius and their morphological features have appeared in the "material examined" sections of a few studies, despite the fact that they had not been formally described. For example, Lawrence et al. (2011) used 516 morphological features of both adults and larvae including Ischyomius chevrolati Champion 1916 (listed in family Pythidae in material examined) and Pytho niger Kirby 1837 to elucidate a phylogeny of the entire Coleoptera. The cladogram depicts Ischyomius as sister group to Pytho Latreille and together, these are a sister group to Elacatis Pascoe (presently within Salpingidae) (clade #71). Several larval features of Ischyomius (and Pytho) were used to define these relationships: incisor edge of mandible with 2 or more subapical teeth , vestiture of legs including at least some stout spines (Fig. 12), urogomphi with accessory processes (Figs. 15-16), and sternum IX with basal row of asperities (Fig. 16). Interestingly, the larger clade (#50) that encompasses the group including Ischyomius comprises current members of Lymexylidae, Synchroidae, Oedemeridae, Pyrochroidae, Boridae and Mycteridae.
As a complement to the morphology-based paper by Lawrence et al. (2011), McKenna et al. (2015 used DNA sequence data from eight nuclear genes to elucidate relationships among the suborders and superfamilies of Coleoptera. In both Bayesian and Maximum Likelihood trees, Ischyomius and Pytho (the only exemplars of Pythidae used in this study) were recovered as sister groups, within a monophyletic cluster approximating the "Salpingid Group" of Watt (1987) and Pollock (1994). In both trees, Ischyomius + Pytho is a sister group to a monophylum consisting of three genera of Salpingidae, with Elacatis (Salpingidae) forming the sister group to these five genera.
Of the three genera mentioned by Pollock & Lawrence (1995) as provisionally belonging to Pythidae, only Ischyomius has had its larval stage formally described. Larvae of Osphyoplesius Winkler and Trimitomerus Horn remain unknown and/or undescribed. As such, the key to genera of known pythid larvae in Pollock & Lawrence (1995) requires expansion, as indicated below (note: the first two couplets in the original key separated out larvae of Boridae and Trictenotomidae and are not repeated here):

3.
Dorsal part of tergite 9 with a more or less continuous transverse series of distinct, setigerous pregomphal tubercles (4 or more per side); urogomphal lip (