Chemosensory systems play a central role in host detection, feeding behavior, and habitat selection in hematophagous insects. Here, we performed a comparative evolutionary analysis of chemosensory gene repertoires across 13 species of the Chagas disease vector genus Rhodnius. While gustatory receptors (GRs), ionotropic receptors (IRs), odorant-binding proteins (OBPs), and chemosensory proteins (CSPs) remained globally conserved, odorant receptors (ORs) displayed extensive lineage-specific expansions, tandem duplications, dynamic transcriptomic regulation, and recurrent signatures of positive selection. Major OR expansions were observed in Rhodnius robustus and Rhodnius colombiensis, suggesting increased sensory diversification in ecologically heterogeneous lineages. In contrast, conserved GR1 expression supports the maintenance of ancestral sugar-detection pathways despite hematophagy lifestyle. We further found no evidence of the canonical insect CO₂-associated GRs, suggesting alternative molecular mechanisms for CO₂ perception in Triatominae. Several receptors, including Orco, also displayed shifts in selective constraints between sylvatic and domiciliary species, consistent with sensory remodeling associated with adaptation to domestic habitats. Together, our results identify ORs as the most evolutionarily dynamic component of the Rhodnius chemosensory repertoire and highlight contrasting evolutionary trajectories among chemosensory gene families during ecological diversification and vector adaptation.

This study aimed to assess the risk of human-vector interactions and Trypanosoma cruzi transmission in touristic areas of northeastern Brazil. We analyzed 180 adult Triatoma brasiliensis specimens collected from nine peridomestic and sylvatic ecotopes across the states of Rio Grande do Norte and Paraíba. High T. cruzi infection rates were detected in both ecotopes, indicating sustained parasite circulation. Metabarcoding of blood meals revealed a broader host range than previously documented, including human blood meals in sylvatic touristic areas with high infection rates, raising concerns about vector-mediated transmission in these settings. Given the potential public health implications, our findings were formally communicated to local health and tourism authorities, with specific recommendations to inform and protect visitors in high-risk zones. These results underscore the need for targeted surveillance and vector control strategies in tourist-exposed areas, where interventions must be integrated with efforts to preserve local cultural and ecological heritage.

Isognathotermes similifinitimus Josens & Deligne sp. nov. urn:lsid:zoobank.org:act: 2CC856EC-F98A-4CA6-B7CE-C4458464CA43 Figs 26–31, 33, 64, distribution map: Fig. 47; Table 15 Diagnosis The worker has a finitimus EVA and looks a little more like I. finitimus than I. planifrons. The soldier also looks a little more like I. finitimus than I. planifrons. The imago, however, clearly looks more like I. planifrons. The specimens now assigned to I. similifinitimus sp. nov. were first considered to be close to I. planifrons; besides their mixed morphology, they were found outside the geographic range of I. planifrons. Isognathotermes similifinitimus sp. nov. is a cryptic species that can be distinguished for sure only with molecular tools: it is therefore possible that some of the samples previously identified as I. finitimus (that could not be sequenced) belong in fact to I. similifinitimus. In terms of its ecology and chorology, this species is up to now only known from the Congolese forest south of Kisangani, DRC. Etymology The name similifinitimus (from the Latin similis, ‘similar’ and finitimus from the species I. finitimus) refers to the morphology that is like that of I. finitimus. Material examined Two complete samples from one location. Holotype DEMOCRATIC REPUBLIC OF THE CONGO • soldier; Yoko; 0°17.62′ N, 25°17.77′ E; 8 Jun. 2010; B. Le Ru leg.; study code: DJ 0744; GenBank no PQ679199 (mitogenome); BE RMCA INS.Iso.059457. Paratypes DEMOCRATIC REPUBLIC OF THE CONGO • soldier, worker, ♀ (queen); same data as for holotype; BE RMCA INS.Iso.059939. Other material examined DEMOCRATIC REPUBLIC OF THE CONGO • soldier, worker, ♂ (king), ♀ (queen); Yoko; 0°17.62′ N, 25°17.98′ E; 8 Jun. 2010; B. Le Ru leg.; study code: DJ 0745; GenBank no PQ679249 (mitogenome); BE RMCA INS.Iso.059456. Historical review This species is described here; only two samples are available which may seem too little for describing a new species but both samples are complete (imago, soldier, worker, and mitogenome). It was discovered by one of us (BLR) in an evergreen forest south of Kisangani, DRC. Description Imago COLOUR. Head capsule: well sclerotised, dark, C6–C7; fontanelle concolorous with head capsule. Postclypeus concolorous with head capsule. Antennae C5–C6 without any difference between proximal and distal articles. Thorax: pronotum C6–C7, concolorous with head capsule; meso- and metanotum C6–C7, as pronotum. Legs C3–C4. Abdomen: tergites C6–C7. Sternites appreciably paler in middle (C3–C4) with both sides darker (C4–C6); posterior sternites darker (C4–C6) than anterior. SETATION. Head capsule with some prominent setae, mainly near the eyes, and a higher density of shorter, finer setae everywhere. Labrum and postclypeus with some prominent setae mixed with shorter ones. Antennae with some prominent setae, some more numerous smaller setae and, mainly distally on most articles, a bunch of very fine, bent setae (visible only at high magnification, 50 × or more). Thorax: pronotum with prominent setae mainly on margins and many shorter ones in middle; meso- and metanotum with some fine, pale setae, arranged in a medio-longitudinal strip, visible at 20 ×, with or without two larger setae on metanotum. Legs very pilose, furnished (among numerous fine setae) with some strong setae: 10–15 on the carina of fore coxa and 2 on the ventral side of fore coxa; tibia pilose; fore, mid, and hind tibia furnished with 10–30 strong setae and bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively. Abdomen: tergites with many large and small setae. Sternites with long setae, erect or directed slightly forward, and many long and smaller setae directed backwards. STRUCTURE (measurements in Table 15; Figs 26, 64). Size: the imagines of I. similifinitimus sp. nov. are medium sized in the genus Isognathotermes; as shown in Fig. 26. Head capsule: compound eyes nearly round; fontanelle a tiny round to elongate marking; ocelli nearly round to oval, removed from eyes by a distance about equal to 0.8–0.9 ocellus small diameter. Labrum: cupola shaped. Left mandible with apical tooth always longer and more prominent than first marginal; marginal teeth three in number but second one only suggested by a slight undulation of edge between first and third marginal teeth; premolar tooth with proximal end obscured by molar prominence in dorsal view; molar tooth bearing a rounded molar prominence dorsally and ending posteriorly in a tiny acute apophysis. Right mandible with apical tooth longer and more prominent than first marginal; marginal teeth two in number; first marginal tooth well developed with a sharp tip; second marginal tooth smaller and with a blunt tip; molar tooth bearing a ventral rounded flange and ending posteriorly in a kind of heel. Thorax: pronotum appreciably wider than long and narrower than head width (including the eyes), straight to very weakly sellate with anterior lobe short and very slightly elevated. Fore coxa flanged ventrally resulting in a sharp carina. Soldier COLOUR. Head capsule C4–C6; there is, in one of the two samples, a strong gradient from a darker frons to a paler back (e.g., from C6 to C4) giving the impression that the head capsule is bicolorous (as in Fig. 13). Gulamentum, antennae, and labrum concolorous with head. Mandibles dark (C7–C8) generally with an abrupt clearing on their bases (two palette levels) which is generally the same colour as frons. Thorax, nota C4 and legs paler (C2–C3). Abdomen grey owing to digestive bolus, with a yellow tinge on tergites. SETATION. Head capsule with few scattered setae; on frons a dense bunch of setae surrounds and overhangs fontanelle. Antennae with some prominent setae, more numerous smaller setae and at distal extremity of distal articles, a bunch of very fine, bent setae (visible only at high magnification, 50 × or more). Labrum always with 5–6 large setae on each lobe. Thorax: pro- and mesonotum with some setae mainly located on margins. Legs: fore coxa furnished with 1–2 spines on carina and not any one on ventral side; trochanter with some long setae, and 4–5 lined-up spines; fore, mid, and hind tibia bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively; all tibiae furnished with a row of 7–15 spines. Abdomen: tergites with some large setae, only on their posterior margins. Sternites with long setae, erect or slightly directed forward, often coloured, and smaller setae directed backwards. STRUCTURE (measurements in Table 15; Figs 28, 64). Size: the soldiers of I. similifinitimus sp. nov. are medium sized in the genus Isognathotermes; (Fig. 28). Head capsule (Fig. 64): always clearly sclerotised and appreciably longer than wide. Dorsal view: lateral sides mostly subparallel with a slight narrowing near posterior third or fourth; from antennal sockets sides converge clearly towards bases of mandibles. Upper profile clearly concave; frons: flat without any hump. Gulamentum in ventral view always constricted in its posterior half, with sides of anterior part forming a kind of ear on each side. Antennae: of 15 articles. Labrum: deeply bifurcate and wider than long, with sides lyre-shaped; lobes angular, with translucent to whitish tips; anterior margin concave. Mandibles: sabre-like, regularly curved; inner edges smooth with one distinct but very small marginal tooth, near molar tooth on each mandible; mandibles clearly shorter than head; entire surface of both mandibles smooth and glossy. Thorax: pronotum sellate, clearly narrower than head, with a slight notch in anterior margin and entire posterior margin. Fore coxa flanged ventrally resulting in a sharp carina. Gut: enteric valve seating on left side, best seen in ventral view, situated in posterior half of abdomen. Arrangement of enteric valve cushions showing trilateral symmetry: the odd cushions are 14 to 18% longer than the even cushions, with humps hardly developed, moreover, the pilosity becoming abruptly very dense shows the place where a hump is expected; secondary cushions wide at the upstream end narrowing noticeably downstream with a homogeneous spine scattering. Caecum rather small, best seen in ventral view, near centre of abdomen, lobed (three small lobes). * Abbreviations: see definitions in Material and methods. Worker COLOUR. Head capsule pale (C2–C3). Antennae: proximal articles pale (C2), distal articles two levels darker (C4). Thorax, nota and legs pale (C1–C3). Abdomen grey owing to digestive bolus. SETATION. Head capsule and postclypeus with few, erect scattered setae. Labrum with few, robust scattered setae. Antennae with some prominent setae, some more numerous smaller setae and at distal extremity of distal articles, a bunch of fine, bent setae (visible only at high magnification, 50 × or more). Thorax: nota with some scattered setae. Legs: fore coxa carinated, bearing one fine seta and furnished with 5 spines on carina and 2 on ventral side; fore trochanter with 6–7 spines; fore, mid, and hind tibia bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively and a row of spines. Abdomen: tergites with scattered setae. Sternites with long setae, erect or slightly directed forward, often coloured, and smaller setae directed backwards. STRUCTURE (measurements in Table 15; Figs 30, 64). Size: the workers of I. similifinitimus sp. nov. are, on average, medium sized in the genus Isognathotermes (but with large overlapping with several other species: Fig. 30). Head capsule weakly sclerotised (except mandibles). Antennae: 14.5 articles. Labrum: cupola shaped. Left mandible: apical tooth well developed with a sharp tip when fresh; marginal teeth three in number, first marginal well developed but with a blunt tip even when fresh, second marginal faint (visible as an undulated edge), third marginal with a blunt tip; premolar tooth with its proximal end hidden under molar prominence; molar tooth bearing a rounded molar prominence dorsally and ending posteriorly in a tiny acute apo

Isognathotermes speciosus (Sjöstedt, 1924) Figs 39–41 Revised herein as junior synonym of I. finitimus. Cubitermes speciosus Sjöstedt, 1924b: 256. Cubitermes speciosus – Sjöstedt 1926: 242–243, table 10 m. — Snyder 1949: 164. — Bouillon & Vincke 1971: 269. — Krishna et al. 2013: 1937. — Williams 1966: 102–104, figs 13, 29, 45, 61, 79. — Josens & Deligne 2019: 61. Isognathotermes speciosus – Hellemans et al. 2021: 233. Junior synonym of Isognathotermes finitimus (Schmitz, 1916). Syn. nov. Etymology The epithet speciosus means ‘beautiful, superb’ in Latin. Historical review Sjöstedt (1924b: 256) briefly described the soldier of this species under the name Cubitermes speciosus, mentioning that it is close to the soldier of C. fungifaber but larger. Two years later Sjöstedt (1926: 242–243) provided a first description of alate imago and worker together with additional information on soldier. He included (1926: 218–225) C. speciosus in imagines’ and soldiers’ keys of Cubitermes species and inserted it in a “ fungifaber -Gruppe” of species with a not protruding soldier’s frons. Snyder (1949: 164) catalogued C. speciosus in the sub-family Termitinae. Williams (1966: 102–104) provided a detailed redescription of imago and soldier along with measurements and numerous figures. Bouillon & Vincke (1971: 269) described the enteric valve of C. speciosus as belonging to the “simple type ” without any spatula. Krishna et al. (2013: 1937) housed C. speciosus in the sub-family Cubitermitinae. Josens & Deligne (2019: 42–44) placed this species within the finitimus valve pattern group. Hellemans et al. (2021: 233) placed this species in the restored genus Isognathotermes. One type sample has been examined (see Isognathotermes finitimus, DJ 0061). Krishna et al. (2013: 1937) mention that other syntype samples are deposited in AMNH and PPRI (not examined). As the type material of Isognathotermes speciosus (Sjöstedt, 1924) matches well Isognathotermes finitimus (Schmitz, 1916) and come from the same forested environment, they are from now on considered as synonyms.

Isognathotermes modicus Josens & Deligne sp. nov. urn:lsid:zoobank.org:act: CC5AB255-A7F9-4CA6-A497-9771F6FA6507 Figs 26–31, 50, distribution map: Fig. 53; Table 9 Diagnosis This is a very small species (the smallest with a finitimus EVA), so small that the first sample was initially thought to come from an incipient colony, but a physogastric queen ruled out this possibility. The worker has a finitimus EVA and is among the smallest in the genus Isognathotermes: (WT3L = 1.12–1.20 mm), and its enteric valve is also the smallest (Fig. 31). The soldier is among the smallest in the genus Isognathotermes (SHdL = 2.42–2.66 mm), as small as the smallest I. fungifaber but with a finitimus EVA. It has evenly curved mandibles with a very low acceleration index of the mandible curvature between proximal and distal parts (ln(SMlpR) - ln(SMldR) = 0.96–1.44) as it is the case for some I. planifrons but with different sizes. The imago, however, is among the medium-sized imagines in the genus Isognathotermes (Fig. 26) and rather like those of I. finitimus, I. rectimalatus sp. nov., and I. planifrons. To date, this species is only known from the “forêt des Abeilles”, Gabon. Etymology The epithet modicus means ‘modest’ in Latin. It refers to the small size of this species. Material examined Five samples from one location. Holotype GABON • soldier; Forêt des Abeilles; 0°19′ S, 12°7′ E; 1995; C. Bordereau and A. Robert leg.; study code: DJ 0862; MNHN EP9886. Paratypes GABON • worker, ♂ (king), ♀ (queen); same data as for holotype; MNHN EP9886. Other material examined GABON • soldier, ♀ (queen); Forêt des Abeilles; 0°19′ S, 12°7′ E; 1995; C. Bordereau (?) leg.; study code: DJ P214; MNHN EP9887 • soldier; Forêt des Abeilles; 0°19′ S, 12°7′ E; 1995; C. Bordereau (?) leg.; study code: DJ P216; MNHN EP9888 • soldier, worker; Forêt des Abeilles; 0°35′ S, 12°25′ E; 1995; C. Bordereau and A. Robert leg.; study code: DJ 0858; MNHN EP9884 • soldier, worker, ♂ (king), ♀ (queen); Forêt des Abeilles; 0°35′ S, 12°25′ E; 1995; C. Bordereau and A. Robert leg.; study code: DJ 0859; MNHN EP9885. Historical review This species is described here. It was discovered by C. Bordereau and A. Robert in the so-called “forêt des Abeilles”, Gabon. Description Imago COLOUR. Head capsule: well sclerotised and dark C6–C7; fontanelle concolorous with head. Postclypeus one level paler than head capsule. Antennae C5–C6. Thorax: pronotum concolorous or almost so with head capsule; meso- and metanotum C6 slightly paler than head. Legs C4. Abdomen: tergites C6. Sternites appreciably paler in the middle (C3–C5) with both sides darker (C5–C6) than anterior. SETATION. Head capsule, with some prominent setae set amongst a high density of short, fine setae forming a dense mat. Labrum and postclypeus with some prominent setae mixed with shorter ones. Antennae with some prominent setae, some more numerous smaller setae and, mainly distally on most articles, a bunch of very fine, bent setae (visible only at high magnification, 50 × or more). Thorax: pronotum with prominent setae mainly on margins and shorter ones in middle; meso- and metanotum with some fine, pale setae, arranged in a medio-longitudinal strip, generally visible at 20–40×; sometimes with one or two large setae on posterior lobe. Legs very pilose, furnished (among numerous fine setae) with 7–11 stronger setae on the carina of fore coxa and 1–3 on the ventral side of fore coxa and trochanter; tibia pilose, fore, mid, and hind tibia furnished with 30–40 spines and bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively. Abdomen: tergites with many large and small setae. Sternites with long setae, erect or directed slightly forward, and many long and smaller setae directed backwards. STRUCTURE (measurements in Table 9, Fig. 50). Size: the imagines of I. modicus sp. nov. are, on average, medium sized in the genus Isognathotermes (Fig. 26). Head capsule: compound eyes nearly round; ocelli shortly oval, removed from eyes by a distance equal to 0.9–1.3 ocellus small diameter; fontanelle generally a tiny round or elongate marking. Antennae: shortened to 12 articles by amputation of probably four articles in queens and kings. Labrum: cupola shaped, wider than long. Left mandible with apical tooth medium sized in the genus (Fig. 27) and always more prominent than first marginal; marginal teeth three in number but second one only suggested by an undulation of edge between first and third marginal teeth; only the apical tooth is acute in unworn specimens; premolar tooth almost totally obscured by molar prominence in dorsal view; molar tooth bearing a rounded molar prominence dorsally and ending posteriorly in a tiny acute apophysis. Right mandible with apical tooth always more prominent than first marginal; marginal teeth two in number; first marginal tooth well developed with a sharp tip when fresh; second marginal tooth smaller and with a blunt tip even when fresh; molar tooth bearing a ventral rounded flange and ending posteriorly in a kind of heel. Thorax: pronotum appreciably wider than long and narrower than head width (including the eyes), straight to very weakly sellate with anterior lobe short and very slightly elevated. Fore coxa flanged ventrally resulting in a sharp carina. Gut not studied. * Abbreviations: see definitions in Material and methods. Soldier COLOUR. Head capsule uniformly C5. Antennae and labrum one to two palette levels paler than head capsule. Mandibles dark (C6) with an abrupt clearing on their bases (one levels) which is generally the same colour as frons. Thorax and legs paler than head capsule (C4). Abdomen grey to red-brown owing to digestive bolus. SETATION. Head capsule with few scattered setae; on frons a dense bunch of setae surrounds and overhangs fontanelle. Antennae with some prominent setae, more numerous smaller setae and at distal extremity of distal articles, a bunch of very fine, bent setae (visible only at high magnification, 50 × or more). Labrum with 4–6 large setae on each lobe. Thorax: pro- and mesonotum with a small number of setae mainly located on margins. Legs: fore coxa with at least one fine seta and 2–3 spines on carina and not any spine on ventral side; trochanter generally with some strong lined-up setae, including 3–4 spines; fore, mid, and hind tibia bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively (the latter sometimes weakly developed) and a row of 6–12 spines along their shaft. Abdomen: tergites with some large setae, mainly or only on their posterior margins. Sternites with long setae, erect or slightly directed forward, often coloured, and smaller setae directed backwards. STRUCTURE (measurements in Table 9, Fig. 50). Size: the soldiers of I. modicus sp. nov. are part of the smallest soldiers in the genus Isognathotermes (Fig. 28). Head capsule: always clearly sclerotised; appreciably longer than wide. Dorsal view: lateral sides mostly subparallel with a very slight narrowing near posterior third or fourth; from antennal sockets sides converge towards bases of mandibles; posterior side regularly convex. Slightly concave or straight upper profile; angle between extended mandibles and frons obtuse; frons flat or with a small anterior hump. Gulamentum in ventral view always constricted in its posterior half, with sides of anterior part roundly convex. Antennae: of 14.5–15 articles. Labrum: always deeply bifurcate and wider than long, with lyre-shaped sides; lobes angular, with fine, whitish, or translucent tips; anterior margin concave. Mandibles: sabre-like; inner edges smooth with one distinct but small marginal tooth, near molar tooth on each mandible; mandibles clearly shorter than head; entire surface of both mandibles smooth and glossy. Right mandible slightly more curved than left. Thorax: pronotum sellate, as wide as 56–60% of head width, with straight anterior and posterior margins. Fore coxa flanged ventrally resulting in a sharp carina. Gut: enteric valve seating on left side, best seen in ventral view, situated in second half of abdomen. Caecum rather small, best seen in ventral view, near centre of abdomen, three-lobed. Arrangement of enteric valve cushions showing trilateral symmetry, the odd cushions recognizable as a finitimus EVA. Worker COLOUR. Head capsule pale (C2–C3). Antennae: proximal articles pale (C2), distal articles two levels darker (C4). Thorax, nota, and legs pale (C2–C3). Abdomen grey to red-brown owing to digestive bolus. SETATION. Head capsule and postclypeus with few, erect, scattered setae. Labrum with few, robust scattered setae. Antennae with some prominent setae, some more numerous smaller setae and at distal extremity of distal articles, a bunch of fine, bent setae (visible only at high magnification, 50 × or more). Thorax: nota with some scattered setae. Legs: fore coxa carinated, bearing one fine seta and furnished with 3–5 spines on carina and not any one on ventral side; fore trochanter with 4–6 lined up spines; fore, mid, and hind tibia bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively and a row of 6–12 spines. Abdomen: tergites with scattered setae. Sternites with long setae, erect or slightly directed forward, often coloured, and smaller setae directed backwards. STRUCTURE (measurements in Table 9, Fig. 50). Size: the workers of I. modicus sp. nov. are the smallest workers with a finitimus EVA, of the same size as the smallest workers of I. fungifaber (which have a fungifaber EVA). Head capsule: weakly sclerotised (except mandibles). Antennae: 14.5 articles. Labrum: cupola shaped. Left mandible: apical tooth well developed with a sharp tip when fresh; marginal teeth three in number, first marginal well developed but with a blunt tip even when fresh, second marginal faint (visible as an undulated edge and disappearing in worn mandibles), third marginal with a blunt tip; premolar tooth with its proximal end generally hidden under molar prominence; molar tooth be

Isognathotermes kemneri (Emerson, 1928) Figs 36–37 Revised herein as junior synonym of I. bulbifrons. Mirotermes (Cubitermes) kemneri Emerson, 1928: 509, fig. 54. Cubitermes kemneri – Snyder 1949: 161. Cubitermes zenkeri – Ruelle 1975: 8; 1992: 501. — Krishna et al. 2013: 1945. — Josens & Deligne 2019: 42–44, 60 (incorrect synonym of C. zenkeri). Isognathotermes kemneri – Hellemans et al. 2021: 233. Junior synonym of Isognathotermes bulbifrons (Sjöstedt, 1924). Syn. nov. Etymology In the original description, Emerson (1928: 509) stated: “I take pleasure in naming this species in honor of Dr. N. Kemner, who has done such valuable work on oriental termites and termitophiles.” Historical review Emerson (1928: 509) described this species under the name Mirotermes (Cubitermes) kemneri. He provided description and measurements of the soldier, noting that it is very close to Cubitermes heghi. Snyder (1949: 161) catalogued this species under the name C. kemneri and housed it in the sub-family Termitinae. Ruelle (1975: 8), on the advice of Emerson (in litteris), proposed C. kemneri as a junior synonym of C. zenkeri. This is one of the species that Ruelle (1992: 501) called “forgotten species”. Krishna et al. (2013: 1945) housed C. kemneri in the sub-family Cubitermitinae. Espousing the view of Ruelle 1975 they considered this species as a junior synonym of C. zenkeri. However, C. kemneri cannot be synonymized with C. zenkeri because the types of both species have different EVAs, i.e., of the finitimus and fungifaber groups respectively (Josens & Deligne 2019: 42–44). Hellemans et al. (2021: 233) placed this species in the restored genus Isognathotermes. One type subsample has been examined (see Isognathotermes bulbifrons, DJ 0297). Krishna et al. (2013: 1945) mention that other subsamples are deposited in UMMZ: holotype (soldier), and a paratype subsample in PPRI (not examined). Only two samples in the museum collections bore the name “ C. kemneri ”: the type sample and a misidentified sample (presently assigned to I. fungifaber). Our morphology study (this article) shows that the type matches well I. bulbifrons. Therefore, I. kemneri (Emerson, 1928) is from now on considered a junior synonym of Isognathotermes bulbifrons (Sjöstedt, 1924).

Isognathotermes modestior (Silvestri, 1914) Figs 28, 30–31, 61–63, distribution map: Fig. 45 Revised herein as junior synonym of I. severus. Cubitermes severus var. modestior Silvestri, 1914: 93–94, fig. LV. Cubitermes modestior – Sjöstedt 1926: 238–239. — Snyder 1949: 161. — Krishna et al. 2013: 1927. — Josens & Deligne 2019: 60. Isognathotermes modestior – Hellemans et al. 2021: 233. Junior synonym of Isognathotermes severus (Silvestri, 1914). Syn. nov. Etymology The epithet modestior means ‘smaller’ in Latin. It referred to the smaller size of the variety C. severus modestior when compared to the main form, as underlined by Silvestri in the original description. Historical review Silvestri (1914: 93–94) described this taxon under the name Cubitermes severus var. modestior; it is known from a single nest. He provided a short description of imago, soldier, and worker together with measurements and figures, mentioning that the imagines are like the typical form (= Cubitermes severus), and indeed they are among the smallest individuals of the other I. severus imagines but also that the soldiers and workers are smaller than the main form. Sjöstedt (1926: 238–239) considered that this taxon deserved to be elevated to the species level. He referred to it under the name Cubitermes modestior, housed it in the sub-family Termitinae, and included it in the “ glebae -Gruppe” of small Cubitermes species. Snyder (1949: 161) catalogued this species under the name C. modestior and housed it in the sub-family Termitinae. Krishna et al. (2013: 1927) referred to this species under the name C. modestior and housed it in the sub-family Cubitermitinae. Josens & Deligne (2019: 39–42) placed this species within the fungifaber valve pattern group. Hellemans et al. (2021: 233) placed this species in the restored genus Isognathotermes. Two type subsamples have been examined (see Isognathotermes severus, DJ 0294 and DJ 0341). Krishna et al. (2013: 1927) mention that another subsample is deposited in PPRI (not examined). Since the type material of I. modestior (Silvestri, 1914) comes from a single nest and is included in the smaller specimens of I. severus (from other regions), it is from now on considered a junior synonym of Isognathotermes severus (Silvestri, 1914).

Cubitermitinae species whose workers have enteric valves of the fungifaber and finitimus patterns were previously gathered into the genus Isognathotermes. Here, we revise this genus, combining the morphological and anatomical features of all castes, as well as the species’ chorological, ecological, and phylogenetic (mitogenome) characteristics in an integrative taxonomic approach. In addition to the genetic signature, the best morphological criteria for generic recognition are found in the workers’ EVA (enteric valve architecture) and the spinosity of the fore coxae of the workers, and the best criteria for specific recognition are found in (a) the valve and caecum of the workers, (b) the caecum, dimensions of the head, and curvature of the mandibles of the soldiers, and (c) the dimensions of the head and the relative dimensions of eyes and ocelli of the imagines. After revision, only eight of the 22 known taxa remain valid, 14 known taxa become junior synonyms, and one is considered as incertae sedis; six new morphologically recognizable species, one cryptic species and four new subspecies are described. The mitogenome of 12 out of the 15 valid species have been sequenced. The following described species are considered valid: I. bulbifrons (Sjöstedt, 1924), I. finitimus (Schmitz, 1916), I. fungifaber (Sjöstedt, 1896), I. minitabundus (Sjöstedt, 1913), I. planifrons (Sjöstedt, 1924), I. severus (Silvestri, 1914), I. ugandensis (Fuller, 1923) and I. zenkeri (Desneux, 1904). The following species are described as new: I. acristatus Josens & Deligne sp. nov., I. modicus Josens & Deligne sp. nov., I. magniplanifrons Josens & Deligne sp. nov., I. phallicaecalis Josens & Deligne sp. nov., I. phalloides Josens & Deligne sp. nov., I. rectimalatus Josens & Deligne sp. nov., and I. similifinitimus Josens & Deligne sp. nov. Four new subspecies of I. ugandensis are recognized: I. ugandensis burundii Josens & Deligne subsp. nov., I. ugandensis kenyae Josens & Deligne subsp. nov., I. ugandensis malawii Josens & Deligne subsp. nov., and I. ugandensis ugandensis Josens & Deligne subsp. nov. The following species are considered junior synonyms: I. antennalis (Sjöstedt, 1924), I. banksi (Emerson, 1928), I. comstocki (Emerson, 1928), I. fungifaber var. elongata (Sjöstedt, 1924), I. gaigei (Emerson, 1928), I. gibbifrons (Sjöstedt, 1924), I. heghi (Sjöstedt, 1924), I. kemneri (Emerson, 1928), I. loubetsiensis (Sjöstedt, 1924), I. modestior (Silvestri, 1914), I. schmidti (Emerson, 1928), I. silvestrii (Sjöstedt, 1925), I. speciosus (Sjöstedt, 1924), and I. subarquatus (Sjöstedt, 1926). Are considered as incertae sedis: I. bredoi (nomen nudum), I. congoensis (Emerson, 1928), an “I. aff. planifrons” (undescribed), and an unknown Isognathotermes from Malawi (undescribed). GenBank accessions: see Suppl. file 3 and ‘Material examined’ of each chapter.

Isognathotermes ugandensis malawii Josens & Deligne subsp. nov. urn:lsid:zoobank.org:act: 025368E6-C9F7-46E7-88F5-7D7BE6A1DCAF Figs 65–66, 69 Diagnosis Soldiers and workers do not allow this subspecies to be distinguished from the other three. The imago is close to that of I. u. kenyae subsp. nov. (but their geographical distributions are disjunct); it is, on average, the second largest of the four subspecies, best seen in a PCA (“ugma”, Fig. 65); it is the subspecies with the greatest distance between the apical and the first marginal teeth (IMlAmD = 0.23–0.27 mm vs 0.20–0.25 mm for all three other subspecies, Table 17). It comes from a relatively high altitude (1460–1770 m, average = 1560 m), from Malawi and northern Zambia. Etymology The name malawii (from Malawi) refers to the country of the type locality. Material examined Eleven samples from eight locations; all old museum samples initially identified either as Cubitermes minitabundus (six samples) or C. ugandensis (five samples). Holotype MALAWI • soldier; Dowa-Lilongwe road; 13°39′ S, 33°51′ E; 13 Sep. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ 0699; initially C. minitabundus; NHMUK 13671919. Paratypes MALAWI • soldier, worker, ♀ (queen); same data as for holotype. Other material examined MALAWI • soldier, worker, ♀ (queen); Dedza; 14°22′ S, 34°19.5′ E; 4 Sep. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ 0688; initially C. minitabundus; NHMUK 13671917 • soldier, worker; Dedza; 14°23.5′ S, 34°18.5′ E; 2 Sep. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ 0698; initially C. minitabundus; NHMUK 13671918 • soldier, ♀ (queen); Dowa – Lilongwe road; 13°39′ S, 33°51′ E; 11 Sep. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ 0697; initially C. minitabundus; NHMUK 13671916 • soldier; Mzimba – Kasungu road; 12°16′ S, 33°38′ E; 18 Sep. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ U073; initially C. ugandensis; NHMUK 13672015 • soldier, worker; Nyika Plateau; 10°54′ S, 33°26′ E; 29 Sep. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ 0670; initially C. ugandensis; NHMUK 13672018 • soldier, worker; Nyika Plateau; 10°45′ S, 33°57′ E; 30 Sep. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ 0671; initially C. ugandensis; NHMUK 13672016 • soldier; Vipya Plateau, Ekwendeni; 11°23′ S, 33°54′ E; 20 Oct. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ U072; initially C. ugandensis; NHMUK 13672017. ZAMBIA • soldier; Tunduma; 9°26′ S, 32°51′ E; 5 Aug. 1953; W.A. Sands and W. Wilkinson leg.; study code: DJ U080; initially C. ugandensis; NHMUK 13672009 • soldier, worker; Kasama–Mbala road; 10°2′ S, 31°15′ E; 23 Dec.1969; M.G. Bingham leg.; study code: DJ 0690; initially C. minitabundus; NHMUK 13671923 • soldier, worker; Mbala–Mpulungu road; 8°48′ S, 31°8′ E; 24 May 1970; J.R. Clover and Kistner leg.; study code: DJ 0689; initially C. minitabundus; NHMUK 13671924. Description Imago See the description of the imago of Isognathotermes ugandensis (Fuller, 1923) with the particularity that the imago of I. u. malawii subsp. nov. is the second largest of the four subspecies, best seen in a PCA (“ugma” in Fig. 65, Table 17). This is the subspecies with the greatest distance between the apical and the first marginal teeth (IMlAmD = 0.23–0.27 mm vs 0.20–0.25 mm for all three other subspecies, Table 17). Soldier See the description of the soldier of Isognathotermes ugandensis (Fuller, 1923). Worker See the description of the worker of Isognathotermes ugandensis (Fuller, 1923). Chorology-ecology Isognathotermes ugandensis malawii subsp. nov. was found between altitudes of 1460 and 1770 m (average = 1560 m), in the central Zambezian miombo woodlands in Malawi and northern Zambia. Molecular data No genetic sequence is currently available.

Isognathotermes phallicaecalis Josens & Deligne sp. nov. urn:lsid:zoobank.org:act: 0647CCCF-BF5A-4FEA-9C01-84FF782A06D9 Figs 26–31, 33, 51–52, distribution map: Fig. 53; Table 10 Diagnosis Soldiers and workers of Isognathotermes phallicaecalis sp. nov. and I. phalloides sp. nov. share the same kind of phalloid cecum, extended forward in a finger-like process that can be swollen distally or capped or crowned (Figs 52, 56), which distinguishes them from all other species of Isognathotermes; they have morphologically similar soldiers, with a slight difference in the curvature of the mandibles (Fig. 22); however, they come from different ecosystems: continental evergreen forest in the case of I. phallicaecalis, forest galleries in the case of I. phalloides. The worker of I. phallicaecalis sp. nov. has a finitimus EVA and is among the small workers in the genus Isognathotermes. Its head is, on average, somewhat wider than that of I. phalloides sp. nov. (Fig. 30). The soldier’s EVA can generally be recognized as belonging to the finitimus pattern; on average it has longer mandibles than I. phalloides sp. nov.: SMlL = 2.17 –2.53 mm (vs 1.99–2.36 mm in I. phalloides). The imago is unknown. This species is also defined by its ecology and chorology: to date it has been found only in forested environments of the northern Congo Republic and Cameroon (Fig. 53). Etymology The epithet phallicaecalis from the Greek φαλλός (phallos, phallus) and the Latin caecalis (of the caecum) refers to the caecum phalloid morphology in workers and soldiers. Material examined Twelve samples from three locations. Holotype CONGO REPUBLIC • soldier; Loundoungou; 2°22.827′ N, 17°4.226′ E; 4 Dec. 2017; Y. Roisin leg.; study code: DJ 0529; GenBank nos MN646722 (COI) MN685925 (COII) MN685986 (28S) PQ679196 (mitogenome); BE RMCA INS.Iso.059288. Paratypes CONGO REPUBLIC • soldier, worker; same data as for holotype; BE RMCA INS.Iso.059936. Other material examined CAMEROON • soldier; Dja Rock; 3°20.73′ N, 12°42.84′ E; 30 Mar. 2015; J. Šobotnik leg.; study code: DJ B320; BE RMCA INS.Iso.059287. CONGO REPUBLIC • soldier, worker; Mokabi; 3°8.796′ N, 16°57.869′ E; 8 Dec. 2017; Y. Roisin leg.; study code: DJ 0527; GenBank no PQ679190 (mitogenome); BE RMCA INS.Iso.059290 • soldier, worker; Mokabi; 3°8.795′ N, 16°57.826′ E; 8 Dec. 2017; Y. Roisin leg.; study code: DJ 0528; BE RMCA INS.Iso.059289 • soldier, worker; Loundoungou; 2°22.929′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0759; GenBank no PV564652 (mitogenome); BE RMCA INS.Iso.059296 • soldier, worker; Loundoungou; 2°22.93′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0760; GenBank no PV564657 (mitogenome); BE RMCA INS.Iso.059291 • soldier, worker; Loundoungou; 2°22.93′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0761; GenBank no PQ679202 (mitogenome); BE RMCA INS.Iso.059297 • soldier, worker; Loundoungou; 2°22.93′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0762; BE RMCA INS.Iso.059292 • soldier, worker; Loundoungou; 2°22.93′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0763; BE RMCA INS.Iso.059295 • soldier, worker; Loundoungou; 2°22.93′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0764; BE RMCA INS. Iso.059293 • soldier, worker; Loundoungou; 2°22.93′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0765; BE RMCA INS.Iso.059298 • soldier, worker; Loundoungou; 2°22.93′ N, 17°4.301′ E; Dec. 2018; S. Lenz leg.; study code: DJ 0766; GenBank no PV564655 (mitogenome); BE RMCA INS. Iso.059294. Historical review This species is described here; the imago is still unknown. It was recently discovered by three of us (JŠ, YR & SL) in Cameroon and Northern Congo. Description Imago The imago is unknown. Soldier COLOUR. Head capsule tending towards faded palette (Cf4–Cf5); there is a gradient from a darker frons to a paler back. Antennae and labrum concolorous with or somewhat paler than frons. Mandibles dark (C7–C8) with an abrupt clearing on their bases (C5–C6) which is concolorous with frons. Thorax and legs generally paler (C3–C4) than head capsule. Abdomen grey to red-brown owing to digestive bolus, sometimes with a yellowish tinge on tergites. SETATION. Head capsule with few scattered setae; on frons a dense bunch of setae surrounds and overhangs fontanelle. Antennae with some prominent setae, more numerous smaller setae and at distal extremity of distal articles, a bunch of fine, bent setae (visible only at high magnification, 50 × or more). Labrum always with 3–6 large setae on each lobe. Thorax: pro- and mesonotum with a small number of setae mainly located on margins. Legs: fore coxa bear 1–3 spines on carina and none (rarely one) on ventral side; trochanter with 2–8 spines; fore, mid, and hind tibia bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively and a row of 8–15 spines along their shaft. Abdomen: tergites with some large setae, mainly on their posterior margins. Sternites with long setae, erect or slightly directed forward, often coloured, and smaller setae directed backwards. STRUCTURE (measurements in Table 10; Figs 51–52). Size: the soldiers of I. phallicaecalis sp. nov. are medium sized among of the genus Isognathotermes (Fig. 28). Head capsule: always clearly sclerotised, appreciably longer than wide. Dorsal view: lateral sides mostly subparallel with a clear narrowing near posterior fourth (Fig. 51), from antennal sockets sides converge more or less clearly towards bases of mandibles; posterior side regularly rounded or with a short straight middle part; upper profile concave. Angle between extended mandibles and frons a little obtuse; frons without any or with a week anterior hump. Gulamentum in ventral view always constricted in its posterior half, with sides of anterior part forming an acute widening or a kind of ear on each side. Antennae: of 14.5–15 articles. Labrum: always deeply bifurcate and wider than long, with lyre-shaped sides; lobes angular, with fine, translucent tips; anterior margin concave. Mandibles: sabre-like with a middle-sized curvature in the genus Isognathotermes; inner edges generally smooth with one distinct but very small marginal tooth, near molar tooth on each mandible; mandibles clearly shorter than head; entire surface of both mandibles smooth and glossy. Right mandible slightly more curved than left. Thorax: pronotum sellate, as wide as 59–64% of head width, with generally entire anterior and posterior margins. Fore coxa flanged ventrally resulting in a more or less sharp carina. Gut: enteric valve seating on left side, best seen in ventral view, situated in second half of abdomen. Caecum always rather well developed, best seen in ventral view, near centre of abdomen, as a typical finger-like process, extended forward and generally swollen distally or seeming to be capped (Figs 52, 56). Arrangement of enteric valve cushions showing trilateral symmetry, the odd cushions being 20–25% longer than the even cushions, with a pilosity becoming very dense distally showing the place where a hump would be expected (in comparison with the worker’s EVA). * Abbreviations: see definitions in Material and methods. Worker COLOUR. Head capsule pale (C2–C3). Antennae: proximal articles pale (C2), distal articles two levels darker (C4). Thorax, nota, and legs pale (C1–C3). Abdomen grey to red-brown owing to digestive bolus. SETATION. Head capsule and postclypeus with few, erect, scattered setae. Labrum with few, robust scattered setae. Antennae with some prominent setae, some more numerous smaller setae and at distal extremity of distal articles, a bunch of fine, bent setae (visible only at high magnification, 50 × or more). Thorax: nota with some scattered setae. Legs: fore coxa carinated, bearing one fine seta and furnished with 3–6 spines on carina and 1–2 on ventral side; fore trochanter with 5–7 spines; fore, mid, and hind tibia bearing 3, 2, 2 apical spurs and 0, 2, 0 subapical spurs respectively and a row of 6–15 spines. Abdomen: tergites with scattered setae. Sternites with long setae, erect or slightly directed forward, often coloured, and smaller setae directed backwards. STRUCTURE (measurements in Table 10; Figs 51–52). Size: the workers of I. phallicaecalis sp. nov. are, on average, medium sized among the genus Isognathotermes (Fig. 30). Head capsule: weakly sclerotised (except mandibles). Antennae of 14.5 (rarely 14) articles. Labrum: cupola shaped. Left mandible: apical tooth well developed with a sharp tip when fresh; marginal teeth three in number, first marginal tooth well developed but with a blunt tip even when fresh, second marginal tooth faint (visible as an undulated edge and disappearing in worn mandibles), third marginal tooth with a blunt tip; premolar tooth with its proximal end not hidden under molar prominence; molar tooth bearing a rounded molar prominence dorsally and ending posteriorly in a tiny acute apophysis. Right mandible: apical tooth well developed with a sharp tip when fresh; marginal teeth two in number; first marginal tooth well developed with a sharp tip when fresh; second marginal tooth smaller and with a blunt tip even when fresh; molar tooth bearing a ventral rounded flange and ending posteriorly in a kind of heel. Thorax: pronotum sellate, as wide as 66–71% of head width. Fore coxa flanged ventrally resulting in a sharp carina. Gut: enteric valve seating on left side, best seen in ventral view, situated in second half of abdomen. Arrangement of enteric valve cushions of the finitimus pattern (Fig. 5) with triradial symmetry: the odd PCs, in their downstream part, bear a higher density of rather short bristles on a globular bulge, the latter sometimes weakly developed; supporting bristles are numerous: 18–33 on each side of the odd PCs (Fig. 51); secondary cushions are wide at the upstream end, narrowing noticeably downstream with a homogeneous spine scattering. Caecum always well developed, best seen in ventral view, near centre of abdomen,

Isognathotermes silvestrii (Sjöstedt, 1925) Figs 62–63 Revised herein as junior synonym of I. severus. Cubitermes Silvestrii Sjöstedt, 1925: 54. Cubitermes silvestrii – Sjöstedt 1926: 251, table 10 a1–a2. — Snyder 1949: 164. — Ruelle 1992: 500. — Krishna et al. 2013: 1937. — Josens & Deligne 2019: 61. Isognathotermes silvestrii – Hellemans et al. 2021: 233. Junior synonym of Isognathotermes severus (Silvestri, 1914). Syn. nov. Etymology The species was named in honour of F. Silvestri who collected the type material. Historical review Sjöstedt (1925: 54) briefly described the soldier of this species under the name Cubitermes Silvestrii. One year later Sjöstedt (1926: 251) provided more detailed descriptions together with figures of the soldier, specifying that they were based on specimens from Kakoulima (Guinea) first identified as Cubitermes severus by Silvestri. He housed the species in the sub-family Termitinae, included it (1926: 218–226) in a soldier’s key of Cubitermes species and inserted it in a “ bulbifrons -Gruppe” of species with an upwards humped frons. Snyder (1949: 164) catalogued Cubitermes silvestrii in the sub-family Termitinae. This is one of the species that Ruelle (1992: 500) called “forgotten species”. Krishna et al. (2013: 1937) housed C. silvestrii in the sub-family Cubitermitinae. Josens & Deligne (2019: 39–42) placed this species within the fungifaber valve pattern group. Hellemans et al. (2021: 233) placed this species in the restored genus Isognathotermes. One type sample has been examined (see Isognathotermes severus, DJ 0302). Krishna et al. (2013: 1937) mention that another subsample is deposited in NHRM (not examined). The type of Cubitermes silvestrii came from the same locality as one of the syntypes of Cubitermes severus and our morphology study shows that it matches well I. severus. Therefore, Cubitermes silvestrii (Sjöstedt, 1925) is from now on considered a junior synonym of Isognathotermes severus (Silvestri, 1914).

Blood feeding is a secondary adaptation in hematophagous bugs. Many proteins are secreted in the saliva that are devoted to coping with the host's defense and to process the blood meal. Digestive enzymes that are no longer required for a blood meal would be expected to be eventually lost. Yet, in many strictly hematophagous arthropods, α-amylase genes, which encode the enzymes that digest starch from plants, are still present and transcribed, including in the kissing bug Rhodnius prolixus (Hemiptera, Reduviidae) and its related species, which transmit the Chagas disease. We hypothesized that retaining α-amylase could be advantageous if the bugs occasionally consume plant tissues. We first checked that the α-amylase protein of Rhodnius robustus retains normal amylolytic activity. Then we surveyed hundreds of gut DNA extracts from the sylvatic R. robustus to detect traces of plants. We found plant DNA in 8% of the samples, mainly identified as Attalea palm trees, where R. robustus are usually found. We suggest that although of secondary importance in the blood-sucking bugs, α-amylase may be needed during occasional plant feeding and thus has been retained.

Exploring the dynamics of disease transmission involves an understanding of complex interactions within the eco-epidemiologic framework. In the context of Chagas disease (CD), elements are mainly represented by the interactions among the pathogen, insect vector, host, humans and the environment. We performed quantitative and qualitative analyses on a dataset derived from 98 Triatoma brasiliensis infected by trypanosomatids, which were linked to a CD outbreak in the semi-arid region of northeastern Brazil. We extracted invertebrate-derived DNA (iDNA) from these insects, comprising 18 populations around the outbreak area, each indicative of various strata of anthropogenic influence. Food source (FS) diversity, representing potential parasite reservoirs, was determined through mitochondrial gene (cyt b) sequencing of vertebrates, and parasite genotyping was accessed using fluorescent amplified fragment barcodes (FFLB) of trypanosomatids. We also assessed the residents' awareness of breeding sites for CD vectors in the inspected houses. The quantification of Trypanosoma cruzi was estimated via real-time PCR and is denominated here as the average parasite load (PL) per insect (T. cruzi/intestinal unit). We aimed to address vector-parasite-host-environment interactions that were discussed based on their significance among the components. Notably, among the significant interactions, we observed that the PL in the insects was significantly influenced by FS. Infected insects that fed on the classic reservoir, Didelphis albiventris, and Galea spixii exhibited higher PLs, compared to those that fed on Kerodon rupestris (p < 0.04)—a primary host. While D. albiventris is already recognized as a synanthropic species, we propose that G. spixii may also be undergoing a synanthropic process. Conversely, domestic cats are frequently identified as FS in infected insects from the sylvatic environment, suggesting a possible change in their behavior towards a wild state. Therefore, we propose that neglected anthropogenic actions have facilitated the reciprocal (sylvatic-peridomestic) circulation of T. cruzi—especially noted for TcI because it was predominant in insects found in peridomestic environments. Residents are often unaware of the existence of insect breeding grounds near their homes, particularly when it involves the storage of materials without planning for use, such as piles of tiles, bricks and wood. Although indirect inferences about the interaction among vector-parasite-host-environment are still incipient, we highlight the potential use of vectors as natural samplers of biological and ecological components in transmitting the disease.

The higher classification of termites requires substantial revision as the Neoisoptera, the most diverse termite lineage, comprise many paraphyletic and polyphyletic higher taxa. Here, we produce an updated termite classification using genomic-scale analyses. We reconstruct phylogenies under diverse substitution models with ultraconserved elements analyzed as concatenated matrices or within the multi-species coalescence framework. Our classification is further supported by analyses controlling for rogue loci and taxa, and topological tests. We show that the Neoisoptera are composed of seven family-level monophyletic lineages, including the Heterotermitidae Froggatt, Psammotermitidae Holmgren, and Termitogetonidae Holmgren, raised from subfamilial rank. The species-rich Termitidae are composed of 18 subfamily-level monophyletic lineages, including the new subfamilies Crepititermitinae, Cylindrotermitinae, Forficulitermitinae, Neocapritermitinae, Protohamitermitinae, and Promirotermitinae; and the revived Amitermitinae Kemner, Microcerotermitinae Holmgren, and Mirocapritermitinae Kemner. Building an updated taxonomic classification on the foundation of unambiguously supported monophyletic lineages makes it highly resilient to potential destabilization caused by the future availability of novel phylogenetic markers and methods. The taxonomic stability is further guaranteed by the modularity of the new termite classification, designed to accommodate as-yet undescribed species with uncertain affinities to the herein delimited monophyletic lineages in the form of new families or subfamilies. Here, the authors produce an updated termite classification with genomic scale analyses, highlighting thirteen family-level lineages and resilience of their classification to future termite research.

Blood feeding is a secondary adaptation in hematophagous bugs. A lot of secreted proteins are devoted to cope with the host’s defences an to process the blood meal. In contrast, one can expect that digestive enzymes that became useless were lost during or after this peculiar adaptation. And yet in many strictly hematophagous arthropods, alpha-amylases genes, coding the enzymes that digest starch from plants, are still present and transcribed, including the kissing bug Rhodnius prolixus (Hemiptera, Reduviidae) and its related species, which transmit the Chagas disease. We hypothesized that retaining alpha-amylase could be advantageous if the bugs happen to consume occasionally plant tissues. To this end, we first checked that the alpha-amylase protein of Rhodnius robustus retained normal amylolytic activity. Then we surveyed hundreds of gut DNA extracts from the sylvatic R. robustus to detect traces of plants. We found plant DNA in 8% of the samples, mainly identified as Attalea palm trees, the usual dwelling of R. robustus. We suggest that although maybe rarely of use in a bug lifetime, alpha-amylase may be crucial at some nutritionally critical moments and thus has been preserved.

BACKGROUND Blood feeding is a secondary adaptation in hematophagous bugs that ancestrally feed on plants. Many vector proteins are devoted to cope with the host’s defenses and to process the blood meal. In contrast, one can expect that some digestive enzymes devoted to phytophagous diet were lost during or after this peculiar adaptation. And yet, in many strictly hematophagous arthropods, alpha-amylases genes, coding the enzymes that digest starch from plants, are still present and transcribed, including in the blood-sucking bug Rhodnius prolixus and its related species R. robustus (Hemiptera, Reduviidae, Triatominae). Triatominae bugs are vectors of Trypanosoma cruzi , the causal agent of Chagas’disease. Besides the parasitic human infection by the vector-borne route via an exposition to infected feces, an oral route is documented by the ingestion of contaminated food or juices made from palm fruit trees. METHODOLOGY/PRINCIPAL FINDINGS We hypothesized that retaining alpha-amylase could be advantageous if the bugs happen to consume occasionally plant tissues. To this end, we surveyed hundreds of gut DNA extracts from the sylvatic species R. robustus caught on palm trees to detect traces of plant meals. We found plant DNA in over 8 % of the R. robustus samples, mostly the palm tree Attalea speciosa . Moreover, we showed that the R. robustus alpha-amylase retained normal amylolytic activity. CONCLUSIONS Preserving alpha-amylase function could be an important way of optimally harness plant substrates, and plant feeding could be a way for bridging the gap between two blood meals. Our data indicate that plants are a common and yet underestimated food source in the wild for Triatomine. Author Summary Adaptation to a specific diet is often accompanied by metabolic, behavioral, physiological changes and hence by genetic changes like gene family expansion, gene losses or gains. In blood-sucking insects some adaptive features such as salivary components acting against blood clotting are known. However, it is intriguing that a digestive enzyme, alpha-amylase, which digests starch, is conserved in those animals, because blood does not contain starch nor related glucose polymers. This is the case in the blood-sucking bugs of the Rhodnius genus (Hemiptera, Reduviidae), which are vectors of the Chagas’disease, an important health issue in Latin America. In this study, we evidence for the first time that sylvatic bugs R. robustus also consume plant tissues in the wild. We detected by PCR performed on DNA from digestive tract that a significant number of wild-caught individuals harbored plant DNA, especially from Attalea palm trees, on which they used to nest. We showed that the amylase enzyme is normally active on starch. We suggest plant feeding could be a way for bridging the gap between two blood meals but might not be linked to nutritional distress.

SummaryThe nutritional symbiosis promoted by bacteria is a key determinant for adaptation and evolution of many insect lineages. A complex form of nutritional mutualism that arose in blood-sucking insects critically depends on diverse bacterial symbionts that supplement the diet of their nutrient-poor hosts with B vitamins. For instance, the triatomine bugRhodnius prolixus, one of the main vectors of the Chagas disease in humans, is known to maintain a nutritional symbiosis with the gut symbiontsRhodococcus rhodnii.In this study, we show thatWolbachiasymbionts are also widely distributed in theRhodniusgenus. We have screened a large set ofRhodniusblood-sucking bugs samples belonging to 17 different species and to the three phylogenetic groups,prolixus, pallescensandpictipes. We assembled 13 genomes ofWolbachiainfecting eightRhodniusspecies fromprolixusandpictipesgroups. We demonstrate that theseWolbachiabelong to supergroup F and are closely related toWolbachiainfecting the bedbugCimex lectularius(wCle). Although bedbugs and triatomines are very distantly related hemipteran bugs, the genomes of their respectiveWolbachiawere highly similar, suggesting recent horizontal host switches. We also show thatRhodnius Wolbachiagenomes infecting theprolixusgroup encode intact biotin operon, the hallmark of nutritional symbiosis in bedbugs. This operon is lacking from all the otherWolbachiainfectingR. pictipes. Finally, host genome analyses provide evidence of massiveWolbachia-to-Rhodniusgene transfers in almost samples, providing footprints of past infections that support a widespread and probably ancient symbiotic association betweenWolbachiaand triatomine bugs.Our results suggest that bothWolbachiaandR. rhodniigut symbionts and theirRhodniushost maintain a highly prevalent symbiotic relationship, in which the vertically-inheritedWolbachiahas the metabolic potantial to ensure or complement, the nutritional mutualism provided by the gut symbionts. Specific loss of the biotin operon in some symbiont genomes suggests that the boundaries between obligatory mutualism, facultative mutualism and parasitism inWolbachiaare transient and fluid, supporting a dynamic process of transition and reversion from one state to another.

We provide in this study a very large DNA dataset on Rhodnius species including 36 samples representing 16 valid species of the three Rhodnius groups, pictipes, prolixus and pallescens. Samples were sequenced at low-depth with whole-genome shotgun sequencing (Illumina technology). Using phylogenomics including 15 mitochondrial genes (13.3 kb), partial nuclear rDNA (5.2 kb) and 51 nuclear protein-coding genes (36.3 kb), we resolve sticking points in the Rhodnius phylogeny. At the species level, we confirmed the species-specific status of R. montenegrensis and R. marabaensis and we agree with the synonymy of R. taquarussuensis with R. neglectus. We also invite to revisit the species-specific status of R. milesi that is more likely R. nasutus . We proposed to define a robustus species complex that comprises the four close relative species: R. marabaensis, R. montenegrensis, R. prolixus and R. robustus. As Psammolestes tertius was included in the Rhodnius clade, we strongly recommend reclassifying this species as R. tertius . At the Rhodnius group level, molecular data consistently supports the clustering of the pictipes and pallescens groups, more related to each other than they are to the prolixus group. Moreover, comparing mitochondrial and nuclear tree topologies, our results demonstrated that various introgression events occurred in all the three Rhodnius groups, in laboratory strains but also in wild specimens. We demonstrated that introgressions occurred frequently in the prolixus group, involving the related species of the robustus complex but also the pairwise R. nasutus and R. neglectus . A genome wide analysis highlighted an introgression event in the pictipes group between R. stali and R. brethesi and suggested a complex gene flow between the three species of the pallescens group, R. colombiensis, R. pallescens and R. ecuadoriensis . The molecular data supports also a sylvatic distribution of R. prolixus in Brazil (Pará state) and the monophyly of R. robustus . As we detected extensive introgression events and selective pressure on mitochondrial genes, we strongly recommend performing separate mitochondrial and nuclear phylogenies and to take advantages of mito-nuclear conflicts in order to have a comprehensive evolutionary vision of this genus.

The genome size of five Rhodnius species (R. milesi, R. nasutus, R. neivai, R. prolixus, and R. robustus) and two Psammolestes species (P. coroedes and P. tertius) were estimated using flow cytometry and/or k-mer distributions in genome sequences. Phylogenetic generalized linear mixed models highlighted significant genome size variations among species and between sexes, with R. prolixus showing the largest genome. In this study we provide the first data on female genome size in Triatominae. For five species, female genome size did not differ from males, except for R. robustus, where females had smaller genomes. Genome size estimations based on the k-mer distribution method were less than those estimated from flow cytometry, but both methods exhibited the same pattern of sexual differences. Further genomic studies are needed to infer whether genome size variation could be an adaptive trait in Rhodnius.