First record of Cinara todocola (Hemiptera: Aphididae) on endangered Christmas tree in South Korea: morphology, biology, and global invasion potential

Abstract

Cinara (Cinara) todocola (Inouye, 1936) is newly recorded on needle fir (Abies holophylla) and Korean fir (Abies koreana) (Pinaceae) in South Korea. This marks the first recorded instance worldwide of this species damaging the Korean fir, an endangered Christmas tree. We provide detailed life photos and descriptions of all available morphs of this species to facilitate rapid species identification, including redescriptions of the poorly known apterous viviparous female and alate viviparous female, as well as descriptions of the hitherto unknown morphs of the parthenogenetic and sexual generation (fundatrix, oviparous female, and male). This species exhibits strong host specificity across various fir trees, and its life cycle is holocyclic, with the colony size doubling twice per year (in June and September) in South Korea. For the first time, we conducted scanning electron microscopy research to elucidate the gross morphology and sensilla of representatives of this species. Additionally, climate niche modeling was applied to assess and predict the global invasion risk of this species, revealing that C. todocola has the potential to invade in specific regions of East Asia, Europe, and North America.

Introduction

Trees of the genus Abies (Pinaceae), comprising about 50 species, are evergreen conifers distributed across the Northern Hemisphere¹. Fir trees are widely cultivated as Christmas trees, particularly in Europe and the USA. In North Carolina, USA, 50 million Fraser firs are grown on 30,000 hectares, contributing over 100 million dollars annually to the industry^2,3. South Korea’s fir forests predominantly consist of four species: the native needle fir (Abies holophylla), Korean fir (A. koreana), and Khingan fir (Abies nephrolepis), along with the introduced momi fir (Abies firma) from Japan^4,5. According to the IUCN Red List, the conservation status of A. holophylla is “Near Threatened,” that of A. koreana is “Endangered,” while the status of A. nephrolepis and A. firma is of “Least Concern”⁶.

The genus Cinara Curtis, 1835 (Hemiptera: Aphididae: Lachninae) comprises over 250 species globally, all closely associated with coniferous trees in the Cupressaceae and Pinaceae^7,8,9,10. Cinara species can spread intercontinentally through international plant trade driven by global plantations¹¹. As invasive species, they can cause severe economic damage under favorable climatic conditions^12,13. For instance, Cinara cupressi (Buckton, 1881), or the cypress aphid, is listed among the world’s 100 worst invasive species, damaging coniferous plantings (species of Cupressus and other Cupressaceae) worldwide^12,14,15,16. In 1991, this species caused $27.5 million in losses in southern and eastern Africa, with annual growth reductions valued at $9.1 million¹⁷. Cinara (Cinara) todocola (Inouye, 1936) exhibits strong host specificity to Abies species, causing damage to young branches and the trunk^9,19. It has spread from its type locality in Japan to neighboring countries, including the far eastern part of Russia, Sakhalin²⁰ and North Korea²¹.

Based on previously unrecorded collections of Cinara todocola on A. holophylla and A. koreana in South Korea, this study was designed to gain a detailed understanding of the morphology and ecology of this species with the ultimate goal of facilitating the development of rapid identification and early warning systems and consequently improving management strategies for tree plantations of economic interest or for safeguarding trees where they are threatened.

Results

Morphological identification

Specimens were identified as the todo-fir aphid, Cinara (Cinara) todocola (Inouye, 1936), according to the taxonomic key of Blackman & Eastop⁹. Additional morphological comparisons were made using Inouye’s¹⁸ original description and the paratypes (Fig. 1r).

Fig. 1

Cinara todocola in life: (a) fundatrix; (b) apterous viviparous female; (c) alate viviparous female; (d) oviparae; (e) male; (f) colony on the bark of young branches on Abies koreana; (g) colony on the bark of young branches of A. holophylla; (h) ant’s shelter made by soil and wood scraps; (i) colony under ant’s shelter; (j) colony on a crack in the trunk of A. holophylla; (k) colony outside the ant’s shelter; (l) yellowing of the leaves of the host plant; (m) honeydew produced by aphids; (n) yellowish egg on needles of A. holophylla; (o) yellowish egg turns black; (p) aphids infected by parasitoid in June; (q) aphids infected by parasitoid in October; (r) slides of paratypes of C. todocola.

Cinara (Cinara) todocola (Inouye, 1936)

Tuberolachnus todocolus Inouye, 1936: 131.

Fundatrix: description

Figures 1a and 2a; S Fig. 3 and S Table 1.

Fig. 2

Cinara todocola: (a) fundatrix; (b) apterous viviparous female; (c) alate viviparous female; (d) oviparous female; (e) male.

Colour in life: head brown. Thorax and abdomen dark greenish. Eyes dark red. ANT I and II brown. ANT III light brown with 1/5 distal end brown. ANT IV light brown with 1/2 distal end brown. ANT V and ANT VI light brown with 3/4 distal end brown. Femora dusky brown with darker apical part. Tibiae dark brown to blackish. SIPH black. Morphometric characters: body oval. HW 0.57–0.66 × ANT. ANT 0.25–0.32 × BL. ANT III without secondary rhinaria, shorter than ANT IV + V + VI. ANT IV shorter than ANT V without secondary rhinaria (S Fig. 3e). ANT V longer than ANT VI with one rounded primary rhinarium with 1–2 small rounded secondary rhinaria. ANT VI with PT 0.09–0.15 × BASE, with one rounded primary rhinarium and 6 accessory rhinaria. Other antennal ratios: VI/III 0.38–0.48, V/III 0.45–0.57, IV/III 0.34–0.40. LS III 3.88–4.93 × BD III. Rostrum reaches ABD III–IV. URS 0.67–0.84 × ANT III, 1.54–1.97 × ANT VI and 1.06–1.22 × HT II with 8–12 fine and pointed accessory setae (S Fig. 3d). Hind legs covered by long, fine, pointed setae on dorsal side longer than the width of tibiae at midpoint, 0.06–0.17 mm long on femora and 0.05–0.18 mm long on tibiae (S Fig. 3f). HT I basal length 0.83–0.97 × dorsal, 0.32–0.37 × ventral. HT II 0.60–0.71 × ANT III and 1.40–1.72 × ANT VI. Dorsal side of abdomen covered by long, fine, pointed setae, 0.12–0.17 mm long on ABD I–VII. ABD I–VII with few small scleroites. ABD VIII with a small broken band and few small scleroites with 45–53 setae, 0.9–0.16 mm long. SIPH cones large, rounded, 3.87–4.45 × SIPH pore with 78–91 setae (S Fig. 3b). GP transverse oval with 42–55 fine setae (S Fig. 3c). Cauda semi-circular with many long, fine, pointed setae.

Apterous viviparous female: redescription

Figures 1b and 2b; S Fig. 4 and S Table 1.

Colour in life: head brown. Thorax brown with lighter waxy. Mesothorax and metathorax dark brown and light green paired spinal sclerotic plates. Abdomen dark green with lighter waxy. Eyes dark red. ANT I and II brown. ANT III light brown with distal end brown. ANT IV light brown with 1/2 distal end brown. ANT V–VI light brown with 3/4 distal end brown. Femora and Tibiae dark brown to black. Tarsi brown. SIPH black. Morphometric characters: body elongated oval. HW 0.55–0.65 × ANT. ANT 0.33–0.42 × BL. ANT III without secondary rhinaria, shorter than ANT IV + V + VI. ANT IV shorter than ANT V without secondary rhinaria (S Fig. 4e). ANT V about equal to or a little longer than ANT VI with one rounded primary rhinarium with 1–2 small rounded secondary rhinaria. ANT VI with PT 0.25–0.33 × BASE, with one rounded primary rhinarium and 6 accessory rhinaria. Other antennal ratios: VI/III 0.45–0.57, V/III 0.50–0.67, IV/III 0.39–0.48. LS III 4.27–4.81 × BD III. Rostrum reaches ABD V–VI. URS 0.80–1.00 × ANT III, 1.66–1.95 × ANT VI and 1.34–1.57 × HT II with 16–24 fine, pointed accessory setae (S Fig. 4d). Hind legs covered by long, fine, pointed setae on dorsal side longer than the width of tibiae at midpoint, 0.05–0.13 mm long on femora and 0.04–0.15 mm long on tibiae (S Fig. 4f). HT I basal length 0.82–1.09 × dorsal, 0.33–0.44 × ventral. HT II 0.56–0.69 × ANT III and 1.14–1.36 × ANT VI. Dorsal side of abdomen covered by long, fine, pointed setae, 0.08–0.14 mm long on ABD I–VII. ABD I–VII with a few small pigmented scleroites of various sizes. ABD VIII with a small broken band and few small scleroites with 23–29 setae, 0.07–0.12 mm long. SIPH cones large, longer than wide, surrounded by single, big sclerotic plate, 6.01–6.98 × SIPH pore with 153–172 setae (S Fig. 4b). GP oval with 41–53 fine setae (S Fig. 4c). Cauda semi-circular with many long, fine, pointed setae.

Alate viviparous female: redescription

Figures 1c and 2d; S Fig. 5 and S Table 1.

Colour in life: head brown. Pronotum, mesonotum, and metanotum black. Propleuron, mesopleuron, and metapleuron brown. Abdomen dark green. Eyes dark red. ANT I and II brown. ANT III light brown with 4/5 distal end brown. ANT IV–VI light brown with 3/4 distal end brown. Femora dusky brown with darker apical part. Tibiae dark brown to black. Tarsi dark brown to black. SIPH black. Wing membrane of all wings is fuscous (Fig. 1c). Morphometric characters: body elongated oval. HW 0.47–0.54 × ANT. ANT 0.36–0.44 × BL. ANT III with 15–22 secondary rhinaria, longer than Ant IV + V + VI. ANT IV shorter than ANT V with 3–8 secondary rhinaria (S Fig. 5e). ANT V shorter than ANT VI with one rounded primary rhinarium with 1–4 small rounded secondary rhinaria. ANT VI with PT 0.21–0.27 × BASE, with one rounded primary rhinarium and 6 accessory rhinaria. Other antennal ratios: VI/III 0.34–0.42, V/III 0.40–0.51, IV/III 0.35–0.43. LS III 3.84–4.38 × BD III. Rostrum reaches ABD III. URS 0.51–0.63 × ANT III, 1.39–1.66 × ANT VI and 1.02–1.13 × HT II with 14–25 fine, pointed accessory setae (S Fig. 5d). Hind legs covered by long, fine, pointed setae on dorsal side longer than the width of tibiae at midpoint, 0.09–020 mm long on femora and 0.04–0.23 mm long on tibiae (S Fig. 5f). HT I basal length 0.83–1.12 × dorsal, 0.33–0.46 × ventral. HT II 0.49–0.57 × ANT III and 1.61–1.82 × ANT VI. Dorsal side of abdomen covered by long, fine, pointed setae, 0.09–0.15 mm long on ABD I–VII. ABD I–VII with a few small pigmented scleroites of various sizes. ABD VIII with a small broken band and few small scleroites with 22–31 setae, 0.10–0.17 mm long. SIPH cones large, longer than wide, SIPH cones large, surrounded by single, big sclerotic plate, 5.66–6.80 × SIPH pore with 161–177 setae (S Fig. 5b). GP oval with 48–54 fine setae (S Fig. 5c). Cauda semi-circular with many long, fine pointed setae. Fore wings with media branched twice.

oviparous female: description

Figures 1d and 2c; S Fig. 6 and S Table 1.

Colour in life: head brown. Thorax brown with lighter waxy. Mesothorax and metathorax dark brown and light green paired spinal sclerotic plates. Abdomen brown with ABD I–VI light greenish small paired spinal and side of ABD I–VI light greenish stripe. Eyes dark red. ANT I and II brown. ANT III–V light brown with distal end brown. ANT VI light brown with 3/4 distal end brown. Femora dusky brown with darker apical part. Tibiae dark brown. Fore, middle, and hind tarsi brown. SIPH dark brown. Morphometric characters: body elongated oval. HW 0.50–0.56 × ANT. ANT 0.33–0.39 × BL. ANT III without secondary rhinaria, shorter than ANT IV + V + VI. ANT IV shorter than ANT V without secondary rhinaria (S Fig. 6e). ANT V longer than ANT VI with one rounded primary rhinarium with 1–2 small rounded secondary rhinaria. ANT VI with PT 0.20–0.26 × BASE, with one rounded primary rhinarium and 6 accessory rhinaria. Other antennal ratios: VI/III 0.40–0.50, V/III 0.49–0.59, IV/III 0.36–0.46. LS III 4.02–5.02 × BD III. Rostrum reaches ABD V–VI. URS 0.64–0.82 × ANT III, 1.43–1.77× ANT VI and 1.08–1.40 × HT II with 17–22 fine and pointed accessory setae (S Fig. 6d). Hind legs covered by long, fine, pointed setae on dorsal side longer than the width of tibiae at midpoint, 0.08–0.19 mm long on femora and 0.09–0.19 mm long on tibiae, and very small 493–568 pseudosensoria which are single, rounded and 8-shaped (S Fig. 6f). HT I basal length 0.72–1.10 × dorsal, 0.32–0.40 × ventral. HT II 0.53–0.61 × ANT III and 1.16–1.40 × ANT VI. Dorsal side of abdomen covered by long, fine, pointed setae, 0.10–0.16 mm long on ABD I–VII. ABD I–VII with few small scleroites. ABD VIII with a small broken band and few small scleroites with 27–33 setae, 0.11–0.21 mm long. SIPH cones large, rounded, surrounded by small sclerotic plate 3.51–4.37 × SIPH pore with 117–128 setae (S Fig. 6b). GP oval with 121–159 fine setae (S Fig. 6c). Cauda semi-circular with many long, fine, pointed setae.

Male: description

Figures 1e and 2e; S Fig. 7 and S Table 1.

Colour in life: head dark brown. Thorax dark brown with a greenish tinge. Abdomen dark green. Eyes dark red. ANT I and II brown. ANT III light brown with 4/5 distal end brown. ANT IV–VI light brown with 3/4 distal end brown. Femora dusky brown with darker apical part. Tibiae dark brown. Tarsi dark brown. SIPH black. Wing membrane of all wings is fuscous (Fig. 1e). Morphometric characters: body elongated oval. HW 0.39–0.49 × ANT. ANT 0.48–0.55 × BL. ANT III with 40–51 secondary rhinaria, shorter than ANT IV + V + VI. ANT IV shorter than ANT V with 8–14 secondary rhinaria (S Fig. 7e). ANT V longer than ANT VI with one rounded primary rhinarium with 4–8 small rounded secondary rhinaria. ANT VI with PT 0.20–0.26 × BASE, with one rounded primary rhinarium and 6 accessory rhinaria. Other antennal ratios: VI/III 0.34–0.41, V/III 0.35–0.43, IV/III 0.35–0.40. LS III 3.63–4.27 × BD III. Rostrum reaches ABD V. URS 0.43–0.55 × ANT III, 1.16–1.41 × ANT VI and 0.96–1.10 × HT II with 12–16 fine and pointed accessory setae (S Fig. 7d). Hind legs covered by long, fine, pointed setae on dorsal side longer than the width of tibiae at midpoint, 0.08–0.19 mm long on femora and 0.09–0.26 mm long on tibiae (S Fig. 7bf). HT I basal length 0.85–1.08 × dorsal, 0.32–0.41 × ventral. HT II 0.44–0.52 × ANT III and 1.15–1.33 × ANT VI. Dorsal side of abdomen covered by long, fine, pointed setae, 0.10–0.17 mm long on ABD I–VII. ABD I–VII with a few small pigmented scleroites of various sizes. ABD VIII with a small broken band and few small scleroites with 16–21 setae, 0.09–0.15 mm long. SIPH cones large, with proximal part subdivided into many scleroites, 4.87–5.33 × SIPH pore with 38–49 setae (S Fig. 7a). Cauda semicircular with many long, fine, pointed setae. Fore wings with media branched twice. Parameres are present, dark brown pigmented, clearly visible, separate, divided into lobate parts giving rise to a finger-shaped projection toward the base of the phallus, fine setae, 0.01–0.02 mm long on the entire surface. The basal part of the phallus is finger-shaped, dark brown pigmented, clearly visible, separate, with pointed setae, 0.009–0.015 mm long. The proximal part of sclerotized arms is robust, dark pigmented and ends in round-shaped apices with long fine setae, 0.04–0.05 mm long. The distal part is much thinner and forms the upper slightly curve-shaped structure that surrounds the genital area. The aedeagus is not visible (S Fig. 7b).

Notes on SEM morphology and sensilla of Cinara todocola

General morphological characters of the morphs

Different morphological forms of C. todocola, are similar to each other within the general type—apterous (wingless) and alate (winged). The fundatrix, apterous viviparous female and oviparous female are characterized by an enlarged abdomen which in the fundatrx is the widest in contrast to the head (Fig. 3a). Apterous viviparous females are very similar in body shape but their abdomen is smaller (Fig. 3b) while the oviparous female with a rather oval abdomen is the smallest (Fig. 3c). In contrast to the apterous morphs, the alate viviparous females and the alate male are very similar, with an enlarged thorax and narrower abdomen, which in the alate viviparous female is more rounded (Fig. 3d) than in the male (Fig. 3e). Both types of morphs are also characterized by similar morphology of the head and compound eyes (Fig. 3f–j). In the apterous morphs (fundatrix, apterous viviparous female and oviparous female), compound eyes are well-developed, rounded, with many facetes and well-visible triommatidia on an ocular tubercle (Fig. 3k,l), while in the alate viviparous females and alate males they are larger and more protuberant (Fig. 3m,n). The head of the winged morphs are moreover characterized by well-developed ocelli (Fig. 3i,j,m,n). All morphs are characterized by a densely setose body and appendages, including tarsi (Fig. 3o) and siphunculi sclerites, which in the apterous morphs are rounded (Fig. 3p) and in the alate morphs are elongated oval (Fig. 3q). Numerous setae also cover the cauda, which is rounded in all morphs (Fig. 3r).

Fig. 3

Scanning electron micrographs (SEM) of the general view and general characters of Cinara todocola morphs: (a) fundatrix, (b) apterous viviparous female, (c) oviparous female, (d) alate viviparous female, (e) alate male, (f) head and thorax of the fundatrix, (g) head and thorax of the apterous viviparous female, (h) head and thorax of the oviparous female, (i) head and thorax of the alate viviparous female, (j) head and thorax of the alate male, (k) compound eye of the fundatrix, (l) compound eye of the apterous viviparous female, (m) compound eye and ocellus of alate viviparous female, (n) compound eye and ocellus of alate male, (o) hind tarsus of apterous viviparous female, (p) siphunculus of apterous viviparous female, (q) siphunculus of alate viviparous female, (r) cauda of alate male.

Antennal sensilla

The antennae of C. todocola in all examined morphs bear all of the so far known types of sensilla in aphids (campaniform, trichoid, placoid, and coeloconic sensilla). One campaniform sensillum can be found on the dorsal side of the pedicel, where additionally on the dorso-lateral side one small rhinariolum in the form of a sunken coeloconic sensillum is also visible (Fig. 4a,b). Campaniform sensilla are rounded; their collars are wide with a rather small, rounded, protuberant cap with a pore near the cap edge (Fig. 4c). The coeloconic sensillum of the rhinariolum is in the form of a sunken, conical peg with 6–7 short, bulky projections (Fig. 4d). Type I trichoid sensilla are the most numerous and present on all antennal segments. There are visible as very long, fine, pointed setae in a general view; these arise from their flexible sockets at an angle of about 45°–50° (Fig. 4e,f). The sockets of the sensilla are characteristically indented in the middle (Fig. 4g). Deeper examination of the type I trichoid sensilla revealed their ribbed surface (but on the very basal part they are smooth) (Fig. 4c) as well as very fine, rounded apical ends (Fig. 4h,i). Besides type I trichoid sensilla, type II trichoid sensilla can be found but only on the apical part of the last antennal segment of the examined morphs and they are called apical and subapical setae (Fig. 4j). The type II trichoid sensilla are much shorter but also seems to be more rigid and they arise from rounded (flat or protuberant) sockets (Fig. 4k). They are also ribbed but in contrast to the type I trichoid sensilla, the ribbing starts at the very base of the sensilla (Fig. 4l,m). In the general view, the pointed apices of the type II trichoid sensilla are characterized by jagged, irregular ends (Fig. 4n).

Fig. 4

SEM of antennal sensilla of Cinara todocola I: (a) pedicel of alate viviparous female with type I trichoid sensilla (blue), rhinariolum (pink) and campaniform sensillum (green), (b) pedicel of alate male with type I trichoid sensilla (blue), rhinariolum (pink) and campaniform sensillum (green), (c) ultrastructure of male pedicel campaniform sensillum, (d) ultrastructure of the male pedicel rhinariolum, (e) part of ANT III of apterous viviparous female with type I trichoid sensilla, (f) part of ANT IV of the fundatrix with type I trichoid sensilla, (g) ultrastructure of the sensillum flexible socket of apterous viviparous female, (h) ultrastructure of trichoid sensillum surface of apterous viviparous female, (i) ultrastructure of trichoid sensillum apex of apterous viviparous female, (j) apical end of terminal process with type II trichoid sensilla, (k) structure of type II trichoid sensillum on the terminal process, (l) ultrastructure of type II trichoid sensillum surface, (m) ultrastructure of type II trichoid sensillum sockets, (n) ultrastructure of type II trichoid sensillum apex.

Besides the omnipresent type I trichoid sensilla, some of the flagellar segments also bear multiporous placoid sensilla (large and small). In the apterous morphs only antennal segments V and VI bear this kind of sensilla (rhinaria). In the fundatrix, apterous viviparous and oviparous females antennal segment V always bears one, large multiporous sensillum (primary rhinarium) at the apical end of the segment and one-two small placoid sensilla placed in one row in the distal half of the segment (secondary rhinaria) (Fig. 5a,b). The alate morphs (alate viviparous females and alate males) antennae are characterized first of all by numerous small placoid sensilla (secondary rhinaria), which are also present on antennal segments III and IV (Fig. 5c,d). The small, multiporous sensilla of the apterous morphs (e.g., fundatrix and apterous viviparous female) are rounded and flat, with clearly visible multiporous membrane (Fig. 5e,f). The large multiporous placoid sensilla of these morphs have similar characters but are additionally surrounded by a low, smooth cuticular collar which is lower than the slightly protuberant sensillum (Fig. 5g,h). The multiporous membrane of these sensilla is mostly smooth and rather densely covered with minute pores, which in small placoid sensilla are more rounded (Fig. 5i,j) and in large sensilla are more oval or slightly elongated (Fig. 5k,l). The characters of placoid sensilla on antennal segments III–V are also very similar in the remaining morphs such as the oviparous female (Fig. 5m), alate viviparous female (Fig. 5n) and alate male in which the small placoid sensilla are smaller and more protuberant (sometimes semispherical) (Fig. 5o,p). The last antennal segment of all morphs bears all types of antennal sensilla besides the campaniform ones (Fig. 5q). The base of the segment is characterized by the presence of a group of sensilla in its distal end, and contains one large placoid sensillum (major rhinarium) and accessory rhinaria which are of three morphological kinds: two small multiporous placoid sensilla, and four sunken coeloconic sensilla of two types (Fig. 5q). The group of sensilla known as accessory sensilla lie more or less on the side and under the large, multiporous placoid sensillum with the two small multiporous placoid sensilla in a polar position and the four sunken coeloconic sensilla between them (Fig. 5r,s). The coeloconic sensilla are arranged more or less in a square position, with the type I sensilla located on the inner side near the major rhinarium and while the type II sensilla are located on the outside (Fig. 5r,s). The placoid and coeloconic sensilla lie in shallow cavities and are surrounded by sclerotic collars. The space between the collar and the sensilla is additionally covered by transverse strengthenings structures (Fig. 5s). Small multiporous sensilla on the last antennal segment are mushroom-shaped with a narrower base and their pores are poorly visible (Fig. 5t,u). There are two types of sunken coeloconic sensilla—type II with 11–13 long projections (Fig. 5v) which is additionally exposed and type I with 8–9 short projections, lying deeper inside the cuticle (Fig. 5w).

Fig. 5

SEM of antennal sensilla of Cinara todocola II: (a) ANT V of the fundatrix with type I trichoid sensilla (blue), small multiporous placoid sensilla (orange) and large multiporous placoid sensillum (yellow), (b) ANT V of the apterous viviparous female with type I trichoid sensilla (blue), small multiporous placoid sensillum (red) and large multiporous placoid sensillum (yellow), (c) part of ANT III of alate viviparous female with type I trichoid sensilla (blue) and small multiporous placoid sensilla (red), (d) ultrastructure of Ant V of the male with type I trichoid sensilla (blue), small multiporous placoid sensilla (red) and large multiporous placoid sensillum (yellow), (e) structure of the small multiporous placoid sensillum of the fundatrix ANT V, (f) structure of the small multiporous placoid sensillum of the apterous viviparous female ANT V, (g) structure of large multiporous placoid sensillum on ANT V of the fundatrix, (h) ultrastructure of large placoid sensillum of apterous viviparous female, (i) ultrastructure of porous membrane of the small placoid sensillum of the fundatrix, (j) ultrastructure of porous membrane of the small placoid sensillum of the apterous viviparous female, (k) ultrastructure of porous membrane of the large placoid sensillum of the fundatrix, (l) ultrastructure of porous membrane of the large placoid sensillum of the apterous viviparous female, (m) structure of small multiporous placoid and large multiporous placoid sensilla of ANT V of oviparous female, (n) structure of small multiporous placoid and large multiporous placoid sensilla of ANT V of oviparous female, (o,p) ultrastructure of small multiporous placoid sensilla on ANT III of the male, (q) ANT VI of apterous viviparous female with type I trichoid sensilla on the base (blue), type II trichoid sensilla on the terminal process (violet), large multiporous placoid sensillum (yellow), small multiporous placoid sensilla (green) and sunken coeloconic sensilla of two kinds, (r) structure of ANT VI sensilla of the male, (s) structure of ANT VI sensilla of the fundatrix, (t) structure of small multiporous placoid sensillum on ANT VI of apterous viviparous female, (u) ultrastructure of small multiporous placoid sensillum on ANT VI of apterous viviparous female, (v) ultrastructure of type II sunken coeloconic sensillum, (w) ultrastructure of type I sunken coeloconic sensillum.

Mouthpart sensilla

The labium of C. todocola morphs is mostly covered with type I trichoid sensilla and characterized by long, narrow ultimate rostral segments (URS or IV + V) (Fig. 6a). Sensilla cover the whole length and each side of segment IV (Fig. 6b–d). On the basal part of the segment there is one pair of type II basiconic sensilla (Fig. 6e). The type II basiconic sensilla arise from rounded or slightly indented flexible sockets and, similarly to the trichoid sensilla on the antennae they are smooth at the very basal part and ribbed over the rest of their length (Fig. 6f,g). The border of rostral segment IV and V bears three pairs of long, fine pointed trichoid sensilla (primary setae), one pair on the ventral side, one pair at the lateral side and one pair on the dorso-lateral side (Fig. 6h–j). The last rostral segment bears seven pairs of long, smooth type III basiconic sensilla with slightly rounded apices. The sensilla are located on each side of the segment apical end: three pairs on the ventral side, two on the lateral side and two on the dorsal side (Fig. 6k–m).

Fig. 6

SEM of mouthpart sensilla of Cinara todocola: (a) ventral side of labial segments III-V of the fundatrix with many trichoid sensilla, (b) ventral side of labial segments IV and V of apterous viviparous female, (c) lateral side of labial segments IV and V, (d) dorsal side of labial segments IV and V of apterous viviparous female, (e) type II basiconic sensilla on the border between labial segments III and IV of apterous viviparous female, (f) structure of the type II basiconic sensillum, (g) socket of trichoid sensillum on labial segment IV, (h) ventral side of labial segment V of apterous viviparous female with trichoid sensilla on the border and type III basiconic sensilla on the apical end, (i) lateral side of labial segment V of male with trichoid sensilla on the border and type III basiconic sensilla on the apical end, (j) dorsal side of labial segment V of apterous viviparous female with trichoid sensilla on the border and type III basiconic sensilla on the apical end, (k) ventral side of apical end of labial segment V with type III basiconic sensilla, (l) lateral side of apical end of labial segment V with type III basiconic sensilla, (m) dorsal side of apical end of labial segment V with type III basiconic sensilla.

Wing and leg characters and sensilla

Wings of alate morphs of C. todocola are membranous with standard venation. On the basal part of the fore wing of alate viviparous females and males campaniform sensilla can be found. The campaniform sensilla are separated into two groups; one group is placed very close to the wing articulation and the second one at the beginning of the Sc + R + M + Cu (Fig. 7a,b). Those campaniform sensilla are of a different kind to the sensillum on the antennal pedicel. They are rounded and more protuberant, and their collar is elevated and very often has an indentation and strongly protuberant cap which can be rounded or slightly pointed with a central pore (Fig. 7c,d). The wing membrane over the whole area is generally smooth with some shallow cavities and triangular scale-like elements (Fig. 7e) which are very numerous in the area of membrane edges and pterostigma. The border between the pterostigma and the rest of the wing membrane bears many rather long, fine, pointed trichoid sensilla (Fig. 7f) which are tubular, smooth and arise from protuberant and bulbous flexible sockets (Fig. 7g). The hind wings have very similar characters, with a well-developed claval area. The part of the claval apparatus on the hind wing is composed of many wide or crescent-shaped scale-like elements which become denser in the immediate vicinity of seven tightly adhering and strongly curved hamuli (Fig. 7h). Campaniform sensilla have also been found on the inner side of the trochanters together with the trichoid sensilla, which are very numerous on the legs (Fig. 7i). The ventral side of the trochanter, besides fine and smooth trichoid sensilla, additionally bear one thicker, rigid and pointed sensillum (‘trochanter seta’) whose surface is additionally ribbed (Fig. 7j). The campaniform sensilla present on the trochanters are more similar to those on the antennal pedicel; they are slightly elevated, with a trapezoid collar and slightly protuberant, rounded cap with a small pore, located near the cap edge (Fig. 7k). All C. todocola morphs are characterized by numerous, very long, fine, pointed trichoid sensilla along the femora (Fig. 7l,m) and tibiae (Fig. 7n). The surface of the sensilla is smooth, including those on the ventral side of the first tarsal segment, on which the sensilla are curved near the apical ends. Additionally, there is one shorter, rigid trichoid sensillum (‘sense-peg’) (Fig. 7o). Trichoid sensilla are also located on the second segment of the tarsi, and at their ends, short, fine, pointed parempodia are visible (Fig. 7p).

Fig. 7

SEM of wing and leg sensilla of Cinara todocola: (a) basal part of the wing of alate viviparous female with two groups of campaniform sensilla (green), (b–d) structure and ultrastructure of the campaniform sensilla, (e) fragment of the wing membrane of alate viviparous female with scale-like elements, (f) area of pterostigma of the male wing with many trichoid sensilla (blue), (g) ultrastructure of the trichoid sensillum flexible socket, (h) structure of the claval apparatus with hamuli, (i) trochanter and proximal part of femur with trichoid sensilla (blue), and campaniform sensilla (green), (j) structure of the trichoid sensillum (trochanter setae) on trochanter, (k) ultrastructure of campaniform sensillum on trochanter, (l) trichoid sensilla on femur of fundatrix, (m) trichoid sensilla on femur of apterous viviparous female, (n) trichoid sensilla on tibia of apterous viviparous female, (o) ventral side of first segment of hind tarsus of apterous viviparous female with many trichoid sensilla and one sense peg (blue), (p) distal part of second segment of hind tarsus of apterous viviparous female with two claws and one visible parempodium (empodial seta), orange.

Body surface and dorsal trichoid sensilla

The body cuticle surface and trichoid sensilla of C. todocola are similar between the examined morphs. The head of the apterous viviparous female is characterized by a generally smooth cuticle and covered with numerous long, fine and pointed trichoid sensilla (Fig. 8a). The sensilla arise from flexible sockets which are more elevated and narrower than those on the antennae and legs and are smooth on the very base (Fig. 8b), then ribbed longitudinally over the entire surface (Fig. 8c) and again smooth near the apical end, with a very slightly narrow capitate apex (Fig. 8d). This character of the sensilla apical end was visible after examination in higher magnification, so in a general view the sensilla apices seems to be pointed. We noticed that some of the head trichoid sensilla arise from much lower but also flexible sockets (Fig. 8e). As mentioned above, the head cuticle surface in a general view is smooth, but examination in higher magnifications revealed that a large part of the cuticle surface is covered with minute oval or slightly rounded, flat protuberances, which are distributed rather regularly (Fig. 8f,g). The characters of the abdominal surface of apterous viviparous females C. todocola are very similar to those described on the head, with a generally smooth cuticle which is densely covered with long, fine, pointed (in general view) trichoid sensilla (Fig. 8h). In contrast to the head cuticle, some sensilla on the abdomen arise from oval and irregular scleroites, which are visible as shallow spots on the cuticle (Fig. 8i). Some parts of the apterous viviparous female abdominal cuticle surface were additionally covered with numerous extremely small structures (Fig. 8j), which in higher magnification turned out to very interesting. The abdominal cuticle structures are very small, about one micrometer in diameter, rounded or oval with a central cavity which closely resembles human erythrocytes (Fig. 8k,l). Taking into accounts their structure, and the fact that on the abdominal cuticle no other known openings or pores have been found and that the apterous viviparous females produce wax powder (see Fig. 1b), these structures are most likely wax glands. Our theory that the rounded structures on the abdominal cuticle of the apterae must be wax glands is further supported by the examination of the abdominal cuticle of the fundatrix, which in life does not produce wax (see Fig. 1a). The abdominal surface of the fundatrix is also smooth in general view, with long, pointed (in this view) trichoid sensilla that seem to be slightly stiffer (Fig. 8m). Nowhere on the abdominal cuticle surface of the fundatrix, could we find a wax gland like in the aptera. In the fundatrix the cuticle on almost the whole area of the abdomen is, as mentioned above, smooth in a general view, but closer examination revealed that the fine structure is covered with numerous shallow depressions, making it appear porous (Fig. 8n). The cuticle changes at the end of the abdomen (ABD VII and ABD VIII); here its sculpture is much rougher, in the form of papilla-like outgrowths and denticles (Fig. 8o,p). In the alate viviparous female and alate male, the abdominal surface is heavily wrinkled and covered with much longer trichoid sensilla than in the apterous morphs. Similarly to other morphs, the sensilla of alate viviparous females are tubular, fine, pointed in general view and ribbed over almost their whole length (Fig. 8q,r). On the other hand, the abdominal sensilla of the male arise from rounded scleroites, and the cuticle, besides being wrinkled, is also characterized by numerous outgrowths and denticles (Fig. 8s,t). Trichoid sensilla of the male arise from narrow and elevated flexible sockets, and, in contrast to those of alate viviparous females their surface is smooth (Fig. 8u,v). The abdominal surface of the oviparous female is also characteristic, and, in contrast to the rest of the examined morphs, the generally rough cuticle surface is in fact densely covered with numerous outgrowths that are mostly longitudinal, semicircular or oval in form (Fig. 8w,x).

Fig. 8

SEM of cuticle and body surface sensilla of Cinara todocola: (a) head cuticle of apterous viviparous female with trichoid sensilla, (b) ultrastructure of trichoid sensilla flexible socket, (c) ultrastructure of the trichoid sensillum surface, (d) ultrastructure of the trichoid sensillum apical end, (e) ultrastructure of trichoid sensillum lower socket, (f,g) ultrastructure of the head cuticle surface, (h) abdominal cuticle and trichoid sensilla of apterous viviparous female, (i) scleroite at setal base on the abdomen of apterous viviparous female, (j) abdominal cuticle with wax glands, (k,l) ultrastructure of the apterous viviparous wax glands, (m) generally smooth abdomen of the fundatrix with trichoid sensilla, (n) ultrastructure of the cuticle of fundatrix abdomen, (o) distal part of fundatrix abdomen, (p) ultrastructure of cuticle of distal part of fundatrix abdomen, (q) abdomen of alate viviparous females with trichoid sensilla and wrinkled cuticle, (r) ultrastructure of alate viviparous female abdomen trichoid sensillum, (s) abdomen of alate viviparous females with trichoid sensilla arising from scleroites and wrinkled cuticle, (t) ultrastructure of the male cuticle and scleroite, (u) ultrastructure of the flexible socket of the male abdomen trichoid sensillum, (v) ultrastructure of the male abdomen trichoid sensillum, (w) generally smooth abdomen of the oviparous female with trichoid sensilla, (x) ultrastructure of the oviparous female abdominal cuticle sculpture.

Special characters of sexual morphs

Besides the morphological characters examined and described above, the sexual morphs of C. todocola are characterized by special characters—genitalia of the alate male and scent plaques of the oviparous female. The genitalia of the alate male are located on the ventral side of the end of the abdomen and in general correspond to the scheme described so far for Cinara and other Lachninae (Fig. 9a). Like other perianal structures (cauda and anal plate), the genitalia are densely covered with numerous trichoid sensilla (Fig. 9b). The whole area of the genital capsule is strongly sclerotized, especially the basal part of the phallus and the parameres (Fig. 9c). Both structures, the parameres and the basal part of the phallus, are characterized by short, fine, pointed trichoid sensilla which arise from elevated and rounded flexible sockets (Fig. 9d,e). The aedeagus (when extended) is membranous. The oviparous form is characterized by slightly swollen hind tibiae which bear a large number of small, mostly rounded, oval or slightly irregular scent plaques (Fig. 9f). Scent plaques of the oviparae are elevated; they occur singly (Fig. 9g), very close to each other (Fig. 9h), or doubly, probably created from two single ones (Fig. 9i). Regardless of the size and shape, the surface of the scent plaques cuticle is covered with numerous, mostly rounded, oval or elongated pores of different sizes (Fig. 9j).

Fig. 9

SEM of special characters of sexual morphs of Cinara todocola: (a) lateral side of male perianal structures and genitalia showing cauda (green), anal plate (orange), genital capsule sclerotized edge (blue), parameres (red), basal part of the phallus (yellow) and aedeagus (turquoise), (b) posterior-ventral side of the male genitalia, (c) lateral side of the male genitalia showing parameres and the basal part of the phallus, (d) basal part of the phallus with trichoid sensilla, (e) parameres with trichoid sensilla, (f) fragment of oviparous female hind tibia with trichoid sensilla (blue) and numerous scent plaques (turquoise), (g–i) different shapes of the scent plaques, (j) ultrastructure of the scent plaque porous surface.

Biology

The life cycle of C. todocola in South Korea (Cheongju, Suwon, Pocheon, and Pyeongchan) was observed through direct studies and mounted material. Fundatrices were first seen as adults on April 29 in Cheongju, producing apterous and alate nymphs. Colonies primarily form on young branches, though some occur independently (Fig. 1f,g). In early May, Lasius japonicus (Formicidae) workers approached the colonies, and by end-May, some colonies were inside ant-made shelters in Pocheon. Adult apterous and alate viviparous females appeared by mid-May in all sites. Most colonies live inside L. japonicus shelters during summer in Pocheon (Fig. 1h,i), while others feed on fir tree trunks, protected by Lasius fuliginosus in Pyeongchan (Fig. 1j). The sexual generation appears in early October, with mating from mid-October and oviposition beginning in late October in Pocheon and Pyeongchan. Males and oviparae are sometimes found in ant shelters (Fig. 1m) but usually form colonies at branch tips, where they are approached by Pristomyrmex pungens and Formica japonica (Fig. 1k). Mated oviparae lay 1–3 yellowish eggs on needle undersides (Fig. 1n), which turn black over time (Fig. 1o). Males are not observed after October (S Fig. 2).

Damage

The colony size peaks in mid-June and declines by mid-July. In September, colonies grow again in preparation for winter. A key symptom of aphid infestation is needle yellowing, observed when large colonies form (Fig. 1l). Honeydew from aphids causes black sooty mold on branches and needles (Fig. 1m). No predatory species were found, but aphid mummies infected by parasitoids were discovered on needles in June (Fig. 1p) and October (Fig. 1q). The parasitoids could not be collected as they had already emerged. In Japan, colonies peak in early August and decrease by late September. Aphid infestations reduce the growth of young trees, causing physiological deficiencies. In some cases, weakened trees may die after two to three years of infestation. Protective shelters built by Lasius niger around the trunk enhance aphid survival¹⁹. In contrast, Pauesia momicola (Hymenoptera: Braconidae) effectively controls aphid populations through biological control^22,23.

Climatic niche modeling

All model algorithms exhibited good performance (i.e., AUC > 0.8 and TSS > 0.7) in discriminating between presence and pseudo-absence points (S Fig. 8). Consequently, all base models were integrated to generate an ensemble prediction. Two precipitation-related variables, Bio18 and Bio14, and a temperature-related variable, Bio3, showed relatively high contributions to the final ensemble model (S Fig. 9). The projected final model suggested that the climate of East Asia, including Korea, Japan, and Southeast China, is suitable for C. todocola. Although this species has not been recorded outside of Asia, parts of Europe and Northeast America may also have a climatically suitable environment for C. todocola (Fig. 10).

Fig. 10

Projected climatic suitability for Cinara todocola in geographic space. Higher scores represent higher climatic suitability.

Discussion

Morphology and biology

Inouye^18,19,24 recorded C. todocola on Abies sachalinensis, A. firma, and Abies homolepis in Japan, and on A. sachalinensis in the far eastern part of Russia (Sakhalin). Szelegiewicz²¹ collected C. todocola on A. holophylla in North Korea. Based on their records, C. todocola is inferred to have a holocyclic life cycle, feeding on various Abies species (oligophagous) without migration. Pashchenko²⁵ noted that C. todocola resembles Cinara smaragdina from Siberia but differs in feeding sites and morphology.

Cinara todocola feeds on branches and trunk, ANT V shorter than the diameter of the cone of SIPH, and thorax and abdomen with large dark sclerites (sometimes divided into small sclerites on all abdominal tergites). In contrast, C. smaragdina feeds on shoots between needle bases, ANT V longer than the diameter of the cone of SIPH, and the thorax and abdomen without dark sclerites. According to Wieczorek et al.²⁶, the male genitalia are hook-shaped and lack setae, based on specimens collected from Pinus sp.; however, Korean and Japanese male specimens show a finger-shaped phallus with setae, collected from Abies spp., suggesting the need for re-examination of Wieczorek et al.’s²⁶ specimens for accurate identification. An ecological study of C. todocola in South Korea revealed its impact on host plants and life cycle. This aphid species threatens endangered plant species and poses significant harm to fir forests in South Korea. Therefore, Korean forestry agencies must implement sustainable management strategies based on its life cycle and natural enemies.

Climatic niche modeling

On a global scale, the climatic niche modeling conducted for C. todocola presents insightful projections about its potential distribution, particularly in regions of East Asia, Europe, and North America. The model, based on robust algorithms with high predictive accuracy (AUC > 0.8, TSS > 0.7), indicates that climate variables such as precipitation (Bio18, Bio14) and temperature (Bio3) play significant roles in determining the species’ suitable habitat. These results suggest that areas with similar climatic conditions to those in Korea, Japan, and Southeast China may be prone to C. todocola establishment. While the model offers valuable predictions, it is important to consider the inherent limitations of ecological niche modeling, particularly when extending predictions outside the species’ current range. The model assumes equilibrium between the species and its environment, but such assumptions may not hold when the species is subject to dispersal constraints, such as geographical or ecological barriers²⁷. For example, areas in Europe and North America, though climatically suitable, may present challenges related to geographical isolation, lack of suitable host plants, or competitive interactions with native species. Additionally, the presence of established biological control agents, such as Pauesia momicola in Japan, may further limit the spread of C. todocola in certain regions, complicating the accuracy of global predictions. These uncertainties highlight the need for ongoing monitoring and adaptive management strategies, especially in newly suitable regions. Further research into C. todocola’s ecological interactions, host plant preferences, and dispersal mechanisms is needed to refine predictions and assess its invasive potential. Proactive measures may be necessary to prevent damage to commercial forestry industries, such as Christmas tree plantations. In conclusion, while the climatic niche model suggests the potential spread of C. todocola, its invasion dynamics must be considered in a broader ecological context, including both climate and biotic factors. Kanturski et al.²⁸ reported that closely related species in the aphid genus Eulachnus are mainly distributed in Eurasian regions coinciding with the natural range of pine trees. Their study suggests that climatic conditions may constrain the distribution of Eulachnus. This understanding will guide management strategies and help mitigate risks associated with C. todocola’s spread.

Materials and methods

Collection and identification

In May 2018, an unidentified Cinara species was found on Abies koreana at the Korea National Arboretum in Gyeonggi Province. Subsequent occurrences on A. holophylla were reported in four regions of South Korea (S Fig. 1). Photographs were taken with a Canon 70D camera equipped with extension rings. The aphid samples were preserved in 80% ethanol and slide glass specimens were mounted in Canada balsam, following Blackman and Eastop’s⁹ method. Measurements and digital images were taken using a Leica DMC 5400 (Leica Z16 APO) and a Leica DM 4000B camera (Active Measure version 3.0.3; Mitani Co. Ltd., Japan). Abbreviations used for descriptions are as follows: ANT, antennae; ANT I, ANT II, ANT III, ANT IV, ANT V, BASE, and PT, antennal segments I, II, III, IV, V, base of VI, and processus terminalis of antennomere VI respectively; LS ANT III, length of longest setae of ANT III; BD III, basal articular diameter of ANT III; BL, body length; HW, greatest head width across compound eyes; MaxW, greatest body width; GP, genital plate; HT I, first segment of hind tarsus; HT Ib, basal length of HT I; HT Id, dorsal length of HT I; HT Iv, ventral length of HT I; HT II, second segment of hind tarsus; SIPH, siphunculi; URS, ultimate rostral segment (segment IV + V); ABD I–VIII, Abdominal tergites I–VIII, respectively; FEMORA III, hind femora; TIBIAE III, hind tibiae. Slide specimens are deposited at the College for Agriculture and Life Sciences, Seoul National University Seoul, Korea (SNU) and in the SEM laboratory of the Institute of Biology, Biotechnology, and Environmental Protection, University of Silesia in Katowice (Katowice, Poland).

Material examined

Fundatrix 6, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. koreana, 08.v.2018, leg. M. Lee, [180508-LMH-9] (SNU); 2, South Korea, Gangwon-do, Pyeongchang-gun, Daegwallyeong-myeon, Hoenggye-ri, on A. holophylla, 10.v.2019, leg. M. Lee, [190510-LMH-1] (SNU); 6, South Korea, Chungcheongbuk-do, Cheongju-si, Sangdang-gu, Miwon-myeon, on A. holophylla, 29.iv.2021, leg. M. Lee, [210429-LMH-4] (SNU); 1, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 03.v.2021, leg. M. Lee, [210503-LMH-3] (SNU).

Apterous viviparous female 2, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. koreana, 28.v.2018, leg. M. Lee, [180528-LMH-1] (SNU); 1, South Korea, Gangwon-do, Pyeongchang-gun, Daegwallyeong-myeon, Hoenggye-ri, on A. holophylla, 30.v.2019, leg. M. Lee, [190530-LMH-1] (SNU); 1, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 25.v.2020, leg. M. Lee, [200525-LMH-2] (SNU); 1, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 25.v.2020, leg. M. Lee, [200525-LMH-15] (SNU); 1, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 25.v.2020, leg. M. Lee, [200525-LMH-16] (SNU); 5, South Korea, Gyeonggi-do, Suwon-si, Gwonseon-gu, on A. holophylla, 15.viii.2020, leg. M. Lee, [200815-LMH-1] (SNU); 2, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 17.ix.2020, leg. M. Lee, [200917-LMH-7] (SNU); 1, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 14.vi.2021, leg. M. Lee, [210614-LMH-10] (SNU); 1, South Korea, Gangwon-do, Pyeongchang-gun, Jinbu-myeon, on A. holophylla, 18.vii.2021, leg. M. Lee, [210718-LMH-3] (SNU); 1, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 23.v.2022, leg. M. Lee, [220523-LMH-4] (SNU).

Alate viviparous female 1, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. koreana, 28.v.2018, leg. M. Lee, [180528-LMH-1] (SNU); 4, South Korea, Gangwon-do, Pyeongchang-gun, Daegwallyeong-myeon, Hoenggye-ri, on A. holophylla, 30.v.2019, leg. M. Lee, [190530-LMH-1] (SNU); 2, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 23.v.2022, leg. M. Lee, [220523-LMH-4] (SNU).

Oviparous female 2, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. koreana, 13.xi.2018, leg. M. Lee, [181113-LMH-7] (SNU); 8, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 22.x.2021, leg. M. Lee, [211022-LMH-14] (SNU); 3, South Korea, Gangwon-do, Pyeongchang-gun, Jinbu-myeon, on A. holophylla, 31.x.2021, leg. M. Lee, [211031-LMH-1] (SNU).

Male 9, South Korea, Gyeonggi-do, Pocheon-si, Soheul-eup, on A. holophylla, 22.x.2021, leg. M. Lee, [211022-LMH-14] (SNU).

Scanning electron microscopy

Samples used in this study were collected directly in the field and kept in 80% ethanol for several days. Dehydration of the ethanol-preserved samples was performed using an ethanol series of 80, 90, 96% and two changes of absolute ethanol for 10 min each. From absolute alcohol the samples were transferred to pure chloroform and stored at room temperature for 24 h. Dehydrated and cleaned specimens were dried using the Leica EM CPD 300 auto critical point dryer (Leica Microsystems, Vienna, Austria). Dry samples were mounted on aluminum stubs with double-sided adhesive carbon tape and sputter-coated with 30 nm layer of gold in a Quorum 150 T ES Plus sputter coater (Quorum Technologies Ltd, Laughton, East Sussex, UK). The specimens were imaged by the Hitachi SU8010 field emission scanning electron microscope FESEM (Hitachi High-Technologies Corporation, Tokyo, Japan) at 10 kV accelerating voltage with a secondary electron detector (ESD) in the SEM laboratory of the Institute of Biology, Biotechnology, and Environmental Protection, University of Silesia in Katowice (Katowice, Poland).

Climatic niche modeling

To estimate the climatic niche envelope of C. todocola, we utilized 19 bioclimatic variables with a 10-min spatial resolution provided by WorldClim²⁹. We first assessed the multicollinearity of the bioclimatic variables using correlation analysis. Correlated variables exhibiting a Pearson’s r > 0.85 were filtered out, with one variable retained from each pair. Consequently, nine bioclimatic variables were selected to estimate the climatic suitability for C. todocola: mean diurnal range (Bio2), isothermality (Bio3), mean temperature of wettest quarter (Bio8), mean temperature of driest quarter (Bio9), precipitation of wettest month (Bio13), precipitation of driest month (Bio14), precipitation seasonality (Bio15), precipitation of warmest quarter (Bio18), and precipitation of coldest quarter (Bio19).

Occurrence points for C. todocola were compiled and converted to geocoordinates from various sources (S Table 2). These occurrence points were primarily distributed in Japan, Korea, and Sakhalin Island of Russia, with a total of 29 presence points used to develop a model for C. todocola. We randomly extracted 100 points from the geographic space, following the guidance on the selection of pseudo-absences for accurate and reliable ecological niche modeling suggested by Barbet-Massin et al.³⁰. We created a convex hull using the 29 presence points for C. todocola. Pseudo-absences were then sampled outside of a 2-decimal-degree buffer around the convex hull. We conducted the selection of pseudo-absences ten times (resulting in ten datasets), and each dataset was used to develop a climatic niche model. All pre-processing of occurrence data was conducted using the sp package in R 4.2.2³¹.

An ensemble approach, aggregating the results of multiple modeling algorithms, was employed to develop a climatic niche model for C. todocola. Two statistical regression models, flexible discriminant analysis (FDA) and Multivariate adaptive regression splines (MARS), as well as two machine learning models, gradient boosting machine (GBM) and random forest (RF), were applied as base models for ensemble predictions using the biomod2 package in R 4.2.2^32,33. The occurrence datasets, which were georeferenced with bioclimatic variables, were randomly split into training (70%) and testing (30%) sets and applied to the four modeling algorithms. Thus, we obtained a total of 40 models (ten replicated occurrence datasets × four algorithms). All models were run with default hyperparameter settings provided by biomod2.

Each individual model was evaluated by applying two evaluation metrics—AUC (area under the receiver operating characteristic [ROC] Curve) and TSS (true skill statistics)—to the test data³⁴. The base models with reasonable performance (AUC > 0.8 and TSS > 0.7) were retained and summarized as the mean probability of presence, ranging from 0 to 1000³⁵. Relative importance of bioclimatic variables in the final ensemble model was evaluated using ten permutation procedures of each variable in the final model.