Abstract.

We describe a new species of Cuphodes Meyrick, 1897 (Lepidoptera: Gracillariidae) from South Africa, C. spermotrophus sp. nov., reared from fruits of Rhamnus prinoides L’Hér. (Rhamnaceae). This represents the first confirmed record of a seed-feeding Gracillariidae species from the African continent. The new species is distinguished from all known congeners by external morphology and male and female genitalia. Molecular data (COI barcode) confirm its distinctiveness and support its placement within Cuphodes. We provide diagnostic characters, illustrations of the adult and genitalia, and notes on its biology and host association. The discovery of this species expands the known feeding habits of Cuphodes, previously thought to include only leaf-mining species, and highlights the still largely unexplored trophic diversity of African Gracillariidae.

Introduction

Despite a long tradition of lepidopterological research in southern Africa, the gracillariid fauna of South Africa remains surprisingly underexplored. Many species are still undescribed, and for a large proportion of named taxa, even basic biological information, such as host-plant associations and feeding modes, is lacking (Triberti et al. 2024). This limited knowledge contrasts with the well-recognized ecological and economic importance of the family, which includes numerous species of horticultural concern as well as globally invasive pests (Lopez-Vaamonde et al. 2010).

Currently, 168 species of Gracillariidae are recorded from South Africa, representing nearly all recognized subfamilies except Marmarinae (Vári 1961; De Prins and De Prins 2011–2025; Triberti et al. 2024). Yet more than one-third of these species remain biologically unknown. Even among species with known life histories, our understanding is fragmentary. Host associations span at least 32 plant families, but most gracillariids are highly specialized, typically restricted to a single host family. Only a handful of species have been shown to utilize more than one plant family, underscoring the strong host specificity that characterizes the group.

Gracillariidae are best known as leaf miners, and this life strategy dominates the family. Larvae usually complete development within leaf mines, though some species exit the mine in later instars to make leaf rolls or leaf stacks, or in the final instar to pupate externally. Beyond leaf mining, however, the family exhibits a remarkable range of feeding strategies, including mining of stems and other tissues and even gall induction (Guiguet et al. 2019, 2026). Stem-mining species of Spulerina Vári, 1961, for example, develop beneath the epidermis of young trunks or twigs and pupate within the mine (Kumata 1964). In South Africa, several species of this genus are known, yet their biology remains only partially documented.

More exceptional still are gracillariids that exploit reproductive plant structures. A recent rearing study of a community of frugivorous Lepidoptera in the lowland rainforests of Papua New Guinea recorded six gracillariid species attacking the fruits of plants belonging to 12 different families (Sam et al. 2017). Only one of these species was previously known, Conopomorpha cramerella (Snellen, 1904), the cocoa pod borer, a significant pest in Southeast Asia and Oceania.

Seed- and inflorescence-feeding habits are rare in the family and largely confined to a few genera. Among these, Conopomorpha Meyrick, 1885 and Epicephala Meyrick, 1880 are particularly notable. Conopomorpha larvae typically develop in seeds of tropical fruits of Malvaceae and Sapindaceae (Hu et al. 2011; Sam et al. 2017; Pasam et al. 2024), while Epicephala has attracted considerable attention for its obligate pollination mutualisms with Phyllanthaceae (Kato and Kawakita 2017). In these systems, females actively collect and deposit pollen using a specialized proboscis, while simultaneously ovipositing in flowers, ensuring that their larvae develop in the resulting seeds (Kato et al. 2003; Kawakita and Kato 2009; Kawakita 2010). Such intricate ecological relationships highlight the evolutionary flexibility of the family but are still poorly documented in the Afrotropical region. In South Africa, three Conopomorpha and six Epicephala species are known, yet their host associations and life histories remain largely unresolved (Vári 1961).

Against this backdrop of limited biological knowledge and rare reproductive-tissue feeding strategies, our recent discovery of a seed-feeding gracillariid in South Africa is particularly noteworthy. The species develops in the fruits of Rhamnus prinoides L’Hér. (Rhamnaceae) and displays morphological features that distinguish it from previously known seed-feeding genera. Instead, these characters place it within the genus Cuphodes, whose members are otherwise known primarily as leaf miners on Ebenaceae, Fabaceae, Putranjivaceae and Rhamnaceae (Kawakita et al. 2010; De Prins et al. 2025) (Table 1). Four species of Cuphodes are listed from South Africa (Vári 1961). Two of these are leaf miners on Ebenaceae, while the other have an unknown biology. However, their taxonomy and generic attribution remains uncertain, as the male of the type species, C. thysanota Meyrick, 1897, is still unknown, and the original description is based solely on the female.

Table 1.

Download as

CSV

XLSX

World check list of the species of the genus Cuphodes, with distribution and hostplant.

Species	Author	Distribution	Hostplant
C. acrolitha	(Meyrick, 1908)	Sri Lanka	unknown
C. albomarginata	(Stainton, 1862)	Australia	unknown
C. callimacha	(Meyrick, 1920)	Australia	unknown
C. calycanthae	De Prins et al. 2025	Australia	Diospyros calycantha O.Schwarz (Ebenaceae)
C. didymosticha	Turner, 1940	Australia	unknown
C. diospyri	Vari, 1961	S. Africa	Diospyros natalensis (Harv.) Brenan (Ebenaceae)
C. diospyrosella	(Issikii, 1957)	Japan	Diospyros kaki L.f. (Ebenaceae)
C. dolichocera	Vari, 1961	S. Africa	unknown
C. drypette	De Prins et al. 2025	Australia	Diospyros calycantha (Ebenaceae), Drypetes deplanchei (Brongn. & Gris) Merr. (Putranjivaceae)
C. habrophanes	Turner, 1940	Australia	unknown
C. holoteles	(Turner, 1913)	Australia	unknown
C. lechriotoma	(Turner, 1913)	Australia	unknown
C. leucocera	Vari, 1961	S. Africa	unknown
C. leucoptera	De Prins et al. 2025	Australia	unknown
C. lithographa	(Meyrick, 1912)	Australia	unknown
C. maculosa	Turner, 1940	Australia	unknown
C. melanostola	(Meyrick, 1918)	Africa	Euclea divinorum Hiern (Ebenaceae)
C. microta	(Turner, 1894)	Australia	unknown
C. niphadias	(Turner, 1913)	Australia	Diospyros calycantha (Ebenaceae)
C. pandoxa	(Turner, 1913)	Australia	unknown
C. plexigrapha	(Meyrick, 1916)	India	Cayanus cayan (L.) Huth (Fabaceae)
C. profluens	(Meyrick, 1916)	India, Australia	Vachellia nilotica (L.) P.J.H.Hurter & Mabb. (Fabaceae)
C. pterocola	(Meyrick, 1914)	India	unknown
C. spermotrophus	sp. nov.	S. Africa	Rhamnus prinoides L’Hér (Rhamnaceae)
C. thysanota	Meyrick, 1897	Australia	unknown
C. tridora	Meyrick, 1911	Seychelles	unknown
C. wisteriella	Kuroko, 1982	Japan	Wisteriopsis japonica (Siebold & Zucc.) J.Compton & Schrire (Fabaceae)
C. zophopasta	(Turner, 1913)	Australia	unknown
sp. 1	Kawakita et al. 2010	Southeast Asia	Berchemiella berchemiifolia (Makino) Nakai, Berchemia floribunda (Wall.) Brogn. (Rhamnaceae)
sp. 2	Kawakita et al. 2010	Southeast Asia	Frangula crenata Miq. (Rhamnaceae)
sp. 3	Kawakita et al. 2010	Southeast Asia	Rhamnella franguloides (Maxim.) Weberb (Rhamnaceae)
sp. 4	Kawakita et al. 2010	Southeast Asia	Diospyros kaki L.f., D. japonica Siebold & Zucc., D. lotus L., D. morrisiana Hance ex Walp. (Ebenaceae)
sp. 5	Kawakita et al. 2010	Southeast Asia	Diospyros sp. (Ebenaceae)
sp. 6	Kawakita et al. 2010	Southeast Asia	Diospyros kaki, D. japonica Siebold & Zucc. (Ebenaceae)

This combination of the unusual biology, uncertain taxonomy, and limited regional knowledge makes the present finding of a seed-feeding Cuphodes significant. It not only broadens the known ecological range of Cuphodes but also contributes to a growing recognition that the diversity and life histories of Afrotropical Gracillariidae remain far from fully documented.

Material and methods

Abbreviations and specimens depository

e.l. ex larva, adult emerging date

f.c. fruit collection date

TRB Triberti Paolo, Museo Civico di Storia Naturale, Verona, Italy

SAMC South African Museum, Iziko Museums of South Africa, Cape Town, South Africa

Sampling and rearing

Fruits were collected directly from R. prinoides over the period of one-year (early July 2023 to early July 2024), in the southern Western Cape region of South Africa, between the towns of George and Knysna. They were obtained from 18 individual shrubs, mostly along forest edges. The average height of R. prinoides in this study was 2.85 m, and the average height at which fruits were collected was 1.88 m.

Mature fruits (i.e., ripe, fleshy berries of purple-red coloration) were transported to a sunlit room in the Plants Laboratory of the George Campus, Nelson Mandela University. Fruits were counted, and placed on absorbent tissue paper in rigid plastic rearing pots, which were covered with a breathable polyester mesh. Each pot contained fruits from a single plant. Rearing pots were misted with water every three days to maintain humidity and were regularly checked for the emergence of insects. Some emerged adults were pinned and dried using standard entomological methods, while most were preserved in 95% ethanol. Larvae were never directly observed; however, cocoons were frequently found inside the rearing pots. The date of cocoon formation and subsequent adult emergence was recorded.

After three months, the rearing pots were transferred to the laboratory, where all fruits (n = 471) were inspected under a stereomicroscope (Olympus SZX7). The fleshy pericarp was removed, and the seeds were inspected. A total of 1,197 seeds were examined, corresponding to an average of 2.54 seeds per fruit. Seeds were classified as shrivelled (n = 715), intact (n = 381) or empty (n = 101). For each fruit sample, the length and width of three seeds were measured. Detailed statistical analyses of seed measurements will be published separately in a subsequent paper.

Subsequent to observations in the rearing pots and on the host plant, cocoons were noted to form on the abaxial side of leaves, along the midrib, post fruit emergence. In April 2026, three leaves of R. prinoides with cocoons present were collected from a forest edge near Nature’s Valley, South Africa, to rear more adult specimens.

The spelling of plant names, their generic combinations and authorship were checked with World Flora Online (WFO 2026).

Morphology

For the genital structures, we follow the interpretation of Klots (1970) and Kristensen (2003). Genitalic dissections were cleared by boiling them in 10% KOH for approximately 5 minutes. They were then cleaned and stained using either a 2% chlorazol black E or mercurochrome. Finally, after being passed through ethanol at increasing percentages (up to 95%), they were permanently embedded in Euparal. The genitalia were drawn using a camera lucida attached to a Leitz Diaplan GMBH.

The curated paratype specimens were imaged at SAMC using a Leica LAS 4.9 imaging system, consisting of a Leica Z16 microscope (using either a 2× or 5× objective) with a Leica DFC450

Camera and 0.63× video objective attached; diffused lighting was achieved using a Leica LED5000 HDI dome. The serial images obtained using an automated Z-stepper were combined using the Leica Application Suite V 4.9 software. Image postproduction was achieved using Adobe Creative Photoshop 2026; plates were produced using Adobe Creative Suite Illustrator 2026.

DNA barcoding

Two specimens were pinned, double-mounted, labeled, photographed, and a leg was removed for DNA barcoding. Tissue samples were stored in 96-well plates and sent to the Canadian Centre for DNA Barcoding (CCDB), University of Guelph, Canada. DNA extraction used the solid phase reversible immobilization (SPRI) method, and PCR-amplified samples were sequenced using SMRT sequencing on a PacBio Sequel platform (Pacific Biosciences, Menlo Park, CA, USA).

Sequences were curated automatically by BOLD (Ratnasingham and Hebert 2007) and assigned to Barcode Index Numbers (BINs) using the RESL algorithm (Ratnasingham and Hebert 2013). In addition, 22 Cuphodes barcode sequences from BOLD, covering 14 different BIN’s across named species (two species) were included in the analyses (Suppl. material 1).

All 24 sequences were aligned using MAFFT (Katoh et al. 2002). A maximum likelihood (ML) tree was inferred with IQ-TREE (Nguyen et al. 2015), and ultrafast bootstrap analysis was performed (Hoang et al. 2017). The resulting ML tree was visualized and edit using iTOL (Letunic and Borg 2019).

Complete voucher data and images of the barcoded specimens can be accessed in the public dataset “DS-CUPHO2” at dx.doi.org/10.5883/DS-CUPHO2 in BOLD.

Results

Systematic account

Genus Cuphodes Meyrick, 1897

Cuphodes Meyrick, 1897. Proc. Linnean Soc. New South Wales 22: 299 (key), 314. Type species: Cuphodes thysanota Meyrick, 1897, by monotypy.

Cyphosticha Meyrick, 1907. Proc. Linnean Soc. New South Wales 32: 49 (key), 61. Type species: Gracilaria (sic) pyrochroma Turner, 1894, by original designation.

Taxonomic history.

The genus was described by Meyrick in 1897 on the basis of a female, C. thysanota from Australia. It was compared with Stathmopoda Herrich-Schäffer, 1853, then included in the family Elachistidae, now included in the family Stathmopodidae (De Prins and De Prins 2011–2025). Meyrick probably confused Stathmopoda for the long and bristly scales on the mid and hindlegs and a similar peculiar posture of the adult. Subsequently Vári (1961) redescribed the genus, arguing that it should be included in the family Gracillariidae, near Cyphosticha Meyrick, 1907. For the description of the male characters, he used a species from India, Phrixosceles trochosticha Meyrick, 1908, the type genus of Phrixosceles Meyrick, 1908 which he considered as a synonym of Cuphodes (Fletcher 1929). For females he used both C. thysanota and P. trochosticha. This synonymy was proposed by Vári (1961) based on the similarity of the wing venation and the bristly hairs on the legs. De Prins et al. (2025), studying the Australian fauna, introduced new characters and considered Cyphosticha as a synonym of Cuphodes. They also moved all Australian species of Phrixosceles to the genus Cuphodes, but left the former genus valid with two species, P. trochosticha Meyrick, 1908 (type genus, India) and P. fibulatrix Meyrick, 1922 (Fiji) (De Prins et al. 2025).

Distribution and species richness.

The genus Cuphodes currently comprises 28 species distributed across Australia, Africa and southeastern Asia (De Prins et al. 2025). The nine species with known biology are leaf miners associated with Ebenaceae, Fabaceae, Putranjivaceae and Rhamnaceae. This total excludes Cuphodes paragrapta (Meyrick, 1915) from South America, which upon examination, does not appear to belong to Cuphodes but to an unknown genus. In addition to the described species, six putative species were reported by Kawakita et al. (2010), comprising leaf miners on Ebenaceae and Rhamnaceae from East Asia (Table 1), whose morphology has not yet been described.

DNA.

Cuphodes has frequently been included in molecular phylogenetic studies. Kawakita (2010) identified it as a putative sister taxon to Epicephala, concluding that it is probably one of the closest relatives of that genus. More recently, De Prins et al. (2025) found that mitochondrial data consistently support the monophyly of the Australian Cuphodes species, recovered as sister to Toowoomba De Prins, Sruoga & Zwick, 2025 + (Parectopa Clemens, 1860+Polysoma Vári, 1961), albeit with low to moderate statistical support. In contrast, Cuphodes diospyrosella (Issiki, 1957) and C. wisteriella Kuroko, 1982, included in the analyses of Kawahara et al. (2016) and Li et al. (2022), were recovered as sister to all other Ornixolinae. In those studies, Parectopa was placed either very distantly (Kawahara et al. 2016) or was recovered as strongly polyphyletic (Li et al. 2022).

Australian Cuphodes.

The characters reported by De Prins et al. (2025) for Australian Cuphodes are a shining-white backround with a subtle yellow ochreous pattern on the forewing and exceptionally pilose hindlegs, equipped with erected sharply pointed spiniform scales. In male genitalia subscaphium+gnathos strongly developed; costa of valva very strongly sclerotised; subapical part of valva with a hanging semi-round flap or other sclerotisations of different sizes and shapes; transtilla complete but in case of absence the function of transtilla is taken by very long basal apodemes of valvae or sclerotised basal bow of vinculum. In female genitalia apophyses anteriores either absent or rather short; papillae anales strongly fused and deeply nested in the cuticle of segment VIII; antrum/colliculum sclerotised; signa small; bulla seminalis usually situated at anterior margin of segment VII or in segment VII, small with convoluted ductus seminalis.

South African Cuphodes.

Examination of African species treated by Vári (1961), specifically those with known males (C. melanostola (Meyrick, 1918), C. diospyri Vari, 1961, and C. dolichocera Vari, 1961), reveals important differences in the male genitalia compared with the characters reported by De Prins et al. (2025), particularly in the shape of the valva, which is simply rounded apically and lacks notable sclerotization. In the newly described C. spermotrophus sp. nov., two bristles are present at the apex of tegumen (Figs 1, 2). This feature is characteristic of several genera in other subfamilies of Gracillariidae, such as Spulerina (Acrocercopinae) and Spanioptila (Ornixolinae), and is particularly significant within Lithocolletinae. Among Australian Cuphodes, apical tegumen bristles are prominent in the pyrochroma clade, which includes C. pyrochroma (Turner, 1894), C. pandoxa (Turner, 1913), C. callimacha (Meyrick, 1920), but are apparently absent in the other groups. Notably, such bristles are not visible in the South African Cuphodes illustrated by Vári (1961) and are not mentioned in his descriptions. The wing venation does not differ from that of the type species C. thysanota Meyrick, 1897, in particular due to the absence of R1 and R2 in the forewing (Figs 4, 5).

Figures 1–3.

Cuphodes spermotrophus sp. nov. South Africa, Western Cape, Woodville, Duif street. Male abdomen and genitalia. 1. Lateral view of last segments, paratype, slide TRB4546. 2. Ventral view of the genitalia, holotype, slide TRB4550. 3. Phallus, c. s. Scale bar: 290 μm.

Figures 4, 5.

Cuphodes spermotrophus sp. nov. South Africa, Western Cape, Woodville, Duif street. Wing venation. 4. Forewing; 5. Hindwing. Scale bar: 900 μm.

Adults of the new South African species exhibit a typical resting posture for Cuphodes: the head is held downward, the abdomen raised and supported by the hindlegs, and the fore- and midlegs extended forward and held close together (Fig. 14) (Kawahara et al. 2016).

Based on these morphological and behavioural traits, we provisionally place spermotrophus sp. nov. in Cuphodes, following Vári’s interpretation, until further molecular or morphological data allow a more definitive placement.

Cuphodes spermotrophus Triberti, Swart, Van Noort & Lopez-Vaamonde, sp. nov.

https://zoobank.org/7F78C666-7322-4E64-97C4-E7F3ACB4BD02

Figs 1, 2, 3, 4, 5, 6, 7, 8

Holotype.

• ♂, South Africa, Western Cape, Woodville, Duif street, e.l. 27/ix/2023, CLV49591, GRAEL7889-25, R Swart leg. slide TRB4550♂. Coll. TRB.

Paratypes.

1♂, 7♀♀ (coll. TRB); • 1♂, South Africa, Western Cape, Woodville, Duif street, e.l. 27/ix/2023, R Swart leg., slide TRB4546♂; • 1♀, same locality, f.c. 18/iv/2024, e.l. 11/v/2024, RP306, R Swart leg., slide TRB4545♀, TRB4549♀ [wings]; • 1♀, same locality, f.c. 24/x/2023, e.l. 30/ix/2023, RP180, R Swart leg., slides TRB4547♀, TRB4543♀ [wings]; • 1♀, same locality, f.c. 21/viii/2023, e.l. 18/ix/2023, R Swart leg., slides TRB4553♀; • 1♀, same locality, f.c. 18–27/ix/2023, CLV49590, GRAEL7888-25, RP122, R Swart leg., slide TRB4544♀; • 1♀, South Africa, Western Cape, Garden Route National Park, -33.969966, 23.538344, 202 m, f.c. 22 April 2026, e.l. 23 April 2026, RC Swart leg., Afromontane Forest edge, Pupa on leaf of Rhamnus prinoides, bottom of leaf along midrib, RHAM26-H230426, SAM-LEP-A053741 (SAMC); • 1♀, ditto, except for: e.l. 27 April 2026, RHAM26-H270426, SAM-LEP-A053742 (SAMC); • 1♀, ditto, except for: e.l. 28 April 2026, RHAM26-H280426, SAM-LEP-A053743 (SAMC).

Etymology.

Derived from Greek sperma- (seed) and -tropho (to feed), referring to the seed-feeding habit of the species, a masculine noun in apposition.

Diagnosis.

Ground colour of forewing pattern dark brown with dorsum marked by a thin white line with an irregular internal margin (7A-C); apex with a small ochre-white area with a central dark brown dot (Fig. 7D). This pattern makes this species easily identifiable from other African Cuphodes but it is very similar to the Australian Diphtheroptila glochidiella De Prins, Sruoga & Zwick, 2025, a problematic species that lives on Glochidion (Phyllanthaceae) (De Prins et al. 2025). However, the latter differs in the different apical pattern of forewing and in the female genitalia while male genitalia and wing venation are unknown. Male and female genitalia of the new species show some affinity with C. diospyri from which it differs for the much shorter valva, costo-apical corner protruded and in the female ductus bursae with convolutions. Adults have strongly setose mid and hindlegs (Fig. 6).

Figure 6.

Cuphodes spermotrophus sp. nov. Adult, paratype, SAM-LEP-A053741: A. Lateral view; B. Dorsal view; C. Ventral view.

Figure 7.

Cuphodes spermotrophus sp. nov. Adult, paratype, SAM-LEP-A053742: A. Dorsal view; B. Left wings. SAM-LEP-A053741: C. Right forewing; D. Apex. SAM-LEP-A053742; E. Head, lateral view.

Description of adult.

Forewing length 4,5 mm (Fig. 7C).

Head (Fig. 7E). Vertex and occiput white, with slightly raised scales, frons of the same colour; labial palpus white, curved upwards, almost as long as fore tibiae, first and second segment slightly spotted brown above; maxillary palpus smooth, 4 segmented, about a third as long as the median segment of labial; antenna slightly longer than forewing, with scape and pedicel brown above and white below, pecten present with about ten thin bristles, light ochre filament with barely evident ringing.

Thorax. White, tegula dark brown. Foreleg femur, tibia and tarsomeres (Fig. 6) whitish yellow, brownish above; midleg white, femur and tibia brownish above, tibia with a white area subapically, covered with thin white, pointed, spine-like scales continuing onto the first two tarsomeres, first stained dark brown subbasally, last two stained dark brown; hindleg white, tibia stained dark brown apically, median and apical spurs striped with brown, less densely covered with thin white and pointed scales which continue on first two tarsomeres, terminal tarsomere with fuscous apex.

Wings. Forewing elongated (Fig. 7A-C), dark brown with a sparse sprinkling of white scales: a white dorsal band irregularly margined in dark ending at the apical 2/3 of wing, followed by small, irregular light spots characterising the apical area; apex with a small ochre-white area with a central dark brown dot and a double fringe line, the innermost one with strong blue-green highlights. Cilia brownish along tornus and then lighter. Hindwings uniform greyish brown with light ochre-grey cilia.

Abdomen (Fig. 1). Male abdomen brownish above, brownish ochre on sides with five short oblique dark brown streaks, white below. Segment VI slightly shorter than segment V, normally scaled, anterior sternal margin with a subtriangular projection; segments VII and VIII weakly membranous, bare, each bearing a pair of coremata consisting of long, piliform scales, posterior slightly longer than anterior and about the length of segment VI; segment VII reduced into a thin sclerite, with a pair of sublateral oval androconial plates consisting of thin specialized scales; posterior margin of tergum VIII with a subtriangular projection covered with piliform scales about 2/3 the length of the anterior corema. In females, the pattern of abdomen is lighter.

Male genitalia (Figs 2, 3). Tegumen broad, conical, with rounded apex on which two setae are present about 1/5 as long as posterior coremata, basal joint broad and sclerotised; tuba analis apically microsetose, just longer than tegumen and with a wide subscaphium covered with minute setae, more ventrally the diaphragm presents a densely microsetose protrusion; valva simple, with parallel edges then tapering towards the apex, without appendages, barely visible the sacculus and with a strongly sclerotised costal margin with exception of cucullus, setae of varying length along the entire ventro-apical margin and, to a lesser extent, along the dorsal, transtilla absent; vinculum wide, with large lateral lobes, sutures very evident, with a long saccus about 0.6 length of valva; phallus straight, tubular, about 4× length of valva, without cornuti, phallobase about 2/3 long, inception of bulbus ejaculatorius at 0.2 length of phallus, aedeagus with crest-shaped apex.

Female genitalia (Fig. 8). Papillae anales fused dorsally, apophyses posteriores longer than apophyses anteriores; segment VIII very reduced, apophyses anteriores present; segment VII with a deep concavity along the ventro-apical margin which also involves the intersegmental membrane; sterigma membranous, with a weakly sclerotised rounded area into which a small ostium bursae opens; along the sub-basal margin two thin protrusions covered by long and thin scales; antrum weakly sclerotised, ductus bursae membranous with two convolutions, slightly thickened in its distal third; corpus bursae elongated, sub-cylindrical, without signa; bulla seminalis at sub-posterior part of segment VII, smaller than the corpus bursae, ductus seminalis not perceptible; ductus spermathecae spiraling into 4/5 convolutions before entering the vesica.

Figure 8.

Cuphodes spermotrophus sp. nov. South Africa, Western Cape, Woodville, Duif street. Female genitalia, paratype, slide TRB4544. Scale bar: 290 μm.

Bionomics.

This species represents the first record of a seed-feeding Gracillariidae from Africa. It attacks the berries of Rhamnus prinoides, a shrub or small evergreen trees (Fig. 9) in the Rhamnaceae family that can reach 6–8 m in height. It is found primarily in East Africa, from Ethiopia to South Africa, but also in the central-western regions, including Cameroon, Angola, and the Democratic Republic of Congo. It grows preferably in regions with an altitudinal range of 1,400–3,200 m, on wooded mountain and hill slopes, at the edges of forests, and on the banks of streams, often among rocks. Its flowers develop into fruits/red berries, which, as they mature, produce ripe, dark-purple, and softer berries, which are ready for harvesting and seed extraction (Negasse 2021).

Figure 9.

Habitat of Rhamnus prinoides. South Africa, Western Cape. A mature (centre-right, 2.4 m) and sapling stage (centre-left, 1.5 m) Rhamnus prinoides in a natural forest canopy gap (12.01.2026) (33°56.669’S, 23°30.510’E).

When seeds of R. prinoides were examined, their mean length was 4.76 mm and their mean width was 3.02 mm. In fruits from which C. spermotrophus emerged, a distinct exit hole is visible on the fruit surface, close to the calyx (Fig. 10). Beneath this opening, an empty seed can be observed (Fig. 11). Only a single exit hole was ever observed per fruit. Cocoons were recorded in January, February, March, May and July while adult emergence occurred in September, October, February, April, May and June. Attacked fruits were found on shrubs growing along forest edges, where the host plant grows naturally. The presence of the species can also be confirmed by examining the underside of the leaf midrib, where cocoons are formed after larval emergence from the seed (Fig. 12).

Figures 10–13.

Cuphodes spermotrophus sp. nov. on fruit of Rhamnus prinoides. South Africa, Western Cape. 10. Fruit with larval exit hole next to the calyx. 11. Upon removing the fleshy part of the fruit, below the exit hole, the empty seed is visible (see yellow arrow). 12. Cocoon on the underside of a leaf of the host plant. 13. Cocoon covered by frothy “bubbles”.

Figure 14.

Cuphodes spermotrophus sp. nov. Adult, resting posture. Photo credit: John Roux.

For reasons that remain unclear, the larva enclosed within its cocoon covers it with frothy “bubbles” produced from the anus and expelled through small slits in the cocoon wall, where they remain attached to the external surface (Fig. 13). This unusual behaviour has also been reported in other Cuphodes species (De Prins et al. 2025) as well as in Chileoptilia, Marmara and related genera (Wagner et al. 2000; Vargas and Landry 2005).

Besides C. spermotrophus, the only frugivorous insect reared from R. prinoides was a single adult belonging to the Drosophila suzukii Matsumura, 1931, a recent invader in South Africa (Steyn et al. 2025).

Parasitoids.

In total, only one eulophid (Tetrastichinae) emerged from hundreds of berries of R. prinoides. The occurrence of the eulophid is noteworthy, as nothing is currently known about the parasitoids of Cuphodes. Among genera with feeding habits similar to spermotrophus, only a few species of Conopomorpha are known to be parasitized by Braconidae, Ichneumonidae, Encyrtidae, and Trichogrammatidae (De Prins and De Prins 2011–2025).

Distribution.

The micromoth so far has only been found between George (33°57.337'S, 22°31.765'E) and Nature's Valley (33°58.200'S, 23°32.301'E) in the Western Cape province. The distribution of the hostplant in Africa suggests that this gracillariid might also have a wide distribution but it has just been overlooked due to its cryptic life cycle.

DNA barcoding analysis.

The two barcodes (GRAEL7888-25; GRAEL7889-25) form a unique BIN BOLD:AHB8508 (Suppl. material 1, Fig. 15) at 9.66% from the nearest Gracillariidae barcode in BOLD. Our phylogenetic analysis shows that Cuphodes spermotrophus sp. nov. occupies an isolated branch. Six Rhamnaceae feeding specimens belonging to three Cuphodes species form a single clade suggesting host use conservatism. Cuphodes spermotrophus does not come out in this clade but internal nodes are poorly supported and it is possible that with more sequence data and taxa the Rhamnaceae feeding Cuphodes will be monophyletic (Fig. 15). Another possible scenario is that the new species evolved independently from Ebenaceae feeders. At the current level of resolution, the tree does not allow us to discriminate between these two hypotheses. The still unknown biology of some African Cuphodes may perhaps provide further information.

Figure 15.

Maximum likelihood tree (log-likelihood = −3482.5934 ± 132.3242) based on 24 Cuphodes COI (DNA barcode) sequences. Each specimen is identified by its Process ID (barcode), taxon name, and, when available, the host plant for reared specimens (see also Suppl. material 1). Bootstrap support values ≥ 70% are shown on the branches. The six samples Rhamnaceae-feeding specimens are highlighted in red and represent underscribed species.

Key to the Cuphodes species of South Africa based on external morphological characters

1	Ground colour of forewing whitish with a mixed pattern of yellow ochre and black	2
–	Ground colour of forewing dark brown, edged dorsally in white	C. spermotrophus sp. nov.
2	Forewing with white yellow pattern	*C. dolichocera*
–	Forewing with dark pattern	3
3	Forewing with a blackish dot in middle of costa and at 1/3 of dorsal margin	*C. diospyri*
–	Forewing with irregular dark pattern	4
4	Antenna fuscous	*C. melanostola*
–	Antenna white	*C. leucocera*

Key to the Cuphodes species of South Africa based on male genitalia

Male genitalia of C. leucocera unknown.

1	Phallus with cornuti	*C. dolichocera*
–	Phallus without cornuti	2
2	Valva subrectangular with rounded apex	3
–	Valva subtriangular with pointed apex	C. spermotrophus sp. nov.
3	Valva with almost parallel margins	*C. diospyri*
–	Valva with wavy margins with a subapical constriction	*C. melanostola*

Key to the Cuphodes species of South Africa based on female genitalia

Female genitalia of C. dolichocera unknown

1	Bursa copulatrix with ductus bursae partially or completely sclerotised	2
–	Bursa copulatrix with ductus bursae not sclerotised	3
2	Ductus bursae sclerotised in the distal half	*C. melanostola*
–	Ductus bursae entirely sclerotised	*C. leucocera*
3	Ductus bursae with one or two convolutions before joining corpus bursae	C. spermotrophus sp. nov.
–	Ductus bursae with no convolutions	*C. diospyri*

Discussion

Species of Cuphodes reported from Rhamnaceae (Kawakita et al. 2010) lack detailed biological information; however, generic traits suggest that they are leaf miners. C. spermotrophus represents the first documented case of seed feeding within the genus, and the first known seed-feeding gracillariid in Africa. This is particularly remarkable because Rhamnaceae are rarely recorded as host plants in Ornixolinae. Prior to this study, Rhamnaceae were known as hosts in Ornixolinae only for the North American genus Apophthisis Braun, 1915 (A. pullata Braun, 1915 feeding on Rhamnus lanceolata) and for a small group of three putative Asian Cuphodes species feeding on Frangula crenata, Berchemia floribunda, Rhamnella franguloides, and Berchemiella berchemiifolia (Kawakita et al. 2010).

The wide distribution of its host plant, Rhamnus prinoides, suggests that C. spermotrophus may have a broader geographic range. This plant occurs widely across the African continent, often cultivated around farms, in pure stands, or intercropped with other annual or perennial crops. All parts of the plant are harvested for food, medicinal, or traditional purposes, and it is an important resource for many bird species (Negasse 2021). Its ability to coppice readily after repeated harvest, provided sufficient water, makes it suitable for soil erosion control while also offering economic benefits to smallholder farmers (Negasse 2021).

Finally, the increasing use of Malaise traps in international DNA barcoding campaigns (deWaard et al. 2019; D’Souza et al. 2021; Van Noort 2025) may facilitate detection of C. spermotrophus, as has occurred for other gracillariid species (Triberti et al. 2024). Such monitoring approaches combined with targeted rearing are crucial for documenting the distribution, host associations, and potential ecological impact of species with cryptic larval habits, including seed-feeding leaf miners.

Acknowledgements

We thank Erik van Nieukerken and Scott Miller for their thorough review of an earlier version of the manuscript. Thanks to both Atsushi Kawakita and Issei Ohshima for helpful comments on Rhamnaceae-feeding Cuphodes. Thanks to Hossein Rajaei for hand-carrying specimens. We would also like to thank the team at the Centre for DNA Barcoding in Guelph, especially Rebekah Hest, Spencer Walker and Claudia Steinke for their invaluable assistance in submitting and curating DNA barcode data in BOLD.

This research was partially funded by the National Research Foundation of South Africa (grant number PSTD220324610 and the Rufford Foundation (grant number 48270-2). DNA barcoding costs were covered by InfoBios FEDER project (EX011185) funded by the Region Centre–Val de Loire, France.

References:

De Prins J, De Prins W (2011–2025) Afromoths, online database of Afrotropical moth species (Lepidoptera). World Wide Web electronic publication. https://www.afromoths.net [accessed 28.03.2025]

De Prins J, Hartley D, Sruoga V, Nicholls J, Wallace J, Zwick A (2025) Diversity of Australian Ornixolinae (Lepidoptera: Gracillariidae) with taxonomic and nomenclatural acts within the related taxa (Acrocercopinae and Gracillariinae) based on the evidence of museomics, bionomics, and mitogenomics. Zootaxa 5616(1): 1–340. https://doi.org/10.11646/zootaxa.5616.1.1

deWaard JR, Levesque-Beaudin V, deWaard SL, Ivanova NV, McKeown JTA, Miskie R, Naik S, Perez KHJ, Ratnasingham S, Sobel CN, Sones JE, Steinke C, Telfer AC, Young AD, Young MR, Zakharov EV, Hebert PDN (2019) Expedited assessment of terrestrial arthropod diversity by coupling Malaise traps with DNA barcoding. Genome 62(3): 85–95. https://doi.org/10.1139/gen-2018-0093

D’Souza ML, van der Bank M, Shongwe Z, Rattray RD, Stewart R, van Rooyen J, Govender D, Hebert PDN (2021) Biodiversity baselines: Tracking insects in Kruger National Park with DNA barcodes. Biological Conservation 256: 109034. https://doi.org/10.1016/j.biocon.2021.109034

Fletcher TB (1929) A list of the generic names used for Microlepidoptera. Memoirs of the Department of Agriculture in India. Entomological Series 11: 1–244.

Guiguet A, Ohshima I, Takeda S, Laurans F, Lopez Vaamonde C, Giron D (2019) Origin of gall-inducing from leaf-mining in Caloptilia micromoths (Lepidoptera, Gracillariidae). Scientific Reports 9: 6794. https://doi.org/10.1038/s41598-019-43213-7

Guiguet A, van Nieukerken EJ, Giron D, Gravendeel B, Lopez-Vaamonde C, Ohshima I (2026) Two new species of Caloptilia (Lepidoptera Gracillariidae) from New Caledonia inducing galls on Glochidion billardierei (Phyllanthaceae) and redescription of C. xanthopharella (Meyrick, 1880). ZooKeys 1268: 113–137. https://doi.org/10.3897/zookeys.1268.173885

Hoang DT, Chernomor O, von Haeseler A, MiBQ, Vinh LS (2017) UFBoot2: Improving the ultrafast bootstrap approximation. Molecular Biology and Evolution 35(2): 518–522. https://doi.org/10.1093/molbev/msx281

Hu B, Wang S, Zhang J, Li H (2011) Taxonomy and biology of two seed-parasitic gracillariid moths (Lepidoptera, Gracillariidae), with description of a new species. ZooKeys 83: 43–56. https://doi.org/10.3897/zookeys.83.783

Kato M, Kawakita A (2017) Biology of the obligate pollination mutualism. In: Kato M, Kawakita A (Eds) Obligate pollination mutualism. Ecological Research Monographs 2017: 63–80. https://doi.org/10.1007/978-4-431-56532-1

Kato M, Takimura A, Kawakita A (2003) An obligate pollination mutualism and reciprocal diversification in the tree genus Glochidion (Euphorbiaceae). Proceedings of the National Academy of Sciences of the USA 100(9): 5264–5267. https://doi.org/10.1073/pnas.0837153100

Katoh K, Misawa K, Kuma KI, Miyata T (2002) MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform. Nucleic Acids Research 30(14): 3059–3066. https://doi.org/10.1093/nar/gkf436

Kawahara AY, Plotkin D, Ohshima I, Lopez-Vaamonde C, Houlihan PR, Breinholt JW, Kawakita A, Xiao L, Regier JC, Davis DR, Kumata T, Sohn JC, De Prins J, Mitter C (2016) A molecular phylogeny and revised higher-level classification for the leaf-mining moth family Gracillariidae and its implications for larval host-use evolution. Systematic Entomology 42: 60–81. https://doi.org/10.1111/syen.12210

Kawakita A (2010) Evolution of obligate pollination mutualism in the tribe Phyllantheae (Phyllanthaceae). Plant Species Biology 25: 3–19. https://doi.org/10.1111/j.1442-1984.2009.00266.1

Kawakita A, Kato M (2009) Repeated independent evolution of obligate pollination mutualism in the Phyllantheae-Epicephala association. Proceedings of the Royal Society B. 276: 417–426. https://doi.org/10.11646/zootaxa.5616.1

Kawakita A, Okamoto T, Goto R, Kato M (2010) Mutualism favours higher host specificity than does antagonism in plant-herbivore interaction. Proceedings of the Royal Society, series B Biological Sciences 277: 2765–2774. https://doi.org/10.1098/rspb.2010.0355

Klots AB (1970) Lepidoptera. In: Tuxen SL (Ed.) Taxonomist’s Glossary of Genitalia in Insects (2nd revised and enlarged edn). Munksgaard, Copenhagen, 115–130.

Kristensen NP (2003) Skeleton and muscles: adults. In: Kristensen NP (Ed.) Lepidoptera, Moths and Butterflies (Vol 2). Morphology, Physiology, and Development. Handbook of Zoology 4(36): 39–132. https://doi.org/10.1515/9783110893724.39

Kumata T (1964) Description of a new stem-miner of coniferous trees frm Japan (Lepidoptera: Gracillariidae). Insecta Matsumurana 27(1): 31–34.

Letunic I, Borg P (2019) Interactive Tree Of life (iTOL) v4: recent updates and new developments. Nucleic Acids Research 47, W256–W259. https://doi.org/10.1093/nar/gkz239

Li X, St Laurent R, Earl C, Doorenweerd C, van Nieukerken E, Davis DR, Johns CA, Kawakita A, Kobayashi S, Zwick A, Lopez-Vaamonde C, Ohshima I, Kawahara A (2022) Phylogeny of gracillariid leaf-mining moths: evolution of larval behaviour inferred from phylogenomic and Sanger data. Cladistics 38(3): 277–300. https://doi.org/10.1111/cla.12490

Lopez-Vaamonde C, Agassiz DJL, Augustin S, De Prins J, De Prins W, Gomboc S, Ivinskis P, Karsholt O, Koutroumpas A, Koutroumpa F, Laštůvka Z, Marabuto E, Olivella E, Przybylowicz L, Roques A, Ryrholm N, Šefrová H, Šima P, Sims O, Sinev S, Tomov R, Zilli A, Lees DC (2010) Chapter 11. Lepidoptera. In: Roques A, Kenis M, Lees D, Lopez-Vaamonde C, Rabitsch W, Rasplus JY, Roy D (Eds) Alien terrestrial arthropods of Europe. BioRisk 4(2): 603–668. https://doi.org/10.3897/biorisk.4.50

Meyrick E (1897) Descriptions of Australian Micro-Lepidoptera. XVII. Elachistidae. Proceedings of the Linnean Society of New South Wales 22: 297−435. https://doi.org/10.5962/bhl.part.12726

Meyrick E (1908) Descriptions of Indian Micro-Lepidoptera. VIII. Journal of the Bombay Natural History Society 18(4): 806−832.

Negasse L (2021) A selection of African native trees: biology, uses, propagation and restoration techniques. Addis Ababa, Ethiopia, 621 pp.

Nguyen L-T, Heiko A, Schmidt A, von Haeseler B, Quang M (2015) IQ-TREE: A fast and effective stochastic algorithm for estimating maximum likelihood phylogenies. Molecular Biology and Evolution 32: 268–274. https://doi.org/10.1093/molbev/msu300

Pasam MR, Longchar I, Choudhary JS, Maiti CS, Devi HS, Neog P, Banik S (2024) Biology, morphometric and integrative taxonomy of litchi fruit borer, Conopomorpha sinensis Bradley, 1986: A major pest of litchi (Litchi chinensis Sonn.). Animal Biology 74(4): 415–433. https://doi.org/10.1163/15707563-bja10148

Ratnasingham S, Hebert PDN (2007) BOLD: The Barcode of Life Data System (http://www.barcodinglife.org). Molecular Ecology Notes 7: 355–364. https://doi.org/10.1111/j.1471-8286.2007.01678.x

Ratnasingham S, Hebert PDN (2013) A DNA-Based Registry for All Animal Species: The Barcode Index Number (BIN) System. PLoS ONE 8(7): e66213. https://doi.org/10.1371/journal.pone.0066213

Sam K, Ctvrtecka R, Miller SE, Rosati ME, Molem K, Damas K, Gewa B, Novotny V (2017) Low host specificity and abundance of frugivorous lepidoptera in the lowland rain forests of Papua New Guinea. PLoS ONE 12(2): e0171843. https://doi.org/10.1371/journal.pone.0171843

Steyn L, Jodamus Z, Steyn V, Collop A, Addison M, Bekker G, Esterhuizen N, Karsten M, Addison P (2025) First records of the fruit pest Drosophila suzukii (Matsumura, 1931) (Diptera: Drosophilidae) in South Africa. African Entomology 33(1): e22307. https://doi.org/10.17159/2254-8854/2025/a22307

Triberti P, Staude H, Sharp I, Lopez-Vaamonde C (2024) Exploring the diversity of Gracillariidae (Lepidoptera) in South Africa: host plants, distribution, and DNA barcoding analysis, with the description of nine new species. Zootaxa 5529(1): 1–51. https://doi.org/10.11646/zootaxa.5529.1.1

Van Noort S (2025) The Iziko South African Museum entomology collection; a goldmine of biodiversity data with mobilization challenges for more than 50 million specimens. Biological Journal of the Linnean Society 145(4): 1–10. https://doi.org/10.1093/biolinnean/blaf062

Vargas HA, Landry B (2005) A new genus and species of Gracillariidae (Lepidoptera) feeding on flowers of Acacia macracantha Willd. (Mimosaceae) in Chile. Acta Entomológica Chilena 29: 47–57.

Vári L (1961) South African Lepidoptera. Vol. I. Lithocolletidae. Transvaal Museum Memoir 1961: 1–238.

Wagner DL, Loose JL, Fitzgerald TD, De Benedictis JA, Davis DR (2000) A Hidden Past: the Hypermetamorphic Development of Marmara arbutiella (Lepidoptera: Gracillariidae). Annals of the Entomological Society of America 93(1): 59–64. https://doi.org/10.1603/0013-8746(2000)093%5b0059:AHPTD%5d.0.CO;2

WFO (2026) World Flora Online. http://worldfloraonline.org [accessed on 16.02.2026]

Supplementary material

Supplementary material 1

Metadata of 24 specimens used in the DNA barcoding analysis, with sample and process ID, BIN and accession number

Carlos Lopez-Vaamonde

Data type: xlsx

Explanatory note: Additional collecting and specimen data are accessible in BOLD through the public dataset: DS-CUPHO2. (dx.doi.org/10.5883/DS-CUPHO2).

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.

Download file (21.23 kb)