Abstract
The genus Gibellula (Cordycipitaceae: Hypocreales) comprises species pathogenic on and specific to spider hosts. Here, we report on the occurrence of a novel species infecting cave-dwelling, orb-weaving spiders of the sub-family Metainae (Tetragnathidae) in the British Isles. The new species, Gibellula attenboroughii is described and illustrated and its ecology is discussed; leading to the conclusion that infected spiders exhibit behavioural changes similar to those reported for zombie ants. The hidden diversity of the genus Gibellula in the British Isles is further highlighted based on fungarium records and literature searches. Two spider pathogens, previously assigned to the genus Torrubiella, are now transferred to the genus Gibellula, based on their Granulomanus synasexual morphs, in accordance with the one fungus-one name initiative: Gibellula albolanata comb. nov. and G. aranicida comb. nov.
Citation: Evans HC, Fogg T, Buddie AG, Yeap YT, Araújo JPM (2025). The araneopathogenic genus Gibellula (Cordycipitaceae: Hypocreales) in the British Isles, including a new zombie species on orb-weaving cave spiders (Metainae: Tetragnathidae). Fungal Systematics and Evolution 15: 153–178. doi: 10.3114/fuse.2025.15.07
Keywords: entomopathogenic fungi, Gibellula attenboroughii, host manipulation, new taxa, systematics
INTRODUCTION
Gibellula (Cordycipitaceae: Hypocreales) is a genus specific to and pathogenic on spiders comprising over 30 accepted species (Mendes-Pereira et al. 2023). These authors also noted that 54 and 58 species are listed in Index Fungorum and MycoBank, respectively; highlighting the uncertainties of species delimitation within the genus. The greater majority of these taxa have a sub-tropical to tropical distribution (Evans 1982, Evans & Samson 1987, Samson et al. 1988, Samson & Evans 1992, Evans 2013, Hughes et al. 2016, Shrestha et al. 2019, Kuephadungphan et al. 2020, 2022, Mendes-Pereira et al. 2023, Nyffeler & Hywel-Jones 2024). There are few records of the genus from the British Isles with only a single species, G. aranearum, being recognised in the early published lists of “British entomogenous fungi” (Petch 1932a, b, Petch 1948). Subsequently, Ellis (1956) recorded G. aranearum on spiders in the fens and meadows of Norfolk; describing it as common from June to October which is evidenced by the large amount of material deposited in Herb K. Earlier, Mains (1950) had included G. aranearum as a synonym of G. pulchra – the type species of the genus – and this has been accepted since (Samson & Evans 1992, Roth & Van Vooren 2016, Shrestha et al. 2019, Mendes-Pereira et al. 2023); although G. aranearum, as described by Sydow (1922) on a spider from Papua New Guinea, remains a legitimate species in both Index Fungorum and MycoBank.
There is no doubt that G. pulchra in the British Isles and elsewhere is a species complex, especially since the only sequences available of G. “pulchra” are from Asia. In the phylogenetic trees of the genus published, thus far, with multiple sequences of G. “pulchra” from various localities, these do not cluster together (Mendes-Pereira et al. 2022; 2023). For example, Mendes-Pereira et al. (2023) in their in-depth study of the genus include 10 sequences; four clustering together in two widely separate groups and the other two scattered within the tree. All these originated from the tropics, whilst the type was collected in Italy (Cavara 1894). Clearly, there is a need to sequence the holotype if located, or to designate a neotype from the country of origin, in order to avoid further confusion.
During the BBC Winterwatch television series in 2021, filmed on location in Northern Ireland, a fungal-infected spider was discovered on the ceiling of an abandoned gunpowder store and provisionally identified as G. “pulchra”. Images, sent by the BBC to the senior author, suggested that this was a novel species of the genus Gibellula and, moreover, that the reclusive cave-spider host – subsequently, identified as the orb-weaving spider Metellina merianae (Tetragnathidae: Araneae) – had moved to an exposed situation before death, indicating a behavioural change. These suppositions were included and discussed in the following 2022 Springwatch series after which the specimen was removed and sent for identification.
Morphological and phylogenetic analyses confirmed the taxonomic novelty of the Gibellula on the cave spider and its distinctiveness from the species accepted, thus far. Here, we describe the new taxon based on this specimen, as well as others discovered on two orb-spider species in cave systems on the island of Ireland. A subsequent investigation into the historical records of the genus Gibellula in the British Isles has revealed a hidden diversity, as well as reports of epizootics; the results of which are presented and discussed.
MATERIALS AND METHODS
Field collection
The type specimen was removed with fine forceps from the ceiling of an abandoned gunpowder storeroom at Castle Espie Wetland Centre in Northern Ireland (54°31’45.66”N, 5°41’53.16”W) and transferred to a sterile plastic tube, then air-dried for 4 d, before being sent for examination to CABI UK-Centre at Egham. Further specimens of Gibellula-infected spiders were collected from cave systems on the island of Ireland by a local speleologist (T. Fogg) and despatched in the same way. In addition, images were taken in situ, as well as details of the positions of the infected spiders within the cave system, particularly in relation to the light zones and the cave entrance.
Fungal isolation and DNA extraction
Dried specimens were examined with a dissecting microscope (Nikon SMZ-10, Japan) and sporulating structures were removed with a flamed needle and streaked onto tap-water agar (TWA) and potato-carrot agar (PCA; Ritchie 2002), and incubated in the dark at 20 °C until evidence of colony growth. Hyphae from the colony periphery were then transferred to fresh PCA and malt-extract agar (MEA) plates using a sterile needle.
Genomic DNA was extracted using MicroLYSIS® PLUS (Microzone Ltd., Stourbridge, UK) with a modified thermal cycling lysis profile of 1× (45 min at 65 °C, 2 min at 96 °C, 4 min at 65 °C, 1 min at 96 °C, 2 min at 65 °C and 30 s at 96 °C).
PCR and sequencing
Amplification of the whole internal transcribed spacer region (ITS), partial region of the large subunit of the nuclear ribosomal DNA (LSU), and the translation elongation factor 1-α (TEF1), was undertaken. The primers used were ITS1F (Gardes & Bruns 1993) and ITS4 (White et al. 1990) (ITS), NL1 and NL4 (O’Donnell 1993) or LR0R and LR3 (LSU), TEF1-983F and TEF1-2218R (Rehner & Buckley 2005) (TEF). All PCR amplifications were performed in 20 μL volumes consisting of 1 µL of DNA template, 6 pmol of each primer, 10 µL of MegaMix-Royal (Microzone Ltd.), and made up to 20 µL with Molecular Grade water (Cytiva, Amersham, UK). Visualisation of the PCR amplicons was achieved on a 1.5 % (w/v) agarose gel in 0.5 × TBE [Tris-Borate-ethylenediaminetetraacetic acid (EDTA) buffer] stained with SafeView Nucleic Acid Stain (NBS Biologicals Ltd., UK). All PCR amplicons were then purified using MicroCLEAN (Microzone Ltd.), per manufacturer’s instructions. Purified amplicons were sequenced using the Big Dye™ Terminator v. 3.1 cycle sequencing kit with 10 pmol of the same primers used for PCR, processed separately. Excess unincorporated dye terminators were removed with AutoSeq™ G-50 (Cytiva) resin columns as per the manufacturer’s instructions, and suspended in 16 µL of Hi-Di™ Formamide (ThermoFisher Scientific, Hemel Hempstead, UK), followed by sequencing on an ABI 3130 Genetic Analyzer (ThermoFisher Scientific).
Molecular phylogeny
Nucleotide sequences, from both forward and reverse primers (.ab1 files), were de novo assembled and edited in Geneious Prime v. 2024.0.5 (Kearse et al. 2012). Additional sequences of SSU, ITS, LSU, TEF, RPB1 and RPB2 from related species in the Cordycipitaceae, selected by Araújo et al. (2022) and Mendes-Pereira et al. (2023), were downloaded from GenBank for phylogenetic analyses (Table 1). Each locus was aligned individually using MAFFT (Katoh & Standley 2013) in Geneious Prime v. 2024.0.5 (Kearse et al. 2012) and further concatenated into a dataset composed of 6 292 bp. The dataset consisted of 14 partitions, one for SSU and LSU and three for ITS (ITS1+5.8S+ITS2), TEF, RPB1 and RPB2. A RAxML analysis was performed using RAxML-HPC2 v. 8.2.12 (Stamatakis 2006, 2014) on XSEDE (http://www.phylo.org/), with the GTRGAMMA+I model and 1 000 bootstrap iterations were executed to evaluate the support. Additionally, we have performed Bayesian analysis in MrBayes v. 3.2.6 (Ronquist et al. 2012). The best-fit models for each partition were implemented as partition-specific models within partitioned mixed-model analyses. The substitution matrix, base frequencies, and gamma shape parameter were unlinked between data partitions, and the rate prior was set to variable (prset applyto = (all), ratepr = variable), allowing partitions to have different rates. Markov chain Monte Carlo (MCMC) runs using default parameters resulted in a low proportion of accepted swaps between adjacent Markov chains, typically less than 5 %, and independent runs did not converge after 40 M generations. Therefore, the number of attempted chain swaps was increased to 2 (nswaps = 2); two independent runs were set, each with eight simultaneous chains, temperature increment parameter set to 0.2, for 70 M generations, sampling trees every 1000th generation. The convergence diagnostic was calculated every 10 000th generation. We verified the minimal effective sample sizes (ESS > 200) and the convergence between the runs with Tracer v. 1.7 (Rambaut et al. 2018). The first 50 % of trees from each run were discarded as a burn-in, and the 50 % majority-rule tree with branch lengths and Bayesian posterior probabilities (BPPs) were calculated from the remaining trees. A BPP value above 0.95 was considered significant. New sequences were submitted to GenBank (Table 1). The final Newick output file from the ML analysis, was exported to Dendroscope (Huson & Scornavaca 2012) for tree layout and then edited in Adobe Illustrator.
Table 1.
Species used in this study, voucher information and GenBank accession numbers. The new species is indicated in bold text.
Morphological examination
Macro-morphological characters were based on dried material and photographed using a digital Sanyo D5100 camera. Micro-morphological characters were examined under a compound microscope (Nikon Optiphot-2, Japan), after removing synnemata with a sterilised needle and mounting them in lactophenol cotton blue or lactofuchsin.
Because of the slow establishment and growth rate, colony characteristics were observed over a 3-mo period on two kinds of media (PCA, MEA) maintained in the dark in an incubator at 20 °C.
RESULTS
Phylogenetic analysis
Our recovered topology agrees with previously published work on Cordycipitaceae fungi (Kepler et al. 2017, Araújo et al. 2022, Mendes-Pereira et al. 2023). In Fig. 1, we present the major hypocrealean groups of araneopathogenic fungi, Jenniferia, Hevansia and Gibellula. Our results show that the original specimen (IMI 507230), observed and reported on during the BBC Winterwatch and Springwatch series, groups within the genus Gibellula clustering with other collections on two orb-spider species from cave systems on the island of Ireland (IMI 507600, IMI 507601; Fig. 1), and is named herein as Gibellula attenboroughii sp. nov. Our newly proposed species is strongly supported and falls within the G. pulchra complex, but its relationship with close relative species, such as G. flava, is still speculative.
Fig. 1.
Maximum likelihood tree, obtained with a concatenated dataset of ITS, SSU, LSU, TEF, RPB1 and RPB2, showing the clade containing the major spider pathogenic hypocrealean genera Jenniferia, Hevansia and Gibellula. On the left, the circular phylogeny represents the full dataset used in this study (Cordycipitaceae). The newly proposed species, Gibellula attenboroughii, is highlighted in bold.
Taxonomy
Based on morphology, and supported by the molecular data, a new species in the genus Gibellula is proposed:
Gibellula attenboroughii H.C. Evans & J.P.M. Araújo, sp. nov. MycoBank MB 854973. Figs 2–10.
Fig. 2.
Gibellula attenboroughii, holotype IMI 507230, on orb-weaving spider, Metellina merianae. A, B. On spider host after removal and drying, showing the multiple synnemata. C. Long conidiophores scattered along lower part of synnema. D. Short conidiophores densely clustered in terminal part of synnema. E, F. Dense, white spore columns on short conidiophores at apex (E) and long conidiophores at base (F). Scale bars: A, B = 1 cm; C, D = 50 µm; E = 100 µm; F = 25 µm.
Fig. 3.
Gibellula attenboroughii, holotype IMI 507230, conidiophore morphology. A. Synnema with almost astipitate conidiophores clustered towards the apex. B, C. Conidiophores towards the base with long, roughened stipes and short, smooth neck region. D. Conidia. Scale bars: A = 40 µm; B = 50 µm; C = 25 µm; D = 10 µm.
Fig. 4.
Gibellula attenboroughii, holotype IMI 507230, in culture. A, B. Colony after 6 wk on MEA (A) and on PCA (B). C. Colony after 4 mo on MEA producing clusters of fertile synnemata (arrow) on pseudostroma. D. Detail of synnema with short conidiophores at apex (short arrow) and long conidiophores at base (long arrow), as on host. Scale bars: A, B = 3 mm; C = 1.5 mm; D = 60 µm.
Fig. 5.
Gibellula attenboroughii on Metallina merianae, paratypes in situ. A. On cave ceiling, Tullybelcoo Ground Bridge, County Fermanagh, Northern Ireland. B. Whitefathers’ Caves, County Cavan, Republic of Ireland. C. On moss, Lake Vyrnwy, Powys, Wales (image: D. McNeil). Scale bars: A, B = 1.5 mm; C = 1 mm.
Fig. 6.
Gibellula attenboroughii paratypes, on orb-weaving spider Meta menardi. A. Paratype IMI 507601, in situ, attached to cave wall above River Barran, with multiple white synnemata covering dorsal surface, Whitefathers’ Caves, County Cavan, Republic of Ireland. B. After drying to show multiple creamish synnemata on ventral abdomen. C. Paratype IMI 507603, after drying; note the free conidiophores on the legs and the naked head region with prominent eyes arrow. D. Paratype IMI 507602, after drying, hanging from a thread above river, showing abundant synnematal production on both dorsal and ventral surfaces. Scale bars: A = 4 mm; B–D = 3 mm.
Fig. 7.
Gibellula attenboroughii, micro-morphology on Meta menardi, paratype IMI 507601. A. Detail of upper part of synemma showing tapering apex and densely-crowded, penicillioid conidiophores (compare Fig. 2D). B. Conidiophores showing short, lightly-roughened stipes and simple penicillioid heads. C. Detail of conidiophore head with prominent, smooth neck region. D. Conidial chain. Scale bars: A = 50 µm; B–D = 10 µm.
Fig. 8.
Gibellula attenboroughii, paratype IMI 507601, ex Meta menardi, in vitro. A. Colony after 3 mo on PCA with aspergilliform conidiophores produced directly from the mycelium. B. Chlamydospore-like structures and verrucose hyphae from the subiculum. C. Aspergilliform conidiophore arising from the mycelium. D. Aspergilliform and penicillioid conidiogenous heads on the mycelium. Scale bars: A = 3 mm; B–D = 15 µm.
Fig. 9.
Gibellula attenboroughii, habitat and paratype IMI 507598, on Meta menardi. A. Whitefathers’ Caves, showing the dark zone above the River Barran, County Cavan, Republic of Ireland. B. Entrance and twilight zone. C. Infected spider in situ, on cave ceiling, showing early infection with immature synnemata on compact, white subiculum, the specimen was found immediately above the figure in the centre. D. Dried specimen, showing creamish-yellow subiculum covering ventral abdomen. Scale bars: C = 0.5 mm; D = 0.25 mm.
Fig. 10.
Gibellula attenboroughii in situ on orb spiders and mycoparasite macro-morphology. A. On orb spiders, Metellina merianae (left, short arrow), and Meta menardi (right, long arrow), on cave ceiling, Whitefathers’ Caves; showing differences in synnematal morphology. B. Same site, with Meta menardi completely overgrown by nodulosporium-like mycoparasite, forming a ‘slimy blob’. C. Dried specimens showing the larger Meta menardi on left and smaller Metellina merianae on right. D. Mycoparasite after 1 mo on PCA, forming slimy colonies. Scale bars: A, B = 5 mm; C = 2 mm; D = 10 mm.
Etymology: Named after the broadcaster and natural historian Sir David Attenborough, a pioneer of BBC natural history programmes, who – in his role as controller of BBC 2 – helped to develop the Natural History Unit; leading, indirectly, to the present nature series during which the new species was first discovered.
Typus: UK, Northern Ireland, County Down, Comber, Castle Espie Wetland Centre, on orb-weaving spider Metellina merianae (Tetragnathidae: Araneae), gunpowder store ceiling, 11 Feb. 2022, J. Clark & M. Turley (holotype dried down culture, metabolically inactive, IMI 507230; culture ex-type IMI 507230).
Paratypes: UK, Northern Ireland, County Fermanagh, Tullybelcoo Ground Bridge, on Metellina merianae, cave roof, 19 Jun. 2022, T. Fogg, IMI 507599. Republic of Ireland, County Cavan, Blacklion, Whitefathers’ Caves, on Metellina merianae, cave roof, 19 Jun. 2022, T. Fogg, IMI 507600; County Cavan, Blacklion, Whitefathers’ Caves, on orb-weaving spider Meta menardi (Tetragnathidae: Araneae), cave roof, 19 Jun. 2022, T. Fogg, IMI 507598; County Cavan, Blacklion, Whitefathers’ Caves, on Meta menardi, on cave wall, 17 Oct. 2023, T. Fogg, IMI 507601–507603; County Cavan, Blacklion, Whitefathers’ Caves, on Metellina merianae, T. Fogg, IMI 507604.
Description: Spider body in situ completely covered by a dense, white mycelial mat (Fig. 2A); becoming creamish yellow on drying (Fig. 2B). Synnemata cylindrical, numerous, from all body parts including legs, white to cream, swollen and floccose at the base, up to 250 µm diam (Fig. 2C); tapering towards the tip, 3–7(–10) mm long, 140–200 µm wide, terminal part often swollen and ovoid or club-shaped (Fig. 2D, E). Conidiophores produced along the entire length of the synnemata; scattered at the base with long stipes, 80–120 × 5–8 µm (Figs 2C, 3B); often becoming conspicuously crowded towards the tip and with short stipes, (10–)20–30 × 6–8 µm (Figs 2D, 3A); heads aspergilliform, occasionally with simple penicillioid heads and almost astipitate; distinctly roughened wall; septa conspicuous, 1–2; constricting to a short, smooth neck region or neck indistinct to absent; terminating in a swollen vesicle (Fig. 3B, C). Vesicles spherical to broadly obovoid, (8–)10–12(–13) µm diam. Metulae borne on vesicle, broadly obovoid or broadly ellipsoid, (8–)10–12 × (4–)6–8 µm, bearing multiple phialides. Phialides cylindrical to narrowly clavate, often apically thickened, 7.5–9.5 × 2.5–3.5 µm. A vesicle together with metulae and phialides forming a spherical to ovoid head measuring, (28–)32–40 × 24–40 µm diam; occasionally, much reduced in size and complexity. Conidia hyaline, smooth, ellipsoidal to fusoid, 4–6 × 1.5–2 µm (Fig. 3D), often adhering in long chains and forming compact, white blocks (Fig. 2E, F). Sexual morph and Granulomanus synasexual morph not observed.
In culture: On MEA, slow-growing, 0.7–0.9 cm diam after 6 wk at 20 °C, white, stromatic; turning creamish to pale brown, raised centrally, often furrowed, brown diffusate around periphery, dark brown reverse after 12 wk, 1.8–2.0 cm diam (Fig. 4A); on PCA, slower growing (1.3–1.5 cm diam after 12 wk), dark cream with a greyish brown reverse (Fig. 4B); after 4 mo becoming feathery and irregular at periphery and up to 3.0 cm diam, forming chains of small, hyaline chlamydospores; occasionally developing short synnemata centrally (Fig. 4C), morphologically identical to those on the host with short conidiophores crowded at tip and longer, scattered conidiophores at base (Fig. 4D).
Paratype IMI 507601, ex Meta menardi, on PCA, slow-growing, 0.5-0.7 cm after 6 wk at 20 °C, white to creamish, compact, raised centrally, brown diffusate; dark brown reverse. After 3 mo, 1.8–2.0 cm, raised and cream to buff brown; producing chains of dark red chlamydospores (Fig. 8B) and scattered, typically penicillioid, conidiophores with chains of pale lilac conidia either directly from the mycelium (Fig. 8A), or from rudimentary synnemata. However, atypical aspergilliform conidiophores also occurred sporadically around the periphery (Fig. 8C, D).
Notes: The holotype shows distinct features in which there is a general lack of pigmentation compared to the paratypes on the same spider host, Metellina merianae, which have a dense yellow mycelial covering or subiculum, with yellow synnemata and pale lilac spore heads. The compact conidial columns or blocks of spores are less evident in these paratypes. The paratypes on the much larger spider host Meta menardi, in which such dense spore columns are rarely observed, the conidiophores are much shorter with simple, penicillioid conidiogenous heads, formed on tapering, fir-tree-like synnemata; contrasting with the complex aspergilliform heads produced on the stouter, less tapered synnemata occurring on Metellina merianae (see Figs 3A, 5, 6, 7A). In fact, before the availability of the molecular data, it was considered that these morphological differences merited separation at the species level.
It is posited here that these differences are due to ecotypic variation with the complete absence of light and air movement within the underground gunpowder store leading to loss of pigmentation and the spores adhering in columns. Within the cave system, especially in the threshold zone favoured by Metellina merianae, there would be both diffuse light and air currents to dislodge and disperse the conidia resulting in a decreased incidence of long chains or blocks of spores. The simpler, more open and penicillioid conidiogenous heads usually forming on Meta menardi would lead to less adhesion of the spores and the general absence of spore columns or blocks. In old cultures derived from the paratypes on Meta menardi, rudimentary synnemata bearing short penicillioid conidiophores were observed, as well as long conidiophores with complex aspergilliform heads producing spore blocks arising directly from the mycelium (Fig. 8D).
Ecology
Although Metellina merianae was moved to this genus many years ago (Bristowe 1939), some authors still maintain it in the genus Meta together with M. menardi (Roberts 1995). The latter author even stated that “this move is incorrect; it may well be reversed in the future and is not followed here”. However, the latest molecular phylogenetic study of metaine spiders (Metainae; Tetragnathidae) shows that Meta menardi and Metellina merianae are not congeneric within the sub-family Metainae (Kallal & Hormiga 2018). These two orb-spider species occupy overlapping niches within cave systems and, typically, Metellina merianae inhabits the shallow threshold or twilight zone in and around cave entrances (Novak et al. 2010, Hesselberg et al. 2019), which is where the infected paratypes were found. Meta menardi, however, is more prevalent in the so-called dark zone but has an optimal adaptation to the epigean/hypogean ecotone, according to Novak et al. (2010), so that the two species can and do occur together (Mammola & Isaia 2018). In fact, in one instance, infected Meta menardi and Metellina merianae specimens were found ca. 3 cm apart on a cave wall, some 20 m from the entrance (see Fig. 10A).
Metellina merianae also favours man-made habitats, such as culverts and cellars, and, in the case of the holotype, in a gunpowder store. In all instances, the infected spiders had moved from their concealed lairs or webs and died exposed on the cave roof or wall and the store ceiling. Similarly, this host when infected by Gibellula was also found in an exposed position around the rocky shores of upland lakes in Wales (McNeil 2012). At the time, the fungus was tentatively identified – based on the received images (see Fig. 5C) – as being close to G. leiopus (H.C. Evans, pers. comm., 2012). No specimens were received for examination and none have been deposited officially but it is considered highly probable that this record on Metellina merianae from Wales is the new species G. attenboroughii; having similar densely-packed, white to pale yellow synnemata, although the stipes are distinctly pigmented and tan-coloured towards the base.
Infected specimens of Meta menardi, or the European cave spider, were collected freely exposed on the cave ceiling, typically, within the dark zone (T. Fogg, pers. comm., June 2022; see Figs 6, 9C). Meta menardi, like Metellina merianae, is a sit-and-wait predator – building a similar planar orb-web – but can also adapt to off-web foraging (Novak et al. 2010, Hesselberg et al. 2019). However, the life-style of both spider species in cave systems is cryptic, often concealing themselves in close proximity to their webs. The fact that Gibellula-infected spiders are found in prominent positions on the roof or ceiling of their subterranean habitats indicates a behavioural change, possibly manipulated by the fungus, in which the sporulating cadavers would be exposed to the air currents circulating through the caves promoting the release and subsequent dispersal of the dry spores through the system. In the case of the suspected infection of Metellina merianae by G. attenboroughii in Wales (McNeil 2012, Evans 2013, Hughes et al. 2016), the spiders appear to have moved from the entrances of rock fissures or similar natural or man-made niches, to die on the surrounding or overhanging sphagnum moss, as there are no cave systems in this lakeside locality (H.C. Evans, pers. obs., September 2022).
The ecological significance of the vastly-different sporulating heads – complex and aspergilliform on Metellina merianae; simple and penicillioid on Meta menardi – is difficult to interpret. Undoubtedly, the aspergilliform head produces significantly more spores than the more open penicillioid form and this may relate to the host niche within the cave with Metellina merianae being exposed to a different microclimate – such as drier conditions and increased air movement, especially when the infected host dies around or outside the cave entrance – compared to Meta menardi occupying the deeper recesses of the cave.
Mycoparasites
Mycoparasites were also observed in situ on Gibellula-infected spiders within the cave systems; being described originally as “blobs” (T. Fogg, pers. comm., Fig. 10B). Culture isolates taken from them were identified as close to Nodulisporium (Fig. 10C, D); whilst others were found and subsequently isolated from dried specimens following examination under a stereoscopic microscope. These included engyodontium- and lecanicillium-like isolates, with the latter also producing a torrubiella-like morph in culture.
Historical overview
The entomopathogenic mycobiota of the British Isles has never been studied systematically and most of the knowledge is based on the work of Tom Petch in the 1930s and 1940s, which was covered in his presidential address to the Yorkshire Naturalists’ Union (1932a) and culminated in a list of “British entomogenous fungi” (Petch 1932b), with a subsequent revision (Petch 1948). Later, these lists were updated in a series of entomological publications based on new records which appeared in the literature, as well as in foray reports, but these were catalogued and indexed under the arthropod host rather than the fungal pathogen (Leatherdale 1958, 1962, 1966, 1970). This resulted in a final list of 106 fungal taxa of which 16 species were assigned to spider (Araneae) hosts, with only a single species of Gibellula, G. “aranearum”, being recognised.
Many of the “post-Petch” fungal records on spiders listed by Leatherdale (1970) were based on reports by amateur collectors in the publications of local natural history societies in England; most notably, in East Anglia (Ellis 1956) and the Midlands (Evans 1967). There have been few publications since, with the exception of that by an amateur collector in the parochial newsletter Shropshire Entomology (McNeil 2012), which merited attention because of the “astronomical” number of specimens collected of: “A small but quite spectacular spider pathogen [Gibellula cf. leiopus] which produced 102 specimens from Lake Vyrnwy (mid-Wales) on a visit in December 2011” (McNeil 2012; see Fig. 5C). A similar number of specimens were also collected from the nearby Lake Bala (D. McNeil, pers. comm., 2012). These records are especially intriguing; not least, as it was in the depths of winter in a temperate mountainous area and Gibellula is a genus typically associated with spider hosts in the tropics and sub-tropics (Evans 1982, Evans & Samson 1987, Samson et al. 1988, Samson & Evans 1992, Evans 2013, Hughes et al. 2016, Shrestha et al. 2019, Kuephadungphan et al. 2020, 2022, Mendes-Pereira et al. 2023). The latter authors accepted 31 species in the genus – although, as stated earlier, significantly more are listed in Index Fungorum and MycoBank – many of which have been described in recent years from Asia and a far cry from the eight species listed in the original monograph on Gibellula (Samson & Evans 1992).
The earliest record of Gibellula in the British Isles would appear to be by Cooke (1892), who listed Isaria arachnophila under the common name, “European Spider Isaria”, but without locality details. Petch (1931, 1932c) considered this to be a synonym of Gibellula “aranearum” and he confirmed British records of this species on spiders in the counties of Cambridgeshire, Hampshire, Somerset, Surrey and Worcestershire. In his revised list (Petch 1948), he also included multiple records from Norfolk and Suffolk where one of the hosts was identified as the red-legged spider, Gongylidium rufipes (Linyphiidae). Petch (1948) also listed G. rufipes as a host of Torrubiella albolanata in the Norfolk Broads with the purported asexual state described as Cylindrophora aranearum both of which had been described earlier as new species (Petch 1944). Later, Ellis (1956) expanded on his collections from Norfolk where this species was said to be common in marshes throughout the county making “upwards of ninety collections of this fungus regularly from May to October each year from 1942 to 1956”; adding that he had made recent collections in Bedfordshire and Berkshire. These collections from various broads in Norfolk have been examined in Herb K and the host is always a small spider (2–3 mm) with distinctive reddish legs, matching the description of Gongylidium rufipes, with the ascomata buried in a white subiculum and an absence of Gibellula synnemata (Fig. 11A). The same host-pathogen association also appears to be common in mainland Europe, with Nyffeler & Hywel-Jones (2024) reporting the occurrence of T. albolanata in Denmark and illustrating fresh specimens showing yellow perithecia buried in a snow-white subiculum. Later collections made by Ellis from Cumbria and Devon were also examined but one of the Devon specimens is atypical, with the ascomata buried amongst lilac-coloured Gibellula synnemata rather than a Granulomanus subiculum. Cylindrophora aranearum has since been recognised as belonging to the genus Granulomanus, a synasexual morph of Gibellula (Samson & Evans 1977, de Hoog 1978, Samson et al. 1988). According to Index Fungorum and MycoBank, Torrubiella albolanata is a legitimate or current name; whilst Cylindrophora aranearum and Granulomanus aranearum are considered to be synonyms of Gibellula arachnophila by Index Fungorum. In contrast, MycoBank recognises Granulomanus aranearum as a legitimate species with Cylindrophora aranearum as a synonym.
Fig. 11.
New Gibellula combinations. A. Gibellula albolanata comb. nov., type of Torrubiella albolanata on the red-legged spider, Gongylidium rufipes (Linyphiidae), fen sedge, Norfolk, 1942 (ex Herb K), showing the yellow ascomata (arrow) embedded in a white subiculum. B. Type illustration of Isaria cuneispora on Gnaphosa lucifuga (Gnaphosidae), from France (Boudier 1887), showing the Granulomanus synasexual morph of G. albolanata. C. Gibellula aranicida comb. nov., labelled as Torrubiella aranicida, from Yorkshire, 1936 (ex Petch collection, Herb K), showing the yellow-orange ascomata scattered on the spider legs and body. D. Type illustration of Torrubiella aranicida, on Gnaphosa lucifuga, France (Boudier 1885). E, F. Gibellula aranicida on the funnel-web spider, Coelotes atropos (Amaurobiidae) collected by G. Lyon, under a log, Devon, 2023, dried fungarium specimen, IMI 507605; note the scattered yellow ascomata. F. Close-up of the spider legs taken in situ, showing the white or pellucid ascomata in the fresh specimen (image, G. Lyon), as described by Petch (1944). Scale bars: A = 1 mm; C, E = 2 mm; F = 0.6 mm.
Another Torrubiella species, T. aranicida also occurs on spiders in the Norfolk fens and, like T. albolanata, it lacks the Gibellula asexual morph (Petch 1948, Ellis 1956). In the Petch collection at RBG Kew, the prominent orange-coloured ascomata are scattered superficially on the body and legs of a large spider (10–12 mm) collected “under hanging moss on a cliff face” in Yorkshire (Petch 1948; see Fig. 11C): a significantly bigger spider than Gongylidium rufipes (2–3 mm), the host of T. albolanata (Fig. 11A). The latter species has also been recorded on another Linyphiidae spider, Leptorhoptrum robustum from sewage filter beds in Warwickshire (Duffy 1997). And, more recently, a species close to T. aranicida has been collected on the funnel-web spider, Coelotes atropos (Amaurobiidae), under a log in Devon (G. Lyons, pers. comm. 2023; IMI 507605). This large spider (9–10 mm) has only traces of the Granulomanus synasexual morph on the body, which is characterised by the translucent to pale cream ascomata (Fig. 11F), which turn a yellow orange colour when dried (Fig. 11E). Petch (1944) noted this colour change, describing the fresh ascomata as “pellucid, but dull orange when dry”. In the type diagnosis of T. aranicida – on the spider Aranea lucifuga (= Gnaphosa lucifuga, Gnaphosidae) from France – the ascomata are described as “ochracea vel ochracea-aurantiaca” or yellowish-brown to orange (Boudier 1885; see Fig. 11D). Later, Boudier (1887) returned to the same site but earlier in the season and reported an asexual morph on the same spider host which he described and illustrated as Isaria cuneispora sp. nov. (Fig. 11B). Petch (1944) considered this to be a species of Cylindrophora and, possibly, C. aranearum as associated with Torrubiella albolanata: now accepted as a synonym of Granulomanus (de Hoog 1978). Although it would appear that the Gibellula morph is rarely or never produced in either Torrubiella albolanata or T. aranicida, the genus Gibellula is now considered to be the legitimate generic name in the one fungus-one name classification (Kepler et al. 2017), with Granulomanus and, therefore, Torrubiella as synonyms.
In accordance with the Code protocol, the following new combinations are proposed:
Gibellula albolanata (Petch) H.C. Evans, comb. nov. MycoBank MB 856522.
Synonyms: Torrubiella albolanata Petch, Trans. Brit. Mycol. Soc. 27: 85. 1944.
Cylindrophora aranearum Petch, Trans. Brit. Mycol. Soc. 27: 85. 1944.
Granulomanus aranearum (Petch) de Hoog & Samson, Persoonia 10: 70. 1978.
Gibellula aranicida (Boud.) H.C. Evans, comb. nov. MycoBank MB 856523.
Synonyms: Torrubiella aranicida Boud., Revue mycol., Toulouse 7: 227. 1885.
Isaria cuneispora Boud., Revue mycol., Toulouse 9: 158. 1887.
DISCUSSION
Spiders play a key role in ecosystem functioning (Nyffeler & Birkhofer 2017) and, therefore, their parasites and pathogens should warrant especial attention. However, until recently, their associated mycobiota – the entomogenous, entomopathogenic or, more accurately, araneopathogenic fungi – has largely been ignored. Evans (2013) discussed this “no-man’s land” between mycologists and arachnologists where, historically, there has been little or no attempt by either group of taxonomists to interpret host-pathogen interactions and to identify both the organisms involved. In the last decade, this situation has changed and more emphasis is now being given to host identification and the ecological significance of these fungi, most notably, in relation to the genus Gibellula (Hughes et al. 2016, Shrestha et al. 2019, Kuephadungphan et al. 2020, 2022, Durkin et al. 2021, Mendes-Pereira et al. 2023, Nyffeler & Hywel-Jones 2024).
Within the species in the genus Gibellula described, thus far, the greater majority produce only the asexual morph. In a few species, however, only the sexual morph and the Granulomanus synasexual morph are found. Within the small sample size recorded here from the British Isles, all three morphs are represented: as, for example, in Gibellula albolanata which produces perithecia embedded in a subiculum of Granulomanus and, G. aranicida in which ascospores appear to be the dominant spore-dispersal form; contrasting with G. attenboroughii where spore density is maximised in the aspergilliform Gibellula heads. The evolutionary pressures to either limit or optimise spore production – or, indeed, to opt for asexual vs sexual reproduction – can only be speculated upon, as is the form and function of the Granulomanus synasexual morph. The diversity of the Gibellula morph was highlighted and illustrated, as “Variations on a theme”, by Evans & Samson (1987). Since then, the number of described species has burgeoned, as has the range of Gibellula forms (Samson & Evans 1992, Evans 2013, Shrestha et al. 2019, Kuephadungphan et al. 2020, 2022, Mendes-Pereira et al. 2023). Doubtless, there are many more species to be found with equally complex and bizarre morphologies.
With the benefit of host identification, it has become possible to better understand the spider-pathogen interactions and, crucially, to assess the impact of infection on host behaviour. Earlier studies on ant-fungal interactions led to the concept of zombie-ant fungi (Evans 2002, Evans et al. 2011, Bekker et al. 2014, Hughes et al. 2016, Araújo et al. 2018, Araújo & Hughes 2019). Hughes et al. (2016) provided the first circumstantial evidence that infected spiders also change behaviour and move from their characteristic concealed niches to die in elevated or freely-exposed situations, typically, on the undersides of leaves in the understorey of tropical forests. This behavioural manipulation has since been reported by Arruda et al. (2021), Durkin et al. (2021) and Mendes-Pereira et al. (2022) for Gibellula-infected spiders in the humid tropics and sub-tropics. Here, we posit that temperate cave spiders also alter their behaviour when infected by Gibellula species facilitating the release and dispersal of fungal spores and, in effect, replicating the manipulation of ants by Ophiocordyceps species.
Finally, the data unearthed during the herbarium and literature searches indicate that there is a hidden diversity in the British Isles and that many more species of Gibellula remain to be discovered. Moreover, the ecological impact on their spider hosts could be significant with epizootics being reported in Norfolk and Wales (Ellis 1956, McNeil 2012) – similar to those recorded on spiders in the humid tropics (Evans 1974, 1982, Samson & Evans 1973, 1977, Mendes-Pereira et al. 2022, 2023) – and these have now been analysed in detail by Nyffeler & Hywel-Jones (2024). Thus, their role in spider-population dynamics warrants further study, as does the metabolites they produce which enable them to exploit such a highly-specific ecological niche.
ACKNOWLEDGEMENTS
We wish to thank J. Clark and M. Turley of the Castle Espie Wetland Centre for collecting and sending the type specimen and L. Davis for facilitating access to the RBG Kew Fungarium. Confirmation of the spider identifications by D. Sherwood (NHM, London) is gratefully acknowledged.
Conflict of interest:
The authors declare that they have no conflict of interest.
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