Integrative taxonomy, larval biology and functional morphology of the little known gall-forming coral endoparasite Petrarca (Thecostraca: Ascothoracida)

Abstract

Petrarca is an ascothoracidan endoparasite in scleratinian dendrophyllid corals. Petrarca can stimulate coral growth, forming a gall chamber to house itself inside the coral skeleton. The diversity, molecular phylogeny and feeding ecology of Petrarca are understudied. This is the first study of Petrarca to be based on light and scanning electron microscopy to document the fine-scale external and functional morphology of its trophi and other structures. A combined molecular and morphological approach revealed at least four closely related species of Petrarca, P. goanna, P. morula, Petrarca nozawai sp. nov. and Petrarca rubus sp. nov. in Turbinaria and Astreopora corals in Asia. Carapace shape and fine morphology, ultrastructure of the antennular aesthetasc, morphological characteristics of the trophi and the shape and size of the penis rami are diagnostic characters. Several morphological characters, which are all probably synapomorphies, have been proposed to distinguish the genus Petrarca from other Ascothoracida. Mouthparts morphology of Petrarca are developed for cutting and chewing, rather than for piercing and sucking as in many other Ascothoracida. The external surface of the carapace of Petrarca is ornamented with densely packed secretory papillae used putatively for the chemical dissolution of the substrate necessary for the formation of the gall chamber.

INTRODUCTION

Ascothoracida are a relatively poorly studied group of Thecostraca that exclusively encompasses parasitic crustaceans that range from ecto-, meso- to endoparasites found on cnidarians (Alcyonacea, Antipatharia, Scleractinia and Zoantharia) and echinoderms (Asteroidea, Crinoidea, Echinoidea and Ophiuroidea) (see: Chan et al., 2021). Currently, Ascothoracida comprise 115 described species assigned to two orders (Grygier, 1987a, 1996): Laurida, species of which are parasites of anthozoans, except for those of Waginella Grygier, 1983, which are ectoparasites of crinoids, and Dendrogastrida, parasites of non-crinoid echinoderms. The body of Ascothoracida is covered by a bivalved carapace (Chan et al., 2021). The valves, usually observed as partially fused in attached females, contain the gonads and gut diverticula. In the ground plan, but with significant exceptions, depending on the degree of modification for parasitism, the trunk consists fundamentally of 11 segments (somites): six thoracic segments bearing biramous, natatory thoracopods and five well-developed abdominal segments, including the telson, which supports a pair of unsegmented furcal rami. The first abdominal segment bears a developed penis in males or hermaphrodites, a conspicuous vestige of which persists in females. The antennules are prehensile to subchelate and generally comprise four to six segments, a claw-like finger being developed on the distal one. The labrum surrounds the mouth parts to form an oral cone or pyramid that also often contains an anterior unpaired process or medial languette. Basically, they have three pairs of piercing mouth parts (mandibles, maxillules and maxillae), but that number is often reduced in advanced species.

The life cycle of ascothoracidans varies among species, and the larval stages can comprise free-swimming naupliar larvae or brooded larvae inside the mantle of adults. It potentially includes up to six planktotrophic or lecithotrophic naupliar instars, albeit being one of the few species where the naupliar phase is condensed or even omitted (Høeg et al., 2014), followed by one or two instars of a specialized non-feeding ascothoracid larva that infects the host. The ascothoracidans are largely dioecious, with larger females and smaller, sometimes dwarf cypridiform males (Grygier & Fratt, 1984; Grygier, 1985, 1987b, 1991a, b; Kolbasov, 2007).

Petrarcidae are endoparasites in scleractinian corals and have secondarily evolved hermaphroditism, and represent one of the most advanced group of the order Laurida (Okada, 1938; Grygier, 1983b, 1983c; Fig. 1). Adults of this family are characterized by a bivalved carapace with thick, inflated valves armed with papillae and lacking a brood chamber, five-segmented antennules with feeble armament, shortened mouth legs and uniramous thoracopods and a more or less reduced abdomen (Grygier, 1987a). To date there are 11 species assigned in three genera, PetrarcaFowler, 1889 (eight species), IntrocorniaGrygier, 1983c (two species) and monotypic ZibrowiaGrygier, 1985. They infect different species of azooxanthellate and zooxanthellate scleractinian corals of the family Dendrophylliidae from deep to shallow waters. Petrarcids normally live in pairs inside a chamber in a single spongy gall without an aperture (Fig. 1A–J; Supporting Information, Video S1). Living within a single calice, in the coumella and adjacent septa, they induce skeletal proliferations that are referred to as ‘internal galls’ (Fig. 1C–E, J). Shallow-water dendrophylliid corals form larger, rounded ‘external galls’ in the coenosteum outside the calices (Fig. 1A, B). In comparison with other ascothoracidans, the petrarcids do not brood their embryos between the carapace valves (Grygier, 1985).

Petrarca was established by Fowler (1889) for the abyssal species, described off the coast of Japan, from a depth of 4200 m and found in a solitary coral Fungiacyathus marenzelleri (Vaughan, 1906). This species was also re-described by Grygier (1985) from East Africa and north-eastern Australia. Subsequently, seven congeners were described from different locations and depths, and in different hosts (Grygier, 1981, 1985, 1991a; Grygier & Cairns, 1996; Tachikawa et al., 2020): Petrarca okadaiGrygier, 1981 from HeteropsammiaMilne Edwards & Haime, 1848, north-eastern Australia, Indonesia and East Africa; Petrarca indicaGrygier, 1985 from Flabellum deludensMarenzeller, 1904, Sri Lanka; Petrarca morulaGrygier, 1985; Oken, 1815; Petrarca azoricaGrygier, 1985 from Enallopsammia rostrata (Pourtalès, 1878), North Atlantic; Petrarca sensoriaGrygier, 1991a from Fungiacyathus sp. Sars, 1872, eastern Australia; Petrarca goannaGrygier, 1991a from Turbinaria reniformisBernard, 1896, north-eastern Australia; and Petrarca madreporaeGrygier, 1996 from Madrepora oculataLinnaeus, 1758, Indonesia and Japan.

Grygier (1991a) indicated that: ‘Turbinaria spp. often serve as hosts of Petrarcidae. Numerous examples of petrarcid galls on dried specimens of this coral genus have been found in various museums.’ He provided numerous records of external galls possibly formed by Petrarca in Turbinaria spp. from different localities in the Indo-Pacific (Grygier, 1991a). In particular, though without identification of the parasite, Grygier mentioned such findings in dried Turbinaria spp. in at least three locations in Taiwan (off the south coast of Taiwan and Lanyu/Orchid Island). However, despite all these speculations, only two species, P. goanna and P. morula, were formally described from Turbinaria corals. Moreover, Grygier & Nojima (1995) indicated that these two species may represent ‘the extremes of morphological variability in a single species’ or a third undescribed species of Petrarca that can infect Turbinaria corals.

In the present study, numerous external galls containing Petrarca specimens of more or less similar morphology were found in Turbinaria and AstreoporaBlainville, 1830 corals from various locations in Taiwan and Thailand. An integrative taxonomy approach, which involved molecular and morphological analyses, revealed that these specimens represent at least four species: two previously described, P. goanna and P. morula, and two new species here named Petrarca nozawai and Petrarca rubus, which are described here. In addition to previous descriptions, our study involves for the first time both light and scanning electron microscopy (SEM) to document the fine-scale functional external morphology of Petrarca and to interpret how they form internal galls and feeding modes. We also describe how larvae were released from the galls.

MATERIAL AND METHODS

Sampling and morphological analysis

Live specimens of Petrarca were collected on coral reefs located in the vicinity of Lǜdǎo/Green Island (Taiwan) and Koh Kye Island (Thailand) at depths of 3–20 m. External galls formed by petrarcids (Fig. 1A, B) were removed from the surface of Turbinaria and Astreopora corals (Supporting Information, Video S1) and transported to the Green Island Marine Research Station of Academia Sinica to be placed in aquariums with 23–25 °C sea water. Coral galls were kept in filtered seawater for 1 day and any larvae released from the parasite were collected. Larvae collected were observed under compound light microscopes (Zeiss Axioplan). After larvae were collected, the coral galls were dissected and parasites were removed from the gall chambers (Fig. 1C–I). Live specimens of Petrarca were photographed using Olympus SZ61 dissecting microscope equipped with a Lumix (Panasonic) GH4 camera with a Leica DG Macro-Elmarit 45 mm f2.8 lens (Fig. 1C–I) then subsequently fixed in 95% alcohol, formalin and glutaraldehyde.

Specimens of Petrarca were studied using light and scanning electron microscopy. For light microscopy, to make chitinous structures clearer, the isolated specimens, including holotypes, were treated with a 10% KOH solution heated using a water bath. The carapace of treated specimens was then removed to observe the body morphology. Carapace valves, trunk, dissected antennules, mouthparts and all thoracopods (see Fig. 1J for morphological parts) were mounted in glycerol on glass slides and examined using an Olympus BX 43 light microscope. Line drawings were made using a drawing tube on the same microscope.

For SEM examination of intact and dissected specimens (one to three for each species), specimens were post-fixed in 2% OsO₄ for 2 h, dehydrated in acetone and critical-point dried from CO₂ in a Hitachi HCP-2 critical point dryer. Dried specimens were sputter-coated with platinum-palladium in an Eiko IB-3 Ion Coater and examined by GAK in a JEOL JSM-6380LA scanning electron microscope at Moscow State University. The resulting photographs were touched up using CorelDraw X3 Graphics Suite, which was also used to prepare all the figures presented here.

DNA extraction, amplification and sequencing

Twenty-five specimens of Petrarca collected in Thailand and Taiwan were included in the molecular analysis (Table 1). Total genomic DNA was extracted from the muscle tissue using Qiagen DNeasy Blood & Tissue Kits (Qiagen, CA, USA) according to the manufacturer’s instructions. Partial sequences of mitochondrial DNA markers, cytochrome c oxidase subunit 1 (COI), were amplified by using the following primers: forward (5ʹ-GCT TGA GCT GGC ATA GTA GG-3ʹ) (Roman & Palumbi, 2004) and reversed (5ʹ-TAD ACT TCD GGR TGD CCA AAR AAY CA-3ʹ and 5ʹ-TAA ACT TCA GGG TGA CCA AAA AAT CA-3ʹ) (Folmer et al., 1994; Schubart & Huber, 2006). Aliquots of 3 μL of DNA template were mixed with 22 μL of reaction buffer including 0.5 μL of each primer, 5 μL of Fast-RunTM Taq Master with Dye (Protech Technology Enterprise Co., Ltd, Taipei City, Taiwan) and 16 μL of ddH₂O. Polymerase chain reaction (PCR) reactions were conducted in a DNA Engine Thermal Cycler (Bio-Rad, Richmond, CA, USA), and the products were checked by electrophoresis on 1% agarose gel in 1 × TAE buffer. The DNA sequencing was performed by Genomics Bio Sci & Tech Ltd (New Taipei City, Taiwan). The sequences were assembled and edited in GENEIOUS PRIME 2022.1.1 (https://www.geneious.com). All data are deposited in GenBank (accession numbers: OP117451-OP117475) (see Table S1).

Species delimitation

Sequences of Gorgonolaureus sp. (KM087532) and Dendrogaster tobasuii (MN913413) downloaded from GenBank were included in the analysis as the outgroup. All the sequences were aligned with MAFFT in GENEIOUS PRIME 2022.1.1 (https://www.geneious.com). Neighbour-joining (NJ) and maximum likelihood (ML) were conducted in MEGA X (Kumar et al., 2018). For the NJ method, Kimura 2-parameter (K2P) distance model was used. For the ML method, GTR+I+G was selected according to the results of the model test in MEGA X (Kumar et al., 2018). Both methods were conducted with the bootstrap values estimated from 1000 pseudoreplicates. Bayesian inference (BI) were also conducted by using BEAST v.1.10 (Suchard et al., 2018). GTR+I+G model and Yule process were chosen. Three independent runs with 1 × 10⁷ generations of Markov chain Monte Carlo (MCMC) chains were performed, and the data were sampled every 1000 generations. The run convergence was visually verified in TRACER v.1.7 (Rambaut et al., 2018), and the effective sample size (ESS) of all parameters was > 200. The tree files from the three runs were combined and the first 25% were discarded as burn-in using LogCombiner 1.10 (BEAST package). The final result was produced by using Tree Annotator 1.10 (BEAST package). K2P genetic distances were also calculated in MEGA X (Kumar et al., 2018).

Assemble species by automatic partitioning analysis (ASAP), Poisson tree processes (PTP) model and generalized mixed Yule coalescent (GMYC) were used to confirm the species delimitation (Pons et al., 2006; Zhang et al., 2013; Puillandre et al., 2021). ASAP method builds the species partitions based on pairwise genetic distance (Puillandre et al., 2021). We used the COI alignment file as the input file, and the analysis was conducted with Kimura 2-parameter model (K2P) using ASAP webserver (https://bioinfo.mnhn.fr/abi/public/asap/). The PTP model delimits species based on the number of substitutions between and within species (Zhang et al., 2013). We performed the PTP method with the PTP webserver (https://cme.h-its.org/exelixis/web/software/PTP/index.html). Both maximum likelihood and Bayesian implementation were used with ML tree (see above) for 500 000 MCMC generations and saved every 500 generations. The first 25% of generations were discarded as burn-in. The GMYC method is a likelihood method for delimiting species by using ultrametric trees (Pons et al., 2006). The BI tree (see above) was used and the GMYC method was performed in R 4.1.1 by using the ‘splits’ package (Species Limits by Threshold Statistics) v.1.0-20 (Ezard et al., 2009).

RESULTS

Larval release and morphology

Observation of the galls under stereomicroscopes revealed that naupliar larvae were released from the surface of the external gall (Fig. 1K–M; Supporting Information, Video S2). Naupliar II larvae have six large bifid spines on the carapace, a single naupliar eye, a labrum and three pairs of limbs (Fig. 1O; Supporting Information, Video S1). The body of nauplius contains numerous oil droplets (Fig. 1O; Supporting Information, Video S2).

SYSTEMATICS

Subclass Ascothoracida Lacaze-Duthiers, 1880

 

Order Laurida Grygier, 1987a

 

Family Petrarcidae Gruvel, 1905

 

Genus PetrarcaFowler, 1889

Type species: Petrarca bathyactidisFowler, 1889.

Petrarca goanna Grygier, 1991

 

(Figs 1F, 2–7)

Petrarca goanna Grygier, 1991: 35–38, figs 18, 19.

Material examined:

Four specimens in Astreopora sp., two specimens in Turbinaria mesenterina (Lamarck, 1816) and 12 specimens in Turbinaria sp., 22°40ʹ39.2″N, 121°28ʹ57.2″E, Green Island (Lǜdǎo), Taiwan, 24.04.2015, at 4–20 m depth. Fifteen specimens in Turbinaria frondens (Dana, 1846) and four specimens in Turbinaria sp., 10°41ʹ56.5˝N, 99°24ʹ28.4″E, Koh Khai Island,Chumphon, Thailand, 06–07.07.2019, at 3–4 m depth.

Grygier (1991a) provided a detailed description of this species. We provide additional observations obtained mainly with SEM.

Diagnosis:

Carapace valves of adults with crenulated margins formed by protruding outer ends of five to eight radially directed dorsal and posterior irregular ridges; ridge exterior with irregular lumpy inflations; ventral side of carapace with few papillae; lateral surface of carapace without small papillae. Labrum with short lateral extensions; maxillae mostly exposed; mandibles with 15–25 sharp, simple or bifid teeth; teeth of the maxillules in lower half with spiniform tips and tending to form two rows; six pairs of thoracopods, first thoracopod setiform; penis with relatively small, squarish rami.

Description:

Living specimens pink or crimson-coloured (Fig. 1F); tip of penis often extending out of carapace (Fig. 2C, E, F). Adult (mature) specimens 5.26–6.08 mm long, 4.32–6.15 mm high (Figs 2A–D, 5A); young (juvenile) specimens (Fig. 2E–G) 2.80–3.83 mm long and 2.40–3.33 mm high. Carapace (Figs 2, 5A) roughly ovoid (spherical), valves with short anteriodorsal hinge line, lateral surfaces with five to eight radial ridges with irregular lumpy inflations. These inflations are less developed in the anterior part of adults and in young specimens. Dorsal and posterior margins of valve crenulated due to carapace ridges (Figs 2, 5F). Ventral margin with few inconspicuous papillae (Fig. 5C, D) with central micropore on tip (Fig. 5E).Cuticle on lateral, external and internal surfaces of carapace with dense, polygonal, small swellings or bumps but without papillae, teeth or ctenes (Fig. 5B, G).

Body inflated, crescent-like, enclosed between carapace valves (Fig. 5F). Cephalon with large adductor muscle lying above big oral cone (Fig. 5F) flanked by five-segmented antennules. Thorax with arched dorsal margin but without distinct segmentation, with clusters of rudimentary uniramous thoracopods; cuticle of thorax with ctenoid scales in dorsal part (Fig. 5H). Abdomen with massive first segment bearing long penis and vestigial rear part (Figs 5F, 7D).

Antennules somewhat prehensile, with little armament or external sculpture on two distal segments (Figs 3A–D, 6A–D). First segment irregularly rectangular, narrowing somewhat distally; second segment trapezoidal; third segment almost triangular, narrowing toward lower/ventral margin; fourth segment slightly longer than wide, with slightly curved ventral margin, short distal seta inserted at anteriodorsal corner (Figs 3A, D, 6A, B). Fifth segment rectangular, slightly shorter and narrower than fourth and armed with sensory and grasping structures (Figs 3A–C, 6A, C, D). Short but massive curved claw with smooth concave margin arising from distal end of segment. Three rudimentary setae at base of claw and to each side; tiny pores (three to four) on inner and outer lateral sides. Claw sheathed by large, oval claw guard on posteriodistal corner; claw guard with three vestigial, distal setae with a terminal pore; developed aesthetasc almost half as long as claw guard, inserted closer to its distal end and terminating with four outgrowths, rudimentary seta at base of aesthetasc (Fig. 6A, D). Ventral (postaxial) margin of fifth segment and claw guard with wrinkled cuticle bearing tiny pores (Fig. 6C).

Oral cone prominent, consisting of massive labrum underlaid posteriorly with massive, fused maxillae, unpaired medial languette and paired mandibles and maxillules (Figs 3E–J, 5F, 6E–H). Labrum prow-shaped, with short posteriolateral extensions leaving maxillae largely exposed, exterior with dense ctenoid scales (Figs 3E, 6E). Medial languette with groups of sharp denticles (two to five) along anterior margin and dense basal pores forming ‘pore-field’ (Fig. 6F). Mandibles (Figs 3F, G, 6G) elongated, outer/upper margin with hillock and clusters of dense, thin setae in middle part, cutting edge straight, with 15–21 sharp, simple or rarely bifid teeth and one to two small spinules in middle part. Maxillules (Figs 3H, I, 6H) with sclerotized, triangular distal parts, inner margin with 17–20 teeth, teeth in upper half with often blunt tips, while those in lower half smaller, irregular, with spiniform tips and tending to form two irregular rows. Fused maxillae (Fig. 3J) ending in a pair of short, rounded lobes, with dense ctenoid scales on lateral surfaces (Fig. 7B), distal ends covered with sclerotized, thick and wrinkled cuticle without denticles, pores or setules (Fig. 7A).

Thoracopods 2–6 (Figs 4B, 5F) are present in all examined specimens, uniramous, unsegmented, grouped in cluster and arranged in an unorderly sequence in lateral view. Thoracopods 2–4 are conical with wide bases, while thoracopods 5 and 6 are shorter and narrower. Cuticle of thoracopods 2–6 has dense and long ctenoid scales (Fig. 7C). Batteries of numerous ampuliform seminal receptacles (Fig. 4B) associated with thoracopods 2–5 (~7, 20, 20 and 11 receptacles, respectively). Setiform thoracopod 1 (Fig. 4A) present in all studied specimens.

Long, thick, terminally bifid penis originates from large first abdominal segment (Figs 4D, 5F, 7E). Long shaft of penis supports two small, squarish rami about 190–200 µm long (Figs 4D, 7E). Distal half of penis, including rami, bearing numerous short but wide conical setae. Rest of abdomen vestigial, apparently consisting of two segments (Figs 4C, 5F, 7D). Posterior end of abdomen with cleavage that may represent rudimentary furcal rami (Fig. 7D).

Remarks:

The morphology of our specimens generally corresponds to that of the individuals described by Grygier (1991a), but differs in having rare ventral papillae on the carapace (absent in Grygier’s specimens) and presence of all six pairs of thoracopods (inconsistent number of thoracopods in Grygier’s material). However, these details could have been overlooked in the type material. This is the first finding of Petrarca in scleractinian corals of the genus Astreopora.

Petrarca morula Grygier, 1985

 

(Figs 1G, 8–13)

Petrarca morulaGrygier, 1985: 1036–1038, fig. 6, Grygier & Nojima, 1995: 93–96, fig. 18 (probably another species).

Material examined:

Four specimens in Turbinaria bifronsBrüggemann, 1877, 22°40ʹ39.2″N, 121°28ʹ57.2″E, Green Island (Lǜdǎo), Taiwan, 24.04.2015, at a depth of 4–20 m.

Here we provide additional descriptions based on SEM analysis that supplements Grygier’s (1985) original description.

Diagnosis:

Carapace spherical, with large lumpy inflations, without radial ridges; carapace margin not crenulated; ventral side of carapace with large conical papillae, lateral surface with small papillae. Mandibles with 10–14 sharp, simple teeth; maxillules with small denticles; six pairs of thoracopods, first thoracopod setiform; penis with relatively small, rounded rami.

Description:

Living specimens pink, with dark inclusions at the anterior part of carapace (Fig. 1G); tip of penis often extending out of carapace (Fig. 8). Adult (mature) specimens 2.13–2.42 mm long (1.2–3.8 mm from the Grygier material), 1.85–1.96 mm high (0.9–4.0 mm from the Grygier material), 2.00–2.24 mm wide (1.0–4.6 mm from the Grygier material) (Figs 8, 11A). Carapace (Figs 8, 11A) spherical; valves with conspicuous rounded lumpy inflations (8–14), without radial ridges, dorsal and posterior margins not crenulated; long, conical papillae with a tiny terminal pore on anterior half of ventral margin (Fig. 11A, D). Cuticle on the lateral external and posteriodorsal internal surfaces of carapace with dense, polygonal, small swellings or bumps (Fig. 11B, H). Small volcano-shaped papillae with tiny a terminal pore and microscopic pores inserted between bumps on external surface of carapace (Fig. 11B, C, E). Thin cuticular lining (mantle) of inner surface of carapace with rows of dense ctenoid scales in central part (Fig. 11F, G, I).

Body inflated, curved, enclosed between carapace valves, tip of penis almost adjoins oral cone (Figs 8, 12A). Cephalon with large adductor muscle lying above big oral cone flanked by five-segmented antennules. Thorax with sinusoid arched dorsal margin, without distinct segmentation, with clusters of rudimentary uniramous thoracopods (Fig. 12A). Abdomen with massive first segment bearing long penis and vestigial rear part (Fig. 12A).

Antennules W-shaped, with little armament (external sculpture) on two distal segments (Figs 9A–C, 12B–H). First segment irregularly rectangular, narrowing somewhat distally; second segment rectangular; third segment triangular, narrowing toward lower/ventral margin; fourth segment trapezoid, with slightly curved ventral margin, short distal seta inserted at anteriodorsal corner (Fig. 12C). Fifth segment narrow, rectangular, slightly shorter than fourth and armed with sensory and grasping structures (Figs 9B, C, 12D–H). Short massive curved claw with smooth concave margin. Three rudimentary setae at base of claw and to each side (Figs 9B, C, 12F); tiny pores (five to seven) on inner and outer lateral sides (Figs 9B, C, 12D, E). Claw sheathed by large, oval claw guard (Fig. 12F); claw guard with three vestigial, distal setae with a terminal pore (Fig. 12H); developed subdistal aesthetasc almost half as long as claw guard, terminates with two outgrowths, rudimentary seta with a terminal pore at base of aesthetasc (Fig. 12F, H). Ventral (postaxial) margin of fifth segment and claw guard with wrinkled cuticle bearing tiny pores (Fig. 12G).

Oral cone prominent (Figs 9D, 12A, I); labrum prow-shaped, exterior with dense ctenoid scales (Fig. 12I, J); mandibles elongated, cutting edge straight, with 14 sharp, simple teeth (ten in Grygier specimen) (Fig. 9E); maxillules with sclerotized, triangular distal parts, inner margin with small denticles (Fig. 9F, G); fused maxillae ending in a pair of short, rounded lobes, with dense ctenoid scales on lateral surfaces (Figs 9H, 12I).

Studied specimens with six pairs of thoracopods; thoracopod 1 setiform (Fig. 10A); thoracopods 2–6 (Fig. 10B) uniramous, unsegmented, grouped in cluster and arranged in an unorderly sequence in lateral view; thoracopods 2–4 longer and wider than thoracopods 5 and 6. Cuticle of thoracopods 2–6 with rows of dense and long ctenoid scales (Fig. 13A, B). Batteries of ampuliform seminal receptacles (Fig. 10B) associated with thoracopods 2–5 (~11, 6, 8 and 7 receptacles, respectively).

Long, terminally bifid penis originates from large first abdominal segment (Figs 10D, 12A, 13C). Shaft of penis supports two small, rounded rami about 125 µm long; distal part of penis, including rami, bearing numerous short but wide conical setae. Rest of abdomen vestigial, consisting of two segments bearing rare ctenoid scales and tiny pores; posterior end of abdomen with cleavage with few terminal denticles (Figs 10C, 13D–F).

Remarks:

Grygier & Nojima (1995) indicated that both P. morula and P. goanna may represent ‘the extremes of morphological variability in a single species’. However, along with molecular evidence provided here, there are a number of morphological features that can be used to distinguish these species. Thus, P. morula differs from P. goanna in having numerous long conical papillae on the anterioventral part of carapace vs. only a few inconspicuous ventral papillae in P. goanna; exterior of carapace with small volcano-shaped papillae (absent in P. goanna) but without radial ribs and crenulated margin (present in P. goanna); teeth of the mandible are simple and fewer (simple and bifid teeth present in P. goanna); inner margin of maxillules with small denticles (bigger teeth in P. goanna); subterminal aesthetasc of claw guard terminates with two outgrowths (four in P. goanna).

Petrarca nozawai sp. nov.

 

(Figs 1H, 14–18)

Probably as P. morula in Grygier & Nojima, 1995: 93–96, fig. 18.

Zoobank Registration: urn:lsid:zoobank.org:act:AEC7C3EF-C405-44B0-B069-364D18F6B6A3

Material examined:

Two specimens (including holotype) in Turbinaria mesenterina, 22°40′39.2″N, 121°28′57.2″E, Green Island (Lǜdǎo), Taiwan, 24.04.2015, at 4–20 m depth. Six specimens in Turbinaria sp., 25°7ʹ49″N, 121°51ʹ26″E, Keelung, Taiwan, 17.08.2015, at 4–10 m depth. Glycerin slide of the holotype (no. Mg 1248) with the carapace valves, the dissected mouth parts, antennules and rest of the body proper with thoracopods and penis, and SEM stub of paratype (no. Mg 1250) with the carapace valves and the body proper are deposited in the Zoological Museum of Moscow State University in Moscow, Russian Federation.

Diagnosis:

Carapace roughly ovoid, valves with eight to ten coalesced lumpy inflations, without distinct radial ridges; carapace margin uneven but not crenulated; ventral side of carapace with numerous, large conical papillae, lateral surface with small papillae. Mandible with 15 sharp, simple teeth; maxillules with 13–15 teeth; six pairs of thoracopods, first thoracopod setiform; penis with small, rounded rami.

Etymology:

The species is named after Dr Yoko Nozawa, our colleague and friend, in appreciation of his help with identification of coral hosts for our Ascothoracida studies.

Description:

Living specimens pink (Fig. 1H). Adult (mature) specimens 3.70–4.22 mm long, 3.52–3.60 mm high and 3.2–3.3 mm wide (Figs 14, 17A, G). Carapace (Fig. 14, 17A, G) roughly ovoid, narrowing towards anterior end; each valve with eight to ten coalesced irregular lumpy inflations but without distinct radial ridges; dorsal and posterior margins of carapace uneven but not crenulated; anterioventral margins slightly curved or straight, with numerous, large conical papillae (Figs 14C, E, 17A, D); lateral surfaces and posterioventral margins with small papillae (Fig. 17B, E). Carapace papillae volcano-shaped, terminating at central microscopic pore (Fig. 17C, F). Cuticle of carapace with dense, polygonal, small swellings or bumps (Fig. 17B, E, D).

Body inflated, curved, enclosed between carapace valves, tip of penis close to oral cone (Figs 14F, 17H). Cephalon with large adductor muscle lying above big oral cone flanked by five-segmented antennules. Thorax with sinusoid arched dorsal margin, with hump in posterior part but without distinct segmentation, with cluster of rudimentary uniramous thoracopods (Figs 14F, 17H). Abdomen with massive first segment bearing long penis and vestigial rear part (Figs 14E, F, 17H).

Antennules somewhat W-shaped and prehensile, with little armament of external sculpture on two distal segments (Figs 14F, 15A–D). First segment irregularly rectangular, narrowing somewhat distally; second segment trapezoidal; third segment almost triangular, narrowing toward lower/ventral margin; fourth segment longer than wide, with curved ventral margin, short distal seta inserted at anteriodorsal corner (Fig. 15A, B). Fifth segment rectangular, narrowing towards distal end, shorter and narrower than fourth and armed with sensory and grasping structures, ventral/postaxial margin almost straight, dorsal/preaxial margin concave (Fig. 15C, D). Short, massive curved claw with smooth concave margin arising from distal end of segment. Three rudimentary setae at base of claw and to each side (Fig. 15C, D); tiny pores (three to five) on inner and outer lateral sides (Fig. 15C, D). Claw sheathed by large, hood-shaped, oval claw guard on posteriodistal corner; claw guard with three vestigial, distal setae; developed subterminal aesthetasc half as long as claw guard, terminates with two outgrowths (Fig. 15C, D).

Oral cone prominent (Figs 17H, 18A). Massive prow-shaped labrum with short posteriolateral extensions, leaving maxillae largely exposed, dense ctenoid scales on the exterior (Fig. 18A, B). Mandibles (Fig. 15E) elongated, cutting edge straight, with 15 sharp, simple teeth. Maxillules (Fig. 15F, G) with sclerotized, triangular distal parts, inner margin with 13–16 teeth, teeth in upper half with blunt tips, while those in lower half smaller and spiniform. Fused maxillae (Figs 15H, 18A, B) with dense ctenoid scales on lateral surfaces, distal ends with rounded zones of sclerotized, thick and wrinkled cuticle without denticles, pores or setules.

Six pairs of thoracopods; thoracopods 2–6 uniramous, unsegmented, grouped in cluster and arranged in an unorderly sequence in lateral view (Figs 16B, 17H). Thoracopod 1 setiform (Fig. 16A); thoracopods 2–4 conical, longer and wider than thoracopods 5 and 6. Batteries of ampuliform seminal receptacles (Fig. 16B) associated with thoracopods 2–5 (~14, 11–12, 9–10 and 9 receptacles, respectively). Cuticle of thoracopods 2–6 with dense and long ctenoid scales (Fig. 18D).

Long (~1.2 mm) and massive, terminally bifid penis originates from large first abdominal segment (Fig. 17H). Shaft of penis supports two small, flat and rounded rami about 100 µm long; distal part of penis, including rami, bearing numerous short but wide conical setae (Figs 16D, 18E). Rest of abdomen vestigial, consisting of two indistinct segments; posterior end of abdomen with shallow cleft (Figs 16C, 18C).

Remarks:

The new species Petrarca nozawai sp. nov. is similar to the other two congeners, P. morula and P. goanna in utilizing Turbinaria corals as hosts. These species are characterized by the large lumpy inflations of the carapace, which are absent in other species of Petrarca. Petrarca nozawai differs from P. morula in having ovoid but not a spherical carapace and larger teeth on the inner margin of maxillules (small denticles in P. morula). Petrarca nozawai is distinguished from P. goanna: (1) by the absence of radial ribs and the crenulated margin of the carapace; (2) by numerous large, long conical papillae on the anterioventral part of the carapace; and (3) by simple and fewer teeth on the mandible (simple and bifid teeth present in P. goanna).

Petrarcarubus sp . nov.

 

(Figs 1I, 19–25)

 

Zoobank Registration: urn:lsid:zoobank.org:act:BA716B70-D5B8-49A1-AF02-670876B5E887

Material examined:

Four specimens (including holotype) in Turbinaria bifrons and Turbinaria sp., 22°40′39.2″N, 121°28′57.2″E, Green Island (Lǜdǎo), Taiwan, 24.04.2015, at 4–20 m depth. Glycerin slide of the dissected holotype (no. Mg 1249) with the carapace valves, antennules, mouth parts, and rest of the body with thoracopods and penis, and SEM stub of paratype (no. Mg 1251) with the carapace valves and the body are deposited in the Zoological Museum of Moscow State University in Moscow, Russian Federation.

Diagnosis:

Carapace roughly ovoid, carapace valves of adults with crenulated margins formed by protruding outer ends of six to seven radially directed dorsal and posterior irregular ridges; ridge exterior with irregular lumpy inflations; ventral side of carapace with numerous, large conical papillae, lateral surface with small papillae. Mandibles with 20 sharp, simple and bifid teeth; teeth of the maxillules in lower half with spiniform tips and tending to form two rows; six pairs of thoracopods, first thoracopod setiform; penis with relatively small, rounded cylindrical rami.

Etymology.

From Latin rubus, raspberry (Rubus idaeus L.), referring to the shape and colour of the carapace in living individuals of the new species resembling ripe raspberries. It is a noun in apposition.

Description:

Living specimens pink or crimson-coloured (Fig. 1I). Adult (mature) specimens 4.35–5.71 mm long, 4.43–5.65 mm high (Figs 19A–C, 22A); young (juvenile) specimen (Fig. 19D) 1.52 mm long and 1.54 mm high. Carapace roughly ovoid in adults (Figs 19A–C, 22A) and spherical in young specimen (Fig. 19D); valves with short anteriodorsal hinge line; lateral surfaces with six to seven radial ridges with irregular lumpy inflations. These inflations are less developed in the anterior part of adults and in young specimens. Dorsal and posterior margins of valve crenulated due to carapace ridges (Figs 19A–C, 22A, E). Ventral margin curved, with numerous, large conical papillae (Fig. 22C) with a central micropore at the tip (Fig. 22D). Cuticle on the lateral external and internal surfaces of carapace with dense, polygonal, small swellings or bumps, with papillae on the external side and tiny pores on the internal side (Fig. 22B, F).

Body inflated, curved, enclosed between carapace valves, tip of penis close to oral cone (Fig. 22E). Cephalon with large adductor muscle lying above big oral cone flanked by five-segmented antennules. Thorax with arched dorsal margin, with small hump in posterior part but without distinct segmentation, with clusters of rudimentary uniramous thoracopods (Fig. 22E). Abdomen with massive first segment bearing long penis and vestigial rear part (Fig. 22E).

Antennules somewhat prehensile, with little armament (external sculpture) on two distal segments (Figs 20A, 23A). First segment irregularly rectangular, narrowing somewhat distally; second segment rectangular; third segment almost triangular, narrowing toward lower/ventral margin; fourth segment slightly longer than wide, with slightly curved ventral margin, short distal seta inserted at anteriodorsal corner (Figs 20A, D, 23A, B). Fifth segment rectangular, narrowing towards distal end, shorter and narrower than fourth and armed with sensory and grasping structures, ventral/postaxial margin uneven, dorsal/preaxial margin concave (Figs 20B, C, 23A, C–I). Short, massive curved claw with smooth concave margin arising from distal end of segment. Three rudimentary setae at base of claw and to each side (Figs 20B, C, 23E); tiny pores (six to eight) on inner and outer lateral sides (Figs 20B, C, 23C, D). Claw sheathed by large, hood-shaped, oval claw guard on posteriodistal corner (Figs 20B, C, 23A); claw guard with three vestigial, distal setae each with a terminal pore (Fig. 23H); developed subterminal aesthetasc almost half as long as claw guard, terminating with three long outgrowths and rudimentary subterminal seta; rudimentary seta at base of aesthetasc (Fig. 23F, G). Ventral (postaxial) margin of fifth segment and claw guard with wrinkled cuticle bearing tiny pores (Fig. 23I). Oral cone prominent, consisting of massive labrum underlaid posteriorly with massive, fused maxillae, unpaired medial languette and paired mandibles and maxillules (Figs 20E–I, 22E, 24). Massive prow-shaped labrum with short posteriolateral extensions, leaving maxillae largely exposed, dense ctenoid scales on the exterior (Fig. 24A). Mandibles (Figs 20F, G, 24B–D) elongated, outer/upper margin with dense, thin setae in the middle, cutting edge straight, with 20 sharp, simple or rarely bifid teeth and three small spinules(or reduced setae?) in the middle. Maxillules (Figs 20H, 24E–H) with sclerotized, triangular distal parts, inner margin with ~27 teeth, teeth in upper half with blunt tips, while those in lower half smaller, irregular, with spiniform tips and tending to form two irregular rows; dense basal pores forming ‘pore-field’ (Fig. 24H). Fused maxillae (Figs 20I, 24I–K) with dense ctenoid scales on lateral surfaces (Fig. 24K), distal ends with rounded zones of sclerotized, thick and wrinkled cuticle without denticles, pores or setules (Fig. 24J).

Six pairs of thoracopods; thoracopods 2–6 uniramous, unsegmented, grouped in cluster and arranged in an unorderly sequence in lateral view (Figs 21B, 22E). Thoracopod 1 setiform (Figs 21A, 25A); thoracopods 2–4 conical, longer and wider than thoracopods 5 and 6. Batteries of ampuliform seminal receptacles (Fig. 21B) associated with thoracopods 2–5 (~18–20, 20–22, 12 and 9 receptacles, respectively). Cuticle of thoracopods 2–6 with dense and long ctenoid scales (Fig. 25B).

Long (~0.8–1.0 mm) and massive, terminally bifid penis originates from large first abdominal segment (Fig. 22E). Shaft of penis supports two relatively small, rounded cylindrical rami about 200–250 µm long; distal part of penis, including rami, bearing numerous short but wide conical setae (Figs 21D, 25D). Rest of abdomen vestigial, consisting of two indistinct segments; posterior end of abdomen with shallow cleft (Figs 21C, 25C).

Remarks:

The new species Petrarca rubus is similar to the other three congeners, P. morula, P. goanna and P. nozawai, in utilizing Turbinaria corals as hosts. These species are characterized by the large lumpy inflations of the carapace, which are absent in other species of Petrarca. Moreover, P. goanna and P. rubus represent a complex of cryptic species and can only be distinguished by a few ultrastructural details: (1) the presence of numerous large, long conical papillae on the anterioventral part of carapace in P. rubus (only a few in P. goanna) and papillae on the external lateral surface of carapace (absent in P. goanna); (2) the ultrastructure of antennular aesthetasc having three long terminal outgrowths in P. rubus instead of four in P. goanna; and (3) rounded cylindrical rami of penis in P. rubus but squarish rami in P. goanna.

Petrarca rubus differs from P. morula and P. nozawai in having radial ribs and a crenulated margin of the carapace valves and the presence of bifid teeth on the mandibles.

MOLECULAR ANALYSIS

Gene tree and genetic distance

COI gene trees inferred by NJ, ML and BI were consistent (Fig. 26). All the Petrarca samples were clustered into four groups with high bootstrap support, which were consistent with morphology results. Among them, P. goanna was a sister-group to P. rubus, and P. morula was a sister-group to P. nozawai. The K2P genetic distances within species ranged from 0.0018 to 0.0131 (Table 1). The distances between species ranged from 0.0542 to 0.2030 (Table 2). Among them, the distances between P. goanna and P. rubus and between P. morula and P. nozawai are the closest (0.0542 and 0.0734, respectively, Table 2).

Species delimitation

Both delimitation results of the PTP model (maximum likelihood and the highest Bayesian supported solution) assigned all the samples into four partitions, which were consistent with the morphology results of our Petrarca species (Fig. 26). However, the results of ASAP were ambiguous. In the result with the highest ASAP-score (score: 1.50), P. goanna and P. rubus were merged into the same partition (Fig. 26). Petrarca morula and P. nozawai were also merged into the same partition in the result with the second highest ASAP-score (score: 2.00) (Fig. 26). All the samples were assigned into four partitions in the result with the third highest ASAP score (score: 3.00) which were consistent with four Petrarca species (Fig. 26). The result of GMYC also showed lumping of morphologically identified species, including P. goanna + P. rubus and P. morula + P. nozawai (Fig. 26).

DISCUSSION

Cryptic diversity and morphological unity of the genus Petrarca

Molecular phylogenetic methods widely applied in taxonomic studies have revealed numerous cryptic forms in marine invertebrates, forming so-called complexes of cryptic species. Although this is the first taxonomic study of Ascothoracida involving molecular analysis, the possible presence of cryptic forms within ascothoracidans has been indicated previously (Grygier & Nojima, 1995; Kolbasov et al., 2019, 2021). For example, genera SynagogaNorman, 1888,, SessilogogaGrygier, 1990 and Waginella, which represent more basal and generalized forms of ascothoracidans, are characterized with similar morphology between congeners (Kolbasov & Newman, 2018; Kolbasov et al., 2019, 2020). Thus, two geographically isolated species of Synagoga, S. arabesqueKolbasov, Petrunina, Ho & Chan, 2019 from Taiwan and S. grygieriKolbasov & Newman, 2018 from Macaronesia, can be morphologically distinguished only by the number of setae on the fifth antennular segment in males and by the armament on the inner side of the furcal ramus in females. Grygier & Itô (1995), who studied two species of Waginella, W. metacrinicola (Okada, 1926) and W. axotremata Grygier, 1983, found in different locations, concluded that only ‘the size and shape of the carapace of brooding females remain the most useful taxonomic characters to distinguish these two nominal species’. Thus, it appears that W. axotremata and probably W. metacrinicola represent complexes of cryptic species, which need to be revised using a combined morphological and molecular approach (Kolbasov et al., 2021).

In 1995, Grygier & Nojima described specimens of Petrarca from Amakusa (Japan) having morphological similarities to both P. goanna and P. morula. They suggested, ‘it is possible that these two lots from Amakusa represent an undescribed species, or that P. morula and P. goanna represent the extremes of morphological variability in a single species, while most of the present specimens are intermediate’. Grygier (1991a) studied morphological variability in P. okadai and found out that ‘carapace morphology and armament, occurrence and size of antennular armament elements, number of thoracopods (although observations of the last are sometimes difficult in whole mounts), and penis setation provide a warning against giving such features much weight in diagnosing species of Petrarca’. While such features as the gross aspects of the carapace armament and details of the mandibles and maxillules armament seem to be the best for species discrimination.

Obviously, application of molecular phylogenetic methods in taxonomic studies of Ascothoracida helps to distinguish the morphological characters that determine different species from those characters that reflect morphological variability in a single species. Thus, our molecular studies revealed that Turbinaria corals are utilized by at least four species of Petrarca, which represent closely-related congeners. Moreover, P. goanna and P. rubus represent a complex of cryptic species characterized by only slight morphological differences and PTP species delimitation results. According to our analysis, the following characters can be identified as discriminatory for Petrarca species: (1) shape and fine morphology of the carapace; (2) ultrastructure of the antennular aesthetasc; (3) fine morphology of mouthparts; and, probably, (4) shape and size of the penis rami.

Morphological characteristics of the carapace remain one of the most taxonomically important features in Petrarca. Thus, species inhabiting Turbinaria corals have carapace valves with conspicuous lumpy bumps and a crenulated margin, while other species have more or less smooth carapace valves. The carapace in Petrarca is normally roughly spheroidal, ovoid or dome-shaped. However, in P. madreporae, carapace valves have developed posterior lobes, a characteristic of another petrarcid genus, Zibrowia, so the likelihood of it being in this genus is high (Grygier & Cairns, 1996). Species of Petrarca are characterized by numerous large conical papillae located on the ventral/anterioventral side of the carapace and smaller papillae scattered over the entire lateral surface. The pattern of distribution of these papillae varies between species, but P. goanna has only a few conical papillae on the ventral side and no papillae on the lateral surface.

The morphology (ultrastructure) of the antennular aesthetasc located on the claw guard also has high taxonomic values. This aesthetasc may represent a modified and reduced proximal sensory process of the antennules found in generalized ascothoracidans, including the basal petrarcid genus Introcornia. It may have a varying number of terminal outgrowths/setae (when described): one in P. madreporae, two in P. bathyactidis and P. morula, three in P. rubus and P. okadai, and four in P. goanna. The armament on the cutting edge of the mandibles and maxillules also varies between species of the genus. Thus, the mandibles may possess only simple teeth (P. morula and P. nozawai), simple and bifid teeth (P. goanna, P. okadai, P. rubus and P. sensoria) or simple, bifid and multifid teeth (P. azorica and P. madreporae). The cutting (inner) edge of maxillules may be unarmed (P. madreporae), armed with small denticles (P. indica and P. morula) or with blunt upper teeth and/or sharp multifid lower teeth (P. azorica, P. goanna, P. okadai, P. nozawai, P. rubus and P. sensoria). Penis rami may be rectangular/square (P. azorica and P. goanna), triangular (P. bathyactidis and P. okadai) or rounded (P. indica, P. madreporae, P. morula, P. nozawai, P. rubus and P. sensoria).

The monophyly of the genus Petrarca, could be established based on the fine morphological details revealed with SEM. Several potentially synapomorphic morphological characters (some shared with other petrarcids) have been previously proposed to distinguish the genus Petrarca from other Ascothoracida: (1) a carapace with soft swollen valves with external papillae lacking brooding chamber and posterior lobes; (2) antennules with reduced armament and large, hood-like claw guard with a subapical aesthetasc; (3) reduced thoracic segmentation; (4) uniramous, unsegmented thoracopods; (5) large first abdominal segment bearing massive penis with two short terminal rami; and (6) a reduced and partially fused rest of the abdomen (segments 2 and 3) and reduced furcal rami. Based on our morphological revision of the genus Petrarca, the following discriminatory characters can be added to the six proposed above: (1) external papillae of carapace with a central terminal micropore; (2) fourth antennular segment with short anteriodistal seta; (3) fifth antennular segment with an array of lateral micropores, wrinkled cuticle with a number of micropores along the ventral margin and three tiny setae at the base of the claw; (4) claw guard with three short terminal setae (sensilla) and a rudimentary seta at the base of the aesthetasc; (5) mandible not piercing, with masticating cutting edge; and (6) six pairs of thoracopods.

Morphology of Petrarca and host–parasite relationships

Petrarcids are endoparasites, known to induce the formation of galls in the skeletal part of the host corals. Therefore, the development, distribution and structure of galls are often discussed (Grygier, 1985, 1991a; Grygier & Cairns, 1996; Tachikawa et al., 2020). According to Tachikawa et al. (2020), ‘The absence of an aperture in most petrarcid galls implies a lack of access to any source of food other than the host coral and supports the common view that the petrarcids relationship with their hosts is principally parasitic.’ On the other hand, the authors stressed that the presence of petrarcids (i.e. P. madreporae in Madrepora oculata) leads to the hypertrophy of solitary corallites and they ‘therefore, have an increased ability compared to uninfected ones to obtain food and to protect the colony and these interactions may be ‘at least partially mutualistic’ (Tachikawa et al., 2020). However, according to our data in Turbinaria corals, the corallites in galls of petrarcids are not greatly enlarged (Fig. 1B), and thus their ability to obtain food should be similar to others. Thus, the mutualistic relationship of petrarcids with their hosts is unlikely.

Fowler (1889) described the first species of Petrarca (P. bathyactidis), noting that ‘judged by the contents and appearance of the alimentary canal of Petrarca, it seems probable that its relation to its host is not a case of true parasitism, but it sucks in the nutritive fluid produced by the digestion and maceration of the food of Bathyactis, and does not derive sustenance directly from the tissues of the polyps’. But following Grygier (1985), we conclude that the morphology of the mouthparts evidences that they are developed for chewing and cutting, rather than for piercing and sucking, as in many Ascothoracida. The true diet of these parasites can only be established with stable isotope analysis.

Being entirely endoparasitic, all petrarcids induce formation of galls. Living in coral galls, each individual forms an internal chamber containing the parasite (Fig. 1C). There are three possible ways to create such a chamber: (1) mechanical burrowing, (2) chemical dissolution or (3) a combination of the two. These three ways are observed in burrowing barnacles living in calcareous substrates (Kolbasov, 2009). Thus, the acrothoracicans (Cirripedia: Acrothoracica) share mechanical excavation (order Cryptophialida) and combine mechanical excavation plus chemical dissolution (order Lithoglyptida). The rows of massive multifid ctenoid scales in the opercular area form a burrowing apparatus (Kolbasov, 1999, 2009). These scales are often associated with specialized opercular papillae with a terminal pore or tiny pores that may secrete carbonic anhydrase used to dissolve the substrate (Turquier, 1968; Kolbasov, 2009). Another burrowing thoracican barnacle, Lithotrya dorsalis (Ellis & Solander, 1786), probably uses chemical dissolution of the substrate. These barnacles possess calcite burrowing spicules, which are densely distributed on the peduncle, and their organic component is penetrated by duct-like structures ending in pores (Dineen, 1988). The external surface of the carapace in Petrarca is ornamented with densely packed secretory papillae, each terminating with a pore and numerous cuticular tiny pores. At the same time, carapace valves are lacking any ctenoid scales, which could be used for mechanical excavation. Therefore, it is likely that petrarcids are using chemical dissolution of the substrate to create and enlarge the chamber of the gall.

[Version of record, published online 3 May 2023; http://zoobank.org/urn:lsid:zoobank.org:pub:A2863AB5-855B-4549-89EC-A57C46140CD9]

ACKNOWLEDGEMENTS

We thank the collaborators of the Laboratory of Electronic Microscopy of Moscow State University and Dr Andrey I. Lavrov for assistance in SEM studies. Thanks also to Rola Chang who edited the English of this manuscript. We additionally thank the two anonymous referees and the editors Maarten Christenhusz and Shane Ahyong for comments on the manuscript during the review process.

FUNDING

For GAK and ASS this work was financially supported by the Russian Foundation for Basic Research (grant 21-54-52003 MNT_a). BKKC is supported by the Taiwan Russia bilateral grants from National Science and Technology Council (NSTC), Taiwan (NSTC-110-2923-B-001-003-MY3).

DATA AVAILABILITY

The data underlying this article are available in the GenBank Nucleotide Database at https://www.ncbi.nlm.nih.gov/genbank/, and can be accessed with accession numbers OP117451-OP117475. Video data of larval release from gall is available in Dryad: https://datadryad.org/stash/dataset/doi:10.5061/dryad.2z34tmpr4

REFERENCES

SUPPORTING INFORMATION

Additional supporting information may be found in the online version of this article on the publisher’s website.

Table S1. Species information of GenBank accession numbers: OP117451-OP117475

Video S1. In situ observation of the coral galls housing Petrarca.

https://datadryad.org/stash/dataset/doi:10.5061/dryad.2z34tmpr4

Video S2. Larval release of Petrarca from the gall surface.

https://datadryad.org/stash/dataset/doi:10.5061/dryad.2z34tmpr4