Onyx disparamphis sp. n. (Nematoda, Desmodorida) from South Korea with a taxonomic review of the genus

A new free-living marine nematode Onyx disparamphis sp. n. (Nematoda, Desmodorida) is described from sandy littoral of Jeju Island, South Korea. The new species differs from all other Onyx species by the unusual amphideal fovea morphology in males (elongated loop). O. disparamphis relates to O. balochinensis, and O. brevispiculatum by having simple non-double terminal pharyngeal bulb and relatively small and straight, non-sigmoid supplementary organs, but differs from them by smaller body length, shorter cephalic setae, smaller terminal pharyngeal bulb, smaller spicules, number of supplementary organs and tail shape expressed as ratio tail length/anal diameter. The genus Onyx is revised with updated genus diagnosis, and an annotated list of 23 valid species is presented. Onyx ferox is considered species inquirenda because the species is known only from a sole immature female specimen, while within Onyx, the males provide the most important distinguishing characters such as enlarged and complicated amphids, supplementary organs and copulatory spicules. For species identification, a pictorial key consisting of illustrations of simplified icons of male heads and posterior body sections, as well as a table of the most important morphometric and numerical characters are provided. Geographical distribution and habitat specifity of Onyx species is analysed briefly.


INTRODUCTION
As part of the study of meiofauna and nematodes on the intertidal sandy littoral of the Jeju Island (South Korea), we have found a number of new nematode species which are partly already published (Jeong, Tchesunov & Lee, 2019;Jeong, Tchesunov & Lee, 2020;Tchesunov, Jeong & Lee, 2020;Tchesunov, Jeong & Lee, 2021). Here, we report on a new Onyx species common on this beach.
Nematode genus Onyx has been established by N.A. Cobb (1891) for a species found by him in so called Amphioxus-sand in the Bay of Naples, Italy. Cobb marked single axial spear attached to the dorsal side of the pharynx as the prominent trait of Onyx perfectus, which provided a ground to consider an evident kinship with the genus Dorylaimus.
This relationship was later recognized as superficial, and Onyx was taken as a relative of Metachromadora (Filipjev, 1918: 214, at that time, Chromadoridae, Spilipherini) and then within Desmodorinae (Filipjev, 1934). Thereafter, Onyx has a stable position in the nematode system as a genus of the order Desmodorida, family Desmodoridae and subfamily Spiriniinae.
Onyx is a globally distributed, well-defined genus within the the family Desmodoridae which is mainly found in shallow coastal sediments. Species of Onyx are usually well recognizable owing to their bold and distinct structural features. Consequently, there are limited nomenclatural problems or bynonyms within the genus. However, the number of species grows, especially by the exploration of tropical meiofauna, that leads to increasing complexity in species identification. Coupled with new species description, several reviews of the genus Onyx were suggested (Blome & Riemann, 1994;Nasira, Rehmat & Shahina, 2011;Armenteros, Ruiz-Abierno & Decraemer, 2014;Huang & Wang, 2015). Increase in number of species requires periodical taxonomic revisions with proper adjustment of species composition and construction of improved keys for species identification.
Here, we propose description of a new species together with revised species list and pictorial key for species identification.

MATERIALS & METHODS
The nematodes were collected and studied in frame of a project on exploration interstitial fauna of sandy beaches of South Korea. The site of sampling is a large intertidal sandy beach Shinyang Seopjikoji at the south-eastern point of Jeju Island (Fig. 1).
The quantitative sediment samples were initially fixed by neutralized 5% formol on filtered sea water. Meiofauna including nematodes was separated from sediment using method of centrifugation with Ludox (Burgess, 2001), then postfixed with 70% ethanol stained with Rose Bengal. Nematode specimens were picked up under a stereomicroscope and placed in Syracuse glass with mixture of glycerine, ethanol and distilled water at a ratio of 1:29:70. After slow evaporation of ethanol and water during two days at 40 • in an oven the nematodes become completely dehydrated as described by Seinhorst (1959). Nematode specimens were mounted in permanent glycerin slides within bee-wax-paraffin glass and with glass beads as separators. Specimens were studied, measured, pictured and drawn in the optical microscope Leica DM 5000 equipped with IC measure v.2.0.0.161 software and digital camera Leica DFC 425C. For scanning electron microscopy (SEM), specimens fixed in formalin in filtered sea water were then dehydrated in a graded series of ethanol-acetone solutions. Specimens were critical point-dried with carbon dioxide. Dried specimens were mounted on stubs, coated with gold-palladium mixture, and examined with a CAMScan S-2.
Type specimens are deposited in National Institute of Biological Resources (South Korea).

Nomenclatural acts
The electronic version of this article in Portable Document Format (PDF) will represent a published work according to the International Commission on Zoological Nomenclature (ICZN), and hence the new names contained in the electronic version are effectively published under that Code from the electronic edition alone. This published work and the nomenclatural acts it contains have been registered in ZooBank, the online registration system for the ICZN. The ZooBank LSIDs (Life Science Identifiers) can be resolved and the associated information viewed through any standard web browser by appending the LSID to the prefix http://zoobank.org/. The LSID for this publication is: 2FA0F335-DF23-4824-A3DC-A04DD46289BD. The online version of this work is archived and available from the following digital repositories: PeerJ, PubMed Central SCIE and CLOCKSS.  Tchesunov, 2014) Desmodoridae, Spiriniinae. Cylindrical body with broad rounded cephalic region and conical tail. Cuticle thin, fine but distinctly annulated, without lateral differentiation. Amphideal fovea spirally coiled in one to several turns or modified; the fovea often shifted to apical surface of the head. Buccal cavity with long spear-like dorsal tooth directed anteriorly. Terminal pharyngeal bulb mostly elongate, may be double with lens-like thickened internal cuticular lining or the lining not thickened. Numerous midventral precloacal supplementary organs tubular and in most species S-shaped. Tail conical.

Etymology
The species name reflects strong sexual dimorphism in amphideal fovea outline.

Description
Males. Body cylindrical, anterior end rounded truncated, tail conical (Figs. 1A, 4A). Cuticle thin, fine but distinctly cross annulated, without a lateral differentiation. Numbers of cuticular annules within 10 µm varies along the body: 14 annules within 10 µm at the level of the long amphideal branch and 18 less distinct annules within 10 µm at the midbody. Apical region of the cephalic region not annulated but finely longitudinally striated, with sharp border between longitudinal striation and annulation (Figs. 6B, 6C). Inner labial sensilla not evident. Outer labial sensilla as six minute papillae (1-1.5 µm). Four long cephalic setae situated apically and directed anteriorly. There first, anterior crown of eight subcephalic setae located just posterior to the cephalic setae at the level of the anterior Scale bars 100 µm. pores distributed along the body; which look like a minute hole in the centre of a smooth circular spot on the cuticle (Fig. 3A, arrow). Amphids shifted onto the apical area of the cephalic region. Anterior end of the amphideal fovea is located on apical surface close to the mouth opening; the fovea turns dorsally and runs on into the long dorsal arm of the fovea extended far hindward as Somatic cuticle not widened around the mouth. Cheilostoma shaped as a truncate cone with longitudinal rugosity. Long and narrow pharyngostoma armed with a long dorsal tooth provided with a conical pointed cuticular arrowhead (corona). The tooth is adherent dorsally to the pharynx tissue at two thirds of its length. Pharyngostoma is surrounded by inflated pharyngeal tissue with fine transversal striation. Middle part of the pharynx slender; posterior part of the pharynx is formed as an elongate terminal bulb with muscular cross striation. Internal cuticular lining of the bulb with muscular cross striation not thickened and seemingly not modified. Midgut slender, filled with orange pigment inclusions.   axis. Series of 14-19 equal midventral precloacal supplementary organs. Supplements consist of three constituents, (1) surface cuticular pit with cuticularized walls, (2) core within the pit, head of the core bears a longitudinal ridge with a papilla, (3) short internal straight cuticular tube extending from the pit obliquely inward (Figs. 4B, 6D-6E). Tail conical, with caudal glands and a terminal spinneret. Caudal gland cell bodies hardly discernible, visibility limited within tail, but may have seemingly pushed out to preanal region in some specimens. Females. Amphideal fovea spirally coiled in three turns and situated entirely on the cephalic apex close to the mouth opening (Figs. 3C, 6C).
Ovaries paired, antidromously reflected, both situated to the left of the intestine in all female specimens studied (Fig. 1B).

Diagnosis
Body length 695-1014 µm, index a 20-40.5, index c' 2.5-4.34. Cephalic setae 6.5-14.2 µm long. Two subsequent crowns of eight subcephalic setae similar in length to the cephalic ones. Amphideal fovea shows strong sexual dimorphism: in females, the amphideal fovea spirally coiled in three turns and located entirely on the head apex, while in males, the fovea turns dorsally from the aperture on the apex, then extended hindward as elongated loop. Dorsal tooth 27-40 µm along the ventral side. Terminal bulb of the pharynx elongated, 40-60 µm long, with faint internal lining. Spicules arcuate, 33-40 µm long. Midventral precloacal supplementary organs 14-19 in number, consist of flat cap and internal short and almost straight cuticular tubes. Tail conical, c' 2.4-4.3.

Relationships
Onyx disparamphis sp. n. differs immediately from all other Onyx species by the peculiarly very long loop of the amphideal fovea of males. O. disparamphis shares simple nondouble terminal pharyngeal bulb with thirteen other Onyx species (Table 2)

Ecological remarks
Onyx disparamphis is a common, but not very numerous species on the Shinyang beach, being the 12th most abundant and comprising 1.6% of total nematode abundance of the beach. O. disparamphis is distributed across the whole intertidal sandy beach from lower to upper horizon, with some increase in numbers at middle and upper horizons. No obvious confinedness to a certain layer in vertical sediment column is observed, evidently because of uniformity of conditions (granulometry) within a sediment depth 0-20 cm. Microscopic examination of gut content does not reveal any evident particles or identifiable remnants. The intestine of the individuals studied either appear empty or with spherical drops. The long and strong dorsal tooth may protrude from the mouth as it is shown for Onyx sagittarius by Gerlach (1950, Abb. 5c). We suppose that Onyx disparamphis and its congeners could pierce covers of some food items (e.g., protists) and suck out the liquid matter by muscular bulb pumping.

Pictorial key for identification Onyx species
The present pictorial key for identification of 23 valid species of Onyx is constructed based on principles of Platt (1984), who first introduced such keys in marine nematology. The key consists of two components, (1) a set of species icons or pictorial key (Figs. 7-8), and (2) a table of the most important morphometric and numerical characteristics (Table 2). Most valid species are known on both males and females, and only two species, O. monstrosum and O. orientalis are described based only on males. Only males are used for pictures since they provide more distinctly perceiving features (such as amphids) on the head and much more features (such as copulatory and supplementary organs) on the posterior body. The table 2 includes characteristics of only males, because females are not known for two species and data on females are often less complete in comparison with males in other descriptions based on two sexes. On Figs. 7-8, the heads are arranged in such an order that species having the largest and most conspicuous amphideal fovea are located at the top of the key; by scanning the icons from top to bottom, the amphids and head setae become gradually smaller and shorter.

Spatial distribution of Onyx species
Like most other marine nematode superspecies taxa, the genus Onyx shows a worldwide distribution (Fig. 9); however, most species are confined with warm waters, and none occurs in Arctic and Antarctic (with possible exception O. ferox sp. inq. in Subantarctic). Nine species are recorded in tropical areas (balochiensis, blomei, cangioensis, cobbi, dimorphus, mangrove, orientalis, paradimorphus, perfectus); eight species in subtropical regions (adenophorus, brevispiculatum, cannoni, cephalispiculus, macramphis, perfectus, potteri, sagittarius), nine species in temperate regions (disparamphis, littorale, minor, monstrosum, perfectus, rizhaoensis, rugatus, sagittarius, septempapillatus). Surprisingly, there were no Onyx species recorded along the east coasts of the Americas, but this is likely due to insufficient information on meiofauna in this specific region. A cluster of six co-occuring species within a limited area was found in the CanGio mangrove habitat (South Vietnam) in silty sediment at a depth 0.5 m.
All the Onyx species are confined with coastal shallows, from intertidal zone to upper sublittoral of several tens of meters depth; no species are recorded from the deep sea. The majority of species inhabits coarse sands (16 of 24 species), often on high energy beaches. The remaining eight species were found on silts, most of them (six) in mangrove milieu.