1. INTRODUCTION

The family Chromadoridae Filipjev, 1917 is a widely distributed group of predominantly free-living marine nematodes, with some representatives also inhabiting freshwater environments, making it one of the larger families of nematodes (Decraemer and Smol 2006;Venekey et al. 2019). This family has been systematically reviewed by Lorenzen (1994) and Tchesunov (2014), with recent re-evaluations by Venekey et al. (2019), who classified it into five subfamilies, 37 valid genera, and 395 recognized species. Within Chromadoridae, the genus PtycholaimellusCobb, 1920 com- prises 24 species recorded worldwide (Huang and Gao 2016;Shih et al. 2022;Leduc and Zhao 2023), most of which are marine, with the exception of two species- P. pandispiculatus (Hopper 1961) and P. ponticus (Filipjev 1922)-found in brackish waters (Venekey et al. 2019). This genus is distinguished by unique morphological traits, including a cuticle with two longitudinal rows of larger dots, a distinct collar demarcating the head from the body, a pronounced S-shaped dorsal tooth, and notable double pharyngeal bulbs at the posterior (Jensen and Nehring 1992;Huang and Gao 2016;Venekey et al. 2019).

To date, seven Ptycholaimellus species have been documented in the Northwestern Pacific region. These include P. adociusDashchenko and Belogurov, 1984, found in sponge habitats in Posjet Bay, Sea of Japan; P. areniculusThanh et al., 2012, and P. brevisetosusThanh et al., 2012, discovered in shallow sandy sediments (1-2 meters deep) in northern Vietnam; P. longibulbusWang, An, and Huang, 2015, identified in intertidal muddy sediments on Ximen Island, East China Sea; P. luoyangShih, Chen, and Guo, 2022, discovered in intertidal mud sediment form a mangrove wetland in the Luoyang River estuary, Fujian Province in China; P. ocellatusHuang and Wang, 2011, found in intertidal muddy sediments and algae along the Lianyungang coast in Jiangsu Province, Yellow Sea, China; and P. pirusHuang and Gao, 2016, collected from intertidal silt off Dongshan Island, East China Sea. Despite this, no morphological taxonomic studies of the genus Ptycholaimellus have been conducted in Korean waters.

Currently, 94 species of free-living marine nematodes have been reported in Korea. Of these, 44% (41 species) are from the East Sea, 28% (26 species) are from Jeju Island, and only 12 species have been recorded in the South Sea (Jung et al. 2023;Kim and Jeong 2023;Lee et al. 2023;Hong et al. 2024;Lim et al. 2024). This study aims to describe a new record of Ptycholaimellus species collected from intertidal sediments along the South Sea of Korea, marking the first taxonomic report of P. ocellatus in the region. The report provides detailed taxonomic descriptions, illustrations, and micrographs using differential interference contrast (DIC) and scanning electron microscopy (SEM).

2. MATERIALS AND METHODS

2.1. Sample collection and preparation

Sediment samples were gathered from intertidal mudflats on the southern coast of Korea. To induce osmotic shock, the collected samples were initially rinsed with tap water for one minute. The samples were then passed through a 67 μm mesh sieve to remove larger debris and preserved in 5% formalin. In the laboratory, the Ludox HS40 flotation method was used to extract meiobenthic organisms from the sediment (Burgess 2001). Extracted specimens were filtered again using a 67 μm mesh and subjected to three rounds of centrifugation to further isolate the target organisms. Nematodes were carefully selected under high magnification using a Pasteur pipette and a dissecting microscope (LEICA 205 C; Wetzlar, Germany), and preserved in a 3% glycerol solution for subsequent analysis.

2.2. Morphological analysis

Specimens were mounted between two coverslips using a wax-ring method and placed on HS glass slides for detailed observation (Shirayama et al. 1993). Microscopic examinations were conducted with an Olympus BX53 microscope (Tokyo, Japan), and images were taken using a LEICA DM2500 LED microscope equipped with a LEICA K5C color CMOS camera. Morphological details and measurements were documented using cellSens Standard software. Anatomical line drawings were created with a drawing tube attached to the LEICA DM2500 LED microscope and digitally refined using Adobe Illustrator on a Wacom Cintiq 22 tablet.

2.3. Scanning Electron Microscopy (SEM) imaging

For high-resolution imaging, specimens were fixed in 5% buffered formalin and rinsed twice in distilled water (each for 30 minutes) to remove residual fixatives. The specimens were freeze-dried using an FDU- 1200 cooling stage (EYELA, Tokyo, Japan). After drying, they were mounted on aluminum stubs and coated with a gold/palladium layer using a sputter coater. The prepared specimens were examined with a desktop mini-SEM (SEC SNE-3200M; Suwon, Korea) to capture high-magnification images of their surface structures.

3. SYSTEMATIC ACCOUNTS

Phylum Nematoda Potts, 1932

Class Chromadorea Inglis, 1983

Order Chromadorida Chitwood, 1933

Family Chromadoridae Filipjev, 1917

Genus PtycholaimellusCobb, 1920

Ptycholaimellus ocellatusHuang and Wang, 2011 (Figs. 1-3; Table 1)

Ptycholaimellus ocellatusHuang and Wang, 2011, p. 2192, Figs. 1, 2.

Material. Two male specimens (MABIK NA00158732, MABIK NA00157563) are stored at the Marine Biodiversity Institute of Korea (MABIK) in Seocheon. Additionally, one male specimen (KIOST NEM-1-2794) is preserved at the Bio-Resources Bank of Marine Nematodes (BRBMN) located at the East Sea Research Institute, Korea Institute of Ocean Science & Technology (KIOST).

Locality and Habitat. Three specimens were collected on May 28, 2020, from an intertidal mudflat at Jangseon Beach, Daeseo-myeon, Goheung-gun, Jeollanam- do, Korea (34°44ʹ59.00ʺN, 127°15ʹ02.00ʺE).

Measurements. Refer to Table 1 for detailed measurements and morphometrics.

Description. Males. The body is cylindrical, gradually tapering towards both ends (Figs. 1A, 3A). The pharynx narrows abruptly at approximately 40% of its length from the anterior end (Figs. 1B, 3E), and SEM images reveal a relatively thin cuticle at this point (Fig. 2A). The annulated cuticle begins around 5-6 μm from the anterior end (Fig. 2B) and shows a lateral differentiation, measuring 3-3.5 μm in width, protruding on both sides of the body (Fig. 2D). This lateral differentiation is bordered by large, longitudinally arranged dots (Fig. 3C, F), extending from the head to the tail. Each annulation measures 1-1.5 μm in width, and no somatic setae are observed on the body surface.

The head is distinctly separated from the body by a constriction (collar) emerging from a groove measuring 2-3 μm in depth (Figs. 1C, 2B, 3B). Both the inner and outer labial sensillae are not clearly discernible, while four cephalic setae, about half the diameter of the head, are attached to the collar. Amphidial organs are not visible, though SEM images show fine transverse lines across the body at the expected position of the amphids (Fig. 2C).

The lips display weakly sclerotized folds, and the buccal cavity houses a prominent cuticularized S-shaped hollow dorsal tooth (Figs. 1C, 2C, 3B). The dorsal portion of the buccal cavity is encircled by pharyngeal muscles. The anterior part of the pharynx is cylindrical, broadening posteriorly and transitioning into the pharyngeal bulb, which features two pairs of protoplasmic interruptions (Fig. 3E). The posterior pharyngeal bulb constitutes approximately 26-27% of the total pharyngeal length, with a diameter of 20-21 μm. The nerve ring is positioned at approximately 58- 59% of the pharyngeal length (Fig. 1B). The cardiac region is short and enveloped by intestinal tissue. The ventral gland cell is large and sac-like, measuring 125- 141 μm in length and reaching a maximum width of 15-21 μm, with an ampulla connected to a fine duct that traverses the collar (Figs. 1A, 3A).

The tail measures approximately 3.8-3.9 times the anal body diameter, transitioning from a conical to a cylindrical form, ending with a prominent, elongated spinneret (Figs. 2E, F, 3G). The reproductive system is monorchic, with the testis positioned on the right side of the intestine. The spicules are arcuate, measuring 40-42 μm in length, with a broad proximal end and a tapered distal tip. The gubernaculum, measuring 17- 18 μm, runs parallel to the distal portion of the spicules (Figs. 1D, 3D).

Female. Unknown.

Distribution. China, Korea.

Remarks. The genus Ptycholaimellus was initially established by Cobb in 1920, with P. carinatus from Indonesia designated as the type species. In 1955, Gerlach suggested Ptycholaimellus as a subgenus of Hypodontolaimus, but Inglis (1969) and Decraemer and Coomans (1978) later recognized it as a valid genus. A comprehensive review by Jensen and Nehring (1992) evaluated the genus and underscored the significance of various distinguishing traits, acknowledging 14 species at that time (Venekey et al. 2019). Currently, there are 24 valid species of Ptycholaimellus reported globally (Huang and Gao 2016;Shih et al. 2022;Leduc and Zhao 2023), with Huang and Gao (2016) providing a recent identification key for all valid species.

Ptycholaimellus ocellatusHuang and Wang, 2011, was first described from intertidal muddy sediments and algae along the coast of Lianyungang, Jiangsu Province, China. This species is characterized by the presence of ocelli near the anterior end, an abruptly tapering cervical region, lateral differentiation in the cuticle with two longitudinal rows of elongated dots, and a prominent ventral gland with a large ampulla (Huang and Wang 2011). To date, only two species within the genus, P. ocellatusHuang and Wang, 2011, and P. adociusDashchenko and Belogurov, 1984, are known to possess anterior ocelli.

In this study, specimens collected from intertidal sediments along the Korean coast exhibited significant similarity to the original description of P. ocellatus by Huang and Wang (2011). These similarities included the general body shape, the presence of ocelli, the sharply narrowing cervical region, lateral cuticular differentiation, and the prominent ventral gland. However, slight variations were observed, such as a relatively longer body length in males (918-937 μm in the Korean specimens compared to 772-828 μm in the original) and a marginally shorter tail in males (De Man’s ratio ‘c’ of 10-11 in the Korean specimens versus 8.1-8.5 in the original). Additionally, while the original description notes an abrupt narrowing of the cervical region at approximately one-third of the pharynx length, this feature was observed at around 40% of the pharynx in the Korean specimens. Despite these minor differences, the alignment of key morphological characteristics strongly supports the identification of the Korean specimens as P. ocellatus.