INTRODUCTION

The study of the fauna of free-living nematodes in the water bodies and watercourses in Vietnam began about 15 years ago in order to develop a database on the aquatic fauna of Vietnam. In recent years, large-scale studies on the fauna of this taxonomic group have been performed. The data on their fauna in fresh water bodies and mangroves are summarized in the works of Gagarin and Gusakov (Gusakov and Gagarin, 2017; Gagarin, 2018).

This study aims to give an illustrated description of two species of free-living nematodes new to science.

MATERIALS AND METHODS

Twenty-four samples of nematodes were collected in March 2017 by colleagues from the Institute of Ecology and Biological Resources of the Vietnam Academy of Sciences and Technologies (Hanoi, Vietnam) in the bottom sediments of an artificial pond for shrimp aquaculture. This water body is located in the Quảng Ninh Province on an island in the South China Sea off the coast of Vietnam. Its area is 1000 m2. The bottom is overgrown by dense thickets of grass Halophila beccarii Ascherso, 871 and Ruppia maritima L., 1953. Material was also collected in the estuaries of the Van Uc River and Ien River in the mangroves. Samples were taken at a depth of 0.3–0.7 m using a cylinder with a diameter of 3.5 cm, a length of 10 cm, and washed through a sieve with a mesh diameter of 0.08 mm. Samples were fixed with hot (60–70°C) 4% formalin solution. The sample was then placed in 200-mL containers, a Ludox TM50 solution (1 : 1) was added, and the samples were centrifuged five times for 40 min. The nematodes were transferred to pure glycerol according to the Seinhorst method (Seinhorst, 1959) and then mounted in a drop of glycerol on a glass slide and sealed with a paraffin (wax) ring. A Nikon Eclipse 80i light microscope equipped with DIC contrast, a Nikon DS-Fil digital camera, and a PC equipped with NIS-Elements D 3.2 software for analysis and documentation were used for measuring, species definition, photographing, and drawings the nematodes.

In the samples, nematode species new to science were discovered. The description and illustrations of the two species are presented below.

RESULTS

Description of new species. Order Plectida Malakhov, 1982. Family Leptolaimidae Örley, 1880. Genus Leptolaimoides Vitiello, 1971.

Leptolaimoides brevicaudatus Gagarin sp. n. (Figs. 1, 2).

Fig. 1.
figure 1

Male (a, b, d) and female (c, e) Leptolaimoides brevicaudatus sp. n.: (a) general view, (b) head, (c) body in the vulva area, and (d, e) tail.

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Fig. 2.
figure 2

Male (a, c, d, e, g, i, j) and female (b, f, h, k) Leptolaimoides brevicaudatus sp. n.: (a, b) general view, (c, d) anterior end of the body, (e, f) head, (g) body in the cardium area, (h) body in the vulva area, (i) body in the cloaca area, and (j, k) tail.

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Material. Holotype ♂, slide no. ND 4.2.19; paratypes: 4♂♂, 1♀. The holotype preparation is stored in the collection of the Nature Museum of the Vietnam Academy of Sciences and Technologies (Hanoi, Vietnam). Paratype preparations are stored in the collection of nematodes at the Institute of Ecology and Biological Resources of the Vietnam Academy of Sciences and Technologies (Hanoi, Vietnam).

Location. The nematodes were found in two locations. Three ♂♂ were found in Northern Vietnam, Quảng Ninh Province, Hà Đông, artificial pond for shrimp aquaculture (21°16ʹ77ʹʹ N, 107°23ʹ92ʹʹ E). Depth 0.5 m, silt with turf, water salinity 20‰. One ♂ and one ♀ were also found in Northern Vietnam, Hai Phong Province, estuary of the Van Uc River, (20°40′28″ N, 106°41ʹ44ʹʹ E). Mangroves, depth 1.1 m, silt, salinity 6‰.

Description. The morphometric characteristics of the holotype and paratypes are given in Table 1.

Table 1.   Morphometric characteristics of Leptolaimoides brevicaudatus sp. n.
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Male. The body is thin, short. The cuticle is finely annulated. Somatic bristles and pores not found. A lateral area in the form of two small ridges stretches along the entire body. The width of the lateral area is 4.0–4.5 μm. The anterior end of the body is narrowed. The width of the labial area is 2.3–2.7 times smaller than the body diameter at the cardium area. The labial area is not isolated from the rest of the body. External labial sensillae barely visible, in the form of small papillae. Four head sensillae in the form of larger papillae. Cheilostoma is relatively large. Pharyngostoma in the form of a narrow tube, 21–23 μm in length. The pharynx is muscular, slightly expanding to its base. Cardium in the form of a long muscular tube, 20–24 μm in length. Renette, its pair and ducts not detected. Amphid fovea in the form of a narrow long cylinder, 16–18-μm length, separated from the anterior end of the body at a distance of 6–9 μm.

Seminal glands are paired, opposed. The anterior testis is straight, located on the right side from the middle intestine; posterior testis short, bent. The spicules are thin, curved, with a head. The spicules are 1.8–2.0 times longer than the body width in the cloaca area. The gubernaculum has the form of a simple plate, 8–10 μm in length, with a curved caudal process of about the same length. Supplementary organs not found. Caudal setae not found. Caudal glands are present, but poorly visible. The spinneret is small, triangular.

Females. Similar to males in general morphology. The structure of the anterior end of the body and the cuticle structure are similar to that in males. The cuticle is finely annulated. Lateral areas stretch along the entire body as two ridges. The anterior end of the body is narrowed. Labial sensillae in the form of small, barely noticeable papillae. Four head sensillae in the form of larger papillae. Narrow tubelike pharyngostoma of 20-μm length. The pharynx is muscular, expanding to its base. The cardium is long, muscular, and does not extend into the lumen of the middle intestine. The reproductive system is didelphic, amphidelphic; ovaries are antidromic, short. Bends of the ovaries reach the vulva. The anterior ovary is located on the right from the intestine, the posterior one, on the left from the intestine. The vulva is equatorial, in the form of a transverse gap. The vagina is short, straight. The uterus is extensive. Each uterus has one rounded spermatheca. The tail is elongated-conical. The caudal glands are barely visible. The spinneret is small, triangular.

Differential diagnosis. Recently, the genus Leptolaimoides Vitiello, 1971 comprises 12 valid species (Holovachov, 2015; Hoang Lai-Phu et al., 2009). In regards to the body size, Leptolaimoides brevicaudatus sp. n. is close to L. thermastris (Lorenzen, 1966), L. cangioensis Hoang Lai-Phu et al., 2009, and L. asiaticus Gagarin, Nguyen Vu Thanh, 2005. It differs from the first species by a shorter and less slender tail (c = 7.1–9.0, c' = 3.8–6.3 versus c = 4.0–4.7, c' = 7.5 in L. thermastris), longer amphid fovea (16–20 μm in length versus 8–12 μm in L. thermastris), and longer spicule (22–25-μm length versus 13 μm in L. thermastris) (Lorenzen, 1972); it differs from L. cangioensis by a shorter and less slender tail (с = 7.1–9.0, c' = 3.8–6.3 versus с = 4.0–6.5, с' = 7.8–10.0 in L. cangioensis), longer amphid fovea (16–20 μm length versus 5–7 μm in L. cangioensis), simply structured lateral areas (in L. cangioensis they are areolated), a longer spicule (22–25 μm in length versus 14–15 μm in L. cangioensis), and by the absence of supplements in males (there are two supplements ahead cloaca in L. cangioensis males); it differs from L. asiaticus by a shorter and less slender tail (с = 7.1–9.0, с' = 3.8–6.3 versus с = 3.7–6.2, с' = 7.5–11.4 in L. asiaticus), longer spiculee (22–25 μm in length versus 15–17 μm in L. asiaticus), and the absence of supplements in males (there are three supplements ahead cloaca in L. asiaticus) (Gagarin, Nguyen Vu Thanh, 2005).

Etimology. Species name means “short-tailed” or “with short tail.”

Order Monhysterida Filipjev, 1929. Family Xyalidae Chitwood, 1951. Genus Paramonhystera Steiner, 1916.

Paramonhystera affinis sp. n. (Figs. 3, 4).

Fig. 3.
figure 3

Female (a, b, c, d) and male (e, f) Paramonhystera affinis sp. n.: (a) general view, (b) head, (c) body in the vulva area, (d, e) tail, (e) spicule, and (f) gubernaculum.

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Fig. 4.
figure 4

Male (a, c, d, f, h, i, j, l) and female (b, e, g, k) Paramonhystera affinis sp. n.: (a, b) general view, (c) anterior end of the body, (d, e, f) head, (g) body in the vulva area, (h, i) body in the cloaca area, (j, k) tail, and (l) tail terminus.

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Material. Holotype ♂, slide no. PL 3.2.12; paratypes: One ♂, four ♀♀. The holotype and paratype preparation of the male is stored in the collection of the Nature Museum of the Vietnam Academy of Sciences and Technologies (Hanoi, Vietnam). Paratype preparations of females are stored in the collection of nematodes at the Institute of Ecology and Biological Resources of the Vietnam Academy of Sciences and Technologies (Hanoi, Vietnam).

Location. Northern Vietnam, Quảng Ninh Province, artificial pond for shrimp aquaculture (20°48ʹ84ʹʹ N, 106°56ʹ02ʹʹ E). Depth 0.3 m, silty sand, water salinity 24‰. Samples were taken in March 2016.

Description. The morphometric characteristics of the holotype and paratypes are given in Table 2.

Table 2.   Morphometric characteristics of Paramonhystera affinis sp. n.
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Male. The body is thin and relatively short. The anterior end of the body is narrowed. The cuticle is finely annulated, thin. The cuticle thickness in the middle part of the body is ~1.0 μm. Somatic setae rare and short, 1.0–1.5-μm length. Six internal labial sensillae in the form of small papillae. Six external labial sensillae and six head sensillae in the form of thin bristles 11-μm-length; they are arranged in one circle. Cheilostoma is relatively large. Faringostoma in the form of a funnel. Amphid fovea in the form of a circle 5 μm in diameter (30% of the body width at this level) are located 10 μm from the anterior end of the body. The pharynx is muscular, only slightly expanding to its base. The cardium is small, surrounded by three oval formations having a glandular structure. Renette, its ducts and pore not found. One testis, straight, located to the right of the middle intestine. The spicules are thin, slightly curved, with small heads. Their length exceeds the body width twofold in the area of the cloaca. Two gubernaculums in the form of a tube or “socket” cover the distal ends of the spicules. The apical ends of the gubernaculums are armed with clawlike formations. The gubernaculum is about half the length of the spicules. Precloacal complementary organs not found. The tail is long, consisting of two parts. It is conical at the base, then thin, flagellum-shaped. Flagellum length reaches 75% of the total tail length. Caudal setae short. The tail terminus is armed with three long setae. Caudal glands and spinneret are present.

Females. Similar to males in general morphology. The cuticle is finely annulated. The inner labial sensillae is in the form of papillae. Six external labial sensillae and six head sensillae in the form of setae of the same size; they are arranged in one circle. Cheilostoma large; pharyngostoma in the form of a funnel. Amphid fovea in the form of a circle with a diameter of ~4 μm located 8–11 μm from the anterior end of the body. The pharynx is muscular, slightly expanding to its base. There are three oval formations having a glandular structure around the cardium. The length of the rectum is smaller than the width of the body in the anus area. The ovary is single, anterior, and straight; it is located to the left of the intestine. The vulva is postequatorial, in the form of a transverse fissure. The vagina is short, tilted to the anterior end of the body. The uterus is extensive. One female carries one egg (19 × 18 μm) in the uterus. Posterior uterus and postvulvar cell not found. The tail is long and consists of two parts. Its anterior segment is conical; the posterior one is thin, in the shape of a flagellum. Flagellum reaches 68–72% of the total tail length. The tail terminus carries three long bristles. Caudal glands and spinneret are present.

Differential diagnosis. Recently, genus Paramonhystera Steiner, 1916 comprises 24 valid species (Gagarin, Nguyen Thi Thu, 2008; Bezerra, 2018). Paramonhystera affinis sp. n. is the closest morphologically to the two species: P. parabütschlii (Timm, 1961) Gagarin, Nguyen Thi Thu, 2008 and P. levicula (Lorenzen, 1972) Lorenzen, 1973. The first species is described from the Bay of Bengal, one male (Timm, 1963). P. affinis sp. n. differs from this specimen by a slender body (a = 37 versus a = 22 in P. parabütschlii), larger number of the head setae (6 versus 4 in P. parabütschlii), longer external labial setae (10–12 μm in length versus 7–8 μm in P. parabütschlii), and shorter spicules (43 μm in length versus 50 μm in P. parabütschlii) (Timm, 1963). It differs from P. levicula by a larger number of external labial setae (six versus four in P. levicula); shorter and sparse somatic setae, especially on the anterior end of the body and on the tail (their length is ~1 μm versus 3–5 μm in P. levicula); the vulva is located much far from the anterior end of the body (V = 58.6–60.3% versus 53% in P. levicula); and there are shorter spicules (43 μm in length versus 64 μm, and twofold versus fourfold body diameter at cloaca area) (Lorenzen, 1972).

Etymology. Species name means “similar.”