Surface ultrastructure of the advanced third-stage larvae of Gnathostoma nipponicum

The surface ultrastructure of advanced third-stage larvae (AL3) of Gnathostoma nipponicum was studied using scanning electron microscopy. The larvae were recovered from the grass snake Rhabdophis tigrina in the Republic of Korea. Parasites had a globular head bulb with a pair of lips at the anterior end and 2 labial papillae and an amphid on each lip. The head bulb was characteristically armed with 3 transverse rows of hooklets, averaging 36, 38, and 43 in number, increasing posteriorly. A total of 213-232 minute unidentate cuticular spines were present along the entire length of the larvae, forming the transverse striations. Two pairs of cervical papillae were located between the 8th and 12th transverse striations, and a pair of body papillae was seen laterally on the posterior third of the body. A pair of caudal phasmids was recognized near the posterior extremity. The surface ultrastructure of AL3 of G. nipponicum is unique compared with that of other species.     

The Jeju weasel, Mustela sibilica quelpartis, a new definitive host for Gnathostoma nipponicum Yamaguti, 1941

Adult gnathostomes were discovered in the stomach of the Jeju weasel, Mustela sibilica quelpartis, road-killed in Jeju-do (Province). Their morphological characters were examined to identify the species. Total 50 gnathostome adults were collected from 6 out of 10 weasels examined. In infected weasels, 4-6 worms were grouped and embedded in each granulomatous gastric tumor, except 1 weasel. Male worms were 25.0×1.4 mm in average size, and had a tail with pedunculate papillae, a spicule, and minute tegumental spines. Females were 40.0×2.5 mm in average size, and had a tail without tegumental spines. Pointed and posteriorly curved hooklets were arranged in 8-10 rows on the head bulb. Tegumental spines were distributed from behind the head bulb to the middle portion of the body. The spines were different in size and shape by the distribution level of the body surface. Fertilized eggs were 65.5×38.9 μm in average size, and had a mucoid plug at 1 pole. These gnathostomes from Jeju weasels were identified as Gnathostoma nipponicum Yamaguti, 1941. By the present study, it was confirmed for the first time that G. nipponicum is distributed in Jeju-do, the Republic of Korea, and the Jeju weasel, M. sibilica quelpartis, plays a crucial role for its definitive host.     

Larval Gnathostoma spinigerum Detected in Asian Swamp Eels,Monopterus albus,Purchased from a Local Market in Yangon,Myanmar

The present study was performed to determine the infection status of swamp eels with Gnathostoma sp. larvae in Myanmar. We purchased total 37 Asian swamp eels, Monopterus albus, from a local market in Yangon in June and December 2013 and 2014. All collected eels were transferred with ice to our laboratory and each of them was examined by the artificial digestion technique. A total of 401 larval gnathostomes (1-96 larvae/eel) were detected in 33 (89.2%) swamp eels. Most of the larvae (n=383; 95.5%) were found in the muscle. The remaining 18 larvae were detected in the viscera. The advanced third-stage larvae (AdL₃) were 2.3-4.4 mm long and 0.25-0.425 mm wide. The characteristic head bulb (0.093 × 0.221 mm in average size) with 4 rows of hooklets, muscular long esophagus (1.025 mm), and 2 pairs of cervical sacs (0.574 mm) were observed by light microscopy. The average number of hooklets in the 1st, 2nd, 3rd, and 4th rows was 41, 45, 48, and 51, respectively. As scanning electron microscopic findings, the characteristic 4-5 rows of hooklets on the head bulb, a cervical papilla, tegumental spines regularly arranged in the transverse striations, and an anus were well observed. Based on these morphological characters, they were identified as the AdL₃ of Gnathostoma spinigerum. By the present study, it has been confirmed for the first time that Asian swamp eels, M. albus, from Yangon, Myanmar are heavily infected with G. spinigerum larvae.     

Detection of Gnathostoma spinigerum Third-Stage Larvae in Snakeheads Purchased from a Central Part of Myanmar

To examine the infection status of freshwater fish with Gnathostoma spp. larvae in Myanmar, we purchased 15 snakeheads, Channa striatus, from a local market in a suburban area of Naypyidaw, the new capital city. Two larval gnathostomes were collected using an artificial digestion technique, and observed by a light microscope and a scanning electron microscope. The size of an intact larva was 2.65 mm long and 0.32 mm wide. The characteristic morphology of the larvae included the presence of a long esophagus (0.80 mm long), 2 pairs of cervical sacs (0.43 mm long), and a characteristic head bulb with 4 rows of hooklets. The number of hooklets in the 1st, 2nd, 3rd, and 4th row was 45, 48, 50, and 52, respectively. Based on these morphological characters, the larvae were identified as the advanced 3rd-stage larvae of Gnathostoma spinigerum. This is the first report of detection of G. spinigerum 3rd-stage larvae in the central part of Myanmar. Our study suggests that intake of raw meat of snakehead fish in Myanmar may result in human gnathostomiasis.     

Discovery of larval Gnathostoma nipponicum in frogs and snakes from Jeju-do (Province), Republic of Korea

A survey was performed to find out the intermediate hosts of Gnathostoma nipponicum in Jeju-do (Province), the Republic of Korea. In August 2009 and 2010, a total of 82 tadpoles, 23 black-spotted pond frogs (Rana nigromaculata), 7 tiger keelback snakes (Rhabdophis tigrinus tigrinus), 6 red-tongue viper snakes (Agkistrodon ussuriensis), and 2 cat snakes (Elaphe dione) were collected in Jeju-do and examined by the pepsin-HCl digestion method. Total 5 gnathostome larvae were detected in 3 (50%) of 6 A. ussuriensis, 70 larvae in 3 of 7 (42.9%) R. tigrinus tigrinus, and 2 larvae in 2 of 82 (8.7%) frogs. No gnathostome larvae were detected in tadpoles and cat snakes. The larvae detected were a single species, and 2.17 × 0.22 mm in average size. They had characteristic head bulbs, muscular esophagus, and 4 cervical sacs. Three rows of hooklets were arranged in the head bulbs, and the number of hooklets in each row was 29, 33, and 36 posteriorly. All these characters were consistent with the advanced third-stage larvae of G. nipponicum. It has been first confirmed in Jeju-do that R. nigromaculata, A. ussuriensis, and R. tigrinus tigrinus play a role for intermediate and/or paratenic hosts for G. nipponicum.     

Divisispiculimermis mirus n. gen., n. sp. (Mermithidae: Nematoda) Parasitizing Midges in Córdoba,Argentina

Divisispiculimermis mirus n. gen., n. sp., a mermithid parasitizing larvae of Chironomus sp. in the Cajón o Grande Stream, Córdoba, Argentina, is described. The new genus differs from all other mermithid genera in having paired spicules which are separated and divided into proximal and distal sectors. The other diagnostic characters of the genus are medium size, nematodes with the cuticle appearing smooth (lacking cross fibers under the light microscope); head separated from the rest of the body by a slight constriction at the level of the amphids, six cephalic papillae, mouth papillae absent, mouth opening posterior to level of cephalic papillae; six hypodermal chords at midbody; weakly S-shaped vagina; postparasitic juvenile with a tail appendage.     

Plerocercoids of Nybelinia surmenicola (Cestoda: Tentacularidae) in Squids,Todarodes pacificus,from East Sea,the Republic of Korea

A visceral helminth of the squid, Todarodes pacificus, is reported from the East Sea, the Republic of Korea. Total 39 squid samples were purchased from a fish market in Jumunjin-eup, Gangneung-si (City) from August 2014 to July 2015 and were examined for helminth parasites with naked eyes and under a stereomicroscope after opening the abdominal cavity with a pair of scissors. Whitish larval worms were mainly found in the stomach and abdominal cavity of the squid. They were detected in 25 (64.1%) out of 39 squids examined, and the infection density was 7 larvae per infected squid. Spatula-shaped larvae were 8.2×2.0 mm in average size, round to slightly flattened anteriorly, with round hatching posteriorly, and had characteristic 4 tentacles with numerous hooklets in the scolex. The larvae were identified as the plerocercoid stage of Nybelinia surmenicola by their morphological features. This finding represents a new host record and the first report of N. surmenicola infection in T. pacificus squids from the east coast of Korea.     

Nematode cysts and larvae found in Achatina fulica Bowdich, 1822

This study describes the morphology of the nematode cysts and larvae found in Achatina fulica (giant African snail) in Brazil. Sixty snails were collected in Mesquita, Rio de Janeiro State. Fourteen of the snails were naturally infected. The cysts were spherical, pink colored and measured 0.97 to 1.57 mm in diameter. In the majority of cases they had a single larvae involved in amorphous material. A total of 222 encysted larvae were recovered. Of these, 30 were utilized in the morphological study. The length of the larvae varied from 2.57 to 5.8 mm and they were classified as small - up to 3.5 mm; medium - from 3.53 to 4.5 mm; and large - greater than 4.52 mm. The average length of the larvae in the three groups was 2.85, 3.87 and 5.23 mm, respectively. The larval cuticle was white, shiny and transversally striated until the posterior end of the body. At the anterior end there is a mouth with three lips, with amphids and papillae, followed by a muscular esophagus with average length of 0.61 mm, terminating in an esophageal bulb and having a nerve ring in the middle third of the esophagus, and an intestine with an opening near the posterior end. The tail begins from this opening and has two types of ends: short and abrupt or long and gradually tapering. The difference in the tail end can suggest sexual dimorphism, although no primordial reproductive structures were observed. These characteristics were not sufficient to identify the larvae, so there is a need for further study.     

Morphological Differences Between Radopholus citrophilus and R. similis

SEM observations of the external morphology of populations of Radopholus citrophilus and R. similis revealed several diagnostic differences. The cloaco-spicular orifice on males of R. citrophilus had three to seven genital papillae (anterior hypoptygmata), whereas males of R. similis were either smooth or had one or two shorter genital papillae (anterior hypoptygmata). Females of R. citrophilus had four annules in the region of the vulval opening, but R. similis had five annules in the same region. The labial disc and lateral lips appeared to be of diagnostic significance, but these areas were more susceptible to artifacts due to fixation. An unknown population of Radopholus from Puerto Rico with a chromosome number of n = 4 was morphologically similar to R. similis. These morphological differences provide additional support that R. citrophilus and R. similis are distinct species.     

Two New Species of Aphelenchoididae (Nemata) from Declining Red Pines in Maryland

Aphelenchoides resinosi n. sp. and Ektaphelenchus joyceae n. sp. are described and illustrated from red pines of the Allegheny plateau of Maryland, USA. The new species were found in trees infested with Bursaphelenchus xylophilus. Primary diagnostic characters of A. resinosi females are constriction of the head, basal stylet knobs, tong postuterine sac, two incisures in the lateral field, and conical tail four to five anal body widths long with a simple terminal mucro. Diagnostic characters of the males are two pairs of subventral caudal papillae and spicule shape: Primary diagnostic characters of E. joyceae females are a slight constriction of the head, six similar lips, conical tail, and short postuterine sac. Diagnostic characters of the males are spicule size and shape, a single row of spermatocytes, and one pair of caudal papillae. Within-tree distributions of A. resinosi and E. joyceae are presented. A total of 70% of both red-needled and chlorotic-needled trees in the study were positive for A. resinosi and E. joyceae. Branch hierarchy was related to the percentage of samples positive for A. resinosi.     

Infection Status of Gnathostoma spinigerum Larvae in Asian Swamp Eels,Monopterus albus,Purchased from Local Markets in Cambodia

Present study was performed to know the infection status of Gnathostoma sp. larvae in swamp eels from Cambodia. We purchased total 30 Asian swamp eels, Monopterus albus, from local markets in Pursat and Takeo Provinces and Phnom Penh on May and November 2017 and May 2018. All collected eels were transferred to our laboratory with ice and each of them was examined by artificial digestion method. A total of 15 larval gnathostomes (1–5 larvae) were detected from 55.6% (5/9) swamp eels in Pursat Province. No larval gnathostomes were found in 21 swamp eels in Takeo Province and Phnom Penh. The advanced third-stage larvae (AdL₃) detected were 2.575–3.825 (3.250) mm in length and 0.375–0.425 (0.386) mm in width. They had the characteristic head bulb (av. 0.104×0.218 mm) with 4 rows of hooklets, long muscular esophagus (1.048 mm), and 2 pairs of cervical sacs (0.615 mm). The number of hooklets in 4 rows on the head bulb was 41, 44, 47, and 50. In scanning electron microscopy, characteristic features were 4 rows of hooklets on the head bulb, cervical papillae, tegumental spines regularly arranged in transverse striations, and anus. The larval gnathostomes were identified as AdL₃ of Gnathostoma spinigerum based on the morphological characters. By the present study, it has been confirmed that G. spinigerum larvae are infected in Asian swamp eels, M. albus, in Pursat Province, Cambodia.     

Scanning Electron Microscope Studies of Steinernema anomali Kozodoi, 1984

This paper presents SEM micrographs of portions of the male, female, and infective-stage juvenile of Steinernema anomali. Included are micrographs of the cephalic and caudal region, spicules, and gubernaculum of the male, the cephalic and vulval region of the female, and the cephalic region of the infective-stage juvenile. Males have six labial and four prominent cephalic papillae and small amphids. There are 11-14 pairs and one single genital papillae; of these, 6-9 pairs are preanal and subventral, one pair preanal, lateral, one pair adanal, and three pairs postanal. Spicules have a short head, a long blade, and a reduced shaft. The distal end is enlarged and bears a dorsal aperture. Gubernaculum much shorter than spicules; cuneus of gubernaculum short and bifurcate anteriorly. Females have six labial and four cephalic papillae and small amphids. Vulva with a thickened posterior lip. Infective juveniles have a smooth head, prominent amphids, and four cephalic papillae. Labial papillae, if present, are not evident.     

Description of Sclerorhabditis miniata n. sp. and First Description of Male of Diploscapter coronatus Cobb, 1913 (Nematoda: Rhabditidae)

A population of Sclerorhabditis miniata n. sp. is described and illustrated from Poonch district of Jammu and Kashmir State, India. The new species is characterized by small body size, with an annulated cuticle, offset labial region, crown shaped, strongly sclerotized lips, thin lateral lips, membranous, stegostom without glottoid apparatus, cheilostom rod shaped, sclerotized, spicules free, strong and thick, gubernaculum simple, bent proximally, bursa open, peloderan with seven pairs of bursal papillae in 1+1/1+1+2+1 pattern. The males of Diploscapter coronatus are described for the first time. They are usually smaller than the females and have labial region similar to females. Spicules separate, with a small dorsal velum, gubernaculum simple, almost straight, bursa open, pseudopeloderan with seven pairs of bursal papillae in 1+1/1+2+1+1 pattern.     

Emendation of the Genus Paurodontella Husain &Khan, 1968 and Description of P. auriculata n. sp. from Western Canada (Nematoda: Paurodontidae)

Males and females of Paurodontella auriculata n. sp. are described and illustrated and the genus emended. The species is characterized by a short, robust body of 355-525 μm, a stylet length of 7-8 μm, lateral field with 5-6 incisures, and a conoid, attenuated tail 1.5 times the vulva-anus distance. Primary characters used in emending the genus are asymmetrical stylet knobs, absence of lateral lips, a deep slit-like amphid dividing the submedian lips to base of head, where open lateral sides are covered by a large auriform cuticular flap, and variable length of the basal esophageal bulb stem and male bursa.     

Paramphistomum cervi: Surface topography of the tegument of adult fluke

Adult Paramphistomum cervi or rumen fluke are pear-shaped, slightly concave ventrally and convex dorsally. The worm measures about 5-13 mm in length and 2-5 mm in width across the mid-section. As observed by scanning electron microscopy (SEM), the tegumental surface in all part of the body, appears highly corrugated with transverse folds alternating with grooves and is spineless. At high magnification, the surface of the fold is composed of microfolds or ridges separated by microgrooves or pits. Corrugations and invaginations of the ventral surface are also more extensive than on the dorsal surface of the body. Both anterior and posterior suckers have thick rims covered with transverse folds without spine. The genital pore is situated at the anterior third of the body. There are two types of sensory papillae on the surface: type 1 is bulbous in shape, measuring 10-15 μm in diameter at the base with nipple-like tips, and type 2 has a similar shape and size and also a short cilia on top. These sensory papillae usually occur in large clusters, each having between 5 and 20 units depending on the region of the body. Clusters of papillae on the ventral surface and around the anterior suckers tend to be more numerous and larger in size. The dorsal surface of the body has the least number of papillae.     

Surface ultrastructure of Pygidiopsis summa (Digenea: Heterophyidae) adult flukes

A scanning electron microscopic study was performed on the surface ultrastructure of Pygidiopsis summa (Digenea: Heterophyidae) adults. Metacercariae were collected from gills and muscles of mullets (Mugil cephalus) caught in a known endemic area, and adult flukes were harvested from dogs after 8 weeks of experimental infection. The worm was calabash form with its posterior part broader than the anterior part. Tegumental spines were densely distributed over the body surface, except on the suckers and genital apparatus, and around the excretory pore. Well differentiated spines were observed on the anterior half of the body, with 14-16 tips ventrally, and 19-20 tips dorsally. On the oral sucker, three pairs of type I sensory papillae (uni-ciliated knob-like swellings) and one pair of type II sensory papillae (aciliated round-swellings) were observed on the anterior and posterior parts of the lip, respectively. On the lip of the ventral sucker, one pair of type II sensory papillae was distributed only on its posterior part. Sperms were seen emerging from or entering into the genital apparatus. The results showed that the surface ultrastructure of P. summa was unique among the heterophyid trematodes, especially in digitation of tegumental spines and in distribution of sensory papillae on oral and ventral suckers.     

Xiphinema llanosum and Trophurus vultus, Two New Plant Nematode Species from Pasture Soils in Colombia

Xiphinema llanosum n. sp. and Trophurus vultus n. sp. are described and illustrated from grass soils in Llanos Orientales, Colombia. Xiphinema llanosum is a bisexual species. The female body length is 2.3-2.7 mm, odontostyle 86-96 μm, and odontophore 58-65 μm long; vulva at 42-47%; anterior ovary is absent; the anterior uterus and oviduct are similar to the posterior branch but slightly reduced; and the tail is dorsally convex-conoid with a blunt hemispherical terminus. Male body length is 2.06-2.96 mm; spicules are 40-44 μm long; and four (rarely three or five) anterior ventromedian supplementary papillae are present. Trophurus vultus females are 0.52-0.67 mm long; vulva at 56-60%; stylet is 10.5-13.5 μm long; isthmus is as long as the basal esophageal bulb; the tail is subclavate, 1.6-2.2 times anal body width long; and the terminal cuticle thickness is about one-sixth of the tail length.     

Identification of Meloidogyne Species on the Basis of Head Shape and,Stylet Morphology of the Male

Head shape and stylet morphology of males of 90 populations of M. arenaria, M. hapla, M. incognita, and M. javanica from geographic regions of the world were compared by light microscopy (LM). In addition, stylets of one population each of M. arenaria, M. incognita, and M. javanica and three different chromosomal forms of M. hapla race A and two of race B were excised and examined with a scanning electron microscope (SEM). Differences among species occurred in both head and stylet morphology. Head morphology differed in size and shape of the head cap, annulation of the head region, and width of the head region relative to the first body annule. Differences in stylets occurred in size and shape of the cone, shaft, and knobs. All populations of M. hapla, except one, had similar head morphology, but stylet morphology was different between cytological races A and B. Populations of M. javanica varied with respect to the presence of head annulations. Head shape and stylet morphology of males are recommended as additional characters useful in the identification of root-knot nematodes.     

Tegumental ultrastructure of juvenile and adult Echinostoma cinetorchis]

The tegumental ultrastructure of juvenile and adult Echinostoma cinetorchis (Trematoda: Echinostomatidae) was observed by scanning electron microscopy. Three-day (juvenile) and 16-day (adult) worms were harvested from rats (Sprague-Dawley) experimentally fed the metacercariae from the laboratory-infected fresh water snail, Hippeutis cantori. The worms were fixed with 2.5% glutaraldehyde, processed routinely, and observed by an ISI Korea DS-130 scanning electron microscope. The 3-day old juvenile worms were elongated and ventrally curved, with their ventral sucker near the anterior two-fifths of the body. The head crown was bearing 37-38 collar spines arranged in a zigzag pattern. The lips of the oral and ventral suckers had 8 and 5 type II sensory papillae respectively, and between the spines, a few type III papillae were observed. Tongue or spade-shape spines were distributed anteriorly to the ventral sucker, whereas peg-like spines were distributed posteriorly and became sparse toward the posterior body. The spines of the dorsal surface were similar to those of the ventral surface. The 16-day old adults were leaf-like, and their oral and ventral suckers were located very closely. Aspinous head crown, oral and ventral suckers had type II and type III sensory papillae, and numerous type I papillae were distributed on the tegument anterior to the ventral sucker. Scale-like spines, with broad base and round tip, were distributed densely on the tegument anterior to the ventral sucker but they became sparse posteriorly. At the dorsal surface, spines were observed at times only at the anterior body. The results showed that the tegument of E. cinetorchis is similar to that of other echinostomes, but differs in the number and arrangement of collar spines, shape and distribution of tegumenal spines, and type and distribution of sensory papillae.