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.

First reported case of intraocular Gnathostoma spinigerum in Cambodia.

A live intraocular nematode was identified from a 37 year-old man presented with iritis, pain, redness, lacrimation, swelling, vision loss and intermittent blindness during many hours per day of the left eye. By using slit lamp examination, a worm was removed from iris in an ophthalmology outpatient department setting and sent to the Medical Microbiology Laboratory, Institut Pasteur du Cambodge. Gnathostoma spinigerum was identified, based on its typical morphology via microscopic examination. Based on our diagnosis, the patient was treated by oral albendazole and responded well to this therapy.

Description of advanced third-stage larvae of Gnathostoma lamothei Bertoni-Ruiz et al. 2005 (Nematoda: Gnathostomatidae) from experimental hosts and contributions to its life cycle.

The advanced third-stage larvae (AdvL(3)) of Gnathostoma lamothei was obtained from experimental hosts. Frogs Lithobates heckscheri and snakes Nerodia fasciata pictiventris were compatible hosts allowing optimal larval development. AdvL(3) are 4,487.94 µm long, have two lateral cervical papillae between rows 10 and 16 and an excretory pore at row 23. The average counts of the cephalic bulb hooklets from the four rows are 39.3, 43.3, 44.2, and 47.3. Larvae show an esophagus that represents 40 % of the body width. These findings indicate that amphibians and reptiles could be involved as G. lamothei natural hosts; nevertheless, their role as etiological agents of human gnathostomiasis is uncertain. This paper reports for the first time the taxonomic description of G. lamothei AdvL(3) obtained from experimental hosts and contributes to the understanding of its life cycle.

Ultrastructural changes in the advanced third larval stage of Gnathostoma binucleatum Almeyda-Artigas, 1991 (Nematoda: Spirurida) following albendazole and albendazole sulfoxide in vitro exposure.

We report here the ultrastructural changes in the third advanced larval stage (AL3) of Gnathostoma binucleatum Almeyda-Artigas, 1991, induced by in vitro treatment with albendazole and the metabolite albendazole sulfoxide. During the whole period of the experiment, the controls remained active and vital throughout the 108-h exposure period. The primary site of action of both drugs appears to be the mitochondria and myofibrils of the polymyarian musculature. Degenerative changes in the secretory portion of the esophagus and somatic musculature were clearly observed. These degenerative changes, leading to complete muscular and glandular necrosis, were interpreted as the consequence of drug-induced blockade of metabolic energy production. Cuticular disruption had also been observed in the experimental groups. We observed a dramatic reduction in the number of mitochondria and muscle fibers, as well as the formation of large autophagic vesicles as result of an irreversible degenerative phenomenon. The passive cutaneous elimination of larvae in human infections is probably a consequence of parasite immobility, due to the effects of this antihelminthic drug.

Gnathostoma binucleatum: pathological and parasitological aspects in experimentally infected dogs.

Lesions and antibody kinetics produced by inoculation of Gnathostoma binucleatum larvae into dogs are described, as well as the morphology of the recovered parasites. In four out of five infected bitches parasite phases were found in the stomach. Only one bitch eliminated eggs and adult parasite phases in feces. In this bitch, the prepatency period lasted 22 weeks and the patency period 14 weeks. Necropsy results showed a copiously vascularized 8-cm diameter fibrous nodule lodged in the greater curvature of the stomach. Two bitches that eliminated no eggs showed 1- to 2-cm diameter nodules on the gastric wall, with five juvenile phases in each. One bitch that eliminated no eggs and exhibited no gastric nodules showed juvenile parasites on the gastric wall. Results confirm dogs as definitive hosts of this parasite. New data on the pathological and parasitological aspects of canine gnathostomosis are presented.

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.

[Gnathostoma larvae investigation in fresh water fishes in the valley of Chinese Chang River and the morphological characteristics of the larvae].

An investigations were carried out on the prevalence of larval gnathostomes in fresh water fishes in southeast area of the Yang-tze Valley, the people's Republic of China, from October to November. The fishes were collected from five districts which were from Shanghai, Chenchiang, Nanching, Chiuchiang and Nanchang, especially the Shanghai district involved Kunshan, Tienshanhu, Chingpu and Nanhui areas. Species of the fishes collected were Channa argus (110), Siniperca chuatssi (24) and Silurus asotus (39). A total number of fishes examined were 137. Muscles of the fishes were each dissected into small pieces and sliced. These slices were examined under a dissecting microscope. The fish viscera from 4-5 individuals were mixed, homogenized and digested by the artificial gastric juice at 37 degrees C overnight. As a result, four encysted larvae were recovered from the muscles of four fish of C. argus. 34 larvae in total were obtained by the digesting. Those larvae mainly recovered from the viscera. The morphological examination showed those 38 larvae could divide into three groups by their body lengths such as 0.58-0.86 mm (group A), 1.12-2.61 mm (group B) and 4.86 mm (group C). Group A consisted of 5 which belong to the early third-stage parasitologically which usually lives the first intermediate host. The majority (32) belongs to group B. Unfortunately, only one larva was obtained in group C which belongs to the advanced third-stage which usually lives in the second intermediate host. Light and scanning electron microscopy revealed that group A and B had the characteristics of G. hispidum and group C had those of G. spinigerum. Next, we investigated fishes near the tow big lakes, Hongtze-hu and Tai-hu situated along the river. In these lakes 12 genera, 12 species fishes (533 individuals) were captured. From these fishes 18 ganathostome larvae collected. Most of the larvae obtained from C. argus and Monopterus albus. All these larvae belongs group B which identified as G. hispidum. In conclusion, many fishes have only G. hispidum of stage of the early third in their viscera.

Intrahepatic growth and maturation of Gnathostoma turgidum in the natural definitive opossum host, Didelphis virginiana.

Gnathostoma turgidum is a gastric nematode parasite of opossums found in the Americas. We recently found that G. turgidum juveniles appear in the liver of the opossums where they become mature adults and almost synchronously move to the stomach during certain months of the year, suggesting the importance of the liver for the growth and maturation of this species in the final hosts. In this study we attempted to detect G. turgidum larvae in the liver of opossums, Didelphis virginiana that are the natural final hosts. The results show that tiny (<3mm in length) third stage larvae (L3) appeared in the liver of opossums around November and December. Also in the liver, we found large L3 of up to about 10mm in length together with juveniles and mature adults from February to March. In spite of their length, large L3 have 4 rows of hooklets, and their gonads remained undeveloped. Morphological features of the small and large L3 of G. turgidum are described including scanning electron microscope images. The seasonal switching of the several growth stages of G. turgidum from small L3 to adult worms in the liver and eventual migration to the stomach in opossums suggests the unique feature of G. turgidum utilizing the liver as the maturation site.

Ultrastructural effects of albendazole on the body wall of Gnathostoma spinigerum third stage larvae.

This study investigated the effects of albendazole (ABZ) on the ultrastructure of Gnathostoma spinigerum advanced third-stage larvae. Two groups of experimentally infected mice received 60 or 90 mg/kg ABZ orally once a day for 21 consecutive days. Both groups had damage to the body walls of the worms, especially to the non-contractile part of the muscular layer. The severity of the damage was dose related, the higher the dose, the greater the damage. The body wall of the ABZ treated larvae demonstrated a decrease in the number of mitochondria in the non-contractile muscular part, especially in the internal surface of the sarcolemma. Some mitochondria developed large vacuoles, and became distorted and degenerated. The nuclei degenerated and had irregular shapes and the number of glycogen granules decreased. The present study demonstrates the structural damage induced by the toxic effects of ABZ and increases our knowledge of the mechanism of action of ABZ against G. spinigerum.

Discovery of an endemic area of Gnathostoma turgidum infection among opossums, Didelphis virginiana, in Mexico.

Gnathostomosis, caused by Gnathostoma binucleatum, is a serious public health issue in Mexico. Although 2 other Gnathostoma spp., G. turgidum and G. lamothei, have been found in wild animals, their natural life cycle or their relation to human disease remains unclear. While we were conducting an epidemiological survey on Gnathostoma spp. in Sinaloa State, Mexico, we found an endemic area for G. turgidum in common opossums, Didelphis virginiana, located in Tecualilla, Sinaloa. The species identification was carried out by morphological and molecular biological methods. This is the first record of an endemic area for G. turgidum infection in opossums, D. virginiana, in the Americas.

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.

Estuarine fish and turtles as intermediate and paratenic hosts of Gnathostoma binucleatum in Nayarit, Mexico.

Human gnathostomosis is a severe public health problem in the State of Nayarit, Mexico. Between 1995 and 2005, the registration of human cases numbered 6,328, which makes it one of the largest focal points of the disease in the country. The present study determined the presence of natural hosts of Gnathostoma binucleatum larvae at the Laguna de Agua Brava in Nayarit, Mexico. A total of 5,450 fish and 247 turtles were sampled. Muscular tissue was ground and observed against the light using a 100-W lamp to identify advanced third-stage larvae. The estuarine species Cathorops fuerthii, Pomadasys macracanthus, Mugil curema, and Dormitator latifrons were found positive for presence of larvae, and annual prevalence was 4.8, 1.83, 2.16, and 4.0%, respectively. The species Oreochromys aureus and Chanos chanos were negative. The species of estuarine turtles Kinosternum integrum and Trachemys scripta were positive with annual prevalence of 79.1 and 52.5%, respectively. The criteria of identification of the Gnathostoma species were: mean number of nuclei in intestinal larval cells (2.3), larval morphometry with optic microscopy, larval morphometry with scanning electron microscopy, and number and sequence of ribosomal deoxyribonucleic acid of adult parasites obtained from experimental infection in dogs. The estuarine fish Pomadasys macracanthus and Mugil curema are reported as intermediate hosts for the first time and likewise the estuarine turtle Kinosternon integrum as a paratenic host.

Larval Gnathostoma hispidum detected in the red banded odd-tooth snake, Dinodon rufozonatum rufozonatum, from China.

A total of 205 larval gnathostomes were collected from 18 (22.5%) of 80 red banded odd-tooth snakes, Dinodon rufozonatum rufozonatum, which had been smuggled from China and confiscated at Customs in Busan, Republic of Korea. In order to identify the species, some of the larvae were observed by a light microscope and a scanning electron microscope (SEM). The larvae were 2.18 x 0.29 mm in average size, and had a pair of lips at the anterior end, a muscular esophagus, 2 pairs of cervical sacs, and brownish intestines. The head bulb was characteristically equipped with 4 rows of hooklets; the average number of hooklets in each respective row was 38.6, 40.5, 41.5, and 43.7. In SEM views, the mouth evidenced a pair of lateral lips of equal size in a half-moon shape. Each lip featured a couple of labial papillae and a small amphid located between the 2 papillae. The hooklets on the head bulb had single-pointed, posteriorly-curved tips. The cuticular spines were larger and more densely distributed on the anterior part of the body, and decreased gradually in size and number toward the posterior body. On the basis of these morphological characteristics, the larvae were identified as the third stage larvae of Gnathostoma hispidum.

Intravitreal gnathostomiasis and review of literature.

Intravitreal gnathostomiasis is a rare disease. To our knowledge, only five case reports are available in the literature wherein the parasites were surgically removed from the vitreous cavity and the species were identified. In this report, we document the clinical features of intravitreal gnathostomiasis along with the route of entry and parasitic migration pattern in ocular tissue, surgical management, and electron microscopic features of the retrieved advanced third-stage larva of Gnathostoma spinigerum (aL3) for a 48-year-old Indian woman with excellent postsurgical visual recovery.

A new species of Gnathostoma (Nematoda: Gnathostomatidae) in Procyon lotor hernandezii from Mexico.

Gnathostoma lamothei n. sp., inhabiting the stomach of Procyon lotor hernandezii Wagler, 1831, in Tlacotalpan, Veracruz State, and Rio Sapo, Oaxaca, Mexico, is described. This new species differs from all other congeners by having the posterior half of the body surface covered by rows of tiny round bosses instead of spines, or lacking ornamentations. Sequences of the ITS2 of the ribosomal DNA of G. lamothei n. sp. are compared with sequences of other species of the genus recorded in Mexico; they show a wide divergence (<50%) with Gnathostoma binucleatum Almeyda-Artigas, 1991, and Gnathostoma turgidum Stossich, 1902, and high similarity with Gnathostoma sp. I sequence (99.2%). On the basis of morphometric traits and sequences, previous records of Gnathostoma sp. I (=Gnathostoma procyonis of Almeyda-Artigas et al., 1994, not Chandler, 1942, and Gnathostoma neoprocyonis nomen nudem) in Mexico are referred to as the new species.

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.

Case report: Ocular gnathostomiasis in northwestern Mexico.

A 42-year-old woman from Culiacan in the Sinaloa State of Mexico presented with a four-year history of migratory, pruritic, painful swellings of the face. Palpebral edema with conjunctival erythema developed when lesions developed near the eye. Routine eye examination showed a mobile worm in the anterior chamber. Following surgical removal, the parasite was identified by light and electron microscopy as an advanced third-stage larva of Gnathostoma sp. This prevalence of this helminthiasis may be increasing in areas where eating freshwater raw fish ("cebiche") is customary.

Morphology of Gnathostoma spp. isolated from natural hosts in Sinaloa, Mexico.

Gnathostomosis is an emerging public health problem in Sinaloa, Mexico, where an increasing number of human cases have been diagnosed since 1989. The present study was carried out to determine the presence of the parasite in other natural hosts from the area. Birds, fish, opossums and raccoons were captured from local dams and lagoons. The flesh from bird and fish specimens was ground and examined under a 100 W light bulb. Larvae were processed for light and electron microscopy. A total of 368 advanced stage 3 (AL3) larvae were found in 300 ichthyophagous birds, with Egretta alba exhibiting the highest infection rate. A total of 4,156 fish were examined, of which six species were infected with AL3 larvae: Arius guatemalensis (blue sea catfish), Dormitator latifrons (Pacific fat sleeper), Gobiomorus sp. (fat sleeper), Oreochromis sp. (Nile tilapia), Cichlasoma beani (Sinaloan cichlid or green guapote) and Eleotris picta (spotted sleeper). Twenty larvae from birds were used to infect domestic cats and dogs. Young adult worms were recovered from the stomach of a cat with a 17 day infection and from a dog with a 35 day infection. Larvae exhibited four rows of hooklets on the head bulb, whereas the young adults had nine rows of hooklets. The cuticular spines of adult worms along the body evolved from single-pointed, bi- or trifurcated spines. Nuclei were counted in intestinal cells examined in serial sections of larvae recovered from a great heron and a fish, in which a mean of 1.6 nuclei/cell was found, corresponding to data published for Gnathostoma binucleatum. Although the external morphology of both larvae and adults are in agreement with previous descriptions of Gnathostoma spinigerum, the results indicate that natural host infections in Sinaloa may be caused by either G. spinigerum or G. binucleatum.

Scanning electron microscopic observations on advanced third-stage larva of Gnathostoma spinigerum after in vitro exposure to albendazole sulphoxide.

Gnathostomiasis is the parasitic disease caused by the migration of an advanced third-stage larva of Gnathostoma spinigerum. To date, albendazole is claimed to be the effective drug in preventing the reoccurrence of migratory swelling in patients. After being exposed to 1 and 2 micrograms/ml albendazole sulphoxide (AlbSO) in vitro, the parasites moved deteriorately, however, no dead larva was found even exposed to these concentrations for 21 consecutive days. The topographical alterations after 21 days of albendazole sulphoxide exposure are described using a scanning electron microscope. The marked changes in surface morphology were observed in both neck and body regions. The tegumental surface on the neck region was swollen and covered with fuzzy materials, whereas, the spines on the posterior region of the body were dislodged. These changes would probably lead to reduction of intermittent cutaneous migratory swelling in human gnathostomiasis patients.