Tribocytic Organ Research Articles

Tegumental topography and molecular characterisation of two trematodes (Platyhelminthes: Digenea) from Clarias gariepinus (Burchell, 1822) in Kenya

The discoveries of new taxonomic features of digenean species through the application of contemporary techniques, such as scanning electron microscopy (SEM) and molecular analysis are still growing. Two species of trematodes Glossidium pedatum and Tylodelphys mashonensis from the intestine and vitreous humour of Clarias gariepinus were recovered from Lake Ol’Bolossat, Kenya. The two endo-helminths were prepared for morphological examination using SEM and molecular characterisation. Additional morphological features were observed for G. pedatum such as domed papillae in the anterior extremity and a protruding cirrus which was unarmed, laterally folded and with a blunt tip as the first such observation for the genus and led to additional characteristics of the diagnosis of the genus. Tylodelphys mashonensis was characterised by a round oral sucker and tribocytic organ rounded with rows of papillae symmetrically arranged. The molecular analyses using ribosomal marker 28S large subunit (LSU) rDNA and mitochondrial (mtDNA) cytochrome c oxidase subunit 1 (cox1) for both G. pedatum and T. mashonensis confirmed the identity of the species and their phylogenetic relationship within the subclass Digenea. This study provides the first mitochondrial (mt)DNA sequence for G. pedatum and also extends the geographical record of two parasites to Kenya.

Intestinal Flukes Recovered from a Herring Gull, Larus argentatus, in the Republic of Korea.

Trematode specimens were collected from the intestine of a herring gull, Larus argentatus, which was found in a critical condition on the shore of a small island (Yubu-do, Seocheon-gun, Chungcheongnam-do) located at the western coast of the Korean peninsula. Total 11 specimens of intestinal flukes, including 3 Cryptocotyle lingua (Heterophyidae), 1 Himasthla alincia (Echinostomatidae), 5 Cardiocephaloides medioconiger (Strigeidae), and 2 Diplostomum spathaceum (Diplostomidae), were recovered. C. lingua was morphologically characterized by the presence of a large ventrogenital apparatus and 2 obliquely tandem testes. H. alincia had an elongated body and a head collar equipped with 31 collar spines. C. medioconiger had a bisegmented body and a voluminous copulatory bursa containing the seminal vesicle and ejaculatory duct. D. spathaceum also had a bisegmented body and its vitellaria extended up to the anterior border of the tribocytic organ. It is of note that C. lingua is potentially zoonotic that can occur in birds and humans. Three of them, i.e., C. lingua, C. medioconiger, and D. spathaceum, are new trematode fauna in Korea. Studies on trematode fauna of migratory birds should be continued in Korea.

Austrodiplostomum ostrowskiae n. sp. (Digenea: Diplostomidae: Diplostominae) from the Double-crested Cormorant, Phalacrocorax auritus (Phalacrocoracidae) from the Galveston, Texas Area of the Gulf of Mexico, U.S.A.

Approximately 1,350 specimens of an undescribed species of Austrodiplostomum (Diplostomidae: Diplostominae) were collected from 10 double-crested cormorants, Phalacrocorax auritus (Phalacrocoracidae), from the Galveston, Texas, U.S.A., area. Austrodiplostomum compactum is synonymized with Austrodiplostomum mordax, the type species in the genus, leaving A. mordax as the only species remaining. Austrodiplostomum ostrowskiae n. sp. can be distinguished from A. mordax by having a small but distinct hindbody, shorter ceca, a uterus and vitelline fields that extend into the tribocytic organ, smaller eggs, and an anterior testis that is about the same width as the posterior testis in A. ostrowskiae n. sp. Five specimens of the neotropical cormorant, Phalacrocorax brasilianus, were also examined from the Galveston, Texas, U.S.A., area during the study, but this host species was not found to be infected with a species of Austrodiplostomum.

Neodiplostomum leei n. sp. (Digenea: Neodiplostomidae) from chicks infected with metacercariae from the grass snake Rhabdophis tigrina.

Neodiplostomula from the European grass snake Rhabdophis tigrina, in the Republic of Korea, were thought to represent 2 species: Pharyngostomum cordatum (large-sized neodiplostomula), an intestinal trematode of cats, and Neodiplostomum seoulense (small-sized neodiplostomula), an intestinal trematode of humans and rodents. The present study describes N. leei n. sp. (Digenea: Neodiplostomidae) on the basis of adult flukes recovered from the small intestines of chicks experimentally infected with small-sized neodiplostomula from the grass snake. The new species differs from N. seoulense in terms of the more extensive distribution of vitellaria, the more severe bilobation of 2 testes (each having a dumbbell shape), the absence of a genital cone, and the smaller size of the tribocytic organ and eggs. Neodiplostomula of the new species and N. seoulense are almost indistinguishable. The new species develops into an adult in chicks, whereas N. seoulense matures in mice and rats. Neodiplostomula of the new species migrate to the liver in mice and rats, without maturing. Results show that there are at least 3 species of neodiplostomid trematodes in the grass snake in the Republic of Korea, i.e.. P. cordatum, N. seoulense and N. leei n. sp.

Revision of Bursacetabulus (Diplostomidae: Diplostominae) with the Proposal of Bursatintinnabulus n. gen., and Description of Bursatintinnabus bassanus n. sp. and Bursacetabulus morus n. sp. from Northern Gannet, Morus bassanus (AVES), from the Texas Gulf Coast

During a study of digeneans of shorebirds from the Texas gulf coast, 2 undescribed species of diplostomes were found in northern gannet, Morus bassanus. Bursatintinnabulus n. gen. (Diplostomidae) is established with reassignment of Bursacetabulus macrobursus Dronen et al., 1999, as type species, and the second species in the proposed genus is described, Bursatintinnabulus bassanus n. sp. Generic diagnosis of Bursacetabulus Dronen et al., 1999, is emended to include a conical hindbody, an inconspicuous pouchlike or conspicuous well-developed tribocytic organ, and digitiform vitellaria distributed mainly in the hindbody with processes extending into the tribocytic organ and ventrolaterally in the hindbody to the level of the testes. Bursacetabulus morus n. sp. is described as the second species in that genus. In Diplostominae, Bursacetabulus and Bursatintinnabulus n. gen. are most similar to Tylodelphys Diesing 1850, but can be distinguished by having smooth testes and vitellaria that extend ventrolaterally into the hindbody to the level of the posterior testis; and the absence of a genital cone, an acetabulum, and a prepharynx. Bursatintinnabulus n. gen. is different from Bursacetabulus and all other genera of Diplostominae by a well-developed, bell-like skirt surrounding the bursa.

Localization of worm antigen in Neodiplostomum seoulense by immuno-electronmicroscopy.

The localization of worm antigen of Neodiplostomum seoulense was examined by immuno-electronmicroscopic observation. Not only the immunized serum of mice with crude worm extract of N. seoulense but also serum of infected mouse were reacted to the worm section. Using immunized serum as primary antibody, the gold particles were deposited on the rough endoplasmic reticulum of the cell of tribocytic organ, spermatozoa in the seminal vesicle, microvilli of the caecum and vitelline follicle. Using infected serum, gold particles were deposited only on the vitelline follicle prominently. This finding suggested that the tribocytic organ, seminal vesicle, caeca and vitelline follicles may play a role of antigen to immunized serum with crude worm extract of N. seoulense, whereas the vitelline follicle, to the infected serum.

Proterodiplostome Parasites (Digenea, Proterodiplostomidae) of the Caiman, Caiman crocodilus yacare (Reptilia, Crocodylia) in the Pantanal mato-Grossense, Brazil, with the description of two new species

Two new species are described from the caiman. Caiman crocodilus yacare. Proterodiplostomum breve n. sp. differs from all other species in the genus by the following chacacteristics: (1) the paraprostate gland is shorter and club-shaped; (2) the genital cone is, in average, eight times longer than that of P. medusae; (3) the genital atrium is larger and without pseudosuckers; (4) the oral sucker and pharynx are longer; and (5) there are larger numbers of papillae surrounding the tribocytic organ (40) against 20 in P. longum, 16 in p. tumidilum, 8 in P. ophidum, and 16-18 in P. medusae. Proterodiplostomum globulare n. sp. differs from all the other species in the fenus by the following characteristics: (1) from P. tumidilum, P. lomgum, P. medusae, and P. breve n. sp. for the absense of pseudosuckers or muscular bunches in the inferior wall of the genital atrium; (2) the shape of the paraprostate gland, which is globular and not cylindrical as in P. longum, P. tumidilum, P. medusae, and P. ophidum; (3) the size of the tribocytic organ 201-407 long, 183-495 wide, while is 138-270 long, 102-292 wide in P. medusae, and 138-270 long, 255 wide in P. ophidum; (4) the number of papillae in the tribocytic organ (18-20) in P. globulare and 16-18 in P. medusae, and 8 in P. ophidum. Specimens belonging to six other species of proterodiplostomes are recorded for the first time infecting the caiman, C. c. yacare in the Pantanal Mato-grossense, Brazil, namely: Proterodiplostomum medusae, P. tumidilum, Cystodiplostomum hollyi, Prolecithodiplostomum constrictum, Paradiplostomum abbreviatum, and Herpetodiplostomum caimancola.

Activities of acid phosphatase and non-specific esterase are present in the tribocytic organ and the caecum of Fibricola seoulensis.

In order to know the enzyme activities of Fibricola seoulensis, an intestinal trematode of human and rodent in Korea, the enzyme histochemical method is applied. Activities of acid phosphatase (E.C.3.1.3.2) and non-specific esterase (E.C.3.1.1) were present in microvilli and glandular cells of tribocytic organ and the epithelium of the caecum.

Transmission electron microscopic ultrastructure of the tegument of Fibricola seoulensis

An electron microscopic study was performed to observe the ultrastructure of the tegument of F. seoulensis. The outer surface of the tegument was covered with a trilaminated plasma membrane. The electron-dense cytoplasmic layer was 2.5 microns wide in the anterior portion and contained numerous vacuoles, mitochondria and granular materials in its matrix. The basement layer was 330 nm wide or so, and its numerous extensions protruded into the cytoplasmic layer. The sensory organ was composed of a small vesicle of 1.7 x 1.1 microns in dimensions, which possessed a cilium of 1.2 x 0.19 micron in size. The pharynx was composed of the epithelial layer of about 0.5 micron wide, well developed muscle layer and basement layer. The tegument of the oral sucker was composed of a cytoplasmic layer of 0.4-0.5 micron width, a narrow basement layer, a well developed muscle layer and tegumental cells. Some kinds of secretory granules that seemed to be originated from the cells of the oral sucker were observed in the parenchymal portions of the adjacent cells. The tribocytic organ consisted of numerous microvilli. The microvilli were 5 nm wide and heptalaminated. Two types of secretory granules originated from the gland cells of tribocytic organ were observed in the tegument and parenchyme. The tegumental cells were irregular in shape, and of which nuclei were multifarious.

Transmission electron microscopic findings of the tribocytic organ of Fibricola seoulensis.

We observed ultra-structure of the tribocytic organ of Fibricola seoulensis with transmission electron microscope. Microvilli are observed on the surface of the tribocytic organ. Below the muscle layer, we can find three types of cells. Type I cell has electron lucent cytoplasm with a few granules, while type II cell shows electron dense cytoplasm and the particulate granules. Type III cell's electron dense cytoplasm possesses many granules. Of the above three cells, Type I and II cells are believed as tegumental cells. Type III cells are thought as glandular cells specific to the tribocytic organ. This finding can also explain the secretory function of the tribocytic organ of F. seoulensis.

Localization and isozyme patterns of phosphatase in Fibricola seoulensis

The present study was carried out to investigate the localization and isozyme patterns of acid phosphatase and alkaline phosphatase in metacercariae and in adults of F. seoulensis by enzyme-histochemistry method and electrophoresis. Acid phosphatase showed a strong activity at pH 5 in the intestinal caecum of adults, but showed no reactions in the non-substrate control and in the inhibitor-treated control. Alkaline phosphatase showed a strong activity at pH 8 in the intestinal caecum and the tribocytic organ of adults, and in the intestinal caecum and in the genital anlagen of metacercariae. In non-denature PAGE, ten bands of protein fraction from the extracts of metacercariae and twenty-two bands from adults were detected. In denature PAGE, two protein bands having molecular weights of 192 kDa and 123 kDa were detected in the metacercariae, but absent from adult stage. In adults, protein fractions of 27.5 kDa, 24.5 kDa, 21.4 kDa, 18 kDa, 16 kDa and 15 kDa were detected. In non-denature PAGE, isozymes of acid phosphatase showed the most strong activity at pH 5, whereas no activity was shown at pH 2 and pH 7. One isozyme band (62 kDa) of AcPase was recognized in metacercariae and 4 isozyme bands (95 kDa, 85 kDa, 73 kDa and 62 kDa) in adults.

A scanning electron microscope study on the tegument of Proalarioides kobayashii Park, 1940 (Trematoda)

A SEM study was performed on the surface of adult P. kobayashii Park, 1940, recovered from the snake, Elaphe rufodorsata. The anterior part of the worms was cup-shape and equipped with oral, ventral suckers, pseudosuckers, and tribocytic organ, and the posterior one was finger-like and round-ended. The tegument of the anterior body was covered with 3-4 pointed small spines on the mid-ventral surface and 1-2 pointed ones on the lateral surface. Sensory papillae such as type II, dome-shape ones, and papillae with an opening were distributed over the ventral surface of the anterior portion. The round tribocytic organ was bearing small stout spines laterally, whereas the surface which comes in contact with the host tissues consisted of numerous long fibrillar fibers. The lip of the oral sucker contained type II papillae. Lateral margin of the anterior body revealed type III papillae.

Histochemical findings of the tribocytic organ and tegument of Fibricola seoulensis

The tribocytic organ and tegument of Fibricola seoulensis were examined histochemically for the detection of carbohydrates, mucosubstances, amyloid, collagen and alkaline phosphatase. The surface, secretes, gland cells of the tribocytic organ, and the tegument of the worms were positive to periodic acid Schiff (PAS) and PAS with diastase stain but negative to other stains. It was inferred that the tribocytic organ and tegument of F. seoulensis comprise neutral mucopolysaccharides, which may take a protective role against host enzymes. The surface and secretes of the tribocytic organ, and the tegument of the worms were also positive to double bridge PAP for alkaline phosphatase. This fact suggests that they may play a role as both self protective and host tissue lytic functions.

Systematic Analysis of the Diplostomidae and Strigeidae (Trematoda)

A systematic analysis of the genera in the Diplostomidae and Strigeidae was made using the Proterodiplostomidae as the outgroup. The Proterodiplostomidae was the family with the greatest preponderance of primitive characters and its monophyly was supported by the unique paraprostate gland. However, no character state supported the monophyly of the Diplostomidae sensu Dubois, 1970. That paraphyletic taxon was composed of 3 monophyletic groups: the pseudosuckerless Neodiplostomidae n. fam. had the most primitive character states of the 3 and its monophyly was based on characters in the neascus/neodiplostomulum metacercariae; the Bolbophoridae n. fam., with pseudosuckers, had its monophyly supported by characters present in the newly named prodiplostomulum metacercaria; and the emended Diplostomidae, also with pseudosuckers, had the most derived states and its monophyly was supported by characters present in the diplostomulum. The presence of pseudosuckers united the Bolbophoridae n. fam., the emended Diplostomidae, and the Strigeidae as a monophyletic assemblage. The Strigeidae had the most derived characters of these 3 taxa and its monophyly was supported by characters in the tetracotyle and the cup-shaped forebody and bilobed tribocytic organ of the adult. In general, the adult stages of these strigeoid families showed very conserved morphology and it was the metacercariae that possessed the innovations. The conserved adult morphology was typical of what one might expect if the intramolluscan stages were analyzed. Thus, the data were concordant with the view that the mollusc and vertebrate definitive host were the original hosts to the Digenea and that the second intermediate host and metacercaria were more recently intercalated. More specifically, the phylogeny of these groups suggested that ancestral bisegmented strigeoids originally infected reptiles, they subsequently radiated into birds with which they coevolved extensively, and on 5 separate occasions they radiated into mammals. The radiations into mammals were, by all available evidence, preceded by second intermediate host shifts from fish to amphibians.

Growth and development of Fibricola seoulensis metacercariae in tadpoles.

In order to observe the growth and development of Fibricola seoulensis metacercariae, the tadpoles of Rana nigromagulata were experimentally infected with the cercariae. The metacercariae of various developmental stages were recovered from the tadpoles after 2 to 65 days of infection. They were prepared for morphological observation, and were given orally to mice to observe their infectivity. The following results were obtained. All of the tadpoles exposed to the cercariae were observed to harbour the larvae in their abdominal cavity. The young metacercariae of 2 days after infection were 121.1 micrometer long and 63.3 micrometer wide. They grow linearly for the first 14 days to be 262.0 micrometer long and 166.4 micrometer wide. Thereafter, no more growth recognized until 65 days. The larvae of 2 days old were similar with cercarial body and had 2 suckers, a pharynx, two ceca and a primordium of germ cells but no tribocytic organ. On the 8th day, they had tribocytic organ, and their morphology resembled that of mature metacercariae. The metacercariae younger than 10 days could not infect the mice. Only the metacercariae older than 14 days had infectivity. The recovery rate increased by the age of metacercariae from 19.0% in 14 days old to 70.0% in 40 days old. Above findings indicate that the tadpole is indispensible for metacercarial development and it needs at least 2 weeks for maturation. The tadpole is a pivotal host in the life cycle of F. seoulensis for connection between the snail and the frog.

A histopathological study on the intestine of mice and rats experimentally infected by Fibricola Seoulensis.

A histopathological study was carried out on the duodenum of mice and rats experimentally infected by F. seoulensis. Each mouse was infected wit 500 metacercariae and killed after 1, 2, 3 days, 1 and 2 weeks from infection. Each rat was given 1,000 metacercariae and was examined after 1, 2, and 4 weeks from infection. The duodenal tissue sections of mice and rats were stained with hematoxylin and eosin, and PAS stained for the rats of 1 week group. The pathological findings are summarized as below. There were no differences in mucosal findings between the mice and the rats, adn between the location of duodenum, 1 and 5 cm distal to the pylorus. Each worm embraced a villus exclusively with its foliate forebody which was inserted into the intervillous spaces. The fluke pinched villous epithelia using its oral and ventral suckers. The tribocytic organ destroyed the villous epithelia deeply up to the stroma after 3 days from infection. Apparent villous changes were observed in the mice after 3 days from infection. Villous changes were shortening, widening, blunting or fusion. The villous stroma showed edema, microscopic hemorrhage, capillary congestion, dilatation of lymphatics and inflammatory cell infiltration. The cells were lymphocytes, plasma cells, eosinophils and giant cells. Rarely submucosal and transmural inflammation was encountered.

Tegumental Changes of Alaria marcianae (Trematoda) during Migration in the Domestic Cat

Mesocercariae of Alaria marcianae were removed from snake paratenic hosts and fed to domestic cats. The cats were necropsied at specified intervals, the parasites recovered, and studied by SEM. During the prepulmonary phase of migration in cats, mesocercariae recovered from the stomach wall, abdominal cavity, diaphragm, liver, and heart underwent no tegumental modification compared to those fixed immediately after removal from the paratenic host. The mesocercaria resembled the body of an enlarged cercaria. The oral sucker was encircled by 14 or 15 rows of simple, posteriorly directed spines. Similar spines were also located along the margins of the venter and equatorially at the level of the acetabulum. This spinal pattern left two regions, one anterior and one posterior to the acetabulum, devoid of spines. Uniciliate papillae were found in a distinct pattern associated with the spined areas. In the pulmonary phase, the mesocercariae transformed into diplostomula. Earlike lappets on either side of the oral sucker developed with subsequent differentiation of microvilliform processes on their ventral sides. Body spines divided to give rise to three-pronged serrations. A depression on the venter between the acetabulum and posterior end of the body gave rise to the tribocytic organ. Stout, densely-clustered spines arose de novo on this organ. A rudimentary cylindrical hindbody gave the diplostomulum a distinct scoop-shaped appearance. Worms coughed up from the lungs and swallowed were later found in the duodenum. This postpulmonary phase of migration was a quick event and young worms in the duodenum were indistinguishable from those in the lungs. Within 4 days, the diplostomula in the duodenum matured to adulthood.(ABSTRACT TRUNCATED AT 250 WORDS)

Studies on intestinal trematodes in Korea X. Scanning electron microscopic observation on the tegument of Fibricola seoulensis

A scanning electron microscopic study was performed to observe the tegumental surface of adult Fibricola seoulensis. The adult worms were collected from the small intestine of mice 5 days to 3 weeks after experimental infection with the metacercariae. The metacercariae were obtained from the viscera of the snakes, Natrix tigrina lateralis, by artificial digestion technique. The results were as follows: The tegument of anterior body was covered with cobblestone-like cytoplasmic processes and that of posterior body showed finger-like processes. The posterior body had 4-5 large transverse wrinklings which formed many discontinued shallow rugae. The entire surface of anterior body was regularly arranged with the spines of which tips diverged into 3 to 4 points. They were densely packed in anterior mid-median portion of dorsal surface where appeared a few spines indented upto 5 points. Farther laterally and posteriorly from this portion, the pointed spines were more sparse and became single tipped and extended to anterior one-third of posterior body. The posterior surface of oral sucker was armed with 50-60 spines having 2-3 tips and ventral sucker also covered with such spines. On anteriormost dorsal surface arranged 60-70 spade-shaped spines. The tribocytic organ was armed with many stout recurved pile-like spines arranged radially. There were 3 types of sensory papillae. The ciliated knob-like (Type I) papillae were almost bilaterally symmetrical in ventral and dorsal surfaces of anterior body, and abundant especially around bases of oral and ventral suckers, tribocytic organ, and in lateral margins of anterior body. About 24 non-ciliated round swellings (Type II) were observed around each lip of oral and ventral suckers. The plate-1ike elevated papilla without cilium (Type III) was found to distribute only in posterior body. These 3 types of papillae seem to be tangoreceptive and/or rheoreceptive in function when their morphology and distributions are considered.

Demonstration of the existence of a marine cycle in the strigeides: Cardiocephalus longicollis Szidat, 1928 (Trematoda: Strigeidae)

During their investigations on parasitism of fishes and birds in the lagoon of Brusc (Var), the authors have discovered the life cycle of Cardiocephalus longicollis. Cercariae of pharyngeate furcocercous type develop in sporocysts in the digestive gland of the marine prosobranch Nassa corniculum. The cercaria is identified as Cercaria nassae (Dolgikh, 1965) from Nassa reticulata in Black sea; it closely resembles Cercaria nassa (Martin, 1945) from Ilyanassa obsolseta (syn. Nassa obsoleta) at Woods Hole (U.S.A.). Several intermediate hosts have been related; they are the marine fishes Diplodus annularis, D. vulgaris, Boops salpa, Pagellus mormyrus (Sparidae) and Belone belone (Scombresocidae). Metacercariae of tetracotyle type were found in the cavity of optical lobes of the brain. Experimental contamination of fishes: Potamoschistus microps, Blennius sp., Mugil cephalus and Syngnathus abaster has been negative; that of Diplodus annularis and D. sargus, positive. It allowed to follow the development from cercaria to tetracotyle. In Provence natural definitive hosts are Larus argentatus and L. ribidundus; contaminations of these birds with metacercariae experimentally raised in Diplodus annularis and D. vulgaris were successfull. Some observations are pointed out especially the development of the parasite in the definitive host, the organisation of tribocytic organ and the geographical distribution of the parasite.

Uptake and Incorporation of Glucose, Tyrosine, Leucine, and Thymidine by Adult Philophthalmus megalurus (Cort, 1914) (Trematoda), as Determined by Autoradiography

Uptake and Incorporation of Glucose, Tyrosine, Leucine, and Thymidine by Adult Philophthalmus megalurus (Cort, 1914) (Trematoda), as Determined by Autoradiography