Redescription and life cycle of the monorchiid Postmonorcheides maclovini Szidat, 1950 (Digenea) from the Southwestern Atlantic Ocean: Morphological and molecular data

The adult monorchiid, Postmonorcheides maclovini Szidat, 1950, digenean parasite of the Patagonian blennie Eleginops maclovinus (Cuvier) (Eleginopidae) from Puerto Deseado (47° 45′ S, 65° 55′ W), Argentina, was characterized and its life cycle elucidated. P. maclovinus is the only species of the genus Postmonorcheides, proposed by Szidat (1950) from Tierra del Fuego province (~ 54° S), Argentina. This digenean uses the Patagonian blennie as definitive host, and the intertidal bivalve Lasaea adansoni (Gmelin) (Lasaeidae) as both first and second intermediate hosts (metacercariae encyst inside sporocysts), being the first record of this clam as intermediate host of trematode parasites. The cercaria may, in addition to encysting in the sporocyst, emerge and presumably infect other intermediate hosts. This is the second report of a monorchiid species with metacercariae encysting inside the sporocyst. Adults were found parasitizing the fish stomach, pyloric caeca and intestine with a prevalence of 100%; sporocysts with cercariae and/or metacercariae were found parasitizing the gonad of the bivalve with a prevalence of 2.78%. The cercariae possess a well-developed tail and eye-spots are absent. The ITS1 sequence from the adult digeneans found in the Patagonian blennie, identified as P. maclovini, was found to be identical to the ITS1 sequences obtained both from sporocysts containing cercariae and encysted metacercariae found in L. adansoni.     

The life cycle of Monorchis parvus (Digenea: Monorchiidae) demonstrated by developmental and molecular data

Cercaria cerastodermae I, a digenean parasite of Cerastoderma edule, was recorded for the first time in the Atlantic Ocean off the Iberian peninsula. Sporocysts were present in the hemolymph of the digestive gland, gonad, gills, and foot of the mollusc. Most of the cercariae present within sporocysts were encysted as metacercariae. The corresponding adult stages were obtained after experimental infection of several Diplodus sargus artificially reared in fish farms and that had previously been protected against natural infections. Numerous adult specimens of Monorchis parvus were collected in Diplodus annularis along the French Mediterranean coast. Comparison of wild and experimental adults allowed the adult stage of Cercaria cerastodermae I to be identified as M. parvus. Another monorchid, Monorchis monorchis, a parasite of Spondyliosoma cantharus, was found in the same Mediterranean area and compared with M. parvus. Additionally, ITS1 nuclear ribosomal DNA sequences of C. cerastodermae I and of the adults collected in naturally infected D. annularis and S. cantharus were obtained. Sequence data indicate that C. cerastodermae I corresponds to the adult of M. parvus found in D. annularis and is clearly distinct from M. monorchis found in S. cantharus.     

Morphological and molecular differentiation of Clinostomum complanatum and Clinostomum marginatum (Digenea: Clinostomidae) metacercariae and adults

The separation of Clinostomum complanatum Rudolphi, 1814 and Clinostomum marginatum Rudolphi, 1819 has long been unclear. Recent data confirm the validity of the junior species, C. marginatum , by ～ 1% differences in its 18S rDNA sequences. We collected adults and metacercariae of C. complanatum and C. marginatum and found reliable morphological differences in the genital complex at both developmental stages. In addition, we identified basic morphometrics (distance between suckers, body width) in metacercariae that may be useful for discriminating the species. The morphological differences were supported by the comparison of sequences of internal transcribed spacers of ribosomal DNA and of the mitochondrial gene cytochrome c oxidase I (COI) from 39 specimens. In 36 specimens, the average divergence between the species was 7.3% in ITS and 19.4% in COI sequences. Two specimens from North America and 1 from Europe had sequences that did not allow them to be clearly allied with either species.     

The distribution and incidence of larval trematodes in the freshwater fauna of the Wentloog level, South Wales

The Wentloog level, in South Wales, is a low-lying coastal plain dissected by a network of drainage channels. The latter contain a rich aquatic invertebrate fauna which harbour a wide range of larval digenetic trematodes. Few of the host species examined were not infected with larval trematodes. Miracidia together with their associated sporocysts, rediae and cercariae were more host specific than metacercariae but many of the miracidia which developed in pulmonate molluscs could do so in more than one genus. Metacercarial host specificity is less rigid, and usually species are only restricted to members of one phylum although several were found which could develop in two or more phyla. Most larval stages showed some host preference when they occurred in several host species and this was attributed to ecological or physiological factors or to mechanical barriers. Seasonal variation of infection rate occurs in all larval stages: infection with developing cercariae showed peaks during June and August, but metacercarial infection rates varied according to the host. In molluscs there may be two peaks of infection with metacercariae (as in the cercarial stages), but for many arthropods infections show a gradual increase through the year. The factors mainly responsible for these seasonal changes are considered to be the life-cycle of the intermediate hosts and the environmental temperature, the latter acting on the rate of development of the trematodes. Rainfall, and patterns of egg production of adult trematodes in the definitive hosts are also important.     

Comparative life histories of two diplostomid trematodes, Ornithodiplostomum ptychocheilus and Posthodiplostomum minimum

A lack of comparative studies limits our understanding of interspecific variation in parasite life histories, especially for species that incorporate asexual and resting stages into their life cycles. Ornithodiplostomum ptychocheilus and Posthodiplostomum minimum are sympatric diplostomid trematodes that share the same first (pond snail, Physa gyrina) and second (fathead minnow, Pimephales promelas) intermediate hosts within lakes in Alberta, Canada. Interspecific differences in the body sizes of adults, cercariae, and metacercariae were evaluated from parasites recovered from laboratory-raised chickens, snails, and minnows, respectively. Differences in adult worm fecundity and cercariae production were estimated from chickens and snails exposed to known numbers of parasite larvae. Posthodiplostomum minimum adults, metacercariae, and eggs were 97, 86, and 5% larger than those of 0. ptychocheilus, respectively. The average numbers of eggs produced per day by P. minimum was 3 times higher than for O. ptychocheilus, and P. minimum produced approximately twice as many cercariae per day. Although the larger of the 2 species had higher egg and cercariae production, conclusions regarding covariation between adult worm size and the reproductive rates of other life cycle stages require further studies on related species.     

Gymnophallid digenean Parvatrema duboisi uses Manila clam as the first and second intermediate host

To identify the metacercariae of a gymnophallid trematode in the Manila clam Ruditapes philippinarum from the Ariake Sea, experimental infection and molecular analysis were conducted. Based on the morphology of adult worms obtained from experimentally infected mice, the parasite was identified as Parvatrema duboisi. Comparison of internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) sequences for metacercariae and sporocysts from Manila clams and adult worms collected from wild wigeon Anas penelope showed minor differences ranging from 0 to 0.8%. These data strongly suggest that in the Ariake Sea, the parasite has a lifecycle using the Manila clam as the first and second intermediate hosts and wigeon as the definitive host.     

范尼道佛吸虫的生活史研究

范尼道佛吸虫的生活史包括毛蚴、胞蚴、尾蚴、囊蚴和成虫5个阶段。它的第一中间宿主为淡水壳菜,第二中间宿主为多种小型鲤形目和鲇形目鱼类,终末宿主是鳜。文中对这种吸虫的胞蚴、尾蚴、囊蚴和成虫的形态特征以及尾蚴的行为特征进行了观察和描述。在用人工方法成功地将尾蚴感染青后,对不同发育阶段的囊蚴在鱼体内的形态变化和器官发育进行了观察。       

Experimental Life History and Biological Characteristics of Fasciola gigantica (Digenea: Fasciolidae)

This study was conducted to investigate the life history, morphology, and maturation of larval stages and adult worms of Fasciola gigantica in experimental mice. Lymnaea auricularia rubiginosa was used as the intermediate host, and Oryza sativa was used for encystment of the metacercariae, while Mus musculus was used as the definitive host for maturation study. Fresh eggs from the gall bladder of water buffaloes fully developed into embryonated ones and hatched out at days 11-12 after incubation at about 29ºC. Free-swimming miracidia rapidly penetrated into the snail host, and gradually developed into the next larval stages; sporocyst, redia, and daughter redia with cercariae. Fully-developed cercariae were separated from the redia and shed from the snails on day 39 post-infection (PI). Free-swimming cercariae were immediately allowed to adhere to rice plants, and capsules were constructed to protect metacercariae on rice plants. Juvenile worms were detected in intestines of mice at days 3 and 6 PI, but they were found in the bile duct from day 9 PI. Juvenile and adult flukes were recovered from 16 mice experimentally infected with metacercariae, with the average recovery rate of 35.8%. Sexually mature adult flukes were recovered from day 42 PI. It could be confirmed that experimentally encysted metacercariae could infect and develop to maturity in the experimental host. The present study reports for the first time the complete life history of F. gigantica by an experimental study in Thailand. The obtained information can be used as a guide for prevention, elimination, and treatment of F. gigantica at environment and in other hosts.     

Larval trematodes (Digenea) of planorbid snails (Gastropoda: Pulmonata) in Central Europe: a survey of species and key to their identification

A survey of the larval stages (cercariae and metacercariae) of trematodes (Digenea) found in planorbid snails in Central Europe (Austria, Czech Republic, south-east Germany, Hungary, Poland and the Slovak Republic) is presented based on a study of 7,628 snails of 12 species examined between 1998-2006. A total of 34 trematode larval stages, comprising cercariae of 28 species and metacercariae of seven species (one species occurred both as cercaria and metacercaria) of nine families were found in 898 (11.5%) snails of eight species. The dominant cercariae were those belonging to the Rubenstrema exasperatum (Rudolphi, 1819)/Neoglyphe locellus (Kossack, 1910) species complex, Tylodelphys excavata (Rudolphi, 1803) and Echinostoma spiniferum (La Valette, 1855) sensu Nasincová (1992), all from Planorbarius corneus (Linnaeus). Almost the same spectrum of cercariae of the families Echinostomatidae, Plagiorchiidae and Omphalometridae was found in the present study as in previous reports; however, a considerably lower spectrum of cercariae of the families Diplostomidae and Strigeidae was recorded. The most frequent metacercariae were those of Echinoparyphium aconiatum Dietz, 1909, Neoglyphe locellus and Moliniella anceps (Molin, 1859), all occurring mainly in P. corneus. The most heavily infected snail species was P. corneus, followed by Planorbis planorbis (Linnaeus) and Segmentina nitida (Müller). The widest spectrum of trematode species was found in P. planorbis and P. corneus. Forty-two cercariae identified to the species level belonging to 15 families, plus an additional 43 taxa recorded under generic or provisional names, were reported from 11 species of planorbids in previous studies carried out in Central Europe. However, the actual number of trematode species occurring in the planorbid snails is probably much lower, because many, if not most, larval stages reported under provisional names or unidentified to the species level may be conspecific with identified adult forms. A key to the cercariae and metacercariae recorded from planorbids in Central Europe, together with illustrations of those species encountered most frequently in the field, is provided to facilitate identification.     

Individual and population variation in cercariae of bird schistosomes of the Trichobilharzia ocellata species group as revealed with the polymerase chain reaction

The polymerase chain reaction with arbitrary (RAPD-PCR) or specific primers was used to study the population variation and to identify the species in cercariae of schistosomes of the Trichobilharzia ocellata species group (Trematoda, Schistosomatidae). In total, 28 cercariae were obtained from two spontaneously invaded mollusks Lymnaea stagnalis (LS) and L. ovata (LO), which were collected in different water bodies of Moscow. RAPD-PCR was carried out with two arbitrary primers, OPA9 and OPB11, which each detected different levels of individual and among-group variation and revealed considerable genetic differentiation of cercariae from different host mollusks. To check whether the cercariae of the two samples belong to one species, sequencing was performed with a region corresponding to intergenic transcribed spacer 2 (ITS2), which was earlier proposed for cercaria identification in three European species of bird schistosomes of the genus Trichobilharzia (T. franki, T. regenti, and T. szidati). The ITS2 sequences of two LO cercariae were identical, each consisted of 319 bp, and showed 100% homology to the T. franki ITS2 sequence. The ITS2 sequences of two LS cercariae were identical, each consisted of 323 bp, and showed 99.4% homology to the T. szidati counterpart. The causes of genetic variation in cercariae and prospects of using RAPD markers to study different stages of the life cycle in trematodes are discussed.     

Parvatrema duboisi (Digenea: Gymnophallidae) Life Cycle Stages in Manila Clams,Ruditapes philippinarum,from Aphae-do (Island), Shinan-gun,Korea

Life cycle stages, including daughter sporocysts, cercariae, and metacercariae, of Parvatrema duboisi (Dollfus, 1923) Bartoli, 1974 (Digenea: Gymnophallidae) have been found in the Manila clam Ruditapes philippinarum from Aphae-do (Island), Shinan-gun, Jeollanam-do, Korea. The daughter sporocysts were elongated sac-like and 307–570 (av. 395) μm long and 101–213 (av. 157) μm wide. Most of the daughter sporocysts contained 15–20 furcocercous cercariae each. The cercariae measured 112–146 (av. 134) μm in total length and 35–46 (av. 40) μm in width, with 69–92 (av. 85) μm long body and 39–54 (av. 49) μm long tail. The metacercariae were 210–250 (av. 231) μm in length and 170–195 (av. 185) μm in width, and characterized by having a large oral sucker, genital pore some distance anterior to the ventral sucker, no ventral pit, and 1 compact or slightly lobed vitellarium, strongly suggesting P. duboisi. The metacercariae were experimentally infected to ICR mice, and adults were recovered at day 7 post-infection. The adult flukes were morphologically similar to the metacercariae except in the presence of up to 20 eggs in the uterus. The daughter sporocysts and metacercariae were molecularly (ITS1–5.8S rDNA-ITS2) analyzed to confirm the species, and the results showed 99.8–99.9% identity with P. duboisi reported from Kyushu, Japan and Gochang, Korea. These results confirmed the presence of various life cycle stages of P. duboisi in the Manila clam, R. philippinarum, playing the role of the first as well as the second intermediate host, on Aphae-do (Island), Shinan-gun, Korea.     

Asexual reproduction in Pygospio elegans claparède (Annelida, Polychaeta) in relation to parasitism by Lepocreadium setiferoides (Miller and Northup) (Platyhelminthes, Trematoda)

Life-history theory predicts that parasitized hosts should alter their investment in reproduction in ways that maximize host reproductive success. I examined the timing of asexual reproduction (fragmentation and regeneration) in the polychaete annelid Pygospio elegans experimentally exposed to cercariae of the trematode Lepocreadium setiferoides. Consistent with adaptive host response, polychaetes that became infected by metacercariae of trematodes fragmented sooner than unexposed controls. Parasites were not directly associated with fission in that exposed polychaetes that did not become infected also fragmented earlier than controls. For specimens of P. elegans that were not exposed to trematodes, new fragments that contained original heads were larger than those that contained original tails, whereas original head and tail fragments did not differ in size for infected polychaetes. In infected specimens, metacercariae were equally represented in original head and tail fragments and were more likely to be found in whichever fragment was larger. Despite early reproduction, parasitism was still costly because populations of P. elegans exposed to parasites were smaller than controls when measured 8 weeks later and because exposure to cercariae reduced survivorship of newly divided polychaetes. Taken together, my results suggest that early fragmentation is a host response to minimize costs associated with parasitism.     

Haplorchis pumilio and H. taichui in Vietnam discriminated using ITS-2 DNA sequence data from adults and larvae

Samples of Haplorchis taichui and Haplorchis pumilio of different life-stages (cercariae, metacercariae and adults) and from different host species (snail, fish, dog, cat and human) were collected in Nghe An and Nam Dinh Provinces in Vietnam. Samples from Thailand were available for comparison. All adults and metacercariae were initially identified using morphological criteria. Polymerase chain reaction (PCR) assays were developed for discriminating between the species. The complete sequence for the nuclear ribosomal internal transcribed spacer-2 (ITS-2) was obtained from one adult and one metacercaria of H. taichui and three adults and three metacercariae of H. pumilio from Vietnam. Sequences from cercariae from three different snails clustered with those of H. pumilio. Intra-individual variation in the ITS-2 region was detected by sequencing of cloned PCR products. These are the first sequences from Vietnamese Haplorchis spp. to be reported and demonstrate that H. taichui and H. pumilio can be identified unambiguously from any life-stage, including the cercarial stage that is difficult to identify using morphology. Discrepancies in the literature are discussed and examples of apparent misidentification highlighted. The data provide a resource to assist in taxonomic studies on heterophyids, in the design of probes for diagnosis and for field surveys to identify infection in snails.     

Life history and life cycles: production and behavior of trematode cercariae in relation to host exploitation and next-host characteristics

Life history trade-offs affect trematode parasites reproducing inside their 1st intermediate hosts. Within the constraint of the effect on host survival, parasite production of cercariae is subject to a size-numbers trade-off. Within each cercaria, resources must be partitioned between host-seeking and subsequent developmental functions. Three species of microphallid trematodes with the same 1st intermediate host (the gastropod Littorina saxatilis) were investigated. Maritrema arenaria periodically released many small cercariae. Microphallus similis released fewer, 15% larger, cercariae without periodicity. Microphallus similis cercariae were strong swimmers, moving toward the dark and downward in turbulent water, whereas Ma. arenaria cercariae remained suspended. Maritrema arenaria cercariae, although smaller in body and tail size, were produced at an average daily volume nearly twice that of M. similis. These differences are interpreted as transmission adaptations related to mobility and predictability of the 2nd intermediate host. Microphallus similis, with a mobile and less predictable crab host, adopted a 'bethedging' prolonged production of fewer cercariae by less intensive host exploitation, each cercaria having a high allocation to host-seeking behavior. Maritrema arenaria, with predictable sessile barnacle hosts, produced less mobile but potentially longer-lived cercariae in larger numbers. Microphallus piriformes metacercariae remain in the gastropod host. The number of M. piriformes metacercariae increased in larger hosts. The 3 species differed in the number of sporocysts and (meta)cercariae per sporocyst within the gastropod but not in the within-host volume of parasites. Variation in host exploitation and life history appeared adaptive for transmission to the next host.     

Molecular Description of Macroorchis spinulosus (Digenea: Nanophyetidae) Based on ITS1 Sequences

We performed a molecular genetic study on the sequences of 18S ribosomal RNA (ITS1 region) gene in 4-day-old adult worms of Macroorchis spinulosus recovered in mice experimentally infected with metacercariae from crayfish in Jeollanam-do Province, Korea. The metacercariae were round, 180 μm in average diameter, encysted with 2 layers of thick walls, but the stylet on the oral sucker was not clearly seen. The adult flukes were oval shape, and 760-820 μm long and 320-450 μm wide, with anterolateral location of 2 large testes. The phylogenetic tree based on ITS1 sequences of 6 M. spinulosus samples showed their distinguished position from other trematode species in GenBank. The most closely resembled group was Paragonimus spp. which also take crayfish or crabs as the second intermediate host. The present study is the first molecular characterization of M. spinulosus and provided a basis for further phylogenetic studies to compare with other trematode fauna in Korea.     

Clarification of Cercaria sevillana (Digenea: Microphallidae) life cycle using morphological and molecular data

Cercaria sevillana is the cercaria larval stage that infects the gonads and the digestive gland of its first intermediate host, Nassarius reticulatus. In this study the decapodous crustacean Carcinus maenas was used to determine if it would serve as second intermediate host in the parasite's life cycle. The latter hypothesis was based on the knowledge that C. maenas is the second intermediate host of several other digenean species. After dissection, it was possible to observe encysted metacercariae in the antennal glands of the green crab. After biochemical excystment, the metacercariae were processed for light and scanning electron microscopy. The morphological features observed led us to conclude that this species was a microphallid fluke, and it was identified as Gynaecotyla longiintestinata. To establish a possible relationship between C. sevillana and this metacercariae, the ITS1 region was analyzed. Thus, DNA was extracted from C. sevillana and from the cysts isolated from the antennal glands. The ITS1 region was amplified and sequenced, and the alignment clearly demonstrated that the cercaria and the metacercariae belonged to the same species, G. longiintestinata.