Evolutionary avenues for, and constraints on, the transmission of frog lung flukes (Haematoloechus spp.) in dragonfly second intermediate hosts

Metacercariae survival patterns and their distribution in second intermediate odonate hosts were examined for 4 species of frog lung flukes. Surveys of aquatic larvae and recently emerged teneral dragonflies and damselflies indicated that prevalence and mean abundance of Haematoloechus spp. metacercariae were significantly lower in teneral dragonflies than larval dragonflies, while there was no significant difference in prevalence or mean abundance of Haematoloechus spp. metacercariae among larval and teneral damselflies. Experimental infections of dragonflies indicated that metacercariae of Haematoloechus coloradensis and Haematoloechus complexus were located in the head, thorax, and branchial basket of dragonflies, whereas metacercariae of Haematoloechus longiplexus and Haematoloechus parviplexus were restricted to the branchial basket of these hosts. Metacercariae of H. coloradensis, H. complexus, and H. longiplexus infected the head, thorax, and abdomen of damselflies, but these insects were resistant to infection with H. parviplexus. Subsequent metamorphosis experiments on experimentally infected dragonflies indicated that most metacercariae of H. longiplexus were lost from the branchial basket during metamorphosis, but most metacercariae of H. coloradensis, H. complexus, and H. parviplexus survived dragonfly metamorphosis. These observations suggest that the observed ecological host specificity of H. longiplexus in semiterrestrial leopard frogs may be due to few metacercariae of H. longiplexus reaching these frogs in a terrestrial environment. Because of the uncertain validity of Haematoloechus varioplexus as a distinct species from its synonym H. parviplexus, their morphological characters were reevaluated. The morphological data on H. varioplexus and H. parviplexus indicate that they differ in their acetabulum length and width, ovary shape, testes length, and egg length and width. Experimental infections of plains leopard frogs, northern leopard frogs, and bullfrogs with worms from bullfrogs indicate that the synonymy of H. parviplexus with H. varioplexus is not warranted, and that these flukes are distinct species, i.e., H. parviplexus in bullfrogs and H. varioplexus in plains leopard frogs and northern leopard frogs.     

Haplorchis infections in intermediate hosts from a clonorchiasis endemic area in Meinung,Taiwan, Republic of China

Snails and freshwater fish were examined from four ponds in the Meinung township in which Clonorchis sinensis was known to be endemic 18 years ago. No metacercariae were found in 478 Tilapia nilotica, whereas of 451 Ctenopharyngodon idellus examined, 16.2%, 3.3% and 0.9% were found to be infected with Haplorchis pumilio, H. taichui and Clonorchis sinensis, respectively. In addition, there were some unidentified metacercariae in 12.0% of Ctenopharyngodon idellus examined. Overall, no positive correlation between infection rates and sizes of infected fish was shown. Six species of snails were collected in this survey and two frequently-occurring snails, Melanoides tuberculata and Thiara granifera were commonly infected with H. pumilio. Reasons for the prevalence of Haplorchis species and the absence of Clonorchis sinensis in fish and snail hosts in a previously reported endemic area for human clonorchiasis are discussed.     

Small frogs get their worms first: the role of nonodonate arthropods in the recruitment of Haematoloechus coloradensis and Haematoloechus complexus in newly metamorphosed northern leopard frogs, Rana pipiens, and woodhouse's toads, Bufo woodhousii

Studies on the life cycles and epizootiology of North American frog lung flukes indicate that most species utilize odonates as second intermediate hosts; adult frogs become infected by ingesting odonate intermediate hosts. Newly metamorphosed frogs are rarely infected with these parasites, predominantly because they are gape-limited predators that cannot feed on large intermediate hosts such as dragonflies. We examined the role of the frog diet and potential intermediate hosts in the recruitment of the frog lung fluke, Haematoloechus coloradensis, to metamorphosed northern leopard frogs (Rana pipiens), Woodhouse's toads (Bufo woodhousii), and bullfrogs (Rana catesbeiana) from western Nebraska. Because of the uncertain validity of H. coloradensis as a distinct species from Haematoloechus complexus, morphological characters of both species were reevaluated and the life cycles of both species were completed in the laboratory. The morphological data on H. coloradensis and H. coimplexus indicate that they differ in their oral sucker to pharynx ratio, uterine loop distribution, and placement of vitelline follicles. However, in terms of their life cycles, both species are quite similar in their use of physid snails as first intermediate hosts, a wide range of nonodonate and odonate arthropods as second intermediate hosts, and leopard frogs and toads as definitive hosts. These results indicate that H. coloradensis and H. complexus are generalists at the second intermediate host level and might be able to infect newly metamorphosed leopard frogs and toads by using small nonodonate arthropods more commonly than other frog lung fluke species. Comparisons of population structure of adult flukes in newly metamorphosed leopard frogs indicate that the generalist nature of H. coloradensis metacercariae enables it to colonize young of the year leopard frogs more commonly than other Haematoloechus spp. that only use odonates as second intermediate hosts. In this respect, the generalist nature of H. coloradensis and H. complexus at the second intermediate host level is an avenue for the colonization of young of year frogs.     

Survey for zoonotic liver and intestinal trematode metacercariae in cultured and wild fish in An Giang Province, Vietnam

Although Vietnam has a high risk of fishborne zoonotic trematode (FZT) infections for humans, little information exists on the epidemiology of these infections in the country's fish. Because of the importance of cultured catfish and snakehead production in An Giang province, a major production area in the Mekong Delta of Vietnam, a survey for FZTs was carried out in randomly selected fish farms between June 2005 and March 2006. For comparison, wild fish from the same area were also surveyed. A total of 852 cultured fish from 4 districts were collected and examined by pepsin digestion to determine their FZT infection status. In Tra catfish, the prevalence of all types of metacercariae was 2.6%, of which the prevalence of Haplorchis pumilio was 0.7%. The overall prevalence of metacercariae in wild fish was 30.6%, of which 10.3% harbored zoonotic species: H. pumilio (2.8%) and Procerovum sp. (5.6%). The prevalence of Opisthorchis metacercariae, which were diagnosed as O. viverrini, was 1.9%. No metacercariae were found in cultured snakehead fish, although wild-caught snakehead fish had a FZT prevalence of 10.3%: 5.1% were O. viverrini 2.6% H. pumilio and 2.6% were Procerovum sp. These are the first reports of H. pumilio, Procerovum sp., and O. viverrini metacercariae in Vietnamese fish. These results indicate that consumption of improperly prepared fish represents a significant risk of acquiring FZTs in this south Vietnam region.     

Zoonotic Trematode Metacercariae in Fish from Phnom Penh and Pursat,Cambodia

A survey was performed to investigate the infection status of freshwater fish with zoonotic trematode metacercariae in Phnom Penh and Pursat Province, Cambodia. All collected fish with ice were transferred to our laboratory and examined using the artificial digestion method. In fish from Phnom Penh, 2 kinds of metacercariae (Opisthorchis viverrini and Haplorchis yokogawai) were detected. O. viverrini metacercariae were positive in 37 (50.0%) of 74 fish in 11 species (average no. metacercariae/fish, 18.6). H. yokogawai metacercariae were detected in 23 (57.5%) of 40 fish in 5 species (average no. metacercariae/fish, 21.0). In fish from Pursat Province, 5 kinds of metacercariae (O. viverrini, H. yokogawai, Haplorchis pumilio, Centrocestus formosanus, and Procerovum sp.) were detected; O. viverrini metacercariae (n=3) in 2 fish species (Henicorhynchus lineatus and Puntioplites falcifer), H. yokogawai metacercariae (n=51) in 1 species (P. falcifer), H. pumilio metacercariae (n=476) in 2 species (H. lineatus and Pristolepis fasciata), C. formosanus metacercariae (n=1) in 1 species (H. lineatus), and Procerovum sp. metacercariae (n=63) in 1 species (Anabas testudineus). From the above results, it has been confirmed that various freshwater fish play the role of a second intermediate host for zoonotic trematodes (O. viverrini, H. yokogawai, H. pumilio, C. formosanus, and Procerovum sp.) in Cambodia.     

Holostephanus metorchis (Digenea: Cyathocotylidae) from chicks experimentally infected with metacercariae from a fish, Pseudorasbora parva, in the Republic of Korea

Holostephanus metorchis (Digenea: Cyathocotylidae) is a parasite of birds, transmitted by freshwater fishes. H. metorchis adults were recovered from chicks experimentally infected with metacercariae collected from freshwater fishes, Pseudorasbora parva. The metacercariae were oval, surrounded with thick fibrous capsules. In adult flukes, the holdfast organ occupied the ventral concavity, and the anterior testis did not reach the level of the ventral sucker. Based on these morphological characteristics, these flukes were identified as H. metorchis.     

Clonorchis sinensis and Opisthorchis viverrini: development of a mitochondrial-based multiplex PCR for their identification and discrimination

We report a single, one-step PCR approach for detection and discrimination of Clonorchis sinensis and Opisthorchis viverrini in different life-stage forms (adults, metacercariae, and eggs) from fish intermediate hosts and from infected patients. Primers designed for species-specific PCR, amplifying portions of the mitochondrial (mt) genome, were also suitable for a multiplex PCR. The latter was a single, one-step reaction under high stringency conditions, using simultaneously 2 pairs of primers (1 pair for C. sinensis--product size 612 bp, and 1 pair for O. viverrini--product size 1357 bp). Assays using serially diluted templates demonstrated that as little as 0.78 ng of genomic DNA of either species could yield amplicons. Genomic DNA extracted from different life-stage forms including adult worms (of both species), eggs (of O. viverrini), eggs possibly of several trematode species (collected from patients infected with C. sinensis in Vietnam) and mixed metacercariae of common trematodes (collected from fishes in the C. sinensis endemic areas), yielded specific bands of the correct size and their identity was confirmed by sequence analysis. The multiplex PCR approach described here proved to be a species-specific, sensitive and fast tool for accurate diagnosis of clonorchiasis and/or opisthorchiasis, permitting the detection of their metacercariae in infected fishes or adult/eggs from patients in endemic areas.     

Multivariate analyses of morphometrical features from Apatemon gracilis (Rudolphi, 1819) Szidat, 1928 and A. annuligerum (v. Nordmann, 1832) (Digenea: Strigeidae) metacercariae

Only two species of the strigeid subgenus Apatemon (Apatemon) sensu Sudarikov (1959), Apatemon gracilis (Rudolhi, 1819) and A. annuligerum (von Nordmann, 1832), are known to infect British fishes. A. annuligerum is poorly characterised, with few life-cycle data available, since only the adult and metacercarial stages have been described. Both of these life-stages demonstrate only minor morphological differences from those of A. gracilis. The multidimensional approach of principal components analysis (PCA) was employed to determine the intra- and inter-specific morphological variation that exists for metacercariae of these species. Analyses revealed that metacercarial morphometrics cannot confidently discriminate between Apatemon gracilis and A. annuligerum specimens. Indeed, levels of intra-specific variation present between A. gracilis metacercariae originating from different piscine hosts were as marked for certain populations as those between the two nominal species. Individual populations of A. gracilis metacercariae differed most in the relative dimensions of their internal organs, whereas variation between the two nominal species was primarily in length-related variables. A. gracilis specimens originating from Welsh and Scottish stoneloach represented two populations with the most comparable total body lengths, yet clustered separately. Conversely, A. gracilis metacercariae infecting rainbow trout and salmon parr differed markedly in size, but demonstrated minimal composite morphological variation. Much, if not all, of the variation described between populations and species of parasites can be attributed to host or to metacercarial location within a particular host. These findings and the lack of strong life-cycle data for A. annuligerum place some doubt on its validity as a species discrete from A. gracilis.     

Prevalence of zoonotic metacercariae in two species of grouper, Epinephelus coioides and Epinephelus bleekeri, and flathead mullet, Mugil cephalus, in Vietnam

Fishborne zoonotic metacercariae have not been reported from brackish water and marine fish from Vietnam waters although these parasites are common in the country's freshwater fish. Both wild-caught and cultured grouper (Epinephelus coioides and Epinephelus bleekeri), and mullet (Mugil cephalus) from brackish and marine waters located in Khanh Hoa province in central coastal Vietnam were examined, and found positive for zoonotic trematode metacercariae. From grouper, Heterophyopsis continua and Procerovum varium were recovered. The prevalence of H. continua ranged from 2.0 to 6.0% and that for P. varium ranged from 11.6 to 15.8%. Mullet were infected with Pygidiopsis summa and H. continua both of these species are new records for Vietnam. The prevalence of P. summa in mullet was generally high, ranging from 17.6 to 75.5%, and was significantly higher than the prevalence of H. continua (2.5 to 32.4%). There were no significant differences in the prevalence of metacercariae between grouper from natural or cultured habitats, indicating that the highest risk of infection occurs in the wild-caught state prior to their placement in culture. Further, there was no difference in metacercarial prevalence between the 2 species of grouper. Infected wild-caught seed were only observed from January to October. Monthly variation in prevalence suggests seasonal variation in mullet infections occurs in this region with the highest transmission taking place from October to December. Basic investigations on the ecology and epidemiology of these intestinal flukes need to be carried out to determine their significance as a public health problem and the aspects of their biology that may be vulnerable to control interventions.     

Infection Status of Zoonotic Trematode Metacercariae in Fishes from Vientiane Municipality and Champasak Province in Lao PDR

The infection status of fishborne zoonotic trematode (FZT) metacercariae was investigated in fishes from 2 localities of Lao PDR. Total 157 freshwater fishes (17 species) were collected in local markets of Vientiane Municipality and Champasak Province in December 2010 and July 2011, and each fish was examined by the artificial digestion method. Total 6 species of FZT metacercariae, i.e., Opisthorchis viverrini, Haplorchis taichui, Haplorchis yokogawai, Haplorchis pumilio, Centrocestus formosanus, and Procerovum varium, were detected in fishes from Vientiane Municipality. The metacercariae of O. viverrini were detected in 50 (49.5%) out of 101 fishes (6 species), and their average number was 154 per fish infected. The remaining 5 species of heterophyid metacercariae were detected in 36.8%, 65.8%, 9.4%, 23.9%, and 5.1% fishes examined, and their average densities were 12, 1,038, 4, 15, and 13 per infected fish, respectively. In fishes from Champasak Province, 3 species of FZT metacercariae, i.e., O. viverrini, H. taichui, and H. yokogawai, were detected. Only 2 O. viverrini metacercariae were found in only 1 Barbonymus schwanefeldi. The metacercariae of H. taichui and H. yokogawai were detected in 60.0% and 50.0% of fishes examined, and their average densities were 47 and 28 per fish infected. By the present study, it has been confirmed that several species of FZT metacercariae are prevalent in fishes from Vientiane Municipality, with P. varium being a new member of FZT in Lao PDR. In comparison, FZT metacercariae are less prevalent in fishes from Champasak Province.     

Skin test for paragonimiasis among schoolchildren and villagers in Namback District, Luangprabang Province, Lao PDR

As a part of a broader effort to determine the status of Paragonimus species infection in Lao PDR, an epidemiological survey was conducted on villagers and schoolchildren in Namback District between 2003 and 2005. Among 308 villagers and 633 primary and secondary schoolchildren, 156 villagers and 92 children evidenced a positive reaction on a Paragonimus skin test. Only 4 schoolchildren out of 128 skin test-positive cases had Paragonimus sp. eggs in their sputum, all of which was collected on 1 day. Several types of crabs, which were identified as the second intermediate host of the Paragonimus species, were collected from markets and streams in a paragonimiasis endemic area for the inspection of metacercariae. Among the examined crabs, only "rock crabs" (Indochinamon ou) harbored Paragonimus sp. metacercariae, and it is speculated that the life cycle of Paragonimus sp. was maintained via rock crabs in Namback District, Lao PDR.     

Fishborne Trematode Metacercariae in Freshwater Fish from Guangxi Zhuang Autonomous Region, China

A survey was performed to investigate the infection status of fishborne trematode (FBT) metacercariae in freshwater fish from Guangxi Zhuang Autonomous Region, China. A total of 307 freshwater fish of 31 species were collected from 5 administrative regions of Guangxi Zhuang Autonomous Region. They were examined by artificial digestion method from July 2003 to August 2004. No metacercariae were detected in fish from Fusui-xian. In fish from Mashan-xian and a market in Nanning, 3 species of metacercariae, Haplorchis taichui, Haplorchis pumilio, and Centrocestus formosanus, were mainly detected. Metacercariae (8 in number) of Clonorchis sinensis were found in 1 Chanodichthys dabryi purchased from a market in Nanning. In fish from Yangshuo, Metagonimus yokogawai metacercariae were detected from all 18 fish species examined. Total 13 C. sinensis metacercariae were found in 3 out of 10 Hemibarbus maculatus from Yangshuo. All 7 Zacco platypus from Yangshuo were infected with 8-112 Echinochasmus perfoliatus metacercariae. In fish from Binyang-xian, H. pumilo metacercariae were mainly detected in all 5 fish species examined, and only 1 metacercaria of C. sinensis was found in a Hemiculter leucisculus. From the above results, it was confirmed that some species of freshwater fish play a role of second intermediate hosts for FBT in Guangxi Zhuang Autonomous Region, China. In particular, 4 species of intestinal flukes, M. yokogawai, H. taichui, H. pumilio, and C. formosanus, were prevalent in fish hosts, whereas C. sinensis metacercariae were detected only in 3 fish species.     

青海高原牛羊双腔吸虫病病原生物学的初步调查

不文报道在青海高原对牛羊双腔吸虫病病原生物学初步调查的结果。从29个流行地区绵羊收集的标本检查结果,发现有四种双腔吸虫,它们是中华双腔吸虫(Dicrocoelium chinensis)、矛形双腔吸虫(D.lanceatum)、客双腔吸虫(D.hospes)和枝双腔吸虫(D.dendriticum)。从三个流行区检查Helicella candacharica及Cathaica przewalskii二种陆地蜗牛,查到四种双腔吸虫成熟尾蚴,它们是中华双腔吸虫、矛形双腔吸虫及二种未定名的双腔吸虫尾蚴,此二种尾蚴是何种双腔吸虫尾蚴,尚待进一步研究。检查流行区中7种蚂蚁是于9月份从乐都的Formica gagates查到双腔吸虫囊蚴,其形态与中华双腔吸虫囊蚴十分相像。     

Molecular link between the metacercariae and adults of four species Haematoloechus (Digenea: Plagiorchioidea), including scanning electron microscopy characterization

Haematoloechus comprises a species-rich genus of trematodes commonly found in the lungs of amphibians across the globe. Of the approximately 70 species reported worldwide, Mexico stands out as a biodiversity hot spot for the genus, with 16 species described. In Los Tuxtlas tropical rainforest, in south-eastern Mexico, three species have been reported, two of them endemic of the frog species occurring in the area. Here, we added the record of a fourth species in the area, and partially elucidated the life cycle of these species using DNA sequence data from the 28S rRNA and the COI genes. Frogs, and a diverse array of species of aquatic insects were sampled in two lakes within the tropical rain forest. Adults and metacercariae of Haematoloechus were obtained. Both developmental stages were linked molecularly contributing to our understanding of their transmission pathways, and probable host-specificity patterns. Each of the four species of Haematoloechus were sampled in a particular species of anisopteran odonate, and therefore were considered as specialists towards their second intermediate host. We validated the presence of H. complexus in the region of Los Tuxtlas because their metacercariae, sampled in the anisopteran dragongly Agriogomphus tumens, matched with sequences of H. complexus from Nebraska and Illinois in USA. We further described the ultrastructure of the tegument of three of the four species of Haematoloechus, sampled from their definitive host (Rana vaillanti, Rana berlandieri), to contribute with characters that can be reliable for species delimitation of the genus.     

A new species of Halocnemum M.Bieb. (Amaranthaceae) from southern Turkey

Halocnemum yurdakulolii Yaprak is described as the second species of the previously monotypic genus Halocnemum. The species is endemic to the Göksu Delta in southern Turkey. The main morphological characteristics that separate H. yurdakulolii from H. strobilaceum (Pall.) M.Bieb. are growth form and spike morphology. Apart from these morphological differences, the species show a clear genetic differentiation. © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society, 2008, 158 , 716–721.     

Ithyoclinostomum yamagutii n. sp. (Digenea: Clinostomidae) in the great blue heron Ardea herodias L. (Aves: Ardeidae) from Mississippi,USA

With only six recognised genera, the family Clinostomidae Lühe, 1901 remains a global research interest of parasitologists and ecologists. Recent efforts have focused on providing molecular data to investigate species diversity, elucidate life-cycles, and make inferences on the group’s evolutionary history. Of the clinostomid genera, the monotypic Ithyoclinostomum Witenberg, 1926 has remained more enigmatic compared to the commonly encountered Clinostomum Leidy, 1856. Recent morphological and molecular evidence from metacercariae suggests a second Ithyoclinostomum species may exist in freshwater cichlids in Central America and Mexico. In a recent survey of great blue herons Ardea herodias L. from commercial catfish production farms in Mississippi, USA, two specimens of an abnormally large (> 20 mm) clinostomid were encountered in the oesophagus of a single bird. These specimens were identified as an Ithyoclinostomum sp. morphologically distinct from the only nominal species Ithyoclinostomum dimorphum (Diesing, 1850). Using morphological and molecular data these adult specimens were confirmed as conspecific with the larval metacercariae previously described from Central America and Mexico and represent the novel species, Ithyoclinostomum yamagutii n. sp.

     

Fishborne Trematode Metacercariae in Luang Prabang,Khammouane, and Saravane Province,Lao PDR

Fishborne trematode (FBT) metacercariae were investigated in fish from 3 Provinces of Lao PDR. Total 242 freshwater fish of 40 species were collected in local markets of Luang Prabang (59 fish of 16 species), Khammouane (81 fish of 19 species), and Saravane (97 fish of 14 species), and each of them was examined by artificial digestion method. Four species of metacercariae (Opisthorchis viverrini, Haplorchis taichui, Haplorchis yokogawai, and Centrocestus formosanus) were detected. O. viverrini was detected in 35 fish (14.5%), and their density was 252 per infected fish (Luang Prabang, 88 metacercariae in 5 fish; Khammouane, 187 in 6 fish; Saravane, 303 in 24 fish). H. taichui was found in 102 fish (42.1%), and their density was 485 per infected fish (Luang Prabang, 260 metacercariae in 38 fish; Khammouane, 1,084 in 23 fish; Saravane, 359 in 41 fish). H. yokogawai was detected in 92 fish (38.0%), and their density was 222 per infected fish (Luang Prabang, 362 metacercariae in 17 fish; Khammouane, 126 in 20 fish; Saravane, 214 in 55 fish). Metacercariae of C. formosanus were found in 8 fish (3.3%), and their density was 3 per infected fish. In the present study, it has been confirmed that FBT metacercariae, in particular, H. taichui, H. yokogawai, and O. viverrini, are highly prevalent in fish from Luang Prabang, Khammouane, and Saravane Province, Lao PDR.     

Brachylaima succini sp. nov. (Trematoda: Brachylaimidae) from Succinea lauta,an amber snail in Hokkaido,Japan

Through a continuous survey of trematodes in land snails of Hokkaido, the northernmost island of Japan, we have discovered four species of the genus Brachylaima (Trematode: Brachylaimidae). Among them, Brachylaima ezohelicis, Brachylaima asakawai, and Brachylaima lignieuhadrae have already been described. Each of the three species is a strict specialist in selecting a particular species of land snail as the first intermediate host. In this report, we propose the fourth species, Brachylaima succini sp. nov., based on ecological, morphological, and phylogenetic considerations. Sporocysts and metacercariae of the new species were found exclusively from Succinea lauta, which is known as an amber snail indigenous to Hokkaido. Phylogenetic trees of nuclear 28S rDNA and mitochondrial cytochrome c oxidase subunit 1 (cox1) demonstrated it to be distinct from the other sympatric species. Although metacercariae of the new species possessed unique morphological characters, adult worms experimentally raised from the metacercariae were similar to those of B. ezohelicis and B. lignieuhadrae. Natural definitive hosts of the new species are unknown, but the existence of common cox1 haplotypes from far-distant localities suggests a possibility that birds are involved as the definitive hosts. Findings of amber snails coinfected with both sporocysts of the new species and Leucochloridium perturbatum also support the involvement of birds.     

Species delimitation methods reveal cryptic diversity in the Hypnea cornuta complex (Cystocloniaceae,Rhodophyta)

The simple and convergent morphologies of many red algae make these species difficult to identify using traditional morphological characters. Many cryptic species have been described in recent years based on molecular datasets, and this has led to the application of an integrative taxonomy approach in species delimitation. Here, we performed several species delimitation methods (mBGD, ABGD, SPN, PTP, GMYCs and GMYCm) based on two different loci (COI-5P and rbcL) in species of the Hypnea cornuta complex. These methods were combined with morphological and phylogenetic data, extensive sampling, analysis of topotype material, and historically relevant herbarium samples. Our findings demonstrate that the groups morphologically assigned to H. cornuta and H. stellulifera consist of five different cryptic species. H. cornuta is a polyphyletic taxon composed of three well-separated lineages, thus requiring sequencing of type or topotype specimens to determine which one is Hypnea cornuta sensu stricto. We have revealed that the distribution of H. stellulifera is limited to Asia, while the Brazilian specimens initially assigned to this species were clarified as a new endemic species: Hypnea cryptica sp. nov. Our results indicated that only an integrative approach combining several lines of evidence, including morphology, nomenclature history, molecular data, biogeography and ecology can correctly solve the taxonomic status of widely distributed cryptic species.     

Foodborne Trematode Metacercariae in Fish from Northern Vietnam and Their Adults Recovered from Experimental Hamsters

The prevalence of foodborne trematode (FBT) metacercariae was investigated in fish from 2 localities of northern Vietnam in 2004-2005. Freshwater fish (9 species) were collected from local markets in Hanoi City (n=76) and Nam Dinh Province (n=79), and were examined for FBT metacercariae using the artificial digestion technique. Adult flukes were obtained from hamsters experimentally infected with the metacercariae at day 8 post-infection. Three (Haplorchis pumilio, Centrocestus formosanus, and Procerovum varium) and 6 (Haplorchis taichui, H. pumilio, C. formosanus, P. varium, Stellantchasmus falcatus, and Heterophyopsis continua) species of FBT metacercariae were detected in the 2 regions, respectively. Overall, among the positive fish species, H. pumilio metacercariae were detected in 104 (80.0%) of 130 fish examined (metacercarial density per infected fish; 64.2). C. formosanus metacercariae were found in 37 (40.2%) of 92 fish (metacercarial density; 14.7). P. varium metacercariae were detected in 19 (63.3%) of 30 fish (Anabas testudineus and Mugil cephalus) (metacercarial density; 247.7). S. falcatus metacercariae were found in all 10 M. cephalus examined (metacercarial density; 84.4). H. continua metacercariae (2 in number) were detected in 1 fish of Coilia lindmani. Morphologic characteristics of the FBT metacercariae and their experimentally obtained adults were described. The results have demonstrated that various FBT species are prevalent in northen parts of Vietnam.