Coexistence and genetic variability of Contracaecum rudolphii A and Contracaecum rudolphii B (Nematoda: Anisakidae) in cormorants, Phalacrocorax carbo sinensis, in the Baltic region

Although numerous genetically isolated entities within Contracaecum rudolphii sensu lato are presently defined, information on the distribution and ecology of these groups is nonetheless in demand. In the present study, information based upon DNA sequence data (restriction analysis and sequencing of rDNA) on the distribution of the species C. rudolphii A and C. rudolphii B in the cormorant Phalacrocorax carbo sinensis from the Baltic region (Poland and Finland) is provided. These data corroborate previous work that identified C. rudolphii A in brackish water regions and C. rudolphii B in fresh water sites, although mixed infections also occur. The 2 species may inhabit the same host specimen. One rare heterozygote of the species was recorded.     

Population biology of Contracaecum rudolphii sensu lato (Nematoda) in the great cormorant (Phalacrocorax carbo) from northeastern Poland

Parameters related to the occurrence, aggregation, and population structure of the nematode Contracaecum rudolphii Hartwich, 1964 , in the great cormorant ( Phalacrocorax carbo ) from northeastern Poland were analyzed. A total of 491 birds of different ages (adults, immatures, and nestlings) was examined; the cormorants studied were taken from both fresh- and brackish water habitats. Contracaecum rudolphii were found in stomachs of 454 birds (92.5%); the 46,244 nematodes included third- (L3) and fourth-stage larvae (L4), and sub-adult and adult females and males. The distribution of nematodes in the host population were highly aggregated. The occurrence of C. rudolphii was significantly dependent on the host's age and habitat, as well as on season; the proportion of development stages differed significantly depending on birds' age and season (the latter only in adult birds). The infrapopulations of C. rudolphii in the adult cormorants showed distinct density-dependent correlations: that is, as the infrapopulation size increased, the number of adult females C. rudolphii decreased, and the proportion of larvae and sub-adult females increased. A higher proportion of larvae and sub-adult females characterized the component population structure of the nematodes in the cormorant nestlings, compared with adult birds, probably because of immune system deficiency in the immature birds, coupled with the development of the nematode population. Seasonal changes in the C. rudolphii population, observed in the adult cormorants, were not related to seasonality of the L3 occurrence in food; instead, the changes are believed to have resulted from independent processes of elimination of the oldest nematodes and their replacement by larval stages that subsequently matured.     

Rainbow trout (Salmo gairdneri) stocking and Contracaecum spp

A stocking program with rainbow trout (Salmo gairdneri) at High Rock Lake, Manitoba failed due to infections with large numbers of Contracaecum spp. larvae. Nematode larvae in the intestinal tract, body cavity and musculature made the fish unmarketable. A combination of experimental infections of rainbow trout and pelicans (Pelecanus erythrorhynchos), observations on the behavior of fish-eating birds, and numbers of larval Contracaecum spp. in minnow species led to the following conclusions. The introduction of rainbow trout attracted large numbers of fish-eating birds, particularly pelicans. Concurrent predation by rainbow trout on fathead minnows (Pimephales promelas), five-spined sticklebacks (Culaea inconstans), and nine-spined sticklebacks (Pungitius pungitius), concentrated the parasites. The combined increase in densities of the introduced fish host and fish-eating birds, and the short life cycle of the parasite, increased the numbers of parasites in rainbow trout over a season and in the indigenous minnow species between years. Numbers of larvae in the indigenous minnow species declined when stocking of rainbow trout was stopped and use of the lake by fish-eating birds, particularly pelicans, returned to normal levels.     

Gastrointestinal helminths of fish-eating birds from Chiloe Island, Chile

Of four species of fish-eating birds from Chiloé Island (Chile), three were infected with eight species of helminths. Tetrabothrius sp. was found in Larus scoresbii. Tetrabothrius cylindraceus, Profilicollis antarcticus, Anomotaenia dominicanus, Stephanoprora denticulata, Capillaria sp. and P. antarcticus were found in Larus dominicanus. Contracaecum rudolphii and Corynosoma sp. infected Phalacrocorax olivaceus. With the exception of S. denticulata, C. rudolphii and Capillaria sp., the above helminths are reported for the first time from Chile.     

Ascaridoidea: a simple DNA assay for identification of 11 species infecting marine and freshwater fish,mammals, and fish-eating birds

Eleven species belonging to superfamily Ascaridoidea, which infect marine and freshwater fish, mammals, and fish-eating birds, were analyzed using a PCR-RFLP method. The following species were investigated: Anisakis pegreffi, A. physeteris, and A. simplex (parasites of fish and mammals), Contracaecum osculatum, C. radiatum, and C. rudolphi (parasites of mammals and fish-eating birds), Hysterothylacium aduncum (a parasite of fish), Porrocaecum angusticolle, P. crassum, P. depressum, and P. ensicaudatum (parasites of fish-eating birds). PCR-amplified rDNA regions encompassing ITS1, 5.8S rDNA, and ITS2 produced on templates of genomic DNA isolated from all investigated species were digested with TaqI, AluI, BsuRI, and RsaI endonucleases. Restriction patterns showed that endonuclease TaqI is the most useful enzyme for identification of all investigated species. No variations in restriction patterns within each species were detected. Therefore, we propose that the PCR-RFLP assay described in this report may be used for identification of marine and freshwater parasites from superfamily Ascaridoidea.     

青海湖裸鲤体腔寄生蠕虫群落研究

青海湖裸鲤体腔为三种寄生蠕虫寄生,即裂头绦虫裂头蚴,舌状绦虫裂头蚴和对盲囊缄虫的三期幼虫,这几种蠕虫不同大小的青海湖裸鲤中数量差异很大,将其看作一个组分群落,并根据其发生数量进行模糊聚类和极点排序,结果将其分为二个亚组分群落,即体长小于１５０ｍｍ的青海湖裸鲤体腔蠕虫亚组分群落和体长大于１５０ｍｍ的个体的体腔蠕虫亚组分群落。这与宿主的行为和食性改变有密切关系,舌状绦虫与另两种蠕虫间有显著负协调关系。     

Infection by Contracaecum rudolphii (Nematoda: Anisakidae) in the Neotropic cormorant Phalacrocorax brasilianus,and fishes from the estuary of the Valdivia river,Chile

Seasonal parameters of infection by the nematode Contracaecum rudolphii in the Neotropic cormorant Phalacrocorax brasilianus (Phalacrocoracidae) and their relation with feeding and infection in fishes from River Valdivia, Chile, were determined. The prevalences of infection in birds were similar during seasons, whereas mean intensity and percentages of adult gravid females were higher in spring and summer, respectively. For fishes no seasonal differences were found in infection. Cormorant diet varied seasonally in relation to fish prey consumed and this can be related to seasonal differences in infection parameters of birds. The high infection in birds should be considered as a potential risk for salmoniculture.     

Seasonal occurrence of helminths in the anadromous fish Coilia nasus (Engraulidae): parasite indicators of fish migratory movements

To understand the seasonal migration of the anadromous Coilia nasus , we attempted to identify the parasites infecting C. nasus and determine their seasonal occurrence. From June 2007 to July 2008, a survey of 775 C. nasus individuals from the estuary of the Yangtze River and the coast of the East China Sea revealed more than 7,300 parasites associated with the gills and alimentary tracts of C. nasus . The following 6 helminth taxa were identified, i.e., the monogeneans Heteromazocraes lingmueni and Helciferus tenuis, the digenean Elytrophallus coiliae, the acanthocephalan Acanthosentis cheni , and larvae of the nematodes Anisakis simplex and Contracaecum sp., all of which are marine or brackish-water parasites. The absence of freshwater helminths suggested that the parasites acquired in freshwater may have been accidentally, and easily, lost by the time the fish had reached the estuary and coast. Contrary to seasonal occurrence of the parasites' life cycles, the lowest mean abundance and prevalence of H. lingmueni and A. cheni occurred in August, which suggested the immigration of C. nasus from freshwater to the Yangtze estuary, with lower parasite burdens. The highest mean abundance and prevalence of the nematodes A. simplex and Contracaecum sp. in May and June, and the lowest in August, indicated the arrival of the fish from the coast and the Yangtze River, to the estuary, respectively. These findings suggested that a majority of the fish prepared for spawning migration in the estuary in spring and early summer and returned to the estuary after spawning in the lower and middle reaches of the Yangtze River in late summer.     

Levels of infection of gastric nematodes in a flock of great cormorants (Phalacrocorax carbo) from Lake Biwa, Japan

A high prevalence (86.7%) of various species of nematodes was observed in the stomach of great cormorants living in Lake Biwa, Japan. There were varying numbers of adults belonging to two common genera, Eustrongylides Jagerskiold 1909 (Nematoda: Dioctophymatidae) and Contracaecum Railliet & Henry 1912 (Nematoda: Anisakidae). The first included common adenophorean nematodes comprising a single species, Eustrongylides tubifex and the second comprised ascaroid nematodes that contained four named species: Contracaecum rudolphii Hartwich, 1964, Contracaecum microcephalum Yamaguti, 1961, Contracaecum multipapillatum Drasche, 1882 and Contracaecum chubutensis Garbin, 2008. After the prevalence and intensity of the infection had been noted, both types of nematodes were frequently observed to penetrate the mucosa and intrude into the wall of the glandular stomach, where they caused gross haemorrhage and ulceration. The Eustrongylides sp. was predominantly found in a nodular lesion of the proventricular wall, while Contracaecum spp. were observed either free in the lumen of the proventriculus or, on occasion, deeply penetrating its wall. Of the Contracaecum spp., C. rudolphii was the most prevalent. Grossly, large numbers of nematodes were present in infected stomachs (for C. rudolphii intensity was 1-34 and 3-57 nematodes in male birds and 1-21 and 1-32 in females; for C. microcephalum 1-2 and 1 in male birds and 1-2 in females; for C. multipapillatum 2 in male cormorants and no infection in females; for C. chubutensis 1-2 and 1 in male birds and 1-5 and 1 in females and for E. tubifex 1-5 nematodes in male birds and 2-8 in females). Ulcerative inflammation and hyperaemia were the most common pathological presentations, especially in areas that had been invaded by parasites. Microscopically, varying degrees of granulomatous inflammatory reactions were seen, in addition to degenerated nematodes which appeared to have deeply penetrated mucosal surfaces and were surrounded by fibrous connective tissues.     

Molecular phylogenetics and diagnosis of Anisakis, Pseudoterranova, and Contracaecum from northern Pacific marine mammals

Individual specimens of Anisakis, Pseudoterranova, and Contracaecum collected from marine mammals inhabiting northern Pacific waters were used for comparative diagnostic and molecular phylogenetic analyses. Forty-eight new sequences were obtained for this study of 14 Anisakis taxa, 8 Pseudoterranova taxa, 4 Contracaecum taxa, and 4 outgroup species. Partial 28S (LSU) and complete internal transcribed spacer (ITS-1, 5.8S, ITS-2) ribosomal DNA was amplified by the polymerase chain reaction and sequenced. Sequences of ITS indicated that Pseudoterranova specimens from Zalophus californianus (California sea lion), Mirounga angustirostris (northern elephant seal), Phoca vitulina (harbor seal), Enhydra lutris (sea otter), and Eumetopias jubatus (Steller's sea lion) exactly matched P. decipiens s. str., extending the host and geographic range of this species. Anisakis from northern Pacific marine mammals were most closely related to members of the A. simplex species complex. Comparison of Anisakis ITS sequences diagnosed the presence of A. simplex C in 2 M. angustirostris hosts, which is a new host record. Anisakis specimens from Phocoena phocoena (harbor porpoise), Lissodelphis borealis (Pacific rightwhale porpoise), and E. jubatus included 3 ITS sequences that did not match any known species. Contracaecum adults obtained from Z. californianus were most closely related to C. ogmorhini s.l. and C. rudolphii, but ITS sequences of these Contracaecum specimens did not match C. ogmorhini s. str. or C. margolisi. These novel Anisakis and Contracaecum ITS sequences may represent previously uncharacterized species. Phylogenetic analysis of LSU sequences revealed strong support for the monophyly of Anisakinae, Contracaecum plus Phocascaris, Pseudoterranova, and Anisakis. Phylogenetic trees inferred from ITS sequences yielded robustly supported relationships for Pseudoterranova and Anisakis species that are primarily consistent with previously published phenograms based on multilocus electrophoretic data.     

THE FOOD AND FEEDING HABITS OF GOLDENEYE BUCEPHALA CLANGULA IN GREAT BRITAIN

The food and feeding habits of Goldeneye Bucephala clangula are described, based on the analyses of 51 stomach contents and a brief review of the literature. The type of food taken depends on the type of habitat being used, the availability and size of the food items and on the methods of feeding—though animal material largely predominates.
In estuarine and coastal areas, small crustaceans and in particular Carcinus maenas figured highly in the diet, with some molluscs and small fish. In two brackish-water feeding birds, seeds formed the bulk of the food, some caddis-fly larvae also being taken. In freshwater feeding birds insects predominated, particularly Trichoptera larvae, chironomid larvae and the adults of Corixa spp. Some crustaceans ( Asellus, Gammarus ) and small molluscs were taken and occasionally small fish. A small amount of plant material was taken in the form of seeds, mainly Potamogeton spp.
The relation between Goldeneye and fishing interests is briefly discussed and it is concluded that fish normally form a small part of the diet and it is unlikely that any serious depredation of fish stocks is common.     

Contracaecum rudolphii B: gene content, arrangement and composition of its complete mitochondrial genome compared with Anisakis simplex s.l

In the present study, we sequenced the complete mt genome (14,022 bp) of parasitic nematode Contracaecum rudolphii B and its structure and organization compared with Anisakis simplex s.l. The mt genome of C. rudolphii B is slightly longer than that of A. simplex s.l. (13,916 bp). C. rudolphii B mt genome is circular, and consists of 36 genes, including 12 genes for proteins, 2 genes for rRNA and 22 genes for tRNA. This genome contains a high A+T (70.5%) content. The mt gene order for C. rudolphii B is the same as those for A. simplex s.l., but it is distinctly different from other nematodes compared. The start codons inferred in the mt genome of C. rudolphii B are TTG and ATT. Six protein-coding genes use TAA as a stop codon whereas five genes use T and one genes use TAG as a termination codon. This pattern of codon usage reflects the strong bias for A and T in the mt genome of C. rudolphii B. Phylogenetic analyses using concatenated amino acid sequences of the 12 protein-coding genes, with three different computational algorithms (Bayes, ML and MP), all revealed distinct groups with high statistical support, indicating that C. rudolphii B and A. simplex s.l. is distinct but closely related species. These data provide additional novel mtDNA markers for studying the molecular epidemiology and population genetics of the C. rudolphii B, and should have implications for the molecular diagnosis, prevention and control of anisakidosis in humans and animals.     

Description and genetic characterisation of Hysterothylacium (Nematoda: Raphidascarididae) larvae parasitic in Australian marine fishes

Nematodes belonging to the genus Hysterothylacium (family Raphidascarididae) infect various species of marine fish in both the larval and adult stages. Humans can be accidentally infected upon eating infected seafood. In spite of their importance, relatively little is known of their occurrence and systematics in Australia. An examination of various species of marine teleosts in Australian waters revealed a high prevalence of Hysterothylacium larval types. In the present study, seven previously undescribed Hysterothylacium larval morphotypes (V to VII and IX to XII) were discovered. In total we found 10 different morphotypes and we genetically characterised nine morphotypes identified. A morphological dichotomous identification key has been established to differentiate these morphotypes. Since some larvae of Hysterothylacium from marine fishes cannot be differentiated morphologically from other nematode larvae, such as Paraheterotyphlum, Heterotyphlum, Iheringascaris and Lapetascaris, the first and second internal transcribed spacers (ITS-1 and ITS-2) of nuclear ribosomal DNA (rDNA) of these larvae were characterised to confirm their taxonomic status. This genetic characterisation implied that some distinct morphotypes belong to different developmental stages of the same species. In addition, it revealed that some morphotypes can comprise distinct genotypes. No match was found between ITS-1 and ITS-2 sequences obtained from larvae in the present study and those from adults available in the GenBank, highlighting the lack of knowledge on occurrence of adult nematodes infecting Australian fish.     

Diversity among Field Populations of Bradyrhizobium japonicum in Poland

Genetic structure in field populations of Bradyrhizobium japonicum isolated in Poland was determined by using several complementary techniques. Of the 10 field sites examined, only 4 contained populations of indigenous B. japonicum strains. The Polish bradyrhizobia were divided into at least two major groups on the basis of protein profiles on polyacrylamide gels, serological reaction with polyclonal antisera, repetitive extragenic palindromic PCR fingerprints of genomic DNA, and Southern hybridization analyses with nif and nod gene probes. Serological analyses indicated that 87.5% of the Polish B. japonicum isolates tested were in serogroups 123 and 129, while seven (12.5%) of the isolates tested belonged to their own unique serogroup. These seven strains also could be grouped together on the basis of repetitive extragenic palindromic PCR fingerprints, protein profiles, and Southern hybridization analyses. Cluster analyses indicated that the seven serologically undefined isolates were genetically dissimilar from the majority of the Polish B. japonicum strains. Moreover, immuno-cross-adsorption studies indicated that although the Polish B. japonicum strains reacted with polyclonal antisera prepared against strain USDA123, the majority failed to react with serogroup 123- and 129-specific antisera, suggesting that Polish bradyrhizobia comprise a unique group of root nodule bacteria which have only a few antigens in common with strains USDA123 and USDA129. Nodulation studies indicated that members of the serologically distinct group were very competitive for nodulation of Glycine max cv. Nawiko. None of the Polish serogroup 123 or 129 isolates were restricted for nodulation by USDA123- and USDA129-restricting soybean plant introduction genotypes. Taken together, our results indicate that while genetically diverse B. japonicum strains were isolated from some Polish soils, the majority of field sites contained no soybean-nodulating bacteria. In addition, despite the lack of long-term soybean production in Poland, field populations of unique B. japonicum strains are present in some Polish soils and these strains are very competitive for nodulation of currently used Polish soybean varieties.     

<Emphasis Type="Italic">Contracaecum bioccai</Emphasis> n. sp. from the brown pelican <Emphasis Type="Italic">Pelecanus occidentalis</Emphasis> (L.) in Colombia (Nematoda: Anisakidae): morphology,molecular evidence and its genetic relationship with congeners from fish-eating birds

Contracaecum bioccai n. sp. is described from the brown pelican Pelecanus occidentalis (L.) in northern Colombia (Totumo Marsh) based on 20 enzyme loci studied using multilocus allozyme electrophoresis. Moreover, genetic relationships between the new taxon and related congeners are presented based on allozyme data-sets and sequence analyses (519 bp) of the mtDNA-cox2 gene. Fixed allele differences were found at some of the allozyme loci analysed in comparison with other Contracaecum spp. from pelicans and cormorants [i.e. the sibling species of the C. rudolphii Hartwich, 1964 complex, C. septentrionale Kreis, 1955, C. micropapillatum (Stossich, 1890), C. microcephalum (Rudolphi, 1809) and C. pelagicum Johnston & Mawson, 1942]. The genetic distance, at the allozyme level, between C. bioccai n. sp. and its congeners ranged from D ( Nei ) = 0.80 versus C. septentrionale to D ( Nei ) = 1.40 versus C. micropapillatum. The genetic distance at the mtDNA cox-2 level ranged, on average, from K-2P = 0.12 versus the C. rudolphii species complex to K-2P = 0.15 versus C. micropapillatum. An overall concordant tree topology, obtained from UPGMA and NJ tree analyses inferred from allozyme data, as well as from MP, UPGMA and NJ inferred from mtDNA-cox2 sequence analysis, showed C. bioccai n. sp. as a separated lineage to the other Contracaecum spp. A concordant result was also obtained by PCA analysis based on both the allozyme and mtDNA cox-2 data-sets. All of the tree topologies, derived from the phylogenetic analysis inferred from both allozymes and mtDNA data-sets, were in substantial agreement and depicted C. bioccai as closely related to the sibling species of the C. rudolphii complex (C. rudolphii A and C. rudolphii B) and C. septentrionale. Morphological analysis and a differential diagnosis based on male specimens of C. bioccai, which had been genetically characterised by both allozyme markers and mtDNA sequences analysis with respect to morphologically related congeners, enabled the detection of differences in a numbers of characters, including spicule length, the morphology of the distal end of the spicule and the distribution patterns of the distal caudal papillae.     

A new anisakid species parasitizing the imperial cormorant Phalacrocorax atriceps from the North Patagonian Coast, Argentina

The anisakid species Contracaecum chubutensis n. sp. was found parasitizing the imperial cormorant Phalacrocorax atriceps at 2 different localities, Bahía Bustamante (45 degrees 11'S, 66 degrees 30'W) and Puerto Madryn (42 degrees 47'S, 65 degrees 02'W) on the Patagonian coast. Morphometrical analysis and further studies of adult specimens of C. chubutensis n. sp. and fourth-stage larvae of Contracaecum sp. were done using light and scanning electron microscopy. The possession and disposition of 7 pairs of tail papillae differentiate this new species from C. osculatum, C. radiatum, C. plagiaticium, and C. mirounga. Double-sized spicules separate it from C. variegatum and C. pelagicum. Moreover, the paracloacal papillae disposition and tail shape differentiate this new species from C. travassosi. The presence of bifurcated interlabia separate it from C. ogmorhini and C. margolisi. Three well-marked lip notches, a sharp tail shape, and the of the transversal paracloacal papillae disposition distinguishes C. chubutensis n. sp. from C. magnipapillatum and C. septentrionale. The species here described can be differentiated from C. caballeroi by its bifurcated interlabia, longer spicules, and for possessing more precloacal papillae pairs. It can be separated from C. rudolphii by a thinner interlabia at the base, deeper lip notches, a sharper tail, the disposition of transversal paracloacal papillae, and greater distance between both papillae of the first subventral distal papillae pair. In addition, fourth-stage larvae recovered from the same host and localities were identified and described as Contracaecum sp. Prevalence of adult parasites was 66.7 %, mean intensity was 4.0, and mean abundance was 7.1.     

Population genetics of the hazel hen <Emphasis Type="Italic">Bonasa bonasia</Emphasis> in Poland assessed with non-invasive samples

Despite a severe decrease in the number of hazel hens during the 20^th century, nowadays this grouse species is rather common in the forests of Northeastern and Southern Poland. We used mitochondrial control region and microsatellite markers to examine the genetic variability of Polish populations of hazel hens. We used non-invasively collected faeces to estimate genetic variability within populations, genetic differentiation among populations as well as genetic differentiation between two regions inhabited by two different subspecies of hazel hens. Our results confirm the usefulness of DNA from faeces to obtain reliable information on the population genetics of hazel hens. We found a rather high level of genetic variability in the Polish population. Genetic variability was higher in birds from continuous forests in the South of the country than in birds from fragmented habitats in the Northeast. Genetic differentiation was higher among subpopulations from Northeastern Poland. Additionally, both classes of molecular markers suggested the presence of two distinct genetic groups of birds, corresponding to previously described subspecies. We conclude that the genetic variability of the Polish hazel hen population has been influenced by habitat fragmentation and the history of the population during its post-glacial colonization of Poland from different glacial refugia.     

Infection status of the sea eel (Astroconger myriaster) purchased from the Noryangjin fish market with anisakid larvae]

Although the sea eel (Astroconger myriaster) is suspected as one of the most important fish host for human anisakiasis in Korea, no report has been made on the infection status of the sea eel with anisakid larvae. In the present study, 26 sea eels (Astroconger myriaster) were purchased from the Noryangjin fish market in Seoul, and anisakid larvae were collected from their viscera, muscle, head and skin. The collected larvae were classified by their morphological types. A total of 1,351 anisakid larvae were collected from 15 of 26 fish examined. Among them, 1,269 were recovered from the viscera, 66 from the muscle, and 16 from the head and skin. Morphologically, most of the anisakids were classified into 6 known larval types, Anisakis type I (564 larvae) of Berland (1961), Contracaecum type A (409) and type D (5) of Koyama et al. (1969), Contracaecum type C' (83) and type D' (117) of Chai et al. (1986), and Contracaecum type V (1) of Yamaguti (1935). Remaining 172 specimens were new in the available literature, hence, designated as Contracaecum type A' (new type). The present results revealed that the sea eels caught in the Korean waters are heavily infected with anisakid larvae, not only in their viscera but also in the muscle, and Anisakis type I was the most common among the 7 larval types.