Expression of Hairpin-Enriched Mitochondrial DNA in Two Hairworm Species (Nematomorpha).

Nematomorpha (hairworms) is a phylum of parasitic ecdysozoans, best known for infecting arthropods and guiding their hosts toward water, where the parasite can complete its life cycle. Over 350 species of nematomorphs have been described, yet molecular data for the group remain scarce. The few available mitochondrial genomes of nematomorphs are enriched with long inverted repeats, which are embedded in the coding sequences of their genes-a remarkably unusual feature exclusive to this phylum. Here, we obtain and annotate the repeats in the mitochondrial genome of another nematomorph species-Parachordodes pustulosus. Using genomic and transcriptomic libraries, we investigate the impact of inverted repeats on the read coverage of the mitochondrial genome. Pronounced drops in the read coverage coincide with regions containing long inverted repeats, denoting the 'blind spots' of short-fragment sequencing libraries. Phylogenetic inference with the novel data reveals multiple disagreements between the traditional system of Nematomorpha and molecular data, rendering several genera paraphyletic, including Parachordodes.

Helminth parasites of the lesser great cormorant Phalacrocorax carbo sinensis from two nesting regions in the Czech Republic.

Parasitological examinations of 102 specimens of the lesser great cormorant Phalacrocorax carbo sinensis (Blumenbach) from two nesting regions in the Czech Republic (South Bohemia and South Moravia) were carried out at the Institute of Parasitology, Czech Academy of Sciences (previously the Czechoslovak Academy of Sciences) in the years 1987-1992. In them, a total of 19 species of helminth parasites was found, including Trematoda (11 species), Cestoda (2), Nematoda (4) and Acanthocephala (2), which can be divided into three main groups regarding their host specificity: parasites specific for cormorants (Phalacrocorax spp.) (37%), those parasitic mainly in cormorants (16%) and non-specific parasites (47%). Of the 19 species recorded, 100% were found in South Moravia, but only 47% of these 19 species in South Bohemia. The higher number of helminth species in cormorants from South Moravia and a higher proportion of non-specific species may be associated with the presence of the large Nové Mlýny water reservoir, in addition to better ecological and environmental conditions in this warmer region. Scanning electron microscopical examination of three common nematode species parasitising cormorants, Contracaecum rudolphii Hartwich, 1964, Desmidocercella incognita Solonitsin, 1932 and Syncuaria squamata (von Linstow, 1883), revealed some taxonomically important, previously unreported morphological features, such as the cephalic structures, numbers and distribution of male caudal papillae or the shapes of spicules.

[Cuticle ultrastructure of the fresh-water horsehair worm Gordionus alpestris (Villot, 1885) (Nematomorpha)].

The cuticular structure of the horsehair worm Gordionus alpestris (Villot, 1885) was studied under scanning and transmission electron microscopes. Adult worms were collected in the Syuk River near Nickel' Village in the Republic of Adygea (Russia) in June 2013. In the sampling area, the G. alpestris juveniles parasitize diplopods Pachyiulus krivolutskyi Golovatch, 1977. Similarities with other Nematomorpha species with the known cuticle ultrastucture are discussed.

Comparative descriptions of non-adult stages of four genera of Gordiids (Phylum: Nematomorpha).

Freshwater hairworms infect terrestrial arthropods as larvae but are free-living in aquatic habitats as adults. Estimates suggest that only 18% of hairworm species have been described globally and biodiversity studies on this group have been hindered by unreliable ways of collecting adult free living worms over large geographical areas. However, recent work indicates that non-adult cyst stages of hairworms may be the most commonly encountered stages of gordiids in the environment, and can be used for discovering the hidden diversity of this group. Unfortunately, little information is available on the morphological characteristics of non-adult stages of hairworms. To address this problem, we describe and compare morphological characteristics of non-adult stages for nine species of African and North American gordiids from four genera (Chordodes, Gordius, Paragordius, and Neochordodes). Observations were made on the oviposition behavior of adult worms and morphological characteristics were recorded for egg strings, larvae and cysts using light and differential interference contrast microscopy and/or scanning electron microscopy. Our study indicates that three distinct types of oviposition behaviors and three distinct morphological types of egg string, larva, and cysts were present among the four genera of gordiids. Although species identification based on cyst characteristics was not always possible among different species of gordiids, cyst morphology was conserved among some genera and all clades of gordiids. More importantly, our work indicates that gordiid larval morphology can be used for predicting cyst morphology among other gordiid genera. The capability to identify and predict gordiid genera and/or clades based on cyst morphology will be useful for culturing gordiids in the laboratory from field collected cysts and these new techniques will undoubtedly allow others to discover new species of gordiids from around the world.

Endohelminth parasites of the blacktail comber Serranus atricauda (Pisces: Serranidae), from Madeira Archipelago (Atlantic Ocean).

Four different endohelminth parasite taxa were found in the viscera of the blacktail comber Serranus atricauda Günther, 1874 caught in the Madeira Archipelago. Nematodes were the dominant group, represented by 2 different taxa, Hysterothylacium spp. Ward & Magath, 1917 and Procamallanus (Spirocamallanus) halitrophus Fusco & Overstreet, 1978 comb. n. Plerocerci of the trypanorhynch Pseudogrillotia epinepheli (synonym: Grillotia epinepheli) Scholz, Garippa & Scala, 1993, and cystacanths of the acanthocephalan Bolbosoma vasculosum Rudolphi, 1819 were found in the visceral cavity. New host records for P. (S.) halitrophus and P. epinepheli and the extension of the geographic distribution of these 2 parasite species provide evidence of parasite transference between the Madeira Archipelago, the Mediterranean and the Gulf of Mexico. The paucity of the parasite fauna of blacktail comber reflect a combination of fish host selective feeding on particular dietary items and its territorial behaviour.

Influence of site, season, silvering stage, and length on the parasites of the European eel Anguilla anguilla in two Mediterranean coastal lagoons of the island of Corsica, France using indicator species method.

The parasites of 425 European eels, Anguilla anguilla, were studied between 2009 and 2012 in two Mediterranean coastal lagoons of the island of Corsica, France. An indicator value (IndVal) method was used for analysis, which combines measures of fidelity and specificity. Because of its resilience to detect changes in abundance, IndVal is an effective ecological bioindicator. The IndVal method demonstrated that site, season, silvering stage, and length could influence the occurrence of parasite species in European eel. A randomization test identified ten parasite species as having a significant indicator value for site (lagoons differed principally in salinity: oligohaline to polyhaline for the Biguglia lagoon and polyhaline to euhaline for the Urbino lagoon; the digeneans Bucephalus anguillae and Lecithochirium musculus, the cestodes Bothriocephalus claviceps, Proteocephalus macrocephalus, and larvae of Myzophyllobothrium sp., the nematodes Anguillicoloides crassus, and encysted larvae of Contracaecum sp., the acanthocephalan Acanthocephaloides incrassatus, the monogenean Pseudodactyogyrus anguillae, and the copepod Ergasilus gibbus); one parasite species for the spring season (the acanthocephalan A. incrassatus); six parasite species for silvering stage (yellow, pre-silver, silver; the trematodes B. anguillae and Deropristis inflata, encysted larvae of the nematode Contracaecum sp., the acanthocephalan A. incrassatus, the monogenean P. anguillae, and the copepod E. gibbus); and three parasite species for some of the five length classes (the cestode P. macrocephalus, encysted larvae of the nematode Contracaecum sp., and the monogenean P. anguillae). Data for species composition and infection levels should help to improve the management of parasitism in the populations of European eels.

Gastrointestinal helminths of Gentoo penguins (Pygoscelis papua) from Stranger Point, 25 de Mayo/King George Island, Antarctica.

The aim of this work is to contribute to the knowledge of gastrointestinal parasites of the Gentoo penguin (Pygoscelis papua) from 25 de Mayo/King George Island (South Shetlands, Antarctica). Gastrointestinal tracts of 37 fresh dead individuals (21 chicks, 10 juveniles, and 6 adults) were collected from December 2006 to February 2012 and examined for macroparasites. Four adult parasite species were found: one Cestoda species (Parorchites zederi), two Nematoda species (Stegophorus macronectes and Tetrameres wetzeli), and one Acanthocephalan (Corynosoma shackletoni). Two species of immature acanthocephalans, Corynosoma hamanni and Corynosoma bullosum, were found in a single host. This is the first record of Tetrameres wetzeli in Gentoo penguins. The low parasite richness observed could be related to the stenophagic and pelagic diet of this host species which feeds almost exclusively on krill.

Novel techniques for biodiversity studies of gordiids and description of a new species of Chordodes (Gordiida, Nematomorpha) from Kenya, Africa.

We review recent advances in the use of non-adult gordiid cyst stages to locate gordiids over large geographical regions and new culturing techniques which can help overcome current difficulties in nematomorph biodiversity studies. Using these techniques, we collected a new species of gordiid as cysts in aquatic snails (Biomphalaria pfeifferi) from the Lake Victoria Basin, western Kenya, Africa and cultured them in the laboratory. We describe the adult free-living male and female worms using morphological (light and scanning electron microscopy) and molecular data as well as the life cycle, mating and oviposition behavior, egg strings, eggs, larvae, and cysts of this new species. Chordodes kenyaensis n. sp. belongs to a large group of African Chordodes in which simple areoles are smooth or superficially structured less so than "blackberry" areoles but contain filamentous projections. Present among the simple areoles are clusters of bulging areoles, crowned and circurmcluster areoles along with thorn and tubercle areoles. In the laboratory, worms developed and emerged within 53-78 days from three, species of laboratory-reared crickets exposed to cysts of this species. Adult male and female C. kenyaensis n. sp. initiated typical Gordian knots within hours to days of being placed together and males deposited masses of sperm on the cloacal region of females. Females began oviposition within a week of copulating and attached egg strings in a continuous zigzag pattern on small branches or air-hoses but never free in the water column. Larvae hatched within two to three weeks, and cysts developed in laboratory-reared and exposed snails within 14-24 days. Morphological characteristics of egg strings, eggs, larvae and cysts of C. kenyaensis were most similar to other gordiids in the genus Chordodes but differed morphologically from other gordiid genera for which similar information is available.

A New Species of Gordionus (Nematomorpha: Gordiida) from the Rocky Mountains of New Mexico.

The 19 genera within the phylum Nematomorpha contain approximately 350 species. The cosmopolitan genus Gordionus Müller, 1926 contains about 58 species, 6 of which occur in the contiguous United States of America. Recently, 2 new Gordionus species were described from high-altitude streams within the southern Rocky Mountains, near Santa Fe, New Mexico. Here we describe another new Gordionus species, from a high-altitude stream in the southern Rocky Mountains, from near Taos, New Mexico. The sites consisted of temporary puddles and a small human-made stream at 3,175-3,250-m altitude in aspen/pine woodland. Gordionus lokeri n. sp. has 1 areole type, which varies in shape and size between and within body regions. Midbody areoles are elongated, polygonal, or triangular, shingled, with the raised side of the areole serrated. The interareolar space is narrow, containing few bristles. The male cloacal opening is surrounded inside and outside by narrow branching bristles that bifurcate or trifurcate deeply; the furcae then subdivide several times terminally. Adhesive warts are lacking. Genetic data, consisting of partial cytochrome c oxidase subunit I (COI) gene sequences, clearly separated G. lokeri n. sp. from other Nearctic species. This is the third Gordionus species described from high-elevation streams in the Rocky Mountains. It appears that this high-altitude habitat represents the preferential niche for numerous species of this genus, and thus future work should focus on describing gordiid diversity in other parts of the Rocky Mountains.

Helminths parasitizing larval fish from Pantanal, Brazil.

Fish larvae of 'corvinas' (Pachyurus bonariensis and Plagioscion ternetzi) from Sinhá Mariana Lagoon, Mato Grosso State, were collected from March 2000 to March 2004, in order to determine the parasitic fauna of fishes. Larvae from the two species were parasitized by the same endoparasites: Contracaecum sp. Type 2 (larvae) (Nematoda: Anisakidae) in the mesentery and Neoechinorhynchus (Neoechinorhynchus) paraguayensis (Acanthocephala: Neoechinorhynchidae) in the stomach and the terminal portion of the intestine. Statistical analysis showed that there was a significant positive correlation between the standard length of hosts and the abundance of acanthocephalans and nematodes, and that the prevalence of nematodes presented a significant positive correlation with the standard length of the two species of hosts, indicating the presence of a cumulative process of infection. The present study constitutes the first record of nematodes and acanthocephalans parasitizing larval fish, as well as the first record of endoparasites in fish larvae in Brazil. In addition, it lists a new locality and two species of hosts for Contracaecum sp. Type 2 (larva) and N. (N.) paraguayensis.

Ultrastructure of the protonephridial system in Neodasys chaetonotoideus (Gastrotricha: Chaetonotida) and in the ground pattern of Gastrotricha.

The taxon Neodasys has a basal position within Gastrotricha. This makes it very interesting for phylogenetic considerations in this group. To complete the reconstruction of the nephridial system in the stem species of Gastrotricha started earlier, we have studied the whole protonephridial system of Neodasys chaetonotoideus by means of complete sets of ultrathin sections and TEM. In many characters, protonephridia of N. chaetonotoideus resemble those of macrodasyidan gastrotrich species. For example, each of the six protonephridia, arranged in three pairs, consists of three distinct cells that constitute the continuous protonephridial lumen. Especially, the terminal cell of the protonephridia of N. chaetonotoideus shows a striking pattern: The perforation of the filter region is a meandering cleft that is continuous with the seam of the enfolded lumen of that cell. With the results presented here and that of former TEM studies, we give a comprehensive idea of the excretory organs in the ground pattern of Gastrotricha. Moreover, we can elaborate on the hypothesized protonephridial system in the stem species of Bilateria. We suggest that a meandering filtration cleft is a feature of the ground pattern of the Bilateria.

Head sensory organs of Dactylopodola baltica (Macrodasyida, Gastrotricha): a combination of transmission electron microscopical and immunocytochemical techniques.

The anterior and posterior head sensory organs of Dactylopodola baltica (Macrodasyida, Gastrotricha) were investigated by transmission electron microscopy (TEM). In addition, whole individuals were labeled with phalloidin to mark F-actin and with anti-alpha-tubulin antibodies to mark microtubuli and studied with confocal laser scanning microscopy. Immunocytochemistry reveals that the large number of ciliary processes in the anterior head sensory organ contain F-actin; no signal could be detected for alpha-tubulin. Labeling with anti-alpha-tubulin antibodies revealed that the anterior and posterior head sensory organs are innervated by a common stem of nerves from the lateral nerve cords just anterior of the dorsal brain commissure. TEM studies showed that the anterior head sensory organ is composed of one sheath cell and one sensory cell with a single branching cilium that possesses a basal inflated part and regularly arranged ciliary processes. Each ciliary process contains one central microtubule. The posterior head sensory organ consists of at least one pigmented sheath cell and several probably monociliary sensory cells. Each cilium branches into irregularly arranged ciliary processes. These characters are assumed to belong to the ground pattern of the Gastrotricha.

Observations on some unusual cell types in the enigmatic worm Xenoturbella (phylum uncertain).

The inner epithelially organized gastrodermis of the enigmatic simple worms of the genus Xenoturbella contains numerous partly phagocytized cells of two kinds, ciliated cells (PCCs) and muscle cells (PMCs). PCCs and PMCs have features of undifferentiated cells and do not derive from differentiated adult cells. Homology of phagocytized cells to pulsatile bodies in acoel and nemertodermatid flatworms is therefore rejected. The phagocytized cells might represent an hitherto unknown process of regeneration in Xenoturbella. The phagocytized material contains as much DNA as in all mitochondria and nuclei of the living cells. This is probably caused by lack of digestion of nucleic acids. The genome size of Xenoturbella bocki was determined. It has a C-value of about 0.55 pg.

The fluorescent detection of Paradiplozoon sp. (Monogenea: Diplozoidae) attachment clamps' sclerites and integumental proteins.

In the genus Paradiplozoon, few hard structures are present therefore most of the taxonomic studies have focused on the unique ultrastructure of the sclerites. Alcohol-fixed specimens were transferred to BSA-saline for 5 min. before staining indefinitely with 5 microl WGA-TRX, 5 microl SYTO 9 and 5 microl of Calcofluor White M2R. Rhodamine, Fluorescein and DAPI bandpass filters on the microscope enabled selective light wavelength illumination of the three flourochromes by a mercury light source. This method provided an easy and rapid methodology to show the internal sclerites of attachment clamps. It is suitable for alcohol preserved specimens and may have additional applications in other helminth organisms.

Gordius gonzalezi, a new species of horsehair worms (Nematomorpha) from Spain.

A new species, Gordius gonzalezi, is described from the Iberian Peninsula, from the Sierra de los Tormantos in Central Spain. Characteristic for the species is the presence of multiple short spines on the cuticle, not present in this form in any other Gordius species. Additionally, two further specimens are reported, which could not be determined to species level. One male of the genus Gordius is close to G. aquaticus and one female with regular areoles could belong either to the genus Gordius or to Gordionus.

Biochemistry of benzimidazole resistance.

Heavy reliance on the benzimidazole (BZ) anthelmintics since their introduction in the 1960's for the control of gastrointestinal parasites of livestock has led to widespread BZ resistance in target parasite species. The BZs exert their primary action by binding to tubulin, the major protein component of microtubules. This review discusses the biochemistry of the interaction between the BZs and tubulin from mammalian and BZ-resistant and -susceptible parasite sources, exploring aspects of the selective toxicity of these drugs and examining the mechanism of BZ resistance. Although tubulin is a highly conserved protein present in both the host and the parasite, the BZs demonstrate relatively low mammalian toxicity. The selectivity of these drugs can be explained by the much higher affinity of the BZs for tubulin from the parasite at 37 degrees C compared to their affinity for tubulin from the host. This difference in affinity reflects the considerably slower rate of BZ dissociation from parasite tubulin. BZ-resistance in parasitic nematodes is characterised by a loss of high affinity BZ-parasite tubulin interactions and a corresponding increase in lower affinity interactions, although there are still significant differences between BZ-resistant parasite tubulin and tubulin from the host. These differences suggest the potential for the design of new generation BZs active against 'BZ-resistant' parasites.

Oocyte differentiation in Urechis caupo (Echiura): a fine structural study.

The fine structure of oocytes of Urechis caupo is described for seven arbitrary stages ranging from the smallest oocytes (7 mum in diameter) in the coelom to the mature oocytes (115 mum in diameter) in the storage organs. Although most types of cytoplasmic organelles accumulate more or less continuously, yolk granules do not appear until oocytes reach a diameter of 35 mum, and there is stage-specific synthesis of cortical granules in 60-80 mum oocytes. In the nucleus a single nucleolus first appears when an oocyte is 15 mum in diameter. Then a nucleolus satellite, which is about 3 mum in diameter, forms in 30 mum oocytes; this nucleolus satellite later (60-70 mum oocytes) becomes surrounded by 750 nm dense spherical bodies. Large (2-4 mum in diameter) juxtachromosomal spherules occur only in the nuclei of mature oocytes. Microvilli become progressively more numerous and longer until the oocyte reaches a diameter of 90 mum; their tips project 1 mum beyond the fibrous surface coat, which is 2 mum thick when well developed. Near the end of oocyte growth, the microvilli retract into the surface coat leaving their pinched-off tips adhering to the outside of the coat.

Two new marine gastrotrichs of the genus Ptychostomella (Macrodasyida, Thaumastodermatidae) from South Korea.

Two new thaumastodermatid gastrotrichs belonging to the genus Ptychostomella are described from the shallow sublittoral sand bottom in the eastern coast of South Korea. In having the embossed cuticular armature, Ptychostomella orientalis n. sp. is most allied to P. lepidota Clausen, 2000, the only congeneric species so far known as possessing such a cuticular armature. Ptychostomella papillata n. sp. is characterized by the terrace-shaped cuticular protrusion on head region, and the numerous papillae with sensory hair(s). Description of the two new species is prepared with a character comparison table and a key to the species of Ptychostomella. This is the first record of genus Ptychostomella from the North Pacific.

Microscopy and the helminth parasite.

Microscopy has a long and distinguished history in the study of helminth parasites and has made a singularly outstanding contribution to understanding how these complex animals organise their lives and relate to their hosts. Increasingly, the microscope has been used as a powerful investigative tool in multidisciplinary approaches to parasitological problems, placing emphasis on functional correlates rather than anatomical detail. In doing so, microscopy has also uncovered a number of attributes of parasites that are of wider significance in the field of biology. Parasite surfaces have understandably demanded most of the attention of microscopists, largely as a result of the pioneering studies using transmission electron microscopy. Their findings focused the attention of physiologists and immunologists on the tegument and cuticle of helminths and in doing so helped unravel the complex molecular exchanges that are fundamental to understanding host-parasite interactions. Scanning electron microscopy succeeded in augmenting these data by revealing novel microtopographical features of the host-parasite relationship, as well as proving invaluable in helminth taxonomy and in assessing the efficacy of test substances in drug screens. Control of helminth parasites has never been more critical: problems of drug resistance demand urgent action to identify exploitable targets for new generation anthelmintics. In this regard, the neuropeptide signalling system of helminths is envisioned as central to nerve-muscle function, and thereby a crucial regulatory influence on their motility, alimentation and reproduction. The use of immunocytochemistry interfaced with confocal scanning laser microscopy has not only been instrumental in discovering the peptidergic system of helminths and its potential for chemotherapeutic exploitation, but through increasingly sophisticated bio-imaging technologies has continued to help dissect and analyse the molecular dynamics of this and other cellular systems within these important parasites.