Confocal laser scanning microscopy (CLSM) as a new tool for morphological characterisation of both newly collected and museum voucher specimens of the Trypanorhyncha Diesing, 1863 (Platyhelminthes: Cestoda).

Taxonomic issues within Trypanorhyncha, e.g., the inaccurate light microscopic visualisation of the hook patterns, are solvable by confocal laser scanning microscopy (CLSM). We applied CLSM imaging to study Trygonicola macropora (Shipley et Hornell, 1906) and Dollfusiella michiae (Southwell, 1929) from Neotrygon caeruliopunctata Last, White et Séret from Bali, Indonesia. To illustrate the strength and limitations of CLSM, images of Otobothrium cysticum (Mayer, 1842) and Symbothriorhynchus tigaminacantha Palm, 2004, both permanent mounts from a collection, were also processed. The CLSM created image stacks of many layers, and edited with IMARIS Software, these layers resulted in three-dimensional images of the armature patterns and internal organs of both species. BABB (benzylalcohol and benzylbenzuolate) clearing was applied to T. macropora. We conclude that trypanorhynch cestodes stained with Mayer-Schuberg's acetic carmine permanently mounted in Canada balsam are suitable for CLSM, allowing detailed analyses of museum type-material as well as freshly collected and processed worms. BABB resulted in imaging the testes in detail, suggesting other stains to be used for CLSM in trypanorhynch cestode research. Application of CLSM for studies of other cestode groups is highly recommended.

Parasites of pufferfish, Lagocephalus spp. and Torquigener flavimaculosus of the Israeli Mediterranean: A new case of Lessepsian endoparasites.

With the opening of the Suez Canal as a link between the Red Sea and the Mediterranean Sea in 1869, the biogeographical event of the Lessepsian migration has been starting. Aided by beneficial conditions in the new habitat, almost 500 marine species have immigrated and often established themselves in the Mediterranean Sea, including several pufferfish species, with all of them extending their range and becoming important components of the local fauna. The parasitic fauna of these pufferfish has scarcely been examined in the Mediterranean Sea or in their native range, which provides the opportunity to study host-parasite interaction in a new habitat. The present study describes the parasitic fauna in four alien invasive pufferfish species (Lagocephalus guentheri, L. sceleratus, L. suezensis, and Torquigener flavimaculosus) of various sizes and ages on the Israeli Mediterranean coast. The parasite fauna of these species was diverse (Maculifer dayawanensis Digenea; Calliterarhynchus gracilis, Nybelinia africana and Tetraphyllidea larvae Cestoda; Hysterothylacium reliquens, Hysterothylacium sp. and Raphidascaris sp. Nematoda; Trachellobdella lubrica Hirudinea and Caligus fugu and Taeniacanthus lagocephali Copepoda) and consisted of mostly generalist species, most likely acquired in the new habitat, and specialist copepod ectoparasites, having co-invaded with the pufferfish. Additionally, the oioxenic opecoelid digenean Maculifer dayawanensis was found in two pufferfish species. The genus was previously only known from the Indo-Pacific Ocean, representing the eighth reported case of a Lessepsian endoparasite so far. Our results suggest a change in parasite fauna to native Mediterranean species in the pufferfish like previously reported in other Lessepsian migrant predatory fish species and a wider spread of co-invasion of fish endoparasites to the Mediterranean Sea than previously assumed. The study also provides several new host records and the first report for parasites in T. flavimaculosus.

Allopodocotyle palmi sp. nov. and Prosorhynchus maternus Bray & Justine, 2006 (Digenea: Opecoelidae & Bucephalidae) from the Orange-Spotted Grouper Epinephelus coioides (Hamilton, 1822) off Bali, Indonesia, Described Using Modern Techniques.

BACKGROUND: The most convincing species of Allopodocotyle Pritchard, 1966 (Digenea: Opecoelidae) are known overwhelmingly from groupers (Serranidae: Epinephelinae). Six species of Allopodocotyle have been reported, collectively, from species of Cromileptes Swainson, 1839, Epinephelus Bloch, 1793 and Plectropomus Oken, 1817. These are A. epinepheli (Yamaguti, 1942), A. heronensis Downie & Cribb, 2011, A. manteri (Saoud & Ramadan, 1984), A. mecopera (Manter, 1940), A. plectropomi (Manter, 1963) and A. serrani (Yamaguti, 1952). In addition, a not yet fully described and unnamed seventh species, morphologically and phylogenetically close to A. epinepheli, was isolated from the orange-spotted grouper Epinephelus coioides (Hamilton, 1822) off Bali, Indonesia in 2016. An eighth species, again from E. coioides off Bali is described herein. METHODS: Morphological and phylogenetic analyses justify the recognition of A. palmi sp. nov., which is also genetically different from the as yet unnamed congener from the same host and locality. For the first time, 3D confocal laser scanning microscopy was applied to study and distinguish Digenea taxonomically. We introduce the 'Palm pattern', a new simplified way to visualise morphometric differences of related digenean taxa. RESULTS: Allopodocotyle palmi sp. nov. is distinguished from its congeners that infect groupers by its elongate body with a size > 2.7 mm and diagonal testes. The ovary is located mainly, and the anterior testis completely, in the posterior half of the body; the uterine coils are in the fourth eighth of the body. The cirrus-sac is 0.75-1.4 (1.1) mm long, its posterior extremity is well separated from the anterior extent of the vitelline fields, just reaching the anterior border of uterine coils. In addition, Prosorhynchus maternus Bray & Justine, 2006 (Bucephalidae) was isolated from E. coioides, representing the first record in Indonesia and the third record for this fish species. CONCLUSION: The biodiversity research in Indonesia is enhanced with a new species description based on modern and newly applied techniques.

Echinocephalus caniculus n. sp. (Nematoda: Gnathostomatidae Railliet, 1895) from the lesser spotted dogfish Scyliorhinus canicula (L.) (Elasmobranchii: Scyliorhinidae Gill, 1862) off Tunisia, with a key to species of the genus Echinocephalus.

Echinocephalus caniculus n. sp. (Nematoda, Gnathostomatidae Railliet, 1895) was isolated from the spiral valve of the lesser spotted dogfish Scyliorhinus canicula (L.) from the waters off Kalaat El Andalous, North East Tunisia. This new species is mainly characterized by a cephalic bulb armed with 31-39 transverse rows of uncinated hooks, a comparatively long oesophagus, short spicules and the presence of a gubernaculum. The new species differs from its congeners by having four cervical sacs of almost equal length, a higher oesophagus/body length ratio, the arrangement of the caudal papillae, the absence of a medioventral preanal organ and numerous scattered `pores´ limited to the lateral side of the posterior part of the body. This is the first report of a member of the genus Echinocephalus Molin, 1858 from the Tunisian coast, and a new host and locality record for the Gnathostomatidae. A key to the species of Echinocephalus is provided.

Third Way to Interacting p-Form Theories.

We construct a class of interacting (d-2)-form theories in d dimensions that are "third-way" consistent. This refers to the fact that the interaction terms in the p-form field equations of motion neither come from the variation of an action nor are they off-shell conserved on their own. Nevertheless, the full equation is still on-shell consistent. Various generalizations, e.g., coupling them to (d-3)-forms, where three algebras play a prominent role, are also discussed. The method to construct these models also easily recovers the modified three-dimensional Yang-Mills theory obtained earlier and straightforwardly allows for higher derivative extensions.

Redescription of Aspidogaster limacoides Diesing, 1834 (Aspidogastrea: Aspidogastridae) from freshwater fishes of northern Germany.

Aspidogaster limacoides Diesing, 1834 (Aspidogastridae) is redescribed based on light and scanning electron microscopy of specimens from the stomach and intestine of Abramis brama, Rutilus rutilus and Scardinius erythrophthalmus (Actinopterygii: Cyprinidae). The fishes were sampled during 2018 and 2019 at Lake Tollense in Mecklenburg-Western Pomerania, Germany. The prevalence of A. limacoides was highest in R. rutilus (61.7%) followed by Scardinius erythrophthalmus (7.7%) and A. brama (2.9%), while it was absent in Perca fluviatilis from the same lake. The following structures of A. limacoides are described for the first time: a depression on the ventral side of the neck, variations in the number and the arrangement of alveoli, numerous pits scattered all over the body surface, the presence of a few papillae-like structures posterior lateral to the mouth, the number of marginal organs represented by openings of exocrine multicellular glands as shown in histology and the subterminal position of the excretory pore. These characters can be used to distinguish three species of Aspidogaster, namely, A. ijimai, A. conchicola and A. limacoides, suggesting that SEM is a useful and promising tool in differentiating Aspidogaster species. Comparison of molecular data of the ITS1-5.8S-ITS2 regions showed a 94% similarity to A. limacoides from the European part of Russia. Phylogenetic analysis showed that the present specimens clustered in the same clade with A. limacoides sensu stricto, forming a distinct group to the exclusion of congeners.

Nematode Parasites in Baltic Sea Mammals, Grey Seal (Halichoerus grypus (Fabricius, 1791)) and Harbour Porpoise (Phocoena phocoena (L.)), from the German Coast.

PURPOSE: Endoparasitic nematodes of six harbour porpoises Phocoena phocoena and four grey seals Halichoerus grypus, stranded at the eastern coast of the Baltic Sea in Germany in winter 2019, were analysed in order to identify nematode parasites and to compare with recent studies from the same area. METHODS: Endoparasitic nematodes were identified by using both morphological and molecular characters. The successfully obtained sequences of the rDNA marker regions ITS-1, 5.8S, ITS-2 from 29 anisakid and the rDNA marker region ITS-2 of 11 pseudalid nematodes were amplified. RESULTS: Analyses revealed the presence of three parasite species, the anisakid nematode Contracaecum osculatum from grey seals and the pseudalid nematodes Pseudalius inflexus and Stenurus minor from the harbour porpoises. Other anisakid nematodes regularly occurring in the Baltic Sea, e.g. Anisakis simplex or Pseudoterranova decipiens, were not found. CONCLUSIONS: The prevalence of 100% and a severe parasite load in grey seals demonstrated a very high C. osculatum infection of Baltic Sea fish as their regular prey. Prevalence of 33% for parasites in harbour porpoises and minor infection rates, combined with a distinct lack of anisakid nematodes, are typical for the current situation of the porpoise parasite fauna in the Baltic Sea. Invasive parasite species as possible indicators for climate change could not be detected.

Bucephalus damriyasai n. sp. (Digenea: Bucephalidae) from the blacktip trevally Caranx heberi (Bennett) (Perciformes: Carangidae) off Bali, Indonesia.

The new species Bucephalus damriyasai n. sp. is described from Caranx heberi (Bennett) from off Bali, Indonesia. It can be distinguished from other Bucephalus spp. recorded from carangid hosts by its narrow elongate body shape and the relatively long distance between the rhynchus and the vitellarium, as well as other features distinguishing it from individual species. The most similar species are differentiated from B. damriyasai n. sp. as follows: B. carangis Yamaguti, 1970 has a much greater length, the rhynchus is smaller and the cirrus-sac is small, not always reaching to the posterior testis; B. fragilis Velasquez, 1959 is a tiny species, the pre-vitelline distance is short and the caecum is saccular; B. gorgon (Linton, 1905) is much longer and relatively broader, the uterus reaches distinctly anterior to the vitellarium and the rhyncheal tentacles appear more complex; B. labracis Paggi & Orecchia, 1965 is distinctly longer, slightly broader, with a slightly larger rhynchus, and has shorter pre-uterine and pre-mouth distances; B. paraheterotentaculatus Velasquez, 1959 is much longer, relatively rather broad, the rhynchus is said to bear 21 tentacles, the post-testicular region and cirrus-sac reach are longer and the caecum is described as saccular; B. sphyraenae Yamaguti, 1952 is longer, slightly broader, the uterus reaches anteriorly to the vitellarium and the caecum is claviform and oriented anteriorly; B. margaritae Ozaki & Ishibashi, 1934 (syn. B. varicus Manter, 1940) is relatively squat, has shorter pre-vitelline and pre-mouth distances and a longer post-testicular distance and cirrus-sac reach; B. yamagutii Gupta & Singh, 1985 is broader, with a relatively short pre-vitelline distance, the caecum extends anteriorly to the pharynx, but not posteriorly and the rhynchus is said to carry five tentacles. The distinctive features of B. damriyasai n. sp. are compared with those of all other marine Bucephalus spp. in a table. The number of bucephalid trematodes known from Indonesian waters is now 13, two of them await further identification. They have been described from the fish families Carangidae, Platycephalidae, Sciaenidae, Serranidae and Sphyraenidae.

Pseudempleurosoma haywardi sp. nov. (Monogenea: Ancyrocephalidae (sensu lato) Bychowsky & Nagibina, 1968): An endoparasite of croakers (Teleostei: Sciaenidae) from Indonesia.

An endoparasitic monogenean was identified for the first time from Indonesia. The oesophagus and anterior stomach of the croakers Nibea soldado (Lacépède) and Otolithes ruber (Bloch & Schneider) (n = 35 each) sampled from the South Java coast in May 2011 and Johnius amblycephalus (Bleeker) (n = 2) (all Sciaenidae) from Kedonganan fish market, South Bali coast, in November 2016, were infected with Pseudempleurosoma haywardi sp. nov. Prevalences in the first two croakers were 63% and 46%, respectively, and the two J. amblycephalus harboured three and five individuals. All three croakers represent new hosts for this monogenean genus. We provide infection rates, light microscopical observations, 3D confocal microscopical illustrations, and a morphometric comparison with all congeners. The new species differs in body size, the position and shape of the ovary and testes, and especially in the composition of the dorsal anchor complex, with the dorsal bar being anteriorly concave rather than planar or convex as in its congeners. The dorsal and ventral anchors of this new species are the longest in the genus, whereas the male copulatory organ is the smallest. The first DNA sequences for a member of this genus demonstrate the greatest similarity with endoparasitic freshwater monogeneans from African cichlid fishes. This suggests a freshwater origin for these marine endoparasitic monogeneans.

Parasite fauna of Epinephelus coioides (Hamilton, 1822) (Epinephelidae) as environmental indicator under heavily polluted conditions in Jakarta Bay, Indonesia.

The objective of this study was to assess the environmental conditions of a heavily polluted marine habitat using descriptors of fish parasites. Epinephelus coioides from Jakarta Bay as well as off Jakarta Bay was studied for metazoan parasites. Based on 70 fish and considering previous studies (230 fish), an environmental indicator system was designed. Including the recent study, a total of 51 parasite species have been recorded for E. coioides in Indonesian waters. Seven of them combined with five parasitological indices are useful descriptors for the environmental status of marine ecosystems. The results are visualized in a star graph. A significant different parasite infection between nine analyzed localities demonstrates the negative influence of the megacity Jakarta onto the coastal environment. We herewith complete a parasite based indicator system for Indonesian coastal waters, and suggest that it can be used in other marine habitats as well as for further epinephelids.

Microbial Diversity and Parasitic Load in Tropical Fish of Different Environmental Conditions.

In this study we analysed fecal bacterial communities and parasites of three important Indonesian fish species, Epinephelus fuscoguttatus, Epinephelus sexfasciatus and Atule mate. We then compared the biodiversity of bacterial communities and parasites of these three fish species collected in highly polluted Jakarta Bay with those collected in less polluted Indonesian areas of Cilacap (E. sexfasciatus, A. mate) and Thousand Islands (E. fuscoguttatus). In addition, E. fuscoguttatus from net cages in an open water mariculture facility was compared with free living E. fuscoguttatus from its surroundings. Both core and shared microbiomes were investigated. Our results reveal that, while the core microbiomes of all three fish species were composed of fairly the same classes of bacteria, the proportions of these bacterial classes strongly varied. The microbial composition of phylogenetically distant fish species, i.e. A. mate and E. sexfasciatus from Jakarta Bay and Cilacap were more closely related than the microbial composition of more phylogentically closer species, i.e. E. fuscoguttatus, E. sexfasciatus from Jakarta Bay, Cilacap and Thousand Islands. In addition, we detected a weak negative correlation between the load of selected bacterial pathogens, i.e. Vibrio sp. and Photobacterium sp. and the number of endoparasites. In the case of Flavobacterium sp. the opposite was observed, i.e. a weak positive correlation. Of the three recorded pathogenic bacterial genera, Vibrio sp. was commonly found in E. fuscoguttatus from mariculture, and lessly in the vicinity of the net cages and rarely in the fishes from the heavily polluted waters from Jakarta Bay. Flavobacterium sp. showed higher counts in mariculture fish and Photobacteria sp. was the most prominent in fish inside and close to the net cages.