Cryptosporidium spp. and Eimeria spp. (Apicomplexa: Eimeriorina) of freshwater Cyprinid fish species in the Kura River basin in Azerbaijan territory.

This study aims to determine the prevalence of Cryptosporidium and Eimeria spp. oocysts in fish specimens in the river Kura. It was conducted during the 2021-2022 at two sites: Mingachevir reservoir in central Azerbaijan and in Neftchala district where the river finally enters the Caspian Sea through a delta of the Kura River estuary. The diagnosis of oocysts was performed microscopically. Fine smears from the intestine epithelial layers stained by Ziehl-Neelsen for Cryptosporidium oocysts. To identify Eimeria oocysts, each fish's faecal material and intestinal scrapings were examined directly under a light microscope in wet samples on glass slides with a coverslip. Results revealed a prevalence of Cryptosporidium and Eimeria species infections in fish hosts from both territories Rutilus caspicus, Alburnus filippi, Abramis brama orientalis and Carassius gibelio. Of 170 investigated fish specimens, 8.8% (15/170) were infected with Cryptosporidium species oocysts. Eimeria species oocysts were identified in 20.6% (35/170). The presence of Cryptosporidium and Eimeria infections in fish specimens are natural infections. However, their presence in fish species may be attributed to the age of the fish species and water pollution. This is the first report regarding the prevalence of Cryptosporidium oocysts in fish species in Azerbaijan.

Sequence and Haplotype Analyses of Ligula intestinalis in Acanthobrama marmid (Cyprinidae) in Turkey.

PURPOSE: Ligulosis caused by Ligula intestinalis adversely affects the fisheries carried out in the lakes and ponds, causing economic losses in the fish industry. In this study, it was aimed to reveal the molecular characterization of L. intestinalis isolates obtained from woodfish (Acanthobrama marmid) in Keban Dam Lake in Elazig province of Turkey by using mt-CO1 gene sequences and to determine the genetic differences and haplotypes between the isolates. METHODS: In the examination made in terms of L. intestinalis, the intestine of the fish was opened with the help of fine-tipped scissors, the contents were allowed to come out, and the parasites were taken into a petri dish containing phosphate buffered saline (PBS). Then, L. intestinalis plerocercoids were taken into 15 ml falcon tubes containing 70% ethanol and stored at - 20 °C until further analysis. From each isolate, total gDNA was extracted from the plerocercoids. A partial (480 bp) mt-CO1 gene was amplified by PCR and sequenced unidirectionally. The final size of the trimmed sequences was 392 bp for 43 sequences. Sequence and haplotype analyses were performed, followed by phylogenetic analyses. RESULTS: All isolates were confirmed as L. intestinalis by BLAST analysis. In addition, 87 nucleotide mutation positions were determined among 43 CO1 gene sequences. As a result of the haplotype network performed for the mt-CO1 gene region of L. intestinalis isolates; arranged in a star-like configuration with the main haplotype (Hap05), separated from other haplotypes by 1-6 mutation steps, and 29 haplotypes were identified, covering 13.9% (6/43) of the total isolates. Also, 75 variable (polymorphic) sites were determined, 52 of which were parsimony informative sites. CONCLUSIONS: The molecular characterization of L. intestinalis in woodfish (A. marmid) was identified for the first time in Turkey.

DNA metabarcoding reveals spatial and temporal variation of fish eye fluke communities in lake ecosystems.

Eye flukes (Diplostomidae) are diverse and abundant trematode parasites that form multi-species communities in fish with negative effects on host fitness and survival. However, the environmental factors and host-related characteristics that determine species diversity, composition, and coexistence in such communities remain poorly understood. Here, we developed a cost-effective cox1 region-specific DNA metabarcoding approach to characterize parasitic diplostomid communities in two common fish species (Eurasian perch and common roach) collected from seven temperate lakes in Estonia. We found considerable inter- and intra-lake, as well as inter-host species, variation in diplostomid communities. Sympatric host species characterization revealed that parasite communities were typically more diverse in roach than perch. Additionally, we detected five positive and two negative diplostomid species associations in roach, whereas only a single negative association was observed in perch. These results indicate that diplostomid communities in temperate lakes are complex and dynamic systems exhibiting both spatial and temporal heterogeneity. They are influenced by various environmental factors and by host-parasite and inter-parasite interactions. We expect that the described methodology facilitates ecological and biodiversity research of diplostomid parasites. It is also adaptable to other parasite groups where it could serve to improve current understanding of diversity, distribution, and interspecies interactions of other understudied taxa.

Supplemental description of Gyrodactylus pseudorasborae (Gyrodactylidae) parasitic on topmouth gudgeon Pseudorasbora parva (Cyprinidae) in South China.

Based on morphology and ITS sequence data, we identify and supplementally describe Gyrodactylus pseudorasborae Ondracková, Seifertová & Tkachenko, 2023 on the fins of topmouth gudgeon (Pseudoraspora parva) from freshwaters of southern China. The highest similarity (99.57% and 99.47%) to G. pseudorasborae suggested they were the same species. Prevalence and mean intensity were 45% and 2.3, respectively. The gyrodactylid species morphologically resembled G. pseudorasborae recorded from the same host species P. parva in Czech Republic, Ukraine, and Central China. But there were slight morphological differences in the shape and size of the marginal hook. Comparisons of marginal hook sickles of various Gyrodactylus species suggested that G. pseudorasborae and G. parvae were members of the G. wageneri-group. A molecular phylogeny of G. pseudorasborae with related species is presented and discussed within the context of the mechanism of local evolution of these sister species.

First report and molecular characterisation of Rhabdochona hospeti Thapar, 1950 (Thelazioidea: Rhabdochonidae) from tor barb, Tor tor (Hamilton) (Cyprinidae), in Assam, India.

Tor barbs, Tor tor (Hamilton), a cyprinid fish collected from Barak Valley, Assam, India were examined for helminth parasites. A nematode species of the genus Rhabdochona Railliet, 1916 (Rhabdochonidae) was found in the intestine of the fish specimens. The morphology of these nematodes was characterised by the presence of a funnel shaped prostom, head with four submedian sublabia, four cephalic papillae, long and slender posterior glandular oesophagus in both sexes, presence of 14 caudal papillae and two unequal spicules with pointed proximal tips in male specimens, and prominent post-equatorial vulval position, posteriorly directed vagina, uterus with fully scattered eggs in female specimens, and a tail pointed with a small mucron in both sexes. This nematode species was subsequently identified as Rhabdochona hospeti Thapar, 1950, reported herein for the first time in the northeastern India. In addition to its morphological description based on light microscopy, partial 18S and 28S rDNA sequences were generated to assess phylogenetic relationships of R. hospeti with other species of Rhabdochona.

Myxobolus coriumicus n.sp. (Cnidaria: Myxozoa) Causing Scale Disease Associated with Skin Lesions in Indian Cultured Carp, Cirrhinus mrigala in Punjab.

PURPOSE: The motive of the present study was to investigate incidences of myxozoan parasite infection in commercially important fishes and their characterization using morphological attributes and molecular approach. METHODS: The specimens of cultured Cirrhinus mrigala were sampled and various organs were examined. The plasmodia were detected on the scales. The identification of species was based on the myxospore morphology and 18S rDNA sequence analyses. For phylogenetic analysis, maximum likelihood (ML) and Bayesian inference (BI) methods were employed. The SEM and histological studies were performed to assess the damage to the scales. RESULTS: The infected fishes had white patches on the scales and red haemorrhagic lesions on the skin. The plasmodia of Myxobolus coriumicus n. sp. were located on the dorsal surface of the scale towards its exposed part. SEM study indicated that lepidonts were damaged in the infected scales. The myxospores were spherical in shape, 9.0-10.0 × 8.0-9.0 [Formula: see text]m in size, with two ovoid equal polar capsules 3.13-4.0 × 2.03-2.33 [Formula: see text]m in size, having 4-5 sutural edge markings at the posterior-lateral margins of the shell valves and binucleated sporoplasm. BLAST search based on 18S rDNA revealed 93.87% sequence similarity with M. rewensis, (MZ230381). The prevalence of infection was 6.3% and Scale Plasmodium Index (SPI) was 3 indicating heavy infection. CONCLUSION: Morpho-molecular data generated during this study enables us to conclude that the present species, M. coriumicus n. sp. infecting scales of C. mrigala is new to the science causing significant damage to the scales.

Genetic characterisation of four Lamproglena spp. (Copepoda, Lernaeidae) from Africa and the first mitochondrial data.

Females of species of Lamproglena von Nordmann, 1832 are parasitic on the gills of teleost fishes and the 38 nominal species are based on mainly morphological data. Only four of these species have been genetically characterised and no mitochondrial data are available for the genus. The present study aimed to provide representative ribosomal DNA (rDNA) data for two additional species of Lamproglena from Africa: Lamproglena clariae Fryer, 1956 and Lamproglena hoi Dippenaar, Luus-Powell et Roux, 2001, alongside mitochondrial DNA (mtDNA) for these and two other African species, Lamproglena hemprichii von Nordmann, 1832 and Lamproglena monodi Capart, 1944. The four species were collected from Clariidae, Cyprinidae, Alestidae and Cichlidae, respectively. Representative 18S rDNA and 28S rDNA data were obtained for L. clariae and L. hoi, while cox1 mtDNA was obtained for all four species. The respective haplotypes supported the distinctness of all species using all three gene regions investigated. Interestingly, species appeared to be grouped more by geographical origin than host family, with L. hoi more closely related to other African species than to Asian species also collected from cyprinid hosts. Even though the results presented here greatly add to the molecular data available for Lamproglena, there are still 32 (>80%) species for which no genetic data are available. The interpretation of the results presented here is thus preliminary and much more data are required before the phylogeny of this genus, and other members of the family, such as Lernaea Linnaeus, 1758, can be studied appropriately.

Feeding of mussel-associated leeches Hemiclepsis kasmiana on bitterling embryos: Novel interaction between parasites in a shared host.

We investigated an interaction between bitterlings and a parasitic leech Hemiclepsis kasmiana in freshwater mussel hosts. We found that leeches fed on bitterling eggs and embryos; this may exert a considerable negative effect on bitterling fitness. Host choices by females of three bitterling species may be differently affected by the presence of leeches within mussels; Tanakia limbata apparently avoided laying eggs in infested mussels while T. lanceolata and Acheilognathus rhombeus did not. Our novel findings suggest that relationships between the parasitic leech and the host mussel may be context dependent.

FIRST REPORTS OF LIGULA INTESTINALIS AND A SCHISTOCEPHALUS SP. INFECTING SMALL-BODIED FISH IN NEW BRUNSWICK, CANADA.

Morphological characteristics and DNA sequencing were used to identify plerocercoids of a Schistocephalus sp. infecting slimy sculpin (Cottus cognatus) from northern New Brunswick and plerocercoids of Ligula intestinalis infecting blacknose dace (Rhinichthys atratulus) in Fundy National Park (FNP, New Brunswick). To our knowledge, no previous publications documented either cestode from New Brunswick, Canada. Blacknose dace represent a new host record for L. intestinalis. Identifications were made based on the presence or absence of segmentation and sequencing partial nicotinamide adenine dinucleotide dehydrogenase subunit 1 (ND1; mitochondrial DNA) and/or partial cytochrome c oxidase subunit 1 (COI; mitochondrial DNA). Plerocercoids from blacknose dace in FNP were identified as Ligula intestinalis based on >99% nucleotide identity with COI for this species in the NCBI GenBank database. Plerocercoids in slimy sculpin from northern New Brunswick were identified as a Schistocephalus sp. based on high nucleotide identity with congenerics in the NCBI GenBank database. The absence of GenBank entries with sufficient high percent identity to our specimens, and potential species hybrids in this genus, prevents species-level identification of Schistocephalus sp. plerocercoids currently. The absence of previous documentation of these cestodes might reflect recent environmental change promoting the transmission of these parasites that can modulate host fish behavior, induce sterility of host fishes, and contribute to epizootics.

Paradiplozoon cirrhini n. sp. (Monogenea, Diplozoidae), a gill parasite of Cirrhinus molitorella (Cyprinidae, Labeoninae) in South China.

Paradiplozoon cirrhini n. sp. (Monogenea, Diplozoidae) is described from the gills of mud carp, Cirrhinus molitorella (Valenciennes, 1844) (Cyprinidae, Labeoninae), collected in Wuzhou, Guangxi Province, and Conghua, Guangdong Province as part of an ongoing survey of the diplozoid fauna in the Pearl River basin of China. The new Paradiplozoon species is distinguished from congeners by the structure of median plate and its outgrowth sclerites. The ITS2 sequences of the new species differ from all known available diplozoid sequences by 22.04%-38.34%. The new species is the first diplozoid species parasitic on Labeoninae in China. Molecular phylogenetic analyses using rRNA ITS2 placed Paradiplozoon cirrhini n. sp. in a sister position to the other Chinese Paradiplozoon, implying that Labeoninae represents an early and potentially ancestral host group for China Paradiplozoon. We also provided ITS2 sequences for four other diplozoids species, namely P. megalobramae Khotenovsky, 1982, P. saurogobionis (Jiang, et al., 1985) Jiang, Wu & Wang, 1989, Sindiplozoon hunanensis Yao & Wang, 1997, and Sindiplozoon sp., and validated their phylogenetic position. The results confirm that all diplozoid species are spilt into two major clades and show monophyly of Sindiplozoon but paraphyly of Paradiplozoon.

Title: Paradiplozoon cirrhini n. sp. (Monogenea, Diplozoidae), parasite des branchies de Cirrhinus molitorella (Cyprinidae, Labeoninae) dans le sud de la Chine. Abstract: Paradiplozoon cirrhini n. sp. (Monogenea, Diplozoidae) est décrit à partir des branchies de la carpe de vase Cirrhinus molitorella (Valenciennes, 1844) (Cyprinidae, Labeoninae), collectée à Wuzhou, province du Guangxi, et à Conghua, province du Guangdong dans le cadre d'une enquête en cours sur la faune des Diplozoidae du bassin de la Rivière des Perles en Chine. La nouvelle espèce de Paradiplozoon se distingue de ses congénères par la structure de la plaque médiane et ses sclérites d'excroissance. Les séquences ITS2 de la nouvelle espèce diffèrent de toutes les séquences de Diplozoidae disponibles connues de 22,04 % à 38,34 %. La nouvelle espèce est la première espèce de Diplozoidae parasite de Labeoninae en Chine. Les analyses phylogénétiques moléculaires utilisant l'ARNr ITS2 ont placé Paradiplozoon cirrhini n. sp. dans une position sœur des autres Paradiplozoon chinois, ce qui implique que les Labeoninae représente un groupe d'hôtes précoce et potentiellement ancestral pour les Paradiplozoon de Chine. Nous avons également fourni des séquences ITS2 pour quatre autres espèces de Diplozoidae, à savoir P. megalobramae Khotenovsky, 1982, P. saurogobionis (Jiang, et al., 1985) Jiang, Wu & Wang, 1989, Sindiplozoon hunanensis Yao & Wang, 1997 et Sindiplozoon sp. et validé leur position phylogénétique. Les résultats confirment que toutes les espèces de Diplozoidae sont réparties en deux clades majeurs et montrent la monophylie de Sindiplozoon mais la paraphylie de Paradiplozoon.

The parasites of a successful invader: monogeneans of the Asian topmouth gudgeon Pseudorasbora parva, with description of a new species of Gyrodactylus.

Monogenean parasites are often co-introduced with their fish hosts into novel areas. This study confirmed co-introduction of two dactylogyrids, Dactylogyrus squameus Gusev, 1955 and Bivaginogyrus obscurus (Gusev, 1955), and a newly described gyrodactylid species, Gyrodactylus pseudorasborae n. sp. into Europe along with their fish host, the invasive topmouth gudgeon Pseudorasbora parva (Temminck & Schlegel) from East Asia. All three species were observed in the lower Dnieper and middle Danube basin regions and had slightly larger haptoral hard parts than the same parasites in their native range. While dactylogyrids occurred sporadically, we recorded regular infection by G. pseudorasborae n. sp. at relatively high prevalence and abundance. This latter species was observed in both the native and non-native range of topmouth gudgeon, and resembles Gyrodactylus parvae You et al., 2008 recently described from P. parva in China. Both species were distinguished based on genetic analysis of their ITS rDNA sequence (6.6% difference), and morphometric differences in the marginal hooks and male copulatory organ. Phylogenetic analysis of dactylogyrid monogeneans showed that B. obscurus clustered with Dactylogyrus species parasitising Gobionidae and Xenocyprididae, including D. squameus, supporting recent suggestions of a paraphyletic origin of the Dactylogyrus genus. In addition to co-introduced parasites, topmouth gudgeon was infected with a local generalist, G. prostae Ergens, 1964, increasing the number of monogeneans acquired in Europe to three species. Nevertheless, monogenean infections were generally lower in non-native host populations, potentially giving an advantage to invading topmouth gudgeon.

Title: Les parasites d'un envahisseur à succès : les monogènes du goujon asiatique Pseudorasbora parva, avec description d'une nouvelle espèce de Gyrodactylus. Abstract: Les parasites monogènes sont souvent co-introduits avec leurs hôtes poissons dans de nouvelles zones. Cette étude a confirmé la co-introduction de deux Dactylogyridae, Dactylogyrus squameus Gusev, 1955 et Bivaginogyrus obscurus (Gusev, 1955), et d'une espèce de Gyrodactylidae nouvellement décrite, Gyrodactylus pseudorasborae n. sp. en Europe, en même temps que leur hôte poisson, le goujon envahissant Pseudorasbora parva (Temminck & Schlegel) qui vient d'Asie de l'Est. Les trois espèces ont été observées dans les régions du bas Dniepr et du bassin moyen du Danube et avaient des parties sclérifiées haptorales légèrement plus grandes que les mêmes parasites dans leur aire de répartition d'origine. Alors que les Dactylogyridae étaient rares, nous avons enregistré une infection régulière par G. pseudorasborae n. sp. à une prévalence et une abondance relativement élevées. Cette dernière espèce a été observée à la fois dans l'aire de répartition indigène et non indigène du goujon asiatique et ressemble à Gyrodactylus parvae You et al., 2008 récemment décrit chez P. parva en Chine. Les deux espèces ont été distinguées sur la base de l'analyse génétique de leur séquence ITS de l'ADNr (différence de 6,6 %) et des différences morphométriques dans les crochets marginaux et l'organe copulateur mâle. L'analyse phylogénétique des monogènes Dactylogyridae a montré que B. obscurus se regroupait avec des espèces de Dactylogyrus parasitant les Gobionidae et les Xenocyprididae, y compris D. squameus, soutenant les suggestions récentes d'une origine paraphylétique du genre Dactylogyrus. En plus des parasites co-introduits, le goujon asiatique était infecté par un généraliste local, G. prostae Ergens, 1964, portant le nombre de monogènes acquis en Europe à trois espèces. Néanmoins, les infections par les monogènes étaient généralement plus faibles dans les populations d'hôtes non indigènes, donnant potentiellement un avantage à l'invasion du goujon asiatique.

Gyrodactylus sprostonae Ling, 1962 infects an indigenous cyprinid in southern Africa: An expanded description.

Gyrodactylus sprostonae Ling, 1962 is a highly invasive parasite reported across freshwater environments of the northern hemisphere. The taxon was originally described from Carassius auratus (Linnaeus, 1758) and Cyprinus carpio Linnaeus, 1758 in China. This parasite has never been reported in Africa or the southern hemisphere. Recently, this taxon was collected from an indigenous yellowfish, Labeobarbus aeneus (Burchell, 1822), in the Vaal River, South Africa. The present study includes the conclusive identification of the gyrodactylid parasites collected from L. aeneus, including additional taxonomic data, using microscopy and molecular techniques. Microscopy included light microscopy (LM) of whole worms and scanning electron microscopy (SEM) of isolated haptoral sclerites. Additionally, morphometric data were obtained from SEM and compared to that generated using LM. For molecular analysis, the internal transcribed spacer (ITS) region of rDNA was amplified and phylogenetic topologies constructed. The specimens were morphometrically and genetically highly similar to other data for G. sprostonae. Additional point-to-point measurements and ITS rDNA sequences were generated for the taxon, contributing to the morphometric and molecular data for G. sprostonae. The study also includes the first study of the isolated haptoral sclerites of the taxon using SEM, with similar morphometric results to LM. This is the first record of G. sprostonae in the southern hemisphere and from a new, indigenous African host, L. aeneus, indicating host switching to smallmouth yellowfish. Furthermore, these results expand on the knowledge of the distribution of invasive parasites in South Africa, as well as Gyrodactylus species diversity in Africa.

Myxobolus shuifuensis sp. n. (Myxozoa: Myxobolidae) infecting the exotic mrigal Cirrhinus mrigala feral in China.

Myxobolus species represents a group of cosmopolitan metazoan parasites commonly harbored in the farmed and wild fish populations. Here, a species of Myxobolus is found in the kidney of an exotic mrigal Cirrhinus mrigala feral in the Yangtze River and utilized for an integrative characterization. Ellipsoidal myxospores are measured at 15.68 ± 0.8 (13.93-17.11) × 11.42 ± 0.54 (10.34-12.3) × 7.94 ± 0.35 (7.58-8.5) µm in dimension. The polar capsules are pyriform, and unequal in size. The morphological and morphometric characteristics of the present isolates are distinct from those of other congeners. Molecularly, the pairwise comparison based on the SSU rDNA sequence indicates that the present amplicon does not match any sequences available in the GenBank database and shares the highest similarity of 92.12% to Myxobolus pavlovskii (MG520369). Accordingly, we propose a name Myxobolus shuifuensis sp. n. for the present isolates. Phylogenetical trees indicate an apparent host-associated phylogenetic pattern. M. shuifuensis sp. n. groups loosely with other Myxobolus species isolated from Cirrhinus fish. Insead, it forms a sister clade to some myxosporeans belonging to the Thelohanellus genus. This result underpins the species identification and provides evidence for challenging the taxonomic separation among both morphologically comparable genera.

Morphological description and molecular characterisation of Dactylogyrus matlopong sp. n. (Monogenea: Dactylogyridae) from the South African endemic Labeobarbus aeneus (Cyprinidae: Torinae).

Dactylogyrus Diesing, 1850 is the most species-rich genus in Platyhelminthes, with over 900 documented species, that are mostly strictly specific to freshwater cyprinoids. The morphological Dactylogyrus groups afrobarbae-type, carpathicus-type, pseudoanchoratus-type, and varicorhini-type are known to occur in Africa. This study describes a new species of Dactylogyrus of the varicorhini-type from the gills of the endemic smallmouth yellowfish Labeobarbus aeneus (Burchell) from the Vaal River, Free State Province, South Africa. Dactylogyrus matlopong sp. n. is unique among its varicorhini-type congeners mainly by the accessory piece of the male copulatory complex that presents a hook-shaped subunit with defined round base not reaching the male copulatory organ, combined with the presence of a conspicuous medial projection on the anterior margin of the ventral bar. Sequences of the partial 28S and 18S rRNA genes, together with entire ITS1 region, were generated for the first time for a species of Dactylogyrus from South Africa. Concatenated phylogenetic analyses of selected Dactylogyrus spp. showed that these parasites group according to their morphological types.

Different hosts in different lakes: prevalence and population genetic structure of plerocercoids of Ligula intestinalis (Cestoda) in Czech water bodies.

Ligula intestinalis (Linnaeus, 1758) is a tapeworm parasite with a worldwide distribution that uses a wide variety of fish species as its second intermediate host. In the present study, we investigated the prevalence and population genetic structure of plerocercoids of L. intestinalis in five common cyprinoid species, roach Rutilus rutilus (Linnaeus), freshwater bream Abramis brama (Linnaeus), white bream Blicca bjoerkna (Linnaeus), bleak Alburnus alburnus (Linnaeus), and rudd Scardinius erythrophthalmus (Linnaeus), collected in six water bodies of the Czech Republic (Milada, Most, Medard, Jordán, Rímov and Lipno). Of the six study sites, the highest frequency of parasitism was recorded in Lake Medard (15%). The overall prevalence rate among the species was as follows: roach > rudd ≥ freshwater bream > bleak > white bream. Two mitochondrial genes (cytb and COI) were used to compare the population genetic structure of parasite populations using selected samples from the five fish species. The results of the phylogenetic analysis indicated that all populations of L. intestinalis were placed in Clade A, previously identified as the most common in Europe. At a finer scale, haplotype network and PCoA analyses indicated the possible emergence of host specificity of several mtDNA haplotypes to the freshwater bream. Moreover, pairwise Fixation indices (FST) revealed a significant genetic structure between the parasite population in freshwater bream and other host species. Parasite populations in roach not only showed the highest rate of prevalence but also depicted a maximum number of shared haplotypes with populations from bleak and rudd. Our results suggest that recent ecological differentiation might have influenced tapeworm populations at a fine evolutionary scale. Thus, the differences in prevalence between fish host species in different lakes might be influenced not only by the parasite's ecology, but also by its genetic diversity.

First record of trace element accumulation in a freshwater ectoparasite, Paradiplozoon ichthyoxanthon (Monogenea; Diplozoidae), infecting the gills of two yellowfish species, Labeobarbus aeneus and Labeobarbus kimberleyensis.

BACKGROUND: Elevated levels of trace elements in the aquatic environment poses risks to the health of biota and humans. Parasites are important components in ecosystems; responding to changes in the health of aquatic ecosystems and can accumulate trace elements in their tissues to higher levels than their hosts. Monogeneans are an important group of fish ectoparasites being directly exposed to the aquatic environment. METHODS: In this study concentrations of Ti, Fe, Cu, Zn, Rb, Sr and Ag were analysed in the monogenean parasite, Paradiplozoon ichthyoxanthon (by total reflection x-ray fluorescence spectrometry and graphite furnace atomic absorption spectrometry), and the muscle, liver and gills of two host fish species, Labeobarbus aeneus and Labeobarbus kimberleyensis (by inductively coupled plasma -- mass spectrometry). RESULTS: Most striking was the accumulation pattern for Zn in parasites; mean levels of Zn were as high as 1448 and 1652 mg kg-1 dw, respectively, with no significant difference between the two host-parasite groups, leading to bioconcentration factors of approximately 93 (parasite/fish muscle) and 15 (parasite/fish liver). In addition, Fe was accumulated in the parasite to a higher degree compared to the fish hosts' tissues. Cu levels were higher in P. ichthyoxanthon than in the muscle tissue of both host fishes, but lower than liver tissue. CONCLUSION: These findings demonstrate the usefulness of this parasite species as a sentinel organism in aquatic ecosystems it inhabits for Fe and Zn. Other trace elements under investigation were not accumulated higher in the parasite compared to its fish host. Lower Rb levels in the parasite compared to its hosts indicate no biomagnification of this metal. Further investigations are required to determine if similar trends in trace element accumulation occur in other monogeneans. DATA AVAILABILITY: All data generated in the analysis of host and parasite tissues are presented in the manuscript.

Morphological and Molecular Characterization of Myxobolus nkondjockei sp. nov. (Myxozoa: Myxobolidae), A Parasite of Labeo batesii Boulenger, 1911 (Teleostei: Cyprinidae) from Makombè River in Cameroon.

PURPOSE: Myxozoans are economically important group of metazoan parasites, which can cause diseases in a large variety of commercially important fishes. Increased knowledge on molecular features has shown that traditional descriptive characters may be misleading. Combination of both descriptive and molecular features is therefore necessary for an integrated taxonomic assessment. METHODS: Cyprinid Labeo batesii, sampled in the Makombè River at Nkondjock in Cameroon were examined for myxosporeans. Identification of parasite species was based on morphological and molecular sequence analyses of myxospores. Phylogenetic analysis was performed using maximum likelihood (ML) and Bayesian inference (BI) methods. RESULTS: The scales of L. batesii were infected by Myxobolus nkondjockei sp. nov Their mature myxospores are ovoid in frontal view and lenticular in lateral view, with two rounded ends. These myxospores measured 10.3 (10-10.9) µm length and 8.0 (7.3-8.5) µm width. Myxospores have two ovoid and equal sizes polar capsules. They measured 4.5 (4.0-5.0) µm in length and 2.4 (2-2.9) µm in width. Polar tubules were coiled in 4-5 turns perpendicular to the longitudinal axis of the polar capsules. Phylogenetic analysis of the 18S rDNA sequence show clustering of M. nkondjockei sp. n. close to an undetermined species Myxobolus sp. reported infecting gill lamellas of Labeo rohita from India. CONCLUSION: The morphological, molecular and phylogenetic data provided for M. nkondjockei sp. n. are solid basis for further identification of this myxozoan of which pathogenicity probably plays an economic role at culturing the hosts.

Surviving in the fish gut: Comparative inhibitory capacities against the host proteinases in cestodes of the genus Proteocephalus.

Currently, little is known about inhibitory substances enabling tapeworms to settle in fish intestines thereby avoiding proteolysis. Contrary to previous studies with certain host-parasite pairs, this research compares the inhibitory capacities in three tapeworm species of the same genus Proteocephalus from four different fishes (P. torulosus from dace and zope, P. sagittus from stone loach and P. cernuae from ruffe). The tapeworm extracts studied significantly reduced the activity of commercial trypsin (although to a lesser degree than the synthetic inhibitor of serine proteinases PMSF), displaying clear inter-specific variation in worms' inhibitory ability. We also measured the proteolytic activity of the host intestinal mucosa exposed to tapeworm extracts which served as inhibitors. Based on per cent inhibition values, all tapeworm extracts significantly suppressed the mucosal proteolytic activity, with marked differences between certain host-parasite pairs. SDS-PAGE electrophoresis of the incubation media and extracts detected in each tapeworm species 20-36 protein bands with apparent molecular weights from 10-12 to 312.5 kDa, mostly below 50 kDa. The incubation medium and extract of each parasite shared one to six bands ranging from 12 to 35 kDa, depending on its species, with only four bands common for two or more species. The band profiles suggest that in various Proteocephalus species inhibitory capacities against host proteinases can be ensured by different proteins.

Transcriptome analysis of grass carp (Ctenopharyngodon idella) and Holland's spinibarbel (Spinibarbus hollandi) infected with Ichthyophthirius multifiliis.

Ichthyophthirius multifiliis is a protozoan ciliate that causes white spot disease (also known as ichthyophthiriasis) in freshwater fish. Holland's spinibarbel (Spinibarbus hollandi) was less susceptible to white spot disease than grass carp (Ctenopharyngodon Idella). In this study, grass carp and Holland's spinibarbel are infected by I. multifiliis and the amount of infection is 10,000 theronts per fish. All grass carp died within 12 days after infection, and the survival rate of Holland's spinibarbel was more than 80%. In order to study the difference in sensitivity of these two fish species to I. multifiliis, transcriptome analysis was conducted using gill, skin, liver, spleen and head kidney of Holland's spinibarbel and grass carp at 48 h post-infection with I. multifiliis. A total of 489,296,696 clean reads were obtained by sequencing. A total of 105 significantly up-regulated immune-related genes were obtained by Gene Ontology (GO) classification and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis in grass carp. Cluster of differentiation 40 (CD40), cluster of differentiation 80 (CD 80), tumor necrosis factor-alpha (TNF-α), toll-like receptor 4 (TLR-4), interleukin 1 beta (IL-1ß) and other inflammatory-related genes in grass carp were enriched in the cytokine-cytokine receptor interaction pathway and toll-like receptor pathway. In Holland's spinibarbel, a total of 46 significantly up-regulated immune-related genes were obtained by GO classification and KEGG pathway enrichment analysis. Immune-related genes, such as Immunoglobin heavy chain (IgH), cathepsin S (CTSS), complement C1q A chain (C1qA), complement component 3 (C3) and complement component (C9) were enriched in phagosome pathway, lysosome pathway and complement and coagulation concatenation pathway. C3 was significantly up-regulated in gill and head kidney. Fluorescence in situ hybridization (FISH) showed that the C3 gene was highly expressed in gill tissue of Holland's spinibarbel infected with I. multifiliis. A small amount of C3 gene was expressed in the gill arch of grass carp after infected with I. multifiliis. In conclusion, the severe inflammatory response in vivo after infecting grass carp with I. multifiliis might be the main cause of the death of grass carp. The extrahepatic expression of the gene of Holland's spinibarbel might play an important role in the immune defense against I. multifiliis.

Individual copy number variation and extensive diversity between major MHC-DAB1 allelic lineages in the European bitterling.

Polymorphism of the major histocompatibility complex (MHC), DAB1 gene was characterized for the first time in the European bitterling (Rhodeus amarus), a freshwater fish employed in studies of host-parasite coevolution and mate choice, taking advantage of newly designed primers coupled with high-throughput amplicon sequencing. Across 221 genotyped individuals, we detected 1-4 variants per fish, with 28% individuals possessing 3-4 variants. We identified 36 DAB1 variants, and they showed high sequence diversity mostly located within predicted antigen-binding sites, and both global and codon-specific excess of non-synonymous mutations. Despite deep divergence between two major allelic lineages, functional diversity was surprisingly low (3 supertypes). Overall, these findings suggest the role of positive and balancing selection in promotion and long-time maintenance of DAB1 polymorphism. Further investigations will clarify the role of pathogen-mediated selection to drive the evolution of DAB1 variation.