Genetic characterization and phylogenetic relationships of Phyllodistomum parasites in Indian subcontinent: insights from freshwater fish and shrimp hosts.

Phyllodistomum is the large digenean group of fish parasites, with 25 species described so far in the Indian subcontinent. Here, we redescribed two adult species of Phyllodistomum (P. srivastava Rai 1964 and P. parorchium Jaiswal 1957) collected from freshwater fish Heteropneustes fossilis Bloch, 1974 and Glossogobius giuris Ham, 1822, respectively, and an unknown Phyllodistomum metacercaria from shrimp (Macrobrachium dayanum Henderson, 1893). These parasites were genetically characterized using 28S and first and second internal transcribed spacers (ITS1 and ITS2) regions of the nuclear ribosomal DNA and CoxI region of the mitochondrial (mt) DNA to establish the link between metacercaria and adult. Morphologically, both the unknown metacercaria in shrimp and adult Phyllodistomum srivastava in fish, resembled in terms of crenulated margin of hind body, arrangement of diagonal testes, bipartite seminal vesicle, and compact paired vitelline masses. The two adult parasite species, P. srivastava from P. parorchium, were different in terms of shape and size of the body, ratio of suckers, the absence of crenulated margin of hind body, a single chambered seminal vesicle, and deeply lobed paired vitelline masses, in the former species. Comparison of the 28S, ITS, and mtCoxI sequence data suggested P. srivastava and Phyllodistomum metacercaria belong to the same species, and supported the distinction between P. srivastava and P. parorchium. Exploring the potential impact of Phyllodistomum infection on host behaviour and health would be prospective areas for future research.

Characterization of Clinostomum (Digenea: Clinostomidae) spp. in India.

Platyhelminths belonging to the family Clinostomidae (Digenea) have a worldwide distribution and are known to infect piscivorous birds through their intermediate hosts, usually fish species. In the present study, clinostome metacercariae were collected from fish hosts, including Channa punctata (Bloch 1793) (n = 25) and Trichogaster fasciata Bloch and Schneider 1801 (n = 25), from a freshwater system in India. The experimental infection of cattle egrets, Ardea (Bubulcus) ibis Linnaeus 1758, with some of the live metacercariae found in the present study was successful. Live adult parasites were obtained from the buccal cavity of the birds. Both metacercaria and adult specimens were subjected to molecular studies to obtain the sequences of 28S, ITS1, and ITS2 (nuclear rDNA) regions. The parasites were found to belong to three species, Clinostomum giganticum Agarwal 1959; C. piscidium Southwell and Prashad 1918; and Euclinostomum heterostomum (Rudolphi 1809). Phylogenetic analyses of the sequences obtained from the adults and metacercariae established a link between the metacercariae in the fish hosts and adults in the avian host, which is essential to elucidate their partial life cycle and specify morphological characteristics in the metacercarial stage.

Morphological and molecular characterization of six Indian Tetracotyle type metacercariae (Digenea: Strigeidae Railliet, 1919), using ribosomal and mitochondrial DNA.

Strigeids have a cup-shaped fore body, containing a holdfast organ with two lobes and hind body. The species diversity of strigeids remains incomplete, especially in the Indian sub-continent. Here, we described six Tetracotyle type metacercariae (T. muscularis, T. fausti, T. lucknowensis, T. xenentodoni, T. mathuraensis and T. glossogobii) from five fresh water fish, collected at Lucknow (India). Next, we characterized these metacercariae using ribosomal (18S, 28S, ITS2) and mitochondrial DNA (COI) to determine their systematic and phylogenetic position. Molecular identification using inter-specific variation range for all four molecular markers revealed 1.9-4.9% (18S); 3.3-8.8% (28S); 6.8-12.9% (ITS2), and 3.5-9.4% (COI) among six Tetracotyle type metacercariae. In phylogenetic analysis, constructed by neighbour-joining (NJ) and maximum likelihood (ML) methods, T. fausti, T. glossogobii, T. xenentodoni, T. lucknowensis and T. mathuraenis nested as sister groups with the member of strigeids for all four markers used; T. muscularis, however, formed a basal clade. Furthermore, phylogenetic placement states the monophyly of the Tetracotyle type of metacercariae in all the markers (18S, 28S, COI), except ITS2.

Morphological and molecular characterisation of three Indian Neascus-type metacercariae (Digenea: Diplostomidae).

Diplostomid digeneans are important parasites whose larval stages infect freshwater snails and fish as first and second intermediate hosts respectively. Diplostomid digeneans as adults are parasitic in many fish-eating birds and mammals. Our understanding of the species diversity of diplostomid digeneans remains incomplete, especially in the Indian sub-continent. Here, we describe three Neascus-type metacercariae (N. hanumanthai, N. gussevi, and N. xenentodoni) from freshwater fish specimens of Channa punctata (Bloch 1793), Trichogaster fasciata (Bloch and Schneider 1801) and Xenentodon cancila (Hamilton, 1822) respectively, collected in India. Next, we characterised these metacecariae using nuclear (28S and ITS1) and mitochondrial DNA (cox1) to determine their systematic and phylogenetic position. Molecular identification using interspecific variation for all three molecular markers revealed a closer relationship between N. hanumanthai and N. gussevi (1.9%-2.4%) than either of N. hanumanthai and N. gussevi to N. xenentodoni (3.1%-3.7% and 4.4%-4.0% respectively). In phylogenetic analyses, estimated by neighbour-joining (NJ) and maximum likelihood (ML) methods, N. gussevi and N. hanumanthai nested as sister groups of Posthodiplostomum Dubois, 1936 for all three markers used; N. xenentodoni, however, nested as a sister taxa of all other diplostomes when using 28S and ITS1 and nested as a sister taxa of Bolbophorus, Alaria and Neodiplostomum using cox1. These findings suggest that N. hanumanthai and N. gussevi are members of Posthodiplostomum, but that N. xenentodoni belongs to a separate and unknown genus. Similarly, by proteomics, we found that the cox1 protein sequences and structures were similar between N. hanumanthai and N. gussevi but distinct for N. xenentodoni.

Threading: A novel insilico indagation method for genetic characterization of some diplostomoid metacercariae (Digenea:Diplostomidae Poirier, 1886).

The protein encoding zone of Mitochondrial DNA region (inherited from single lineage) seems most suitable and effective for taxonomic, systematic, ecological, evolutionary, DNA barcoding, cryptic species and population studies, exploiting nucleotide/amino acid datasets (1D/2D/3D conformational level). Nowadays, expeditious computerized methods are in trend for analyzing genetic material to demonstrate variations at various levels of protein structures. Structural proteomics have implemented here for genetic identification, differentiation and relationship of species from information rich data of mt COI gene of the family Diplostomidae with inclusion of molecular tools. Various aspects have been utilized herein for re-validation and infallible discrimination of Trematode diplostomoid metacercariae (Tetracotyle lucknowensis Pandey, 1971; T. xenentodoni Chakrabarti, 1970; T. fausti Rai and Pande, 1969; T. muscularius Chakrabarti, 1970 and Diplostomulum minutum Pandey, 1968), the infective stage in the life cycle, causing severe damage to fish host, whose adults are found mainly in fish eating birds and mammals.

A taxonomic revision of some previously described species of monogenoideans found on Indian Siluriform fishes.

A Taxonomic revision of. Bychowskyella bagariusi Sharma, 1983 and Bychowskyella singhi Rajeshwari & Kulkarni, 1983 Jain, 1952 found on Indian Siluriform fishes is being done in the present work. These species were considered as species inquirenda or nomen nudum, (Lim et al. Syst Parasitol 50:159-197, 2001) and thus, their taxonomical clarification was required Among them, B. bagariusi Sharma, 1983 from Bagarius bagarius (Hamilton) is considered as valid and B. singhi Rajeshwari & Kulkarni, 1983 from Wallago attu (Bloch & Schneider), belongs to the genus Thaparocleidus Jain, 1952.

A new species of an interesting amphistome Pseudodiplodiscoides Murty, 1970 in Bellamya bengalensis Lamarck, 1882 from water bodies near district Barabanki, U.P.

A new species of an interesting trematode genus Pseudodiplodiscoides (Murty, Zool Anz 185:312-317, 1970), is described from the gut of the snail Bellamya bengalensis Lamarck, 1882, collected from the fresh water bodies near district Barabanki, U.P. After careful examination of several freshly collected specimens, the genus is validated and another species described under the genus Diplodiscus viz., D. minutus (Saksena et al., Indian J Helminthol 4:37-54, 1987) from Indoplanorbis exustus Deshayes has also been transferred to the genus Pseudodiplodiscoides as P. mimutus (1987). The new species is characterised by the shape and size of non operculated eggs, extension of vitellaria and position of genital pore.

One known and an unknown species of the genus Thaparocleidus Jain, 1952, infecting Sperata aor (Hamilton, 1822): comparison with species from China, on molecular basis.

Sperata aor commonly called as long- whiskered cat fish or "Bada Tengan" in local fish markets harboured one new and one previously known species of genus Thaparocleidus Jain, 1952, along with two species of Cornudiscoides Kulkarni, 1969, infesting gills. Thaparocleidus aori (Rizvi, 1971) Lim, 1996, was earlier described by Rizvi therefore was briefly recorded in the present study, except the egg. The newly found species Thaparocleidus susanae n.sp was characterized by the structure of its peculiar copulatory organ. Phylogenetic relationship of the two species under study, along with 14, reterived from GenBank was established using the sequences of 28S rDNA region (Dactylogyrus Diesing, 1850 taken as an out group).

Microhabitat distribution of some monogenoideans, parasitizing the gills of Wallago attu (Bl. and Sch., 1801) and their seasonal variation.

The parasitic microhabitat distribution of the monogenoideans Thaparocleidus wallagonoius Jain, 1952, Mizelleus indicus Jain, 1957and T. gomtius Jain, 1957 parasitizing the gills of a fresh-water demersal Wallago attu and their seasonal variations were investigated in the present study. The microhabitat preferences of these species were observed. The gill segments and gill areas were the most important factors segregating the monogenoidean parasites on gills, not their abundance. Low niche overlap was found within each of the four gill arches and parasites were segregated in the same microhabitats within each gill arch. The distribution of parasites was independent of the left or right side of gill. The distribution plots in our study indicated that the anterior segment of first and fourth gill arches were more preferred site of infection. Seasonal variation of parasites, recorded during 2006 & 2007, in male and female hosts was also investigated. Due to ectoparasitic nature and high host specificity exhibited by the species of monogenoidea, studies on their seasonal dynamics are very significant as they could contribute significantly to the understanding of their biological adaptations to their environment.

A redescription of Sundanonchus behuri (Agrawal & Singh, 1982) n. comb. (Monogenoidea: Dactylogyridea: Sundanonchidae) from the gills of the freshwater Gangetic leaffish, Nandus nandus (Perciformes: Nandidae) in India.

Urocleidus behuri [Agrawal N., Singh H.S., On a known and three unknown monogenetic Urocleidus Mueller, 1934. Pranikee 1982; 3: 22-34], described from Nandus nandus in India, is reassigned to Sundanonchus Lim & Furtado [ Lim L.H., Furtado J.I., Sundanonchus g.n. (Monogenea, Tetraonchoididae) from two Malaysian freshwater fishes. Fol Parasitol 1985; 32: 11-19]. Sundanonchus triradicatus Lim & Furtado [Lim L.H., Furtado J.I., Sundanonchus g.n. (Monogenea, Tetraonchoididae) from two Malaysian freshwater fishes. Fol Parasitol 1985; 32: 11-19] is determined to be a junior subjective synonym of U. behuri and a new nomenclatural combination, Sundanonchus behuri is proposed. This article provides a redescription of this new combination, S. behuri [ Agrawal N., Singh H.S., On a known and three unknown monogenetic Urocleidus Mueller, 1934. Pranikee 1982; 3: 22-34], including the previously undescribed egg morphology, based on newly collected specimens from N. nandus in India-a new geographical record.

Discriminant analysis: A supportive tool for monogenoidean taxonomy.

Data on the measurements of haptoral parts of 47 parasite species of Bychowskyella (1), Cornudiscoides (2) and Thaparocleidus (3), found on Indian freshwater catfish, were gathered from previously reported studies. Based on six morphometric variables concerning haptors, these species were subjected to discriminant analysis in order to more accurately distinguish their generic placement. This paper describes a comparative study of several classification models. Using the original variables in the study, a simple linear discriminant analysis model was constructed and models using principal components (PC) for discrimination have also been explored. The effectiveness of these models is measured in terms of percentage of correct classification. Quadratic discriminant models using original variables and their principal components provided the highest (95%) correct classification for Bychowskyella (1). Species of Thaparocleidus (3) were correctly classified using dorsal and ventral hard-part measurements (100%) as well as hook measurements. The highest percentage (75%) of correct classification for Cornudiscoides (2) was achieved through a quadratic discriminant model using hook measurements.

The status of Quadriacanthus Paperna, 1961 and Anacornuatus Dubey et al., 1991 (Monogenoidea: Dactylogyridae) with redescription of Q. kobiensis Ha Ky, 1968, new geographical records for Q. bagrae Paperna, 1979 and Q. clariadis Paperna, 1961 from India and a note on speciation in Monogenoidea.

Quadriacanthus Paperna, 1961 is revised and emended. Q. kobiensis Ha Ky, 1968 is redescribed from Clarias batrachus, and Q. bagrae Paperna, 1979 and Q. clariadis Paperna, 1961 are recorded from C. gariepinus, both from the River Gomti, Lucknow. India is a new geographical record for Q. bagrae and Q. clariadis. Anacornuatus Dubey et al., 1991 is considered a junior subjective synonym of Quadriacanthus Paperna, 1961. Quadriacanthus spp. are more specific to their hosts (micro-habitats) than geographical localizations (macro-habitats) suggesting geographical diversification of hosts plays a lesser role in monogenoidean speciation.

Monogeneans from the gills of glassfishes (Teleostei: Perciformes: Ambassidae) in India, with the proposal of Chandacleidus n. g. (Monogenea: Dactylogyridae).

Chandacleidus n. g. (Monogenea, Dactylogyridae) is proposed to include three species collected from the gills of Indian glassfishes (Ambassidae): Chandacleidus recurvatus (Jain, 1961) n. comb. (syn. Urocleidus recurvatus Jain, 1961) from Chanda nama and C. ranga (new host record) is redescribed; and Chandacleidus saiensis n. sp. and C. lucknowensis n. sp., both from Chanda nama and C. baculis, are described. Chandacleidus n. g. is characterised by species possessing: posteriorly united intestinal caeca; overlapping gonads (testis dorsal to ovary); a counterclockwise male copulatory organ; a grooved accessory piece; a dextro-marginal vaginal pore; a haptor with two lateral flaps and armed with dissimilar dorsal and ventral anchor/bar complexes and 14 similar hooks (dissimilar in size); and hook shanks comprised of two subunits.

Spicocleidus namae n. g., n. sp. (Monogenea: Dactylogyridae) on Chanda nama (Hamilton) from Lucknow, India.

Spicocleidus n. g. is proposed for S. namae n. sp., a new monogenean collected from the gills of an ambassid, Chanda nama (Ham.), in the River Sai near Lucknow, Uttar Pradesh, India. The new genus is characterised by a pair of modified dorsal anchors (spikes), the absence of a dorsal bar, alate lateral expansion of the anterior haptor and the post-ovarian position of the testis.

Monogenoids from the gills of spiny eels (Teleostei: Mastacembelidae) in India and Iraq, proposal of Mastacembelocleidus gen. n., and status of the Indian species of Actinocleidus, Urocleidus and Haplocleidus (Monogenoidea: Dactylogyridae).

Mastacembelocleidus gen. n. (Monogenoidea: Dactylogyridae) is proposed to include two species collected and redescribed from spiny eels (Mastacembelidae) in India and Iraq: Mastacembelocleidus bam (Tripathi, 1959) comb. n. (syn. Ancyrocephalus bam Tripathi, 1959) from the gills of Macrognathus pancalus (new host record) and Macrognathus aculeatus (Synbranchiformes: Mastacembelidae) from Lucknow, India; and Mastacembelocleidus heteranchorus (Kulkami, 1969) comb. n. (syn. Urocleidus heteranchorus Kulkarni, 1969) from the gills of Mastacembelus armatus from Lucknow, India, and Mastacembelus mastacembelus (new host record) from the environs of Erbil, Iraq (new locality record). Urocleidus rhyncobdelli Jain, 1959, Haliotrema tandani Agrawal et Singh, 1982 and Urocleidus raipurensis Dubey, Gupta et Agarwal, 1992 are considered junior subjective synonyms of M. bam.

Redescription of some Indian species of Thaparocleidus Jain, 1952 (Monogenea), with aspects of the developmental biology and mode of attachment of T. pusillus (Gusev, 1976).

Thaparocleidus wallagonius Jain, 1952, the type-species of the genus, is redescribed from the type-host and locality. T. gomtius (Jain, 1952), T. seenghali (Jain, 1961), T. indicus (Kulkarni, 1969), T. malabaricus (Gusev, 1976) and T. pusillus (Gusev, 1976) were also redescribed, as their original descriptions are incomplete. The following new synonymies for some Indian species are proposed: Urocleidus bhagwanpurensis Agrawal & Singh, 1982, as a junior synonym of T. wallagonius; U. gorakhpurensis Agrawal & Singh, 1982, Murraytrema gorakhpurensis Agrawal & Singh, 1985 and Ancylodiscoides kulkarnii Agrawal & Sharma, 1988 as junior synonyms of T. seenghali; and U. tandani Agrawal & Singh, 1982, A. meerutensis Singh, Agarwal, Kumar & Vibha, 1997 and Silurodiscoides elongatus Singh, Agarwal, Kumar & Vibha, 1997 as junior synonyms of T. indicus. Data on the developmental biology and mode of attachment of T. pusillus from Mystus bleekeri at Lucknow are included.