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Intraspecific genetic differentiation of the trematode Skrjabinolecithum spasskii and its phylogenetic relationships with other species of the family Haploporidae were studied by comparing the nucleotide sequences of a part of the 28S rRNA gene and the ITS1-5.8S-ITS2 rDNA region. Trematodes were isolated from so-iuy mullet Liza haematocheila fishes collected in rivers of Primorye and flathead grey mullet Mugil cephalus fishes collected in water bodies of Vietnam (27 fishes in total). A phylogenetic analysis showed that S. spasskii is close to species of the genus Capitimitta of the subfamily Waretrematinae. By intraspecific variation of rDNA sequences, trematodes were divided into three groups with tree different genotypes, which had fixed nucleotide substitutions. Genotype I was found in trematodes from fishes collected in Primorye. Genotype II was detected in trematodes from M. cephalus fishes collected in the Tonkin Bay, Cat Ba Island, Vietnam. Genotype III was found in five trematodes from L. haematocheila collected in the Kievka River, Primorye. The genetic distances between genotypes I and III from Primorye were 0.4 and 0.65% by 28S and ITS rDNA sequences, respectively. The lowest genetic distances were observed between genotypes II (Vietnam) and III (Primorye), 0.1 and 0.33% by 28S and ITS rDNA sequences, respectively. Possible causes of genetic differentiation of S. spasskii from different geographic locations and different definitive host species are discussed.

We have determined that S-SAP method (Sequence specific amplification polymorphism) reveals clonal variability in the genomes of larvae of flatworm Himasthla elongata (Trematoda, Echinostomatidae). Being parthenogenetic the larvae were previously considered to be genetically homogeneous. Cloning and sequencing of a -500 bp conservative fragment (B1) from the fragments' pattern has been performed. Sequence analysis of B1 has shown that this fragment has maximum homology with LINE elements from CR1 family of Hydra and sparrow. In situ hybridization (FISH) has detected dispersed distribution of B1. Several other fragments cloned from the same lane of agarose electrophoresis correspond to conservative domain of reverse transcriptase (RT) from CR1 family. Thus, we have shown that 1) cercariae of trematode H. elongata have clonal variability; 2) the S-SAP method allows to obtaining patterns of fragment distribution characteristic of individual cercariae; 3) conservative domain of RT of CR1 family participates in the pattern of polymorphic fragments generation. Identification of the CR1 transcripts in cercariae of H. elongata transcriptome is the aim of the future work. Cloning of the variable fragments from the fragments' pattern is in progress.

The color of the broodsac sporocyst traditionally serves as the main taxonomic criterion for distinguishing of trematodes of the genus Leucochloridium. Broodsacs of L. paradoxum (Cams, 1835) are green, while broodsacs of L. perturbation (Pojmanska, 1969) are brown. We used molecular genetic analysis of sporocyst rDNA for verifying the accuracy of the mentioned morphological criteria. Trematode infected snails Succinea sp. were collected in Vyritsa and Lyuban (Leningrad Province, Russia). Nucleotide sequences of L. paradoxum (n = 18) and L. perturbatum (n = 10) rDNA including transcribed spacers (ITS1 and ITS2) and 5.8 S rRNA gene were received, rDNA fragments of Leucochloridium sp. sporocysts of the same color were identical. The difference in the ITS1 (2.6%) and ITS2 (6.7%) between sequences of L. paradoxum and L. perturbatum was revealed. Specific nucleotide sequences are deposited at the GeneBank.

Haploid complement of Phyllodistomumn umblae ex baikal whitefish from Lake Baikal consists of 8 chromosomes (6 metacentrics and 2 submetacentrics). The significant difference is observed in the ratio of acrocentrics and metacentrics in the haploid set of chromosomes between isolated populations of P. umblae from Baikal region and from Chukotka. The hypothesis is advanced that these chromosome populations may be chromosome races of P. umblae.

Opisthorchiasis is one of the significant naturofocal diseases in Russia. The diagnosis of opisthorchiasis may mask some diseases, such as opisthorchiasis, metorchiasis, and clonorchiasis - biohelminthoses that are induced by various representatives of the family Opisthorchiidae-Opisthorchis felineus/O.viverrini, Metorchis bilis, and Clonorchis sinensis, respectively. Coproovoscopy and serologic methods fail to accurately define the species-specific affiliation; in this connection the identification of opisthorchids, by using DNA diagnostic techniques, becomes urgent. The present paper gives the results of development of DNA diagnosticum, which differentiates parasitic diseases induced by O. felineus and M. bills. The ribosomal RNA gene cluster fragment incorporating internal transcribed spacer 2 (ITS2) was used as a diagnostic marker. A system for diagnosing opisthorchiasis was developed as a multiplex polymerase chain analysis and tested on 37 patients infected with various species of opisthorchids.

Three arbitrary primers produced 114 RAPD markers for 37 cercariae from nine Bucephalus polymorphus sporocysts obtained from three Dreissena polymorpha mollusks, which were collected in two water reservoirs of the Volga basin. Analysis of the RAPD patterns established a unique genotype for each cercaria. The topology of an UPGMA dendrogram did not reliably differentiate the cercaria according to the corresponding sporocysts. However, three groups of genotypes were isolated and corresponded to the host mollusks, indicating that each cercaria clone had a different genotype set. A within-sporocyst variation made the greatest contribution (53.0%) to the total RAPD diversity, while the contributions of within-host and between-host variations to the total diversity were equal (23.5%). Cercariae isolated from two mollusks of the Rybinsk Water Reservoir were more similar to each other than to cercariae from the geographically distant Gor'kovskoe Water Reservoir. Possible causes and distribution specifics of the observed genetic diversity of B. polymorphus are discussed.

A novel transposable element Hemar 1 was identified and characterized in the genome of Himasthla elongata flatworm. This element is a member of mariner family; it belongs to capitata subfamily and highly homologues to turbellarian worm Dugesia tigrina mariner transposon. Hemar 1 is a dispersed repeat and accounts for about 0.01% of H. elongata genome. Identified element Hemar 1 represents useful tool for investigation of functions of the mobile elements in H. elongata genome.

The random amplified polymorphic DNA (RAPD) technique was applied for the studies of genetic heterogeny between several natural populations of trematodes belonging to the Microphallida family. Initially, the metacercariae from the daughter sporocysts infestating Littorina saxatilis and Littorina littorea periwinkles were used. Comparison of the banding patterns obtained for the different metacercariae within one sporocyst gave an unpredicted results. For two of three studied species (M. pygmaeus and M. pseudopygmaeus), the considerable differences in RAPD patterning was detected. According to the classical point of view, the process of cercariae (metacercariae in case of the "pygmaeus" group of microphallides) formation does not include DNA recombination. Because of that, all metacercariae within one single sporocyst should be genetically identical. However, data obtained clearly shows that at least in some cases it is not so. We can hardly believe that such result could be a methodological artifact, for not single difference in a RAPD patterns was recorded between the metacercariae within sporocysts of M. piriformes. Moreover, even the 100 fold dilution of the total DNA used for PCR amplification does not change the banding patterns. Hence, our results can not be explained by slight fluctuations in the DNA concentrations between the samples. The most evident conclusion is that we came across yet strange, but real phenomenon--some degree of genetic difference within the progeny of each of the sporocysts--metacercariae. However, the detailed study is needed to understand and interpret these observations correctly. Amplification of the total DNA extracted from the whole sporocysts (containing metacercariae) never showed any differences in RAPD patterns between the parasites been derived from one infestated snail (local parasite hemipopulation). That allowed us to compare different parasite populations referring the RAPD pattern of one sporocyst from a snail to as a representative of one local hemipopulation. Analysis of the RAPD-loci frequencies showed a considerable genetic differences between the subpopulations of M. piriformes, infestating different paraxenic intermediate hosts--L. saxatilis and L. obtusata. This phenomenon was statistically significant for 2 localities of 3 studied. No heterogeny within populations was recorded for M. pygmaeus. Both M. piriformes and M. pygmaeus are characterized by the genetic differentiation in the microgeographic scale (within the Chupa bay of the White sea, the longest distance between the analyzed localities is 20 km). According to the frequencies of the RAPD-loci, parasites from the sheltered locality differ significantly from the parasites of other two localities exposed to the open sea. For both species the degree of genetic similarity between the populations correlates positively with the distance between the localities. We can suppose that the population structure of microphallids depended mainly upon the population structures of their intermediate hosts, definitive hosts and geographical structure of the areal. However, taking into consideration the low motility of snails, we believe that the distribution, migration and species composition of the definitive hosts play the key role in the genetic structuring of M. pygmaeus and M. piriformes hemipopulations. As an addition, the RAPD analysis of the parasite populations from the Barents Sea (East cost of Murman peninsula) and North Sea (Western cost of Sweden) revealed no significant genetic differences between the worms from those places and from the White Sea. However in case of this macrogeographic comparison, insufficient number of samples does not allow us to draw any final conclusions.

Polymorphism of the genome DNA in different development stages of trematodes by means of the polymerase chain reaction (PCR) with universal primers was investigated. Interspecific variation in Trichobilharzia ocellata (cercaria) was shown in samples from the Moscow area and some regions in Byelorussia (Naroch Lake). Population groupments of T. ocellata are correlated with migration routes of subpopulation groupments of city populations of the mallard duck (Anas platyrhynchus). The cercariae of T. ocellata from rather distant geographical isolates (Moscow and Naroch Lake) differ by PCR pattern in greater extent than groupments of cercariae within Moscow. These distant isolates are considered to be members of separate populations. The cercariae of T. ocellata emitted from infected snails belonging to the different species (Lymnaea auricularia, L. ovata, L. pusilla pusilla) and during definite period (6 days with 24 h analysis) do not differ by the PCR patterns. T. ocellata cercariae emitted from L. auricularia in autumn period (from molluscs of the new generation in "new" summer infection) are found to be different significantly from the "spring" cercariae of the same year and the "autumn" ones of the previous year. The study of the experimentally reproduced life cycle of Opisthorchis felineus revealed an essential difference of cercariae genome. DNA from that in other development stages. This indicates, that genetic reorganisation in the genome of products of parthenogenetic development took place in the parthenita stage.

The genetic variability of helminths belonging to the three classes Cestoda, Nematoda and Trematoda was analysed by means of polymerase chain reaction with randomly amplified polymorphic DNA (RAPD-PCR). Dendrograms of genetic relationships between some representatives of the genus Trichinella, Fasciola hepatica, Cysticerus ovis, Echinococcus granulosus, and Nematodirus spathiger were constructed on the basis of DNA amplification patterns. The formation of ecological and genetic adaptations in the helminth groups studied is discussed.

Population variability of the adhesive apparatus and copulatory organ of Tetraonchus momenteron from young and adult pikes (Esox lucius) from Rybinsk reservoir (Volga river system) and adult pikes from lake Baikal has been studied. Comparison analysis of seven plastic and one meristic characters allowed to discover differences between the groups of monogeneans from young and adult pikes from "rybinsk" population by frequently of the variations of number of the tight coils of copulatory organ support bar. Differences between "rybinsk" and "baikal" populations of T. monenteron were found. The "baikal" monogeneans differ from the "rybinsk" ones by frequently of the copulatory organ phenes and also more large sizes of the chitinoid formations.

The chromosomal apparatus of three species of trematodes of Diplostomum (D. spathaceum, D. indistinctum, D. mergi) and one species Tylodelphys (T. clavata) is described. It has been suggested that karyological investigations can be used for taxonomic purposes. The opinion has been expressed that the number of chromosomes in the karyotype of these trematodes plays no taxonomic role. Of great taxonomic significance are the size and morphology of chromosomes. Differences in karyotypes of trematodes belonging to different species and genera were revealed by these characters. The absence of close dependence between morphology of adult and specific peculiarities of karyotypes in species of the same genus enables in some cases the use of chromosomal apparatus as a most reliable criterion for a differentiation of close species.

Morphology, ecology and chromosomes of P. asperum and P. saginatum were investigated. The both species have a diploid set consisting-of 20 chromosomes with similar morphology and size of bivalents. Their karyotypes are supposed to be identical. According to their morphology and ecology the both species are clearly differentiated. A conclusion is drawn on the distinct status of these trematodes having originated as a result of simpatric speciation.