Electrophoretic comparison of myosin light chains and parvalbumins of trunk and head muscles from two barbel (Barbus barbus) populations

• 1.1. Myosin light chains and parvalbumins have been compared in several trunk and head muscles from small and large size barbels (Barbus barbus) living in river or reared in hatchery.
• 2.2. These proteins isolated from white, red and ventricle fibres exhibit identical electrophoretic characteristics in the four batches.
• 3.3. The slow fibre content and the parvalbumin distribution are generally similar in river and hatchery barbels of the same size but differ between small and large barbels within a population.
• 4.4. Alterations of the mode of fertilization and breeding conditions do not modify the differentiation of myosin and parvalbumins in barbels.

     

Phylogeny and evolution of the genus Barbus in the Iberian Peninsula as revealed by allozyme electrophoresis

Variation of 14 enzyme systems encoded by 31 presumptive loci in different barbel species was studied using starch gel electrophoresis. Eighteen samples representing 11 Barbus tetraploid taxa were taken, including 10 populations from the Iberian Peninsula, six from other parts of western and southern Europe, one from northern Africa and one diploid species as outgroup from Asia Minor. The genetic analysis reassessed of the taxonomic status of the Iberian barbels into two distinct species groups. The first group included B. bocagei, B. comiza, B. graellsii, B. gulraonis, B. microcephalus , and B. sclateri , that aligned with B. callensis from northern Africa and with B. apoensis from Asia Minor; the other group included B. haasi and B. merldlonahs that was related to the European species, B. barbus, B. plebejus and B. peloponnesius . These groups are probably not monophyletic. It is suggested that the isolation of the Iberian Peninsula from Europe since the Oligocene-Miocene may explain the genetic affinities of the Iberian barbels with those of North African rather than with the European group.     

Phylogenetic relationships of Iberian cyprinids: systematic and biogeographical implications.

The phylogenetic relationships among all Iberian endemic cyprinids were inferred using the complete nucleotide sequence of the cytochrome b gene. The inferred molecular phylogeny included representatives from Central European, Asian and North African species, and is highly congruent with previous phylogenies based on osteological characters. Iberian cyprinids were grouped into only five, very speciose lineages (with the exception of the monotypic Anaecypris): Barbus, Luciobarbus, Chondrostoma, Leuciscus and Anaecypris. The existence of such a relatively small number of Iberian cyprinid lineages can be explained by the historical isolation of the Iberian Peninsula. North African and Asian barbels are the sister group of Iberian Luciobarbus, supporting a south-eastern route of colonization of the Iberian Peninsula for this subgenus. With leuciscins, Anaecypris hispanica was considered a relict species as it could not be related to any other Iberian cyprinid. The phylogenetic relationships among the main lineages of Iberian cyprinids based on cytochrome b sequence data supported the traditional division of the Cyprinidae into two subfamilies: Cyprininae and Leuciscinae.     

Functional characterization of parvalbumin from the Arctic cod (Boreogadus saida): similarity in calcium affinity among parvalbumins from polar teleosts

Calcium dissociation constants (KD) were measured as a function of temperature for parvalbumin, a small acidic protein expressed abundantly in fast-twitch muscle, from the Arctic cod (Boreogadus saida) and compared to values previously determined for Antarctic and temperate zone teleosts. Estimates of KD were derived independently from fluorometric titrations and calorimetry. In addition, the primary structure of B. saida parvalbumin was determined. Calcium KDs for parvalbumin from B. saida were fundamentally similar to those for parvalbumins from Antarctic species (6.68+/-0.59 nM and 7.77+/-0.72 nM at 5 degrees C, respectively), but significantly different from temperate zone species (1.35+/-0.28 nM at 5 degrees C). However, estimates of KD for B. saida parvalbumin at 5 degrees C closely matched values for temperate zone fish at 25 degrees C (6.54+/-0.56 nM), recapitulating the prior observation that calcium affinity of parvalbumin is conserved at the native temperature of teleost fish. Full sequence of B. saida parvalbumin was generated using reverse-phase HPLC and RACE-PCR. The Arctic parvalbumin showed 83% homology to a carp parvalbumin. None of the 16 total substitutions between the two parvalbumins resided in the cation binding sites of the protein, indicating that the structural locus of the thermal sensitivity of function lies outside the active regions.     

Expression of myofibrillar proteins and parvalbumin isoforms in white muscle of dorada during development

Several polyacrylamide gel electrophoresis techniques were used to study developmental changes in myofibrillar protein composition and parvalbumin distribution in the myotomal muscle of Brycon moorei . Two myosin LC2 chains and two troponin I isoforms were successively detected. Up to four troponin T isoforms were synthesized. Slow red-muscle myofibrils from adult fish showed no common component (except actin) with larval, juvenile or adult fast white-muscle myofibrils. During growth of B. moorei , two classes of parvalbumin isoforms were sequentially expressed: larval PA I, PA IIa, and PA IIb and adult PA III. In adult fish, the content in Tn T-2 isoform decreased from the anterior to the posterior myomeres, in favour of Tn T-1 and Tn T-4. The parvalbumin content also diminished from the rostral to the caudal muscle. The fast rate of transition from larval to adult isoforms appeared to parallel the extremely fast growth of B. moorei . Sequential expression of these isoforms presumably reflected variations in the contractile properties of the muscle fibres, required by changes in physiological demands of the propulsive musculature.     

Genome‐wide selection footprints and deleterious variations in young Asian allotetraploid rapeseed

Brassica napus (AACC, 2n = 38) is an important oilseed crop grown worldwide. However, little is known about the population evolution of this species, the genomic difference between its major genetic groups, such as European and Asian rapeseed, and the impacts of historical large‐scale introgression events on this young tetraploid. In this study, we reported the de novo assembly of the genome sequences of an Asian rapeseed (B. napus), Ningyou 7, and its four progenitors and compared these genomes with other available genomic data from diverse European and Asian cultivars. Our results showed that Asian rapeseed originally derived from European rapeseed but subsequently significantly diverged, with rapid genome differentiation after hybridization and intensive local selective breeding. The first historical introgression of B. rapa dramatically broadened the allelic pool but decreased the deleterious variations of Asian rapeseed. The second historical introgression of the double‐low traits of European rapeseed (canola) has reshaped Asian rapeseed into two groups (double‐low and double‐high), accompanied by an increase in genetic load in the double‐low group. This study demonstrates distinctive genomic footprints and deleterious SNP (single nucleotide polymorphism) variants for local adaptation by recent intra‐ and interspecies introgression events and provides novel insights for understanding the rapid genome evolution of a young allopolyploid crop.     

Allozyme variation of the pygmy wood mouse Sylvaemus uralensis (Rodentia, Muridae) and estimation of the divergence of its chromosome forms

The genetic divergence between the eastern European, southern European, and Asian chromosome forms of the pygmy wood mouse Sylvaemus uralensis, whose karyotypes differ from one another in the amount of pericentromeric heterochromatin, has been reevaluated using allozyme analysis. In general, Asian S. uralensis living in eastern Kazakhstan, eastern Turkmenistan (the Kugitang Ridge), and Uzbekistan are more monomorphic than European populations of this species. However, the allozyme differences between all chromosome forms of the pygmy wood mouse is comparable with the interpopulation differences within each form and are an order of magnitude smaller than those between "good" species of the genus Sylvaemus. Thus, the chromosome forms of S. uralensis cannot be considered to be separate species. The concept of races as large population groups that have not diverged enough to regard them as species but differ from one another in some genetic characters is used to describe the differentiation of S. uralensis forms more adequately. The currently available evidence suggests the existence of two S. uralensis races, the Asian and the European ones, and two chromosome forms (eastern and western) of the European race. The possible historical factors that have determined the formation of the races of the pygmy wood mouse are considered. According to the most plausible hypothesis, the shift and fragmentation of the broad-leaved forest zone during the most recent glacial period (late Pleistocene) were the crucial factors of the formation of these races, because they resulted in a prolonged isolation of the European and Asian population groups of S. uralensis from each other.     

Phylogeography, genetic structure and diversity in the endangered bearded vulture (Gypaetus barbatus, L) as revealed by mitochondrial DNA

Bearded vulture populations in the Western Palearctic have experienced a severe decline during the last two centuries that has led to the near extinction of the species in Europe. In this study we analyse the sequence variation at the mitochondrial control region throughout the species range to infer its recent evolutionary history and to evaluate the current genetic status of the species. This study became possible through the extensive use of museum specimens to study populations now extinct. Phylogenetic analysis revealed the existence of two divergent mitochondrial lineages, lineage A occurring mainly in Western European populations and lineage B in African, Eastern European and Central Asian populations. The relative frequencies of haplotypes belonging to each lineage in the different populations show a steep East-West clinal distribution with maximal mixture of the two lineages in the Alps and Greece populations. A genealogical signature for population growth was found for lineage B, but not for lineage A; futhermore the Clade B haplotypes in western populations and clade A haplo-types in eastern populations are recently derived, as revealed by their peripheral location in median-joining haplotype networks. This phylogeographical pattern suggests allopatric differentiation of the two lineages in separate Mediterranean and African or Asian glacial refugia, followed by range expansion from the latter leading to two secondary contact suture zones in Central Europe and North Africa. High levels of among-population differentiation were observed, although these were not correlated with geographical distance. Due to the marked genetic structure, extinction of Central European populations in the last century re-sulted in the loss of a major portion of the genetic diversity of the species. We also found direct evidence for the effect of drift altering the genetic composition of the remnant Pyrenean population after the demographic bottleneck of the last century. Our results argue for the management of the species as a single population, given the apparent ecological exchangeability of extant stocks, and support the ongoing reintroduction of mixed ancestry birds in the Alps and planned reintroductions in Southern Spain.     

Haplotype diversity and phylogenetic relationships among the Iberian barbels (Barbus,Cyprinidae) reveal two evolutionary lineages

The phylogenetic relationships and haplotype diversity of all Iberian barbels were examined by analyzing the complete mitochondrial cytochrome b gene sequence (1141 bp) of 72 specimens from 59 Iberian localities. Phylogenetic findings demonstrated a clear distinction between two mitochondrial lineages and confirmed the existence of two previously considered subgenera: Barbus and Luciobarbus: The first subgenus, Barbus, is represented on the Iberian Peninsula by Barbus haasi and Barbus meridionalis. The second subgenus, Luciobarbus, includes the remaining endemic Iberian species: Barbus comizo, Barbus bocagei, Barbus microcephalus, Barbus sclateri, Barbus guiraonis, and Barbus graellsii. Mean haplotype divergence between these subgenera was 10.40%, providing evidence of a clear subdivision within the Iberian barbels. Our results conflict with those reported in a recent study, based on 307 cytochrome b base pairs, that failed to identify any division within the genus Barbus in the Iberian Peninsula. The inclusion of nine further species belonging to this genus (used as outgroups) allowed us to establish a closer relationship of the Iberian species of the subgenus Barbus with other European taxa than with the Iberian Luciobarbus, which was found to cluster with North African, Caucasian, and Greek species. At the population level, no biogeographic structure was shown by specimens of each species (only 5.98% of the variation was attributable to differences among populations of each species). Given the discrete amount of divergence found among the Luciobarbus species, the formation of current hydrographic basins during the Plio-Pleistocene seems to have played a major role in their isolation and evolution.     

Myosin light chain isoforms retain their species-specific electrophoretic mobility after processing, which enables differentiation between six species: 2DE analysis of minced meat and meat products made from beef, pork and poultry

Investigation of protein changes as well as authentication of meat is particularly difficult in processed meat products due to their different composition, complexity and very often inhomogeneity. The aim of this study was to check if the inter-species differences in the expression of myosin light chain (MLC) isoforms observed in raw meat were retained in meat products. MLCs from mixtures of minced meat (16 variants), frankfurters and sausages (15 products) made from cattle, pig, chicken, turkey, duck and goose were analysed by 2DE. Species-specific patterns of MLC isoforms were observed in all the mixtures and processed meat products. Relatively small degradation was observed in the MLCs after processing. Image analysis enabled species identification of the meat in all samples when the content of meat of one species was not lower than 10%. However, it was impossible to differentiate between all the six species under investigation on the basis of individual isoform. It was possible when the combination of all the three isoforms (myosin light chain 1 fast, myosin light chain 2 fast and myosin light chain 3 fast) was analysed. The results evidenced that MLCs have potential to be used as markers in authentication of meat products made from the analysed six species.     

Parvalbumin in mouse muscle in vivo and in vitro

Parvalbumin is a cytosolic calcium-binding protein found in adult fast-twitch mammalian muscle. Using an antibody to paravalbumin, we have shown that its distribution in adult mouse muscles is associated with certain fibre types. It is absent from slow-twitch type 1 fibres, is absent or at low levels in fast-twitch type 2A fibres, but is present at moderate or high levels in fast-twitch type 2B fibres. When adult mouse muscle is cultured with embryonic mouse spinal cord, the regenerated fibres become innervated, express the adult fast isoform of myosin heavy chain and appear histochemically as fast-twitch fibres. We therefore investigated whether these apparently mature fibres also contained parvalbumin. Parvalbumin was not found in any fibres of twenty mature cultures, suggesting that neurotrophic activity in the absence of specific adult nerve activity patterns was insufficient to cause the expression of parvalbumin in the cultures.     

Evolutional and Geographical Relationships of <Emphasis Type="Italic">Bartonella grahamii</Emphasis> Isolates from Wild Rodents by Multi-locus Sequencing Analysis

To clarify the relationship between Bartonella grahamii strains and both the rodent host species and the geographic location of the rodent habitat, we have investigated 31 B. grahamii strains from ten rodent host species from Asia (Japan and China), North America (Canada and the USA), and Europe (Russia and the UK). On the basis of multi-locus sequencing analysis of 16S rRNA, ftsZ, gltA, groEL, ribC, and rpoB, the strains were classified into two large groups, an Asian group and an American/European group. In addition, the strains examined were clearly clustered according to the geographic locations where the rodents had been captured. In the phylogenetic analysis based on gltA, the Japanese strains were divided into two subgroups: one close to strains from China, and the other related to strains from Far Eastern Russia. Thus, these observations suggest that the B. grahamii strains distributed in Japanese rodents originated from two different geographic regions. In the American/European group, B. grahamii from the North American continent showed an ancestral lineage and strict host specificity; by contrast, European strains showed low host specificity. The phylogenetic analysis and host specificity of B. grahamii raise the possibility that B. grahamii strains originating in the North American continent were distributed to European countries by adapting to various rodent hosts. An erratum to this article can be found at     

Allozyme Variation of the Pygmy Wood Mouse Sylvaemus uralensis (Rodentia,Muridae) and Estimation of the Divergence of Its Chromosome Forms

The genetic divergence between the eastern European, southern European, and Asian chromosome forms of the pygmy wood mouse Sylvaemus uralensis, whose karyotypes differ from one another in the amount of centromeric heterochromatin, has been reevaluated using allozyme analysis. In general, Asian chromosome forms in S. uralensis living in eastern Kazakhstan, eastern Turkmenistan (the Kugitang Ridge), and Uzbekistan are more monomorphic than European populations of this species. However, the allozyme differences between all chromosome forms of the pygmy wood mouse are comparable with the interpopulation differences within each form and are an order of magnitude smaller than those between good species of the genus Sylvaemus. Thus, the chromosome forms of S. uralensis cannot be considered to be separate species. The concept of races as large population groups that have not diverged enough to regard them as species but differ from one another in some genetic characters is used to describe the differentiation of S. uralensis forms more adequately. The currently available evidence suggests the existence of two S. uralensis races, the Asian and the European ones, and two chromosome forms (eastern and southern) of the European race. The possible historical factors that have determined the formation of the races of the pygmy wood mouse are considered. According to the most plausible hypothesis, the shift and fragmentation of the broad-leaved forest zone during the most recent glacial period (late Pleistocene) were the crucial factors of the formation of these races, because they resulted in a prolonged isolation of the European and Asian population groups ofS. uralensis from each other.     

Detection and Genetic Characterization of Relapsing Fever Spirochete Borrelia miyamotoi in Estonian Ticks

During the years 2008–2010 I. ricinus and I. persulcatus ticks were collected from 64 sites in mainland Estonia and on the island Saaremaa. Presence of B. miyamotoi was found in 0.9% (23/2622) of ticks. The prevalence in I. persulcatus and I. ricinus ticks differed significantly, 2.7% (15/561) and 0.4% (8/2061), respectively. The highest prevalence rates were in found South-Eastern Estonia in an area of I. persulcatus and I. ricinus sympatry and varied from 1.4% (1/73) to 2.8% (5/178). Co-infections with B. burgdorferi s.l. group spirochetes and tick-borne encephalitis virus were also revealed. Genetic characterization of partial 16S rRNA, p66 and glpQ genes demonstrated that Estonian sequences belong to two types of B. miyamotoi and cluster with sequences from Europe and the European part of Russia, as well as with sequences from Siberia, Asia and Japan, here designated as European and Asian types, respectively. Estonian sequences of the European type were obtained from I. ricinus ticks only, whereas the Asian type of B. miyamotoi was shown for both tick species in the sympatric regions.     

The susceptibility of European tree species to invasive Asian pathogens: a literature based analysis

Alien invasive pathogens have caused numerous disastrous epidemics around the globe during the last two centuries. The frequency of these catastrophes has increased in parallel with the increase of international plant trade. Effective control of the risks requires understanding of factors governing vulnerability of indigenous plants. We tested whether the threat caused by alien pathogens of Asian origin is random among various tree species in Europe or whether it relates to their distribution range. A database including distribution ranges of 75 European tree species and literature-derived information on their susceptibility to invasive forest pathogens (IFPs) of Asian origin was compiled. Analysis on this database indicated that the susceptibility to Asian pathogens is significantly more common among tree species that occur only within Europe than among species with distributional ranges from Europe to Siberia (disease susceptibility percentage, DSP, 52 and 19 %, respectively). Notably, all severely attacked tree species are strictly European while tree species with distribution ranges extending from Europe to Siberia show at most only mild or moderate symptoms of Asian IFPs. Furthermore, the proportion of European broadleaf tree species susceptible to Asian IFPs is significantly higher than that of conifer species. Our results suggest that in Europe, Asian pathogens cause a higher risk to temperate and Mediterranean forests, largely composed of broadleaved species with distributional ranges restricted to Europe, than to boreal forests dominated by conifers distributed to Siberia.     

Dual bioluminescent systems in the anglerfish genus Linophryne (Pisces: Ceratioidea)

Females of the anglerfish genus Linophryne bear barbels containing luminous organs, in addition to an escal light organ. Luminescence has been observed from the barbels of four species of Linophryne , and the morphology of the luminous organs investigated. The barbel light organs do not contain bacteria but complex paracrystalline photogenic granules. The esca contains luminous bacteria. The esca is ectodermal in origin whereas the barbel organs may be derived from the mesoderm.
The possible significance of this unique dual system of luminous organs is discussed.     

Monoclonal antibodies toward scallop (Patinopecten yessoensis) testis and wheat germ calmodulins.

A monoclonal antibody (IM7) toward scallop testis calmodulin and another one (PBE2) toward wheat germ calmodulin were produced. Ca2+ was required for IM7 to react with scallop calmodulin. IM7 reacted with the C-terminal region (Asp78-Lys148) of the calmodulin. As observed on competitive ELISA, IM7 reacted with chicken calmodulin, but not with Euglena gracilis or wheat calmodulin, troponin C, myosin light chains, or parvalbumin. It is assumed that the cluster of Thr143, Thr146, and Ser147 in the C-terminal region acts as the antigenic site. IM7 (and Fab of IM7) inhibited the activities of myosin light chain kinase and cAMP-phosphodiesterase. PBE2 reacted with wheat germ calmodulin irrespective of the presence or absence of Ca2+, the antigenic site being in the N-terminal region (Ala1-Met37). It reacted with wheat and spinach calmodulins, but not with scallop, chicken, or Euglena calmodulin, troponin C, myosin light chains, or parvalbumin. PBE2 had no effect on the activities of myosin light chain kinase and cAMP-phosphodiesterase.     

Expression of myofibrillar proteins and parvalbumin isoforms during the development of a flatfish,the common sole Solea solea: comparison with the turbot Scophthalmus maximus

Developmental changes in myofibrillar protein and parvalbumin isoform composition were investigated in the myotomal muscle of the flatfish Solea solea, characterized by a very brief metamorphic stage. Results were compared with previously obtained data on another pleuronectiform teleost, the turbot (Scophthalmus maximus), displaying prolonged metamorphosis. Electrophoretically measurable changes in myofibrillar proteins and parvalbumins were detected late in the sole, after completion of metamorphosis. In the course of development, muscles showed the usual sequential synthesis of isoforms of the myofibrillar proteins myosin light chain LC2, troponin-T, and troponin-I. An adult parvalbumin isoform (PA III) was found to predominate during sole growth. The two flatfish were characterized by highly species-specific parvalbumin isoforms. Compared with turbot, the profiles of the myofibrillar subunits and parvalbumin isoforms varied little in the course of sole development. The early appearance of adult traits might be correlated with the brevity of metamorphosis of this fish.     

Genetic divergence between Cyprinus carpio carpio and Cyprinus carpio haematopterus as assessed by mitochondrial DNA analysis,with emphasis on origin of European domestic carp

Although common carp is the major fish species in Asian and European aquaculture and many domestic varieties have occurred, there is a controversy about the origination of European domestic common carp. Some scientists affirmed that the ancestor of European domestic common carp was Danube River wild common carp, but others considered it might be Asian common carp. For elucidating origination of European domestic common carp, we chose two representative European domestic common carp strains (German mirror carp and Russian scattered scaled mirror carp) and one wild common carp strain of Cyprinus carpio carpio subspecies (Volga River wild common carp) and two Asian common carp strains, the Yangtze River wild common carp (Cyprinus carpio haematopterus) and traditionally domestic Xingguo red common carp, as experimental materials. ND5–ND6 and D-loop segments of mitochondrial DNA were amplified by polymerase chain reaction and analyzed through restriction fragment length polymorphism (RFLP) and sequencing respectively. The results revealed that HaeIII and DdeI digestion patterns of ND5–ND6 segment and sequences of control region were different between European subspecies C. carpio carpio and Asian subspecies C. carpio haematopterus. Phylogenetic analysis showed that German mirror carp and Russian scattered scaled mirror carp belonged to two subspecies, C. carpio carpio and C. carpio haematopterus, respectively. Therefore, there were different ancestors for domestic carp in Europe: German mirror carp was domesticated from European subspecies C. carpio carpio and Russian scattered scaled mirror carp originated from Asian subspecies C. carpio haematopterus.     

Re-visiting phylogenetic and taxonomic relationships in the genus saga (insecta: orthoptera)

Twelve of the 13 bushcricket species of the Saga genus are bisexuals and diploids, except the parthenogenetic and tetraploid bush cricket, Saga pedo. Despite a continuous research effort stretching through the 1900s, the taxonomic relationships of the Saga species are still disputed. In this study, our primary aim was to reveal natural relationships of the European Saga species and three of their Asian relatives, with special attention to the problematic taxonomy of two subspecies: S. campbelli campbelli and S. c. gracilis. Following a phylogenetic analysis of eight species, a comprehensive study was carried out on the above three taxa by using acoustic and morphometric approaches in parallel. Our phylogenetic data showed that European Saga species evolved from a monophyletic lineage. The geographical transitional species S. cappadocica was positioned between European and Asian lineages supporting the idea that the European Saga lineage originated phylogeographically from the Asian clade. The above results showed better agreement with the morphological data than with earlier ones based either on karyology or acoustic information only. After reviewing our data, we concluded that Saga pedo has most likely evolved from S. c. gracilis and not from S. rammei or S. ephippigera, as proposed by earlier studies. S. c. gracilis shares the same ITS2 haplotype with S. pedo, indicating that the latter could have evolved from populations of the former, probably through whole genome duplication. Based on acoustic and morphometric differences, we propose to elevate the two subspecies, S. campbelli campbelli and S. c. gracilis, to species level status, as Saga gracilis Kis 1962, and Saga campbelli Uvarov 1921. The present work sets the stage for future genetic and experimental investigations of Saginae and highlights the need for additional comprehensive analysis involving more Asian Saga species.