Trout (<Emphasis Type=Italic>Salmo trutta</Emphasis>) mitochondrial DNA polymorphism in the centre of the marble trout distribution area

The marble trout, a lineage of the Salmo trutta complex, is endemic to the Southern Alpine region. Although it is endangered throughout its entire distribution range, population genetic data were lacking for the central area, including the upper Etsch/Adige River system (South Tyrol, Northern Italy). A total of 672 Salmo trutta specimens, comprising phenotypic marble trout and phenotypic brown trout, from 20 sampling sites throughout South Tyrol were analysed by sequencing the complete mitochondrial DNA control region. Thirteen distinct haplotypes were identified, which clustered within three major genetic lineages: the Marmoratus (MA), the Atlantic (AT) and the Danubian (DA) lineage. 41.7% of the investigated individuals carried haplotypes of the MA lineage, 47.9% of the AT lineage and 10.4% of the DA lineage. It is noticeable that AT haplotypes were present at all sampling sites and no “pure” marble trout population with exclusively MA haplotypes was found. This points to a considerable impact of stocking with allochthonous brown trout, given that there is no evidence for natural colonisation by individuals of the AT lineage. However, our data indicate, for at least four localities, a limited gene flow between the native marble trout and hatchery-reared strains. Future conservation and rehabilitation measures will thus have to concentrate on the identification of remnant pure marble trout individuals from such mixed populations. Handling editor: C. Sturmbauer     

Assessing fish metrics and biotic indices in a Mediterranean stream: effects of uncertain native status of fish

Implementation of the Water Framework Directive requires tools for measuring and monitoring the ecological status of aquatic ecosystems. Several indices are in use in the Iberian Peninsula, although there has been little comparison among them. We sampled the fish assemblage and limnological features of the Tordera stream (NE Spain) quarterly from September 2001 to May 2003 to evaluate the usefulness of several fish metrics and to compare habitat quality and biotic indices currently in use. Data for eight biotic and abiotic indices for this and three other Catalan river basins were also compiled in order to analyse the relationships among indices. In the Tordera stream, fish abundance and richness increased with stream order except in the last sampling site that had the lowest fish abundance owing to the effects of drought and water abstraction. Although most indices were positively correlated, some displayed low or null correlations particularly for the Tordera basin which is more affected by water abstraction and less by pollution; a commonly used physico-chemical index (ISQA) was the least correlated. In a regional fish index (IBICAT) under development, the brown trout (Salmo trutta) has been previously considered as introduced in the Tordera basin. Here, we report an old published record that demonstrates that trout was present before 1845 and we argue that its status should be considered as uncertain given the current information available. Whether brown trout is treated as native or introduced to this river basin has profound effects on the results of fish metrics because of its dominance in the upper reaches. We briefly discuss the role of introduced species, particularly in headwater streams, in the development of fish indices. Our study exemplifies the need for careful, basin-specific assessment of native/introduced status in the development of fish metrics. Handling editor: K. Martens     

Detection of ‘long-haired’ Saprolegnia (S. parasitica) isolates using monoclonal antibodies

The ability of five monoclonal antibodies (Mabs) raised against a pathogenic Saprolegnia parasitica isolate from brown trout to detect and differentiate between isolates with bundles of long hairs (S. parasitica) and other Saprolegnia species was determined by means of an indirect immunofluorescence assay. Four of the Mabs used recognized some of the long-haired S. parasitica isolates but also cross-reacted with other Saprolegnia species without bundles of hairs and with Achlya sp. The other Mab (named 18A6) was able to differentiate between the asexual and most of the sexual isolates in the group of long-haired S. parasitica isolates, but did not recognize Achlya sp. or the Saprolegnia species without bundles of hairs, with the exception of S. hypogyna. These results indicate that isolates with bundles of long hairs are closely related with other members of genus Saprolegnia and share several antigens. However, Mab 18A6 seems to recognize an epitope that is expressed mainly in the asexual isolates in the long-haired S. parasitica isolates.     

Managing fish populations under mosaic relationships. The case of brown trout (Salmo trutta) in peripheral Mediterranean populations

Brown trout were collected from 36 locations inthe Iberian Peninsula representing thesouthwestern extreme of this species'distribution in Mediterranean drainages.Allelic distributions among these peripheralpopulations for 26 polymorphic allozyme loci(corrected to remove effects of introgressionfrom exogenous hatchery introductions)indicated a mosaic pattern both within andamong drainages. This distribution wasinterpreted to reflect reticular evolutionaryprocesses involving multiple colonizationepisodes by distinct lineages, secondaryintergradations, and drift. Under thisscenario, management to conserve native genepools differs from that commonly used underhierarchical structures (such as in AtlanticIberian drainages) where geographic patternscould justify regional stocking of indigenousfish in adjacent unsampled areas. Because suchaction risks erosion of native populations inthe present case, remedial efforts should focuson habitat recovery and environmental educationprograms, and harvest sustained by naturalreproduction.     

A large scale survey of Populus nigra presence and genetic introgression from non-native poplars in Switzerland based on molecular identification

Populus nigra is considered a rare and threatened tree species in Switzerland because of dramatic habitat loss owing to river regulations during the last two centuries and because of potential gene introgression from non-indigenous P. deltoides through planted P. x canadensis hybrids. The significance of introgression as an endangerment to P. nigra, however, is controversial. The aims of the present study were (1) to assess how abundant P. nigra trees are in Switzerland and (2) to assess potential gene introgression. We present data from a molecular survey of 1372 putative P. nigra trees from Switzerland, using both chloroplast DNA and nuclear DNA markers. The results show that P. nigra is more abundant in Switzerland than hitherto thought. Furthermore, we detected a low frequency of gene introgression.     

The Plasmodium parasite—a ‘new’ challenge for insect innate immunity

Though lacking adaptive immunity, insects possess a powerful innate immune system, a genome-encoded defence machinery used to confront infections. Studies in the fruit fly Drosophila melanogaster revealed a remarkable capacity of the innate immune system to differentiate between and subsequently respond to different bacteria and fungi. However, hematophagous compared to non-hematophagous insects encounter additional blood-borne infectious agents, such as parasites and viruses, during their lifetime. Anopheles mosquitoes become infected with the malaria parasite Plasmodium during feeding on infected human hosts and may then transmit the parasite to new hosts during subsequent bites. Whether Anopheles has developed mechanisms to confront these infections is the subject of this review. Initially, we review our current understanding of innate immune reactions and give an overview of the Anopheles immune system as revealed through comparative genomic analyses. Then, we examine and discuss the capacity of mosquitoes to recognize and respond to infections, especially to Plasmodium, and finally, we explore approaches to investigate and potentially utilize the vector immune competence to prevent pathogen transmission. Such approaches constitute a new challenge for insect immunity research, a challenge for global health.     

Special traits of biology and of the parasite fauna of juvenile salmonid fish of the genus <Emphasis Type="Italic">Salmo</Emphasis> of the Tornio River system (The Baltic Sea basin)

Juveniles of Atlantic salmon Salmo salar and trout S. trutta populating the system of the Tornio River (the Baltic Sea basin) are investigated. The obtained parasitological data indicate the absence of rigid spatial and food competition between juvenile salmon and trout in the case of their cohabitation.     

Reading the history of a hybrid fish complex from its molecular record

Squalius alburnoides is a widely distributed intergeneric hybrid complex with fish of both sexes, varying ploidy levels and proportions of the parental genomes. Its dispersal routes were here delineated and framed by the reconstruction of the phylogeny and phylogeography of other Squalius with which it hybridizes, based on the available data on the paleohydrographical history of the Iberian Peninsula. Results based on sequences of cytochrome b and beta-actin genes showed that: proto-Squalius pyrenaicus originated at least five species as it dispersed throughout the Iberian Peninsula in the Mio-Pliocene; the S. alburnoides complex likely had a single origin in the bulk of Iberia, in the Upper Tagus/Guadiana area, when hydrographical rearrangements allowed the contact between its ancestors (around 700,000 years ago); interspecific crosses allowed the introgression of mitochondrial and nuclear genes of S. alburnoides in allopatric species/populations of other Squalius and vice-versa; and reconstituted S. alburnoides non-hybrid males may contribute to the replacement of the typical mtDNA of the complex (in the populations where they occur, crosses with females of other Squalius seem to have been especially frequent). A number of dispersal events and colonization routes are proposed.     

A ‘phase-shift’ fusion system for the regulation of foreign gene expression by lambda represser in Gram-negative bacteria

A ‘phase-shift’ translation fusion vector was constructed in which mutually compatible restriction sites BamHI, BelI and BglII are positioned in such a manner that the cut point is in a different reading frame, immediately following the ATG start codon and ribosome-binding site of the λ cro gene. The λ cro gene is expressed from promoter p_R and controlled by a thermosensitive (cI857) λ repressor. The usefulness of the expression vector was demonstrated using a gal gene lacking the ATG start codon and fusing this to the p_R promoter and ATG start codon of the λ cro gene, resulting in cI857-regulated expression of galactokinase. The vector is of general use for foreign gene expression in Escherichia coli when the target gene has a compatible cohesive end (5′-GATC-3′) at the N terminus (provided, for example, by a BamHI linker). The A λ cI857-p_R-cro-galK cassette was cloned into pJRD215, a wide-host-range plasmid and transferred by conjugation to a variety of Gram-negative bacteria. In all cases, thermosensitive regulation of galactokinase could be demonstrated, though the levels of induction varied considerably. These results show that the powerful λ p_R promoter and the efficient A repressor can be used to regulate expression of foreign genes in Gram-negative organisms other than E. coli.     

The ‘philosophical grasp of the appearances’ and experimental microscopy: Johannes Müller’s microscopical research, 1824–1832

Romantic Naturphilosophie has been at the centre of almost every account of early nineteenth-century sciences, be it as an obstacle or as an aid for scientific advancement. The following paper suggests a change of perspective. I seek to read Naturphilosophie as one manifestation among others of a more general concern with the question of how experience enables the subject to acquire knowledge about objects. To illustrate such an approach, I focus on Johannes Müller’s early work. Here one finds two contrasting images of microscopical observation, its set-up, and the observer: the embryological study of 1830 demands a ‘philosophical grasp’ of the appearances. In contrast, the investigations of blood of 1832 are presented as a series of controlled experiments. I argue that an interpretation of this contrast in terms of an appropriation and casting aside of Naturphilosophie is not altogether convincing. Instead, both images of microscopy are manifestations of a more general problem, namely, the problem of exactly how subject and object came together in experience. I show how this concern not only shaped the methodological sensibilities particular to Müller’s embryology and the investigation of bodily liquids but also provided the epistemological principles and the target for his sense-physiological experiments. It bound Müller’s work together with Naturphilosophie and linked Naturphilosophie with other contemporaneous projects in philosophy. All of these enterprises sought to contribute to ongoing debates about how experience allowed the subject to acquire knowledge about the world.     

Genetic structure of brown trout (<Emphasis Type="Italic">Salmo trutta</Emphasis> L.) from the Hardangervidda mountain plateau (Norway) analyzed by microsatellite DNA: a basis for conservation guidelines

The Hardangervidda in southern Norway, the largest mountain plateau in Europe, has thousands of lakes and streams, mainly between 1000 and 1300 m above sea level, where brown trout is the only fish species. To describe the current genetic diversity of brown trout in this area, a total of 863 fish from 20 lakes were genotyped with eleven microsatellites. Most diversity is within lake populations, but diversity among geographical groups and populations within groups was significant, too. Neighbor-joining, principle coordinate analysis and Bayesian clustering show three major geographic groups in accordance with the river systems. Bias was caused by recent stocking in two lakes. Low/no genetic differentiation among some populations indicates that intermixing is common when lakes are well-connected, as was also shown by assignment test. We recommend preserving the genetic diversity of brown trout in this unique area by managing stocking in lake systems according to genetic structure.     

Genetic characterization of brown trout (<Emphasis Type="Italic">Salmo trutta</Emphasis>) populations from the Southern Balkans using mtDNA sequencing and RFLP analysis

In the present study, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay was used to survey variation in a 1450-bp mitochondrial DNA (mtDNA) segment which comprises part of the cytochrome oxidase III (COIII) and ATPase subunit VI genes in 8 brown trout (Salmo trutta L) populations from the southern Balkans. In addition, a 300 bp fragment at the 5′ end of the control region was sequenced from representatives of the populations studied providing the opportunity to assign PCR-RFLP haplotypes into major phylogenetic lineages (i.e. Atlantic, Danubian, marmoratus, Adriatic and Mediterranean). The level of polymorphism found in the 1450 bp segment suggests that this PCR-RFLP assay may be useful for future diagnostic analyses of mitochondrial DNA in brown trout populations. A reduced within-population genetic variability but considerable among-population differentiation was observed. The results are in accordance with previous data on phylogeography of Mediterranean brown trout suggesting that mitochondrial DNA haplotypes are distributed in a mosaic pattern as a consequence of a complex evolutionary history. The present study shows that brown trout populations from the Southern Balkans are highly divergent and possess a unique genetic profile that should be taken into account when establishing conservation management programs. Handling editor: C. Sturmbauer     

The genetic structure of endangered indigenous populations of the amago salmon, <Emphasis Type="Italic">Oncorhynchus</Emphasis><Emphasis Type="Italic">masou</Emphasis><Emphasis Type="Italic">ishikawae</Emphasis>, in Japan

The amago salmon, Oncorhynchus masou ishikawae, is an endemic subspecies of O. masou in Japan. Owing to the extensive stocking of hatchery fish throughout Japan, indigenous populations of O. m. ishikawae are now on the verge of extinction. We examined the genetic effects of stocking hatchery fish on wild populations in the River Koza, Japan, using microsatellite and mitochondrial DNA (mtDNA) markers. For mtDNA, haplotype mt1, which is common in wild populations, was present exclusively in isolated wild populations assumed to be unaffected by previous stocking, while it was never observed in hatchery fish. Genetic diversity was much higher in wild populations in the stocked area, which shared many mtDNA haplotypes with hatchery fish, than in isolated wild populations with haplotype mt1. Pairwise F _ST estimates based on microsatellites showed significant differentiation among the isolated populations with many microsatellite loci monomorphic. Significant deviation from Hardy–Weinberg equilibrium was observed in wild populations in the area subject to stocking, where a Bayesian-based assignment test showed a high level of introgression with hatchery fish. These results suggest that wild populations with haplotype mt1, which became isolated through anthropogenic environmental change in the 1950–1960s, represent indigenous populations of O. m. ishikawae in the River Koza. They have low genetic diversity, most likely caused by genetic bottlenecks following damming and environmental deterioration, while stocking of hatchery fish over the past 30 years apparently had a large impact on the genetic structure of wild populations in the main channel of the River Koza.     

The ‘home advantage’ is necessary for a full winner effect and changes in post-encounter testosterone

Winning aggressive contests can both enhance future winning ability and change post-encounter hormones; however, it remains unclear if the context of a fight also influences such winner effects and hormone changes. We investigated this issue by using California mice (Peromyscus californicus) to test if the effect of residency status is necessary to improve future winning ability and alter post-encounter hormones. Male mice were subjected to an aggressive contest and their blood was collected 45 min after the fight. Upon contest initiation, focal mice had a ‘home advantage’ and three prior winning experiences, only one of these factors, or neither factor. Only individuals with a ‘home advantage’ and prior winning experience showed a full winner effect. Post-encounter changes in testosterone and progesterone resulted from an interaction between residency status and winning experience. These data indicate that in male California mice a ‘home advantage’ is necessary to produce the full winner effect and that residency status helps regulate inter-individual variation in the expression of post-encounter testosterone pulses and progesterone changes. Furthermore, these respective behavioral and physiological phenomena might be modulated in a context-specific manner, in part by the surrounding physical environment.     

The elevated anthocyanin level in the shaded peel of ‘Anjou’ pear enhances its tolerance to high temperature under high light

Pigments, chlorophyll fluorescence, dark respiration, and the antioxidant system in the shaded peel of green ‘Anjou’ pear (Pyrus communis L.) and its bud mutation, red ‘Anjou’, were compared in response to high peel temperature, high light alone or in combination to determine the protective role of anthocyanins under high temperature with or without light. Under high temperature treatment alone, no difference in the maximum quantum yield of PSII (F_V/F_M) was detected between red ‘Anjou’ and green ‘Anjou’; the superoxide dismutase activity and the glutathione pool were up-regulated in green ‘Anjou’ peel but remained unchanged in red ‘Anjou’ peel. Under high temperature coupled with high light, the F_V/F_M of green ‘Anjou’ peel was decreased to a lower value than that of red ‘Anjou’, and significant interaction was detected between temperature and light for both cultivars. Furthermore, the difference in F_V/F_M between red ‘Anjou’ and green ‘Anjou’ under high temperature coupled with high light was significantly larger than that under high light alone, indicating that this larger difference was caused by the interaction between high temperature and high light as no significant difference was detected in F_V/F_M between the two cultivars under high temperature treatment alone at any sampling point. It is concluded that the elevated anthocyanin level in the shaded peel of red ‘Anjou’ does not alter its thermotolerance in the dark, but makes it more tolerant of high temperature under high light.     

A shift in the habitat use pattern of a lentic galaxiid fish: an acute behavioural response to an introduced predator

Despite potentially reducing predation mortality, behavioural responses of native species to introduced predators may still have sub-lethal impacts. In video-recorded laboratory trials, we examined the effects of introduced brown trout, Salmo trutta, on the short-term behaviour of a threatened, lake-dwelling galaxiid fish and confirmed a suspected diel pattern in habitat use by this species. We found that Galaxias auratus followed a distinct diel pattern in the use of complex habitats and open water, which was significantly altered by the presence of brown trout. In trials without the introduced predator, G. auratus used complex habitats (rocks or macrophytes) during the day, and open water during the night. In trials with brown trout present, G. auratus spent significantly less time in open water and rarely ventured out of the macrophytes. However, when given the option of using only rocky substrate or open water, which is the more common situation in the lakes to which this galaxiid is endemic, the fish reduced the amount of time they spent in the open water during the night, but still spent more time in open water than when macrophytes were available. Spending the daylight hours amongst the cover of rocks or macrophytes is most likely an adaptation to reduce the risk of predation by visual predators, and the pattern of reduced use of open water habitats in the presence of brown trout is an acute response to the close proximity of the introduced predator. The difference in the nocturnal use of macrophytes and rocks when trout are present may be related to differences in feeding opportunities or success within these habitats.     

Mitochondrial DNA sequence variability in red-legged partridge, <Emphasis Type="Italic">Alectoris rufa</Emphasis>, Spanish populations and the origins of genetic contamination from <Emphasis Type="Italic">A. chukar</Emphasis>

The analysis of 135 mitochondrial D-loop sequences of the Iberian autochthonous red-legged partridge (Alectoris rufa) from wild population hunting bags from various locations and fowl runs in Spain yielded 37 different haplotypes. Among these, three haplotypes correspond to chukar partridges (Alectoris chukar), indicating genetic introgression from birds illegally introduced for restocking: three individuals carrying such haplotypes where found in natural populations, one appeared among those sampled on a mass reproduction farm and the remaining 10 in another fowl-run. The geographical origin of the contaminating chukar haplotypes could be assigned to the most easterly area of the chukar partridge geographical distribution in China. Molecular diversity parameters in the A. rufa samples indicate a considerable amount of genetic variation. Φ_ST showed significant differences among populations that are not explained by geographical distance alone. Particularly, one northern population (Palencia) shows a certain degree of genetic differentiation that could reflect a previously suggested subspecies division. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.