An improved protocol for the production of AFLPTM markers in complex genomes by means of capillary electrophoresis

The amplified fragment‐length polymorphism (AFLP) technology is a recently introduced method to investigate genomes of different complexity, from microbial to higher organisms. It is applied to purposes as diverse as identification of species, strain and varieties, investigation of genetic diversity within and between populations, simple and complex trait mapping, and construction of linkage and physical maps. This technology has been designed on the use of primers labelled with radioactivity and on AFLP fragment separation on sequencing gel. We show that the original EcoRI/TaqI AFLP protocol does not perform appropriately when transferred to fluorescent labelling and capillary electrophoresis (CE), and propose an improved protocol for the production of high‐quality AFLP markers in fish, rodents and artiodactyles by means of the Beckman‐Coulter CEQ2000 automatic DNA sequencer. In addition, we describe the procedure routinely used in our laboratory to obtain binary matrices from AFLP profiles with the aid of Genographer free‐share software (vers. 1.6.0, J.J. Benham, Montana State University), able to elaborate original fragment data and convert them to standard graphical formats for phylogenetic analyses. Comparison with radioactive AFLPs in goats confirmed the reliability of the protocol developed for CE. In fact, 107 fragments generated by two primer combinations and identified by both techniques were attributed the same scoring. Compared with traditional methods, the use of capillary systems and automated analysis increases data throughput and scoring reliability, decreasing the overall experimental error.     

Weak effects of habitat type on susceptibility to invasive freshwater species: an Italian case study

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Pharmacokinetics of marbofloxacin after a single oral dose to loggerhead sea turtles (Caretta caretta)

The single-dose disposition kinetics of marbofloxacin (MBX) were determined in clinically healthy loggerhead sea turtles (n = 5) after oral (PO) administration of 2 mg kg⁻¹ bodyweight. Marbofloxacin plasma concentrations were determined by DAD–HPLC (LOD/LOQ 0.015/0.05 μg ml⁻¹). Data were subjected to non-compartmental analysis. Following PO administration, marbofloxacin achieved maximum plasma concentrations of 11.66 ± 2.53 mg L⁻¹ at 15.00 ± 3.00 h. The absence of general adverse reactions in the turtles of the study, and the favourable pharmacokinetic properties (long half-life and high maximum plasma concentration) of MBX administered PO at the single-dose of 2 mg kg⁻¹ suggest the possibility of its safe and effective clinical use in loggerhead sea turtles.     

Volatile Organic Compounds,Indole, and Biogenic Amines Assessment in Two Mediterranean Irciniidae (Porifera,Demospongiae)

Solid phase microextraction (SPME) coupled to gas chromatography-mass spectrometry (GC-MS) was employed for the headspace determination of the volatile organic fraction emitted by two of the most common Mediterranean demosponges, Ircinia variabilis and Sarcotragus spinosulus, and of indole and some biogenic amines released by sponges in an aqueous medium. A total of 50/30 µm divinylbenzene/carboxen/polydimethylsiloxane and 75 µm carboxen/polydimethylsiloxane fibers were used for the headspace extraction of low molecular weight sulfur compounds from a hermetically sealed vial containing sponge fragments, while the direct immersion determination of indole and biogenic amines was performed. The biogenic amines were extracted after in-solution derivatization with isobutyl chloroformate. All analytical parameters (linearity, limits of detection, and quantification, precision, and recovery) were evaluated for indole and biogenic amines. SPME-GC-MS proved to be a reliable means of highlighting the differences between molecules released by different sponges, principally responsible for their smell. The combined approaches allowed the identification of several volatile compounds in the headspace and other molecules released by the sponges in an aqueous medium, including indole and the BAs cadaverine, histamine, isobutylamine, isopentylamine, propylamine, 2-phenylethylamine, putrescine and tryptamine. The results obtained represent a further contribution to the picture of odoriferous molecules secreted by sponges.     

The influence of land abandonment on forest disturbance regimes: a global review

Landscape Ecology - Since the nineteenth century, rural areas have experienced progressive abandonment mostly due to socioeconomic changes, with direct and indirect effects on forest disturbance...     

Application of amplified fragment length polymorphism markers to assess molecular polymorphisms in gynogenetic haploid embryos of turbot (Scophthalmus maximus)

Among the variety of cultured marine species, the turbot Scophthalmus maximus is a fish of growing importance in European aquaculture. In this paper, an advanced application of AFLPs to estimate the genetic diversity of haploid gynogenetic families with the aim of obtaining a preliminary genetic map is presented. Ten EcoRI/TaqI primer combinations were tested in four families comprising diploid mothers and their haploid progenies. The amplified fragment length polymorphism (AFLP) analysis revealed an average of 6.8 polymorphic bands per primer combination and a total number of 88 polymorphisms out of 579 fragments. Among various primer pairs, seven combinations were selected in relation to the quality of profiles and number of polymorphic fragments, to be used in the determination of genetic linkage relationship between AFLP markers within the largest haploid family. Co‐migration of non‐homologous fragments was also investigated in one primer combination adding a fourth selective nucleotide to the three used in the classic TaqI AFLP protocol. Surprisingly, a rate of 38.7% of non‐homologous fragments co‐migrating with monomorphic bands was identified, due to the combined effect of homoplasy and the protocol used. Additional polymorphic markers discovered by this protocol were included in the linkage map. The turbot AFLP linkage map comprises 52 AFLP markers distributed in 12 linkage groups. On the basis of this map, turbot expected total genome length sums up to 1225.6 cM. The results confirm the usefulness of AFLPs in revealing genome segregation in haploid turbot progeny.     

Managing European cormorant‐fisheries conflicts: problems,practicalities and policy

This article focuses on transboundary conflicts between great cormorants, Phalacrocorax carbo L., and European fisheries interests. Two races of this species commonly occur in Europe; the ‘Atlantic’ Phalacrocorax carbo carbo and the ‘Continental’ P. c. sinensis. This migratory fish‐eating bird has steadily expanded its European range and many consider this a nature conservation success. However, fishery representatives see the birds as harmful to their business and way of life, considering European Union (EU) legislation to be blocking the most intuitive and effective solution to the cormorant problem, namely co‐ordinated, pan‐European population reduction. This article argues that cormorant‐fisheries conflicts and their resolution and management cannot be characterised as, or reduced to, a simplistic ‘pan‐European’ vs ‘local’ argument. The nature of cormorant‐fisheries conflicts is explored, key issues of cormorant ecology examined, and consideration is given to how impacts on fisheries might be measured. Discussions then focus on whether current policy prevents balanced and constructive solutions or whether better use of available instruments at local‐national levels could actually be a vital step to a sustainable management solution.     

Is the private forest sector adapting to climate change? A study of forest managers in north Wales

? Context

Two-thirds of Britain’s forest area is privately owned. Thus, understanding private forest owners and managers, and their attitudes to uncertainty and change, is essential for the success of climate change adaptation policies.

? Aim

The aims of this study are to (1) assess how beliefs in climate change in the private sector have influenced forest management practices; (2) identify constraints related to changes in species choice and silvicultural systems; (3) analyse the implications for implementing climate change policy in forestry.

? Method

Semi-structured interviews with key informants who provide advice to, or manage woodlands in, the private forest sector in north Wales.

? Results

Woodland managers and some advisers are not generally convinced of a need to adapt. They feel the future is uncertain, more usually in relation to tree disease than to climate change itself. Species choice is the principle focus of adaptation activities and reveals a deep divide in opinion. Commercial advisors look to new exotics but are inhibited by absence of markets, while small-scale owners rely more on native genetic diversity.

? Conclusions

Findings that are likely to apply widely include: the influential role of forest agents in forest management decisions including species choice; lack of confidence in climate change predictions, and in markets; more immediate concerns about tree pests and diseases; demand for leadership from the public sector, and for engagement amongst the private sector. Further research is needed across a wider area to test the variability in relationship between attitudes and behaviours, and local conditions including climate change predictions.     

Inheritance of seed coat colour in broad bean (Vicia faba L.)

Summary The inheritance of the seed coat colours violet, spotted, brown, green, red, black and beige was investigated in a 10×10 diallel cross between broad bean (Vicia faba L. major) lines. Spotted seed colour was dominant over any uniform seed coat colouring. Brown was dominant over black, green and normal (beige colour). Black and red seed parents behaved as recessive in all F₁ progenies. A 3 (coloured): 1 (normal) segregation ratio was observed in the F₂ of crosses of violet, brown, black, red and spotted seed coat parents to nornal seed coloured parents. Green x beige gave a segregation ratio of 9:7 in F₂. When two parents with different seed coat colour were involved in a cross, the F₂ showed a typical digenic segregation ratio thus demonstrating two unlinked and sometimes epistatic loci.Segregation of a multiallelic series at two loci explains all segregation ratios observed for seed coat colour in broad bean.