Vicariance patterns in the Mediterranean Sea: east–west cleavage and low dispersal in the endemic seagrass Posidonia oceanica

Aim The seagrass, Posidonia oceanica is a clonal angiosperm endemic to the Mediterranean Sea. Previous studies have suggested that clonal growth is far greater than sexual recruitment and thus leads to low clonal diversity within meadows. However, recently developed microsatellite markers indicate that there are many different genotypes, and therefore many distinct clones present. The low resolution of markers used in the past limited our ability to estimate clonality and assess the individual level. New high‐resolution dinucleotide microsatellites now allow genetically distinct individuals to be identified, enabling more reliable estimation of population genetic parameters across the Mediterranean Basin. We investigated the biogeography and dispersal of P. oceanica at various spatial scales in order to assess the influence of different evolutionary factors shaping the distribution of genetic diversity in this species. Location The Mediterranean. Methods We used seven hypervariable microsatellite markers, in addition to the five previously existing markers, to describe the spatial distribution of genetic variability in 34 meadows spread throughout the Mediterranean, on the basis of an average of 35.6 (± 6.3) ramets sampled. Results At the scale of the Mediterranean Sea as a whole, a strong east–west cleavage was detected (amova) . These results are in line with those obtained using previous markers. The new results showed the presence of a putative secondary contact zone at the Siculo‐Tunisian Strait, which exhibited high allelic richness and shared alleles absent from the eastern and western basins. F statistics (pairwise θ ranges between 0.09 and 0.71) revealed high genetic structure between meadows, both at a small scale (about 2 to 200 km) and at a medium scale within the eastern and western basins, independent of geographical distance. At the intrameadow scale, significant spatial autocorrelation in six out of 15 locations revealed that dispersal can be restricted to the scale of a few metres. Main conclusions A stochastic pattern of effective migration due to low population size, turnover and seed survival is the most likely explanation for this pattern of highly restricted gene flow, despite the importance of an a priori seed dispersal potential. The east–west cleavage probably represents the outline of vicariance caused by the last Pleistocene ice age and maintained to this day by low gene flow. These results emphasize the diversity of evolutionary processes shaping the genetic structure at different spatial scales.     

Hypocotyl growth of Pinus pinaster seedlings. Changes in the molecular weight distribution of hemicellulosic polysaccharides

Lorences, E. P., Suárez, L. and Zarra, I. 1987. Hypocotyl growth of Pinus pinaster seedlings. Changes in the molecular weight distribution of hemicellulosic polysaccharides.
The changes in the molecular weight distribution of water-soluble hemicelluloses and xyloglucan during hypocotyl growth of intact seedlings of Pinus pinaster Aiton were investigated. The mass-average molecular weight of total polysaccharides of the hemicellulose fraction soluble in 4% KOH dramatically increased during hypocotyl growth while xyloglucan slightly decreased. These phenomena were due to an increase in the degree of polymerization of an arabinogalactan and a slight depolymer-ization in the xyloglucan present in this fraction. In the hemicellulose fraction soluble in 24% KOH, xyloglucan increased its degree of polymerization from day 7 to 10 after which it decreased slightly. The xyloglucan of the hemicellulose fraction soluble in 4% KOH may thus be involved in cell wall loosening which makes cell wall expansion possible during hypocotyl growth.     

Presence of a drifting algal bed in the Mar Piccolo basin,Taranto (Ionian Sea,Southern Italy)

A survey is reported of the drifting algal community in Mar Piccolo, a polluted basin subject to sewage outlets. The key role was played by a few key species, mainly floridean red algae.     

The localization and rate of disappearance of a synaptic vesicle antigen following denervation

Summary A proteoglycan-specific antiserum has been used to monitor the effects of denervation in the electric organ of Torpedo marmorata. The antiserum was produced by injecting a highly purified synaptic vesicle fraction prepared from the electric organs of Torpedo marmorata. Following absorption the serum appears to be specific towards synaptic vesicles. The ultrastructural localization of the antigen determined by immuno-electron microscopy confirmed the specificity of the antiserum and showed that it did not crossreact with the proteoglycans of the basal lamina. The rate of disappearance of the vesicle proteoglycans following denervation was evaluated by means of the antiserum and was compared to the rate of disappearance of other vesicular and nerve terminal-associated markers. The results suggest that degeneration affects the vesicular constituents at varying rates resulting in a progressive disappearance of the entire functional capacity of the synaptic vesicles.     

An immunohistochemical study of synaptogenesis in the electric organ of Torpedo marmorata by use of antisera to vesicular and presynaptic plasma membrane components

Summary Synaptogenesis has been studied in the electric organ of embryonic Torpedo marmorata by use of two antisera directed against components of synaptic vesicles (anti-SV) and presynaptic plasma membranes (ap-anti-TSM), respectively. The anti-SV serum was previously shown to recognize a proteoglycan specific for synaptic vesicles. The ap-anti-TSM serum was raised to plasma membranes of synaptosomes derived from the electromotor nerve terminals and affinity-purified on electric-organ gangliosides. The vesicular antigen was first detectable at the 81-mm stage of development, which is 1–2 weeks earlier than the formation of morphologically mature presynaptic terminals, but is coincident with a rise in choline acetyltransferase levels and the ability of the electric organ to generate discharges. The gangliosidic antigen recognized by the ap-anti-TSM was first detectable on the ventral electrocyte surface at the 93-mm stage of development. This indicates that specific carbohydrate epitopes, not present on the growth cones, are expressed during maturation of the nerve terminal. The nerve terminal components recognized by these sera arose pari passu with neurite coverage of the ventral surface of the electrocyte, reaching a maximum in the adult. In contrast, postsynaptic aggregates of acetylcholine receptor, rendered visible with rhodamine-labeled -bungarotoxin, arose previous to the presynaptic antigens, reaching a maximum surface density at 110 mm and then declining in the adult.     

Evidence of the existence of two different intraneuronal pools from which pharmacological agents can release serotonin

The effect of various drugs on the release of [³H]-serotonin from synaptosomes of reserpine-treated rats was compared with that obtained with synaptosomes of untreated animals.The increase in [³H]-serotonin release induced by d-fenfluramine was virtually abolished by reserpine; the effect of d-norfenfluramine, the main metabolite of fenfluramine, was instead enhanced in synaptosomes of reserpine treated animals. [³H]-serotonin release induced by l-isomers of fenfluramine or norfenfluramine was increased or not affected, respectively, after reserpine treatment.The effects of other drugs, known to activate serotonin mechanisms such as metachlorophenylpiperazine and quipazine, like d-norfenfluramine, were increased by the reserpine treatment.The present data show that [³H]-serotonin can be released by drugs from two pools with different sensitivity to reserpine. The reserpinized synaptosomes could provide useful information on the mechanisms of action of drugs acting on brain serotonin.     

A simplified microassay for serotonin: Modification of the enzymatic isotopic assay

A simplification of the enzymatic isotopic assay for serotonin is described, Serotonin is converted to [³H]melatonin by a two-step reaction: N-acetylation of serotonin using acetic anhydride, followed by O-methylation with the enzyme hydroxyindole O-methyltransferase (EC 2.1,1.4) and S-adenosyl- -[methyl-³H]methionine as methyl donor. The present assay avoids the use of unstable acetylating enzyme, rat liver N-acetyltransferase (EC2.3,1.5). Blank values are lowered considerably and the sensitivity is doubly increased. Two-tenths micromole of serotonin per 30 μl of sample in tissue homogenates can be measured.     

Measurement of Outflow Facility Using iPerfusion

Elevated intraocular pressure (IOP) is the predominant risk factor for glaucoma, and reducing IOP is the only successful strategy to prevent further glaucomatous vision loss. IOP is determined by the balance between the rates of aqueous humour secretion and outflow, and a pathological reduction in the hydraulic conductance of outflow, known as outflow facility, is responsible for IOP elevation in glaucoma. Mouse models are often used to investigate the mechanisms controlling outflow facility, but the diminutive size of the mouse eye makes measurement of outflow technically challenging. In this study, we present a new approach to measure and analyse outflow facility using iPerfusion^™, which incorporates an actuated pressure reservoir, thermal flow sensor, differential pressure measurement and an automated computerised interface. In enucleated eyes from C57BL/6J mice, the flow-pressure relationship is highly non-linear and is well represented by an empirical power law model that describes the pressure dependence of outflow facility. At zero pressure, the measured flow is indistinguishable from zero, confirming the absence of any significant pressure independent flow in enucleated eyes. Comparison with the commonly used 2-parameter linear outflow model reveals that inappropriate application of a linear fit to a non-linear flow-pressure relationship introduces considerable errors in the estimation of outflow facility and leads to the false impression of pressure-independent outflow. Data from a population of enucleated eyes from C57BL/6J mice show that outflow facility is best described by a lognormal distribution, with 6-fold variability between individuals, but with relatively tight correlation of facility between fellow eyes. iPerfusion represents a platform technology to accurately and robustly characterise the flow-pressure relationship in enucleated mouse eyes for the purpose of glaucoma research and with minor modifications, may be applied in vivo to mice, as well as to eyes from other species or different biofluidic systems.     

Viral SPP1 DNA is infectious in naturally competent Bacillus subtilis cells: inter- and intramolecular recombination pathways

A proteolyzed bacteriophage (phage) might release its DNA into the environment. Here, we define the recombination functions required to resurrect an infective lytic phage from inactive environmental viral DNA in naturally competent Bacillus subtilis cells. Using phage SPP1 DNA, a model that accounts for the obtained data is proposed (i) the DNA uptake apparatus takes up environmental SPP1 DNA, fragments it, and incorporates into the cytosol different linear single-stranded (ss) DNA molecules shorter than genome-length; (ii) the SsbA-DprA mediator loads RecA onto any fragmented linear SPP1 ssDNA, but negative modulators (RecX and RecU) promote a net RecA disassembly from these ssDNAs not homologous to the host genome; (iii) single strand annealing (SSA) proteins, DprA and RecO, anneal the SsbA- or SsbB-coated complementary strands, yielding tailed SPP1 duplex intermediates; (iv) RecA polymerized on these tailed intermediates invades a homologous region in another incomplete molecule, and in concert with RecD2 helicase, reconstitutes a complete linear phage genome with redundant regions at the ends of the molecule; and (v) DprA, RecO or viral G35P SSA, may catalyze the annealing of these terminally redundant regions, alone or with the help of an exonuclease, to produce a circular unit-length duplex viral genome ready to initiate replication.     

Characterization of PfiT/PfiA toxin–antitoxin system of Pseudomonas aeruginosa that affects cell elongation and prophage induction

Toxin–antitoxin (TA) systems are small genetic modules usually consisting of two elements—a toxin and an antitoxin. The abundance of TA systems among various bacterial strains may indicate an important evolutionary role. Pseudomonas aeruginosa, which can be found in a variety of niches in nature, is an opportunistic pathogen for various hosts. While P. aeruginosa strains are very versatile and diverse, only a few TA systems were characterized in this species. Here, we describe a newly characterized TA system in P. aeruginosa that is encoded within the filamentous Pf4 prophage. This system, named PfiT/PfiA, is a homologue of the ParE/YefM TA system. It is a type II TA system, in which the antitoxin is a protein that binds the toxic protein and eliminates the toxic effect. PfiT/PfiA carries several typical type II characteristics. Specifically, it constitutes two small genes expressed in a single operon, PfiT inhibits growth and PfiA eliminates this effect, PfiA binds PfiT, and PfiT expression results in elongated cells. Finally, we assigned a novel function to this TA system, where an imbalance between PfiT and PfiA, favouring the toxin, resulted in cell elongation and an increase in virion production.