Exploring the Combined Action of Adding Pertuzumab to Branded Trastuzumab versus Trastuzumab Biosimilars for Treating HER2+ Breast Cancer.

The binding activity of various trastuzumab biosimilars versus the branded trastuzumab towards the glycosylated extracellular domain of the human epidermal growth factor receptor 2 (HER2) target in the presence of pertuzumab was investigated. We employed size exclusion chromatography with tetra-detection methodology to simultaneously determine absolute molecular weight, concentration, molecular size, and intrinsic viscosity. All trastuzumab molecules in solution exhibit analogous behavior in their binary action towards HER2 regardless of the order of addition of trastuzumab/pertuzumab. This analogous behavior of all trastuzumab molecules, including biosimilars, highlights the robustness and consistency of their binding activity towards HER2. Furthermore, the addition of HER2 to a mixture of trastuzumab and pertuzumab leads to increased formation of high-order HER2 complexes, up to concentrations of one order of magnitude higher than in the case of sequential addition. The observed increase suggests a potential synergistic effect between these antibodies, which could enhance their therapeutic efficacy in HER2-positive cancers. These findings underscore the importance of understanding the complex interplay between therapeutic antibodies and their target antigens, providing valuable insights for the development of more effective treatment strategies.

Binding Affinity of Trastuzumab and Pertuzumab Monoclonal Antibodies to Extracellular HER2 Domain.

The binding affinity of trastuzumab and pertuzumab to HER2 has been studied using both experimental and in silico methods. The experiments were conducted using the antibodies in their complete IgG form, as used in clinical therapy, and the extracellular domain of the HER2 protein in solution. This approach provides a precise, reproducible, and reliable view of the interaction between them in physicochemical conditions similar to those found in the tumoral environment. Dynamic light scattering and size exclusion chromatography coupled with tetra detection were utilized to characterize the protein complexes, measure their concentrations, and calculate the equilibrium-free binding energy, ΔGbind. In addition, PRODIGY, a QSAR-like model with excellent predictive ability, was employed to obtain in silico ΔGbind estimations. The results obtained indicate that pertuzumab exhibits a slightly higher binding affinity to HER2 than trastuzumab. The difference in binding affinity was explained based on the contribution of the different interfacial contact (IC) descriptors to the ΔGbind value estimated by the PRODIGY model. Furthermore, experiments revealed that the pertuzumab IgG antibody binds preferentially to two HER2 proteins, one per Fab fragment, while trastuzumab mainly forms a monovalent complex. This finding was interpreted based on a geometrical model that identified steric crowding in the trastuzumab-HER2 complex as compared with the pertuzumab-HER2 complex.

Revealing the transfer pathways of cyanobacterial-fixed N into the boreal forest through the feather-moss microbiome.

Introduction: Biological N2 fixation in feather-mosses is one of the largest inputs of new nitrogen (N) to boreal forest ecosystems; however, revealing the fate of newly fixed N within the bryosphere (i.e. bryophytes and their associated organisms) remains uncertain. Methods: Herein, we combined 15N tracers, high resolution secondary ion mass-spectrometry (NanoSIMS) and a molecular survey of bacterial, fungal and diazotrophic communities, to determine the origin and transfer pathways of newly fixed N2 within feather-moss (Pleurozium schreberi) and its associated microbiome. Results: NanoSIMS images reveal that newly fixed N2, derived from cyanobacteria, is incorporated into moss tissues and associated bacteria, fungi and micro-algae. Discussion: These images demonstrate that previous assumptions that newly fixed N2 is sequestered into moss tissue and only released by decomposition are not correct. We provide the first empirical evidence of new pathways for N2 fixed in feather-mosses to enter the boreal forest ecosystem (i.e. through its microbiome) and discuss the implications for wider ecosystem function.

Melting Temperature Depression of Polymer Single Crystals: Application to the Eco-Design of Tie-Layers in Polyolefinic-Based Multilayered Films.

In this paper, we describe a method for determining polymer compatibility, which will aid in establishing the requirements of polyolefinic materials for the eco-design of multilayer films for mechanical recycling while avoiding the use of reactive tie layers. Our ultimate goal is to define the molecular characteristics of the polyolefinic structural layer that improve compatibility with the tie layer during mechanical recycling. We have investigated the melting temperature depression of single crystals of various polyethylenes embedded in commercial polymeric matrices with various functionalities (ester, acrylate, acetate and methacrylic acid sodium ionomer), which can be potentially used as tie layers. We demonstrate how the concentration and molecular architecture of the matrices affect the melting temperature of the embedded single crystals differently depending on the latter's molecular architecture. The main finding indicates that the tie layers are more compatible with linear polyethylene than with branched polyethylenes. Indeed, our results show that the heterogeneous Ziegler-Natta linear low-density polyethylene is incompatible with all of the tie layers tested. The depression of melting temperatures observed are in excellent agreement with the results obtained by investigating the rheological behaviour and morphological features of solution-mixed blends in which segmental interactions between polymeric chains have been, in theory, maximized. Because Ziegler-Natta linear density polyethylene is one of the most commonly used polymers as a structural layer in multi-layer applications, the findings of this study are useful as they clearly show the unsuitability of this type of polyethylene for recycling from an eco-design standpoint. The specific molecular requirements for polyethylene layers (branching content less than 0.5/100 carbon atoms) can be specified for use in packaging, guiding the eco-design and valorisation of recycled multi-layered films containing this material.

Biofilm mechanics in an extremely acidic environment: microbiological significance.

A variety of natural biofilms were collected from an extremely acidic environment at Río Tinto (Spain). In order to provide insights into the structure-function relationship, the microstructure of the biofilms was explored using low temperature scanning electron microscopy (LTSEM) in combination with rheological analysis. The creep-recovery experiment results have demonstrated the typical behaviour of viscoelastic materials that combine both elastic and viscous characters. The LTSEM visualization and rheological characterization of biofilms revealed that the network density increased in bacterial biofilms and was the lowest in protist Euglena biofilms. This means that, in the latter biofilms, a lower density of interactions exist, suggesting that the whole system experiences enhanced mobility under external mechanical stress. The samples with the highest dynamic moduli (Leptospirillum-Acidiphilium, Zygnemopsis, Chlorella and Cyanidium) have shown the typical strain thinning behaviour, whereas the Pinnularia and Euglena biofilms exhibited a viscous thickening reaction. The Zygnemopsis filamentous floating structure has the highest cohesive energy and has shown distinctive enhanced resilience and connectivity. This suggests that biofilms should be viewed as soft viscoelastic systems the properties of which are determined by the main organisms and their extracellular polymeric substances. The fractional Maxwell model has been found to explain the rheological behaviour of the observed complex quite well, particularly the power-law behaviour and the characteristic broad relaxation response of these systems.

Haloterrigena sp. Strain SGH1, a Bacterioruberin-Rich, Perchlorate-Tolerant Halophilic Archaeon Isolated From Halite Microbial Communities, Atacama Desert, Chile.

An extreme halophilic archaeon, strain SGH1, is a novel microorganism isolated from endolithic microbial communities colonizing halites at Salar Grande, Atacama Desert, in northern Chile. Our study provides structural, biochemical, genomic, and physiological information on this new isolate living at the edge of the physical and chemical extremes at the Atacama Desert. SGH1 is a Gram-negative, red-pigmented, non-motile unicellular coccoid organism. Under the transmission electron microscope, strain SGH1 showed an abundant electro-dense material surrounding electron-lucent globular structures resembling gas vacuoles. Strain SGH1 showed a 16S rRNA gene sequence with a close phylogenetic relationship to the extreme halophilic archaea Haloterrigena turkmenica and Haloterrigena salina and has been denominated Haloterrigena sp. strain SGH1. Strain SGH1 grew at 20-40°C (optimum 37°C), at salinities between 15 and 30% (w/v) NaCl (optimum 25%) and growth was improved by addition of 50 mM KCl and 0.5% w/v casamino acids. Growth was severely restricted at salinities below 15% NaCl and cell lysis is avoided at a minimal 10% NaCl. Maximal concentrations of magnesium chloride and sodium or magnesium perchlorates that supported SGH1 growth were 0.5 and 0.15M, respectively. Haloterrigena sp. strain SGH1 accumulates bacterioruberin (BR), a C50 xanthophyll, as the major carotenoid. Total carotenoids in strain SGH1 amounted to nearly 400 µg BR per gram of dry biomass. Nearly 80% of total carotenoids accumulated as geometric isomers of BR: all-trans-BR (50%), 5-cis-BR (15%), 9-cis-BR (10%), 13-cis-BR (4%); other carotenoids were dehydrated derivatives of BR. Carotenogenesis in SGH1 was a reversible and salt-dependent process; transferring BR-rich cells grown in 25% (w/v) NaCl to 15% (w/v) NaCl medium resulted in depigmentation, and BR content was recovered after transference and growth of unpigmented cells to high salinity medium. Methanol extracts and purified BR isomers showed an 8-9-fold higher antioxidant activity than Trolox or ß-carotene. Both, plasma membrane integrity and mitochondrial membrane potential measurements under acute 18-h assays showed that purified BR isomers were non-toxic to cultured human THP-1 cells.

Assessing global range expansion in a cryptic species complex: insights from the red seaweed genus Asparagopsis (Florideophyceae).

The mitochondrial genetic diversity, distribution and invasive potential of multiple cryptic operational taxonomic units (OTUs) of the red invasive seaweed Asparagopsis were assessed by studying introduced Mediterranean and Hawaiian populations. Invasive behavior of each Asparagopsis OTU was inferred from phylogeographic reconstructions, past historical demographic dynamics, recent range expansion assessments and future distributional predictions obtained from demographic models. Genealogical networks resolved Asparagopsis gametophytes and tetrasporophytes into four A. taxiformis and one A. armata cryptic OTUs. Falkenbergia isolates of A. taxiformis L3 were recovered for the first time in the western Mediterranean Sea and represent a new introduction for this area. Neutrality statistics supported past range expansion for A. taxiformis L1 and L2 in Hawaii. On the other hand, extreme geographic expansion and an increase in effective population size were found only for A. taxiformis L2 in the western Mediterranean Sea. Distribution models predicted shifts of the climatically suitable areas and population expansion for A. armata L1 and A. taxiformis L1 and L2. Our integrated study confirms a high invasive risk for A. taxiformis L1 and L2 in temperate and tropical areas. Despite the differences in predictions among modelling approaches, a number of regions were identified as zones with high invasion risk for A. taxiformis L2. Since range shifts are likely climate-driven phenomena, future invasive behavior cannot be excluded for the rest of the lineages.

Structure and temporal dynamics of the bacterial communities associated to microhabitats of the coral Oculina patagonica.

Corals are known to contain a diverse microbiota that plays a paramount role in the physiology and health of holobiont. However, few studies have addressed the variability of bacterial communities within the coral host. In this study, bacterial community composition from the mucus, tissue and skeleton of the scleractinian coral Oculina patagonica were investigated seasonally at two locations in the Western Mediterranean Sea, to further understand how environmental conditions and the coral microbiome structure are related. We used denaturing gradient gel electrophoresis in combination with next-generation sequencing and electron microscopy to characterize the bacterial community. The bacterial communities were significantly different among coral compartments, and coral tissue displayed the greatest changes related to environmental conditions and coral health status. Species belonging to the Rhodobacteraceae and Vibrionaceae families form part of O. patagonica tissues core microbiome and may play significant roles in the nitrogen cycle. Furthermore, sequences related to the coral pathogens, Vibrio mediterranei and Vibrio coralliilyticus, were detected not only in bleached corals but also in healthy ones, even during cold months. This fact opens a new view onto unveiling the role of pathogens in the development of coral diseases in the future.

Phylogenetic and Functional Substrate Specificity for Endolithic Microbial Communities in Hyper-Arid Environments.

Under extreme water deficit, endolithic (inside rock) microbial ecosystems are considered environmental refuges for life in cold and hot deserts, yet their diversity and functional adaptations remain vastly unexplored. The metagenomic analyses of the communities from two rock substrates, calcite and ignimbrite, revealed that they were dominated by Cyanobacteria, Actinobacteria, and Chloroflexi. The relative distribution of major phyla was significantly different between the two substrates and biodiversity estimates, from 16S rRNA gene sequences and from the metagenomic data, all pointed to a higher taxonomic diversity in the calcite community. While both endolithic communities showed adaptations to extreme aridity and to the rock habitat, their functional capabilities revealed significant differences. ABC transporters and pathways for osmoregulation were more diverse in the calcite chasmoendolithic community. In contrast, the ignimbrite cryptoendolithic community was enriched in pathways for secondary metabolites, such as non-ribosomal peptides (NRP) and polyketides (PK). Assemblies of the metagenome data produced population genomes for the major phyla found in both communities and revealed a greater diversity of Cyanobacteria population genomes for the calcite substrate. Draft genomes of the dominant Cyanobacteria in each community were constructed with more than 93% estimated completeness. The two annotated proteomes shared 64% amino acid identity and a significantly higher number of genes involved in iron update, and NRPS gene clusters, were found in the draft genomes from the ignimbrite. Both the community-wide and genome-specific differences may be related to higher water availability and the colonization of large fissures and cracks in the calcite in contrast to a harsh competition for colonization space and nutrient resources in the narrow pores of the ignimbrite. Together, these results indicated that the habitable architecture of both lithic substrates- chasmoendolithic versus cryptoendolithic - might be an essential element in determining the colonization and the diversity of the microbial communities in endolithic substrates at the dry limit for life.

Solar Radiation Stress in Natural Acidophilic Biofilms of Euglena mutabilis Revealed by Metatranscriptomics and PAM Fluorometry.

The daily photosynthetic performance of a natural biofilm of the extreme acidophilic Euglena mutabilis from Río Tinto (SW, Spain) under full solar radiation was analyzed by means of pulse amplitude-modulated (PAM) fluorescence measurements and metatrascriptomic analysis. Natural E. mutabilis biofilms undergo large-scale transcriptomic reprogramming during midday due to a dynamic photoinhibition and solar radiation stress. Photoinhibition is due to UV radiation and not to light intensity, as revealed by PAM fluorometry analysis. In order to minimize the negative effects of solar radiation, our data supports the presence of a circadian rhythm in this euglenophyte that increases their opportunity to survive. Differential gene expression throughout the day (at 12:00, 20:00 and night) was monitored by massive Illumina parallel sequencing of metatranscriptomic libraries. The transcription pattern was altered in genes involved in Photosystem II stability and repair, UV damaged DNA repair, non-photochemical quenching and oxidative stress, supporting the photoinhibition detected by PAM fluorometry at midday.

Adaptation strategies of endolithic chlorophototrophs to survive the hyperarid and extreme solar radiation environment of the Atacama Desert.

The Atacama Desert, northern Chile, is one of the driest deserts on Earth and, as such, a natural laboratory to explore the limits of life and the strategies evolved by microorganisms to adapt to extreme environments. Here we report the exceptional adaptation strategies of chlorophototrophic and eukaryotic algae, and chlorophototrophic and prokaryotic cyanobacteria to the hyperarid and extremely high solar radiation conditions occurring in this desert. Our approach combined several microscopy techniques, spectroscopic analytical methods, and molecular analyses. We found that the major adaptation strategy was to avoid the extreme environmental conditions by colonizing cryptoendolithic, as well as, hypoendolithic habitats within gypsum deposits. The cryptoendolithic colonization occurred a few millimeters beneath the gypsum surface and showed a succession of organized horizons of algae and cyanobacteria, which has never been reported for endolithic microbial communities. The presence of cyanobacteria beneath the algal layer, in close contact with sepiolite inclusions, and their hypoendolithic colonization suggest that occasional liquid water might persist within these sub-microhabitats. We also identified the presence of abundant carotenoids in the upper cryptoendolithic algal habitat and scytonemin in the cyanobacteria hypoendolithic habitat. This study illustrates that successful lithobiontic microbial colonization at the limit for microbial life is the result of a combination of adaptive strategies to avoid excess solar irradiance and extreme evapotranspiration rates, taking advantage of the complex structural and mineralogical characteristics of gypsum deposits-conceptually called "rock's habitable architecture." Additionally, self-protection by synthesis and accumulation of secondary metabolites likely produces a shielding effect that prevents photoinhibition and lethal photooxidative damage to the chlorophototrophs, representing another level of adaptation.

Eukarya associated with the stony coral Oculina patagonica from the Mediterranean Sea.

Oculina patagonica is a putative alien scleractinian coral from the Southwest Atlantic that inhabits across the Mediterranean Sea. Here, we have addressed the diversity of Eukarya associated with this coral and its changes related to the environmental conditions and coral status. A total of 46 colonies of O. patagonica were taken from Alicante coast (Spain) and Pietra Ligure coast (Italy) and analyzed using denaturing gradient gel electrophoresis (DGGE) of the small-subunit 18S rRNA and 16S plastid rRNA genes, internal transcribed spacer region 2 (ITS 2) analyses, and electron microscopy. Our results show that Eukarya and plastid community associated to O. patagonica change with environmental conditions and coral status. Cryptic species, which can be difficult to identify by optical methods, were distinguished by 18S rRNA gene DGGE: the barnacle Megatrema anglicum, which was detected at two locations, and two boring sponges related to Cliona sp. and Siphonodictyon coralliphagum detected in samples from Tabarca and Alicante Harbour, respectively. Eukaryotic phototrophic community from the skeletal matrix of healthy corals was dominated by Ochrosphaera sp. while bleached corals from the Harbour and Tabarca were associated to different uncultured phototrophic organism. Differences in ultrastructural morphologies of the zooxanthellae between healthy and bleached corals were observed. Nevertheless, no differences were found in Symbiodinium community among time, environments, coral status and location, showing that O. patagonica hosted only one genotype of Symbiodinium belonging to clade B2. The fact that this clade has not been previously detected in other Mediterranean corals and is more frequent in the tropical Western Atlantic, is a new evidence that O. patagonica is an alien species in the Mediterranean Sea.

A Laboratory of Extremophiles: Iceland Coordination Action for Research Activities on Life in Extreme Environments (CAREX) Field Campaign.

Existence of life in extreme environments has been known for a long time, and their habitants have been investigated by different scientific disciplines for decades. However, reports of multidisciplinary research are uncommon. In this paper, we report an interdisciplinary three-day field campaign conducted in the framework of the Coordination Action for Research Activities on Life in Extreme Environments (CAREX) FP7EU program, with participation of experts in the fields of life and earth sciences. In situ experiments and sampling were performed in a 20 m long hot springs system of different temperature (57 °C to 100 °C) and pH (2 to 4). Abiotic factors were measured to study their influence on the diversity. The CO2 and H2S concentration varied at different sampling locations in the system, but the SO2 remained the same. Four biofilms, mainly composed by four different algae and phototrophic protists, showed differences in photosynthetic activity. Varying temperature of the sampling location affects chlorophyll fluorescence, not only in the microbial mats, but plants (Juncus), indicating selective adaptation to the environmental conditions. Quantitative polymerase chain reaction (PCR), DNA microarray and denaturing gradient gel electrophoresis (DGGE)-based analysis in laboratory showed the presence of a diverse microbial population. Even a short duration (30 h) deployment of a micro colonizer in this hot spring system led to colonization of microorganisms based on ribosomal intergenic spacer (RISA) analysis. Polyphasic analysis of this hot spring system was possible due to the involvement of multidisciplinary approaches.

Photosynthetic performance of phototrophic biofilms in extreme acidic environments.

Photosynthesis versus irradiance curves and their associated photosynthetic parameters from different phototrophic biofilms isolated from an extreme acidic environment (Río Tinto, SW, Spain) were studied in order to relate them to their species composition and the physicochemical characteristics of their respective sampling locations. The results indicated that the biofilms are low light acclimated showing a photoinhibition model; only floating communities of filamentous algae showed a light saturation model. Thus, all the biofilms analysed showed photoinhibition over 60 µmol photon m(-2) s(-1) except in the case of Zygnemopsis sp. sample, which showed a light-saturated photosynthesis model under irradiations higher that 200 µmol photon m(-2) s(-1). The highest values of compensation light intensity (I(c)) were showed also by Zygnemosis sp. biofilm (c. 40 µmol photon m(-2) s(-1)), followed by Euglena mutabilis and Chlorella sp. samples (c. 20 µmol photon m(-2) s(-1)). The diatom sample showed the lowest I(c) values (c. 5 µmol photon m(-2) s(-1)). As far as we know this is the first attempt to determine the photosynthetic activity of low pH and heavy metal tolerant phototrophic biofilms, which may give light in the understanding of the ecological importance of these biofilms for the maintenance of the primary production of these extreme and unique ecosystems.

Geomicrobiology of La Zarza-Perrunal acid mine effluent (Iberian Pyritic Belt, Spain).

Effluent from La Zarza-Perrunal, a mine on the Iberian Pyrite Belt, was chosen to be geomicrobiologically characterized along a 1,200-m stream length. The pH at the origin was 3.1, which decreased to 1.9 at the final downstream sampling site. The total iron concentration showed variations along the effluent, resulting from (i) significant hydrolysis and precipitation of Fe(III) (especially along the first reach of the stream) and (ii) concentration induced by evaporation (mostly in the last reach). A dramatic increase in iron oxidation was observed along the course of the effluent [from Fe(III)/Fe(total) = 0.11 in the origin to Fe(III)/Fe(total) = 0.99 at the last sampling station]. A change in the O(2) content along the effluent, from nearly anoxic at the origin to saturation with oxygen at the last sampling site, was also observed. Prokaryotic and eukaryotic diversity throughout the effluent was determined by microscopy and 16S rRNA gene cloning and sequencing. Sulfate-reducing bacteria (Desulfosporosinus and Syntrophobacter) were detected only near the origin. Some iron-reducing bacteria (Acidiphilium, Acidobacterium, and Acidosphaera) were found throughout the river. Iron-oxidizing microorganisms (Leptospirillum spp., Acidithiobacillus ferrooxidans, and Thermoplasmata) were increasingly detected downstream. Changes in eukaryotic diversity were also remarkable. Algae, especially Chlorella, were present at the origin, forming continuous, green, macroscopic biofilms, subsequently replaced further downstream by sporadic Zygnematales filaments. Taking into consideration the characteristics of this acidic extreme environment and the physiological properties and spatial distribution of the identified microorganisms, a geomicrobiological model of this ecosystem is advanced.

Eukaryotic microbial diversity of phototrophic microbial mats in two Icelandic geothermalhot springs.

The composition of the eukaryotic community and the three-dimensional structure of diverse phototrophic microbial mats from two hot springs in Iceland (Seltun and Hveradalir geothermal areas) were explored by comparing eukaryotic assemblages from microbial mats. Samples were collected in July 2007 from 15 sampling stations along thermal and pH gradients following both hot springs. Physicochemical data revealed high variability in terms of pH (ranging from 2.8 to 7), with high concentrations of heavy metals, including up to 20 g Fe/l, 80 mg Zn/l, 117 mg Cu/l, and 39 mg Ni/l at the most acidic sampling points. Phylogenetic analysis of 18S rDNA genes revealed a diversity of sequences related to several taxa, including members of the Bacillariophyta, Chlorophyta, Rhodophyta, and Euglenophyta phyla as well as ciliates, amoebae, and stramenopiles. The closest relatives to some of the sequences detected came from acidophilic organisms, even when the samples were collected at circumneutral water locations. Electron microscopy showed that most of the microecosystems analyzed were organized as phototrophic microbial mats in which filamentous cyanobacteria usually appeared as a major component. Deposits of amorphous minerals rich in silica, iron, and aluminium around the filaments were frequently detected.

Eukaryotic microbial diversity of phototrophic microbial mats in two Icelandic geothermalhot springs

The composition of the eukaryotic community and the three-dimensional structure of diverse phototrophic microbial mats from two hot springs in Iceland (Seltun and Hveradalir geothermal areas) were explored by comparing eukaryotic assemblages from microbial mats. Samples were collected in July 2007 from 15 sampling stations along thermal and pH gradients following both hot springs. Physicochemical data revealed high variability in terms of pH (ranging from 2.8 to 7), with high concentrations of heavy metals, including up to 20 g Fe/l, 80 mg Zn/l, 117 mg Cu/l, and 39 mg Ni/l at the most acidic sampling points. Phylogenetic analysis of 18S rDNA genes revealed a diversity of sequences related to several taxa, including members of the Bacillariophyta, Chlorophyta, Rhodophyta, and Euglenophyta phyla as well as ciliates, amoebae, and stramenopiles. The closest relatives to some of the sequences detected came from acidophilic organisms, even when the samples were collected at circumneutral water locations. Electron microscopy showed that most of the microecosystems analyzed were organized as phototrophic microbial mats in which filamentous cyanobacteria usually appeared as a major component. Deposits of amorphous minerals rich in silica, iron, and aluminium around the filaments were frequently detected (AU)

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Effect of oxidative stress on the growth of magnetic particles in Magnetospirillum magneticum.

Individual magnetosome-containing magnetic mineral particles (MMP) from magnetotactic bacteria grow rapidly such that only a small fraction (5%) of all magnetosomes contain dwarf (< or =20 nm) MMP. Studies of the developmental stages in the growth of MMP are difficult due to the absence of techniques to separate dwarf from mature particles and because the former are sensitive to extraction procedures. Here, O2 stress was used to inhibit MMP expression in Magnetospirillum magneticum strain AMB-1. In addition, defined growth conditions not requiring chemical monitoring or manipulation of the gas composition during growth resulted in the production of cells containing high numbers of dwarf MMP. Cells exposed to different incubation treatments and cells with dwarf MMP were compared to cells with normal MMP with respect to growth, respiration, iron content, and relative magnetite load (RML). The cells were examined by electron microscopy, low temperature magnetometry, X-ray diffraction (XRD), and Mössbauer spectroscopy. In the 0-110 microM O2(aq) range, growth was positively correlated with [O2] and negatively correlated with RML. Most MMP formed during exponential growth of the cells. At 50-100 microM O2(aq) with stirring (150 rpm) and 30% O2 loss during incubation, MMP expression was strongly inhibited whereas MMP nucleation was not. Cells highly enriched (~95%) in dwarf MMP were obtained at the end of the exponential phase in stirred (150 rpm) cultures containing 45 microM O2(aq). Only one dwarf MMP formed in each MMP vesicle and the chain arrangement was largely preserved. O2-stress-induced dwarf MMP consisted of non-euhedral spheroids (~25 nm) that were similar in shape and size to immature MMP from normal cells. They consisted solely of magnetite, with a single domain signature, no superparamagnetic behavior, and magnetic signatures, Fe(II)/Fe(III) ratios, and XRD patterns very similar to those of mature MMP. These results show that O2 stress in liquid cultures amended with an inorganic redox buffer (S2O3 2-/S0) can be used to produce abundant dwarf MMP that are good proxies for studying MMP development.

Effect of oxidative stress on the growth of magnetic particles in Magnetospirillum magneticum

Individual magnetosome-containing magnetic mineral particles (MMP) from magnetotactic bacteria grow rapidly such that only a small fraction (5%) of all magnetosomes contain dwarf (< or =20 nm) MMP. Studies of the developmental stages in the growth of MMP are difficult due to the absence of techniques to separate dwarf from mature particles and because the former are sensitive to extraction procedures. Here, O2 stress was used to inhibit MMP expression in Magnetospirillum magneticum strain AMB-1. In addition, defined growth conditions not requiring chemical monitoring or manipulation of the gas composition during growth resulted in the production of cells containing high numbers of dwarf MMP. Cells exposed to different incubation treatments and cells with dwarf MMP were compared to cells with normal MMP with respect to growth, respiration, iron content, and relative magnetite load (RML). The cells were examined by electron microscopy, low temperature magnetometry, X-ray diffraction (XRD), and Mössbauer spectroscopy. In the 0-110 microM O2(aq) range, growth was positively correlated with [O2] and negatively correlated with RML. Most MMP formed during exponential growth of the cells. At 50-100 microM O2(aq) with stirring (150 rpm) and 30% O2 loss during incubation, MMP expression was strongly inhibited whereas MMP nucleation was not. Cells highly enriched (~95%) in dwarf MMP were obtained at the end of the exponential phase in stirred (150 rpm) cultures containing 45 microM O2(aq). Only one dwarf MMP formed in each MMP vesicle and the chain arrangement was largely preserved. O2-stress-induced dwarf MMP consisted of non-euhedral spheroids (~25 nm) that were similar in shape and size to immature MMP from normal cells. They consisted solely of magnetite, with a single domain signature, no superparamagnetic behavior, and magnetic signatures, Fe(II)/Fe(III) ratios, and XRD patterns very similar to those of mature MMP. These results show that O2 stress in liquid cultures amended with an inorganic redox buffer (S2O3 2-/S0) can be used to produce abundant dwarf MMP that are good proxies for studying MMP development (AU)

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Prokaryotic community structure in algal photosynthetic biofilms from extreme acidic streams in Río Tinto (Huelva, Spain)

Four algal photosynthetic biofilms were collected from the Río Tinto (SW Spain) at four localities: AG, Euglena and Pinnularia biofilms; ANG, Chlorella and Pinnularia biofilms; RI, Cyanidium and Dunaliella biofilms; and CEM, Cyanidium, Euglena and Pinnularia biofilms. Community composition and structure were studied by a polyphasic approach consisting of 16S rRNA analysis, scanning electron microscopy by back-scattered electron detection mode (SEM-BSE), and fluorescence in-situ hybridization (FISH). Acidophilic prokaryotes associated with algal photosynthetic biofilms included sequences related to the Alpha-, Beta-, and Gammaproteobacteria (phylum Proteobacteria) and to the phyla Nitrospira, Actinobacteria, Acidobacteria and Firmicutes. Sequences from the Archaea domain were also identified. No more than seven distinct lineages were detected in any biofilm, except for those from RI, which contained fewer groups of Bacteria. Prokaryotic communities of the thinnest algal photosynthetic biofilms (-100 microm) were more related to those in the water column, including Leptospirillum populations. In general, thick biofilms (200 microm) generate microniches that could facilitate the development of less-adapted microorganisms (coming from the surrounding environment) to extreme conditions, thus resulting in a more diverse prokaryotic biofilm (AU)

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