A novel model for cyanobacteria bloom formation: the critical role of anoxia and ferrous iron

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Gene flow in environmental Legionella pneumophila leads to genetic and pathogenic heterogeneity within a Legionnaires’ disease outbreak

Background

Legionnaires’ disease is a severe form of pneumonia caused by the environmental bacterium Legionella pneumophila. Outbreaks commonly affect people with known risk factors, but the genetic and pathogenic complexity of L. pneumophila within an outbreak is not well understood. Here, we investigate the etiology of the major Legionnaires’ disease outbreak that occurred in Edinburgh, UK, in 2012, by examining the evolutionary history, genome content, and virulence of L. pneumophila clinical isolates.

Results

Our high resolution genomic approach reveals that the outbreak was caused by multiple genetic subtypes of L. pneumophila, the majority of which had diversified from a single progenitor through mutation, recombination, and horizontal gene transfer within an environmental reservoir prior to release. In addition, we discover that some patients were infected with multiple L. pneumophila subtypes, a finding which can affect the certainty of source attribution. Importantly, variation in the complement of type IV secretion systems encoded by different genetic subtypes correlates with virulence in a Galleria mellonella model of infection, revealing variation in pathogenic potential among the outbreak source population of L. pneumophila.

Conclusions

Taken together, our study indicates previously cryptic levels of pathogen heterogeneity within a Legionnaires’ disease outbreak, a discovery that impacts on source attribution for future outbreak investigations. Furthermore, our data suggest that in addition to host immune status, pathogen diversity may be an important influence on the clinical outcome of individual outbreak infections.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-014-0504-1) contains supplementary material, which is available to authorized users.     

Incomplete recovery of plant diversity in restored prairie wetlands on agricultural landscapes

Restoration efforts are being implemented globally to mitigate the degradation and loss of wetland habitat; however, the rate and success of wetland vegetation recovery post‐restoration is highly variable across wetland classes and geographies. Here, we measured the recovery of plant diversity along a chronosequence of restored temporary and seasonal prairie wetlands ranging from 0 to 23 years since restoration, including drained and natural wetlands embedded in agricultural and natural reserve landscapes in central Alberta, Canada. We assessed plant diversity using the following structural indicators: percent cover of hydrophytes, native and non‐native species, species richness, and community composition. Our findings indicate that plant diversity recovered to resemble reference wetlands in agricultural landscapes within 3–5 years of restoration; however, restored wetlands maintained significantly lower species richness and a distinct community composition compared to reference wetlands located within natural reserves. Early establishment of non‐native species during recovery, dispersal limitation, and depauperated native seed bank were probable barriers to complete recovery. Determining the success of vegetation recovery provides important knowledge that can be used to improve restoration strategies, especially considering projected future changes in land use and climate.     

Repeated bouts of eccentrically biased endurance exercise stimulate salivary IgA secretion rate

To determine the salivary secretory immunoglobulin A (sIgA) response to repeated bouts of unaccustomed, downhill running (eccentrically biased) and examine potential protective immunological adaption from a repeated bout effect. Eleven active but untrained males (age: 19.7±0.4 years; VO_2peak: 47.8± 3.6 ml · kg⁻¹ · min ⁻¹) performed two 60 min bouts (Run 1 and Run 2) of downhill running (−13.5% gradient), separated by 14 days, at a speed eliciting 75% of their VO_2peak on a level grade. Saliva samples were collected before (baseline), immediately post exercise (IPE), and every hour for 12 h and every 24 h for 6 days after each run. Salivary sIgA concentration was measured and sIgA secretion rate was calculated. Results were analysed using repeated measures ANOVA (12 h period: 2x14; 24 h intervals: 2x7; p ≤ 0.05) with Tukey post-hoc tests where appropriate. Results are reported as means ± SE. There was a significant (p < 0.0001) interaction effect for sIgA secretion rate, IPE, with higher values after Run 2, as well as a significant (p < 0.01) time effect with elevated levels IPE and between 24 h and 144 h. There was a run effect (p < 0.0001), with the sIgA secretion rate significantly higher after Run 2. Repeated bouts of unaccustomed, eccentrically biased exercise induced alterations in the salivary sIgA secretion rate. This may serve as a protective mucosal adaptation to exercise-induced tissue damage.     

The photochemistry of some main chain liquid crystalline 4,4'-stilbene dicarboxylate polyesters.

Several aspects of the photochemistry and photophysics of four main chain liquid crystalline polyesters with a rigid trans-stilbene 4,4'-dicarboxylate mesogen as chromophore and flexible spacer groups are reported. The three polymers with the longest 'spacer' groups are liquid crystalline at room temperature, two have smectic phases. Chromophore aggregation has a dramatic effect on the photophysics and photochemistry of these polymers. Each of the polymers in poor solvents or as films has greatly perturbed UV-Vis absorption and fluorescence spectra due to aggregation of the stilbene chromophore. These effects are more pronounced upon annealing above the glass transition temperature, T(g), and in the mesophase. Film fluorescence is excitation wavelength dependent and is suppressed at elevated temperatures. The stilbene 'environment' in both films and solution is clearly heterogeneous and energy transfer processes relatively slow. The dominant photochemical reaction upon direct excitation above 300 nm is 2 + 2 photocycloaddition rendering polymer films insoluble. No significant trans-to-cis photoisomerization can be detected upon initial irradiation of the polymer films. There is evidence for the formation of aldehyde and carboxylate functionality upon irradiation in the presence of air. Loss of the aggregate UV-Vis absorption and fluorescence occurs during irradiation. Difference UV-Vis spectra of irradiated films suggest preferential initial consumption of dimeric aggregates. Loss of stilbene UV-Vis absorption upon irradiation above 300 nm can be partly photoreversed upon subsequent 254 nm irradiation. The rate of stilbene chromophore loss from films increased significantly above Tg and in the smectic phase above room temperature.     

Tropomyosin Isoform Expression Regulates the Transition of Adhesions To Determine Cell Speed and Direction

The balance of transition between distinct adhesion types contributes to the regulation of mesenchymal cell migration, and the characteristic association of adhesions with actin filaments led us to question the role of actin filament-associating proteins in the transition between adhesive states. Tropomyosin isoform association with actin filaments imparts distinct filament structures, and we have thus investigated the role for tropomyosins in determining the formation of distinct adhesion structures. Using combinations of overexpression, knockdown, and knockout approaches, we establish that Tm5NM1 preferentially stabilizes focal adhesions and drives the transition to fibrillar adhesions via stabilization of actin filaments. Moreover, our data suggest that the expression of Tm5NM1 is a critical determinant of paxillin phosphorylation, a signaling event that is necessary for focal adhesion disassembly. Thus, we propose that Tm5NM1 can regulate the feedback loop between focal adhesion disassembly and focal complex formation at the leading edge that is required for productive and directed cell movement.Among the different modes of migration that cells adopt, mesenchymal cell migration is dependent on integrin-based adhesion to the extracellular matrix (14), and the cellular mechanisms regulating integrin adhesion formation and turnover (adhesion dynamics) are integral to this process. The fate of integrin adhesions is intimately linked with filaments of polymerized actin (4). At the molecular level, actin filaments are highly dynamic, and this aspect of actin polymer biology provides an important control mechanism by which cells can organize filaments into structures with distinct properties. Tropomyosins are a multi-isoform family of actin-associating proteins that confer isoform-specific regulation of diverse actin filaments (3, 16, 34, 35). The interdependence of integrin adhesions and actin filaments suggests that expression of actin-associated proteins such as the tropomyosins may represent a mechanism for the regulation of adhesion dynamics that determine cell migration.In migrating cells small integrin-based focal complexes form at the periphery of lamellipodial extensions (32). These complexes are characterized by their subcellular distribution, dot-like shape, dependence on Rac activity, phosphorylated paxillin, and association with the network of short, branched actin filaments at the leading edge. The focal complexes are short lived (43) but provide strong traction forces at the leading edge (2) and most likely regulate directional migration (19). Subsets of focal complexes mature into focal adhesions, structures characterized by: Rho GTPase and Rho kinase dependence, dash-like shape, high levels of paxillin and phosphorylated paxillin, and low levels of the actin-binding molecule tensin (43, 44). The focal adhesions play an important role in anchoring bundles of polymerized actin stress fibers, providing the contractile force necessary for the translocation of the cell body during migration. There are at least three distinct classes of stress fibers observed in migrating cells (20, 27). Dorsal stress fibers are inserted into focal adhesions at the ventral surface of the cell. The distal end of the dorsal fibers can associate with a second type of actin fiber, the transverse arcs that run parallel to the leading edge and are not directly connected to focal adhesions. Ventral stress fibers have focal adhesions at either end and can be established following the contraction of two dorsal stress fibers and the associated transverse arc to form one actin bundle (20).Increased ventral stress fibers and focal adhesions are characteristic of nonmotile cells, in contrast, cell migration depends on focal adhesion turnover at the leading edge, allowing the formation of newly protruding regions of membrane and focal complex formation (28, 39). While the precise mechanism of focal adhesion turnover is incompletely understood, activation and phosphorylation of Src kinase, p130Cas, and paxillin (13, 39, 45) have all been implicated in focal adhesion turnover. A biphasic relationship between cell adhesion and cell speed suggests that conditions that alter the turnover rate of focal adhesions (either too much or too little) can reduce cell speed (18, 22).In cells with a fibroblastic phenotype, increased levels of acto-myosin contractility promote focal adhesion transition to fibrillar adhesions (also known as ECM contacts) (6, 7): elongated, thin, central arrays of dots or elongated fibrils that characteristically contain tensin but low levels of phosphorylated paxillin (29, 44, 45) and bind fibrils of fibronectin parallel to actin bundles (23, 29). These adhesions are formed by ligand-occupied fibronectin integrin receptor translocation from focal adhesions along bundles of actin filaments toward the cell center, and the process is dependent on an intact actin cytoskeleton and myosin activity (29). Receptor translocation stimulates matrix reorganization by transmitting cytoskeleton-generated tension through the integrin receptors onto the surrounding matrix (25, 29). The rate of receptor translocation is apparently independent from the rate of cell migration (29). However, the cytoskeletal tension that causes the fibrillar adhesion formation is also reported to decrease paxillin phosphorylation (45). Since phosphorylated paxillin is required for the generation of new focal complexes (45), conditions which switch the balance of adhesion in favor of fibrillar adhesion should presumably result in significantly reduced paxillin phosphorylation, leading to reduced focal adhesion turnover and correspondingly decreased cell migration.The cytoskeletal tropomyosin Tm5NM1 is a broadly distributed isoform (37) that alters cell shape (34), localizes to and promotes stress fibers that are resistant to actin depolymerizing drugs (9), enhances myosin IIA activation and recruitment to stress fibers, and inhibits cell migration (3). Therefore, we hypothesized that Tm5NM1 expression might determine cell migration by coordinating actin-dependent transition toward a predominance of focal adhesions and fibrillar adhesions. Using overexpression, knockdown, and genetic knockout models, we demonstrate that Tm5NM1 inhibits cell migration by promoting selective stabilization of focal adhesions and transition to fibrillar adhesions via the regulation of paxillin phosphorylation.     

Branchiobdellid annelids and their crayfish hosts: are they engaged in a cleaning symbiosis?

Branchiobdellid annelids and their freshwater crayfish hosts are generally thought to have a commensal relationship. Branchiobdellids of the genus Cambarincola exploit their hosts through a variety of mechanisms; however, an effect of branchiobdellids on crayfish has not been conclusively demonstrated. We investigated whether branchiobdellids positively affect the host crayfish Cambarus chasmodactylus in the New River, North Carolina. In a laboratory experiment, we placed 0, 3, or 6 branchiobdellids on C. chasmodactylus and observed a significant effect of branchiobdellid presence on both growth and mortality of host crayfish; crayfish with branchiobdellids exhibited faster growth and lower mortality with increasing branchiobdellid density. A tracer experiment demonstrated that branchiobdellids feed on items found in the branchial chamber of C. chasmodactylus. We hypothesize that such feeding activity by branchiobdellids reduces fouling of crayfish gills by epibionts and particulate matter and could lead to the reduced mortality and increased growth rates observed in the laboratory experiment. Specifically, Cambarincola may improve the ventilatory and excretory fitness of C. chasmodactylus by cleaning gill filaments. Field data support this hypothesis by demonstrating that branchiobdellids are found disproportionately at sites proximal to the branchial chamber in the New River. This study provides evidence that the relationship between C. chasmodactylus and Cambarincola may be a cleaning symbiosis, at least in environments where gill fouling is a problem for C. chasmodactylus.     

Relationship between intracellular PO2 recovery kinetics and fatigability in isolated single frog myocytes.

In single frog skeletal myocytes, a linear relationship exists between "fatigability" and oxidative capacity. The purpose of this investigation was to study the relationship between the intracellular Po(2) (Pi(O(2))) offset kinetics and fatigability in single Xenopus laevis myocytes to test the hypothesis that Pi(O(2)) offset kinetics would be related linearly with myocyte fatigability and, by inference, oxidative capacity. Individual myocytes (n = 30) isolated from lumbrical muscle were subjected to a 2-min bout of isometric peak tetanic contractions at either 0.25- or 0.33-Hz frequency while Pi(O(2)) was measured continuously via phosphorescence quenching techniques. The mean response time (MRT; time to 63% of the overall response) for Pi(O(2)) recovery from contracting values to resting baseline was calculated. After the initial square-wave constant-frequency contraction trial, each cell performed an incremental contraction protocol [i.e., frequency increase every 2 min from 0.167, 0.25, 0.33, 0.5, 1.0, and 2.0 Hz until peak tension fell below 50% of initial values (TTF)]. TTF values ranged from 3.39 to 10.04 min for the myocytes. The Pi(O(2)) recovery MRT ranged from 26 to 146 s. A significant (P < 0.05), negative relationship (MRT = -12.68TTF + 168.3, r(2) = 0.605) between TTF and Pi(O(2)) recovery MRT existed. These data demonstrate a significant correlation between fatigability and oxidative phosphorylation recovery kinetics consistent with the notion that oxidative capacity determines, in part, the speed with which skeletal muscle can recover energetically to alterations in metabolic demand.