Temporal and spatial variability in nearshore bacterioplankton communities of Lake Michigan

The spatial and temporal variability of bacterial communities were determined for the nearshore waters of Lake Michigan, an oligotrophic freshwater inland sea. A freshwater estuary and nearshore sites were compared six times during 2006 using denaturing gradient gel electrophoresis (DGGE). Bacterial composition clustered by individual site and date rather than by depth. Seven 16S rRNA gene clone libraries were constructed, yielding 2717 bacterial sequences. Spatial variability was detected among the DGGE banding patterns and supported by clone library composition. The clone libraries from deep waters and the estuary environment revealed highest overall bacterial diversity. Betaproteobacteria sequence types were the most dominant taxa, comprising 40.2–67.7% of the clone libraries. BAL 47 was the most abundant freshwater cluster of Betaproteobacteria , indicating widespread distribution of this cluster in the nearshore waters of Lake Michigan. Incertae sedis 5 and Oxalobacteraceae sequence types were prevalent in each clone library, displaying more diversity than previously described in other freshwater environments. Among the Oxalobacteraceae sequences, a globally distributed freshwater cluster was determined. The nearshore waters of Lake Michigan are a dynamic environment that experience forces similar to the coastal ocean environment and share common bacterial diversity with other freshwater habitats.     

Modulation of chloride secretory responses and barrier function of intestinal epithelial cells by the Salmonella effector protein SigD

The Salmonella effector protein SigD is an inositol phosphate phosphatase that inhibits phosphatidylinositol 3-kinase-dependent signaling. Because epidermal growth factor (EGF) inhibits chloride secretion via phosphatidylinositol 3-kinase, we explored whether Salmonella infection might modify the inhibitory effect of EGF. As expected, EGF inhibited chloride secretion induced by carbachol in T₈₄ epithelial cells. Infection with wild-type (WT) but not sigD^– mutant S. typhimurium SL1344 decreased CCh-stimulated chloride secretion. Moreover, WT but not sigD^– Salmonella reduced the inhibitory effect of EGF on carbachol-stimulated chloride secretion. Complementation of sigD restored the ability of mutant Salmonella to reverse the inhibitory effect of EGF. EGF-induced EGF receptor phosphorylation was similar in cells infected with either WT or mutant Salmonella, and neither WT nor sigD^– Salmonella altered recruitment of the p85 subunit of phosphatidylinositol 3-kinase to EGF receptor, implying that SigD acts downstream of these signaling events. Furthermore, transepithelial resistance fell more rapidly in cells infected with WT vs. sigD^– Salmonella, indicating an early role for SigD in reducing barrier function, perhaps via activation of protein kinase C. We conclude that the Salmonella bacterial effector protein SigD may play critical roles in the pathogenesis of disease caused by this microorganism. chloride secretion; Salmonella typhimurium; epidermal growth factor     

Evaluating Human Risk from Exposure to Alkylated PAHs in an Aquatic System

Polycyclic aromatic hydrocarbons (PAHs) are a large class of organic chemicals typically found as mixtures in the aquatic environment from natural, petrogenic, and pyrogenic sources. People can be exposed to PAHs through ingestion or dermal contact with contaminated sediments or through ingestion of finfish and shellfish exposed to contaminated sediments. Although more than 100 PAHs have been identified, human exposure and risk are commonly evaluated for 18 individual PAHs. Other PAHs, such as alkylated PAHs, likely contribute to biological activity of environmental PAH mixtures; however, insufficient toxicity data are available to quantify their potential risk. This article presents an initial evaluation of the potential for human health risk from exposure to alkylated PAHs in sediment and fish. Individual alkylated PAHs have been observed to have potentially mutagenic, tumor-promoting, or carcinogenic activity. However, except for 1-and 2-methylnaphthalene, insufficient toxicity data are available to quantify toxicity or cancer risk from exposure to individual alkylated PAHs or mixtures of alkylated PAHs. This article describes a proposed strategy to better understand the potential human health risk from exposure to alkylated PAHs. Implementation of this strategy will contribute to evaluations of human exposure to complex PAH mixtures in the environment.     

Oligomerization of ICP4 and rearrangement of heat shock proteins may be important for herpes simplex virus type 1 prereplicative site formation

Herpes simplex virus type 1 (HSV-1) DNA replication occurs in replication compartments that form in the nucleus by an ordered process involving a series of protein scaffold intermediates. Following entry of viral genomes into the nucleus, nucleoprotein complexes containing ICP4 can be detected at a position adjacent to nuclear domain 10 (ND10)-like bodies. ND10s are then disrupted by the viral E3 ubiquitin ligase ICP0. We have previously reported that after the dissociation of ND10-like bodies, ICP8 could be observed in a diffuse staining pattern; however, using more sensitive staining methods, we now report that in addition to diffuse staining, ICP8 can be detected in tiny foci adjacent to ICP4 foci. ICP8 microfoci contain UL9 and components of the helicase-primase complex. HSV infection also results in the reorganization of the heat shock cognate protein 70 (Hsc70) and the 20S proteasome into virus-induced chaperone-enriched (VICE) domains. In this report we show that VICE domains are distinct but adjacent to the ICP4 nucleoprotein complexes and the ICP8 microfoci. In cells infected with an ICP4 mutant virus encoding a mutant protein that cannot oligomerize on DNA, ICP8 microfoci are not detected; however, VICE domains could still be formed. These results suggest that oligomerization of ICP4 on viral DNA may be essential for the formation of ICP8 microfoci but not for the reorganization of host cell chaperones into VICE domains.     

Assembly of an RNA-protein complex. Binding of NusB and NusE (S10) proteins to boxA RNA nucleates the formation of the antitermination complex involved in controlling rRNA transcription in Escherichia coli

Analytical ultracentrifugation and fluorescence anisotropy methods have been used to measure the equilibrium parameters that control the formation of the core subcomplex of NusB and NusE proteins and boxA RNA. This subcomplex, in turn, nucleates the assembly of the antitermination complex that is involved in controlling the synthesis of ribosomal RNA in Escherichia coli and that also participates in forming the N protein-dependent antitermination complex in lambdoid phage synthesis. In this study we determined the dissociation constants (K(d) values) for the individual binary interactions that participate in the assembly of the ternary NusB-NusE-boxA RNA subassembly, and we showed that multiple equilibria, involving both specific and nonspecific binding, are involved in the assembly pathway of this protein-RNA complex. The measured K(d) values were used to model the in vitro assembly reaction and combined with in vivo concentration data to simulate the overall control of the assembly of this complex in E. coli at two different cellular growth rates. The results showed that at both growth rates assembly proceeds via the initial formation of a weak but specific NusB-boxA complex, which is then stabilized by NusE binding. We showed that NusE also binds nonspecifically to available single-stranded RNA sequences and that such nonspecific protein binding to RNA can help to regulate crucial interactions in the assembly of the various macromolecular machines of gene expression.     

Seaweed processing using industrial single-mode cavity microwave heating: a preliminary investigation

A single-cavity microwave heating system has been designed and fabricated for microwave-assisted extraction of carrageenans from seaweed. The system comprises a single mode (TE101) waveguide fitted with power and temperature controls, together with a continuous-flow-recycle reactor operating at atmospheric pressure. The system has been tested by extraction of E. cottonii and E. spinosum in aqueous organic solvents. Even without purification, the extraction products were found to have virtually identical FTIR and 13C and 1H NMR spectra to the reference samples of kappa- and iota-carrageenan, respectively. The principal advantages of the microwave system are substantial reduction of extraction time and low consumption of organic solvents.     

Nereis diversicolor effect on the stability of cohesive intertidal sediments

The effect of the polychaete Nereis diversicolor on the stability of natural cohesive sediments was investigated in the laboratory. Three densities (450, 600 and 1200 ind m⁻²) of N. diversicolor were used. Sediment shear strength was measured using a cone penetrometer. Sediment erodability was assessed using an annular flume (current velocities from 5 to 55 cm s⁻¹) in which flow velocity was increased incrementally, and water sampled to quantify suspended material in order to derive critical erosion velocity and erosion rates. At low current velocities ( <25 cm s⁻¹), we found N. diversicolor to have a stabilising effect, reflected by an increase of up to 20% in the critical erosion velocity. This is related to an enhancement of ~50% in shear strength, due probably to gallery building activities, responsible for the promotion of lateral compaction, an increase in the area of the sediment–water interface, and enhanced microphytobenthos production. Once the sediment began to erode, the stabilising effect of N. diversicolor reverses, leading to an increase of up to 40% in eroded matter due to compaction, which resulted in the erosion of larger aggregates. The balance between the effect of N. diversicolor on herbivory and microphytobenthos production due to the presence of galleries is discussed. Our results indicate that neither chlorophyll a, nor shear strength nor critical erosion velocity are good indicators of erodability. This underlines the need to include biogeochemical processes in any realistic sediment transport model.