Isolation and identification of strains of Bacteroides fragilis group from the digestive tract of Callithrix penicillata marmosets

Thirty-five strains of the Bacteroides fragilis group were isolated from oral and intestinal samples from 5 wild caught, captive Callithrix penicillata. Nine oral strains of Bacteroides fragilis (7) and Bacteroides distasonis (2), and 26 intestinal strains of Bacteroides fragilis (14) and Bacteroides distasonis (12) were identified.     

Two new harvestmen species (Arachnida: Opiliones) from the Caucasus

Two new harvestmen species of the family Phalangiidae, Rilaena caucasica sp. n. and Rilaena silhavyi sp. n. are diagnosed, illustrated, and described from the Caucasus region. Comparative illustration of the related Rilaena anatolica (Roewer, 1956), R. atrolutea (Roewer, 1915) and R. kelbajarica Snegovaya &; Pkhakadze, 2014 Snegovaya, N. Y., &; Pkhakadze, V. D. (2014): New species of the genus Rilaena (Opiliones, Phalangiidae) from the mount Gyamish, Azerbaijan. Vestnik zoologii, 48, 313–318. doi: 10.2478/vzoo-2014-0037[Crossref] , [Google Scholar] are given.

http://www.zoobank.org/urn:lsid:zoobank.org:pub:7B29FD94-45A2-4E32-A41E-3276E016410B     

Contrasting Lesion Dynamics of White Syndrome among the scleractinian corals Porites spp

White syndrome (WS) is currently the most prevalent disease of scleractinian corals in the Indo-Pacific region, with an ability to exist in both epizootic and enzootic states. Here, we present results of an examination of WS lesion dynamics and show that potentially associated traits of host morphology (i.e., branching vs. massive), lesion size, and tissue deposition rate influence disease severity and recovery. Lesion healing rate was positively correlated with initial lesion size in both morphologies, but the rate at which lesions healed differed between morphologies. New lesions in branching Porites cylindrica appeared less frequently, were smaller and healed more quickly, but were more abundant than in closely-related massive Porites sp(p). The positive association between lesion size and healing rate was partly explained by geometry; branching limited lesion maximum size, and larger lesion margins contained more polyps producing new tissue, resulting in faster healing. However, massive colonies deposited tissue more slowly than branching colonies, resulting in slower recovery and more persistent lesions. Corallite size and density did not differ between species and did not, therefore, influence healing rate. We demonstrated multiple modes of pathogen transmission, which may be influenced by the greater potential for pathogen entrainment in branching vs. massive morphologies. We suggest that attributes such as colony morphology and species-specific growth rates require consideration as we expand our understanding of disease dynamics in colonial organisms such as coral.     

Regulation of maximal open probability is a separable function of Ca(v)beta subunit in L-type Ca2+ channel, dependent on NH2 terminus of alpha1C (Ca(v)1.2alpha)

beta subunits (Ca(v)beta) increase macroscopic currents of voltage-dependent Ca2+ channels (VDCC) by increasing surface expression and modulating their gating, causing a leftward shift in conductance-voltage (G-V) curve and increasing the maximal open probability, P(o,max). In L-type Ca(v)1.2 channels, the Ca(v)beta-induced increase in macroscopic current crucially depends on the initial segment of the cytosolic NH2 terminus (NT) of the Ca(v)1.2alpha (alpha1C) subunit. This segment, which we term the "NT inhibitory (NTI) module," potently inhibits long-NT (cardiac) isoform of alpha1C that features an initial segment of 46 amino acid residues (aa); removal of NTI module greatly increases macroscopic currents. It is not known whether an NTI module exists in the short-NT (smooth muscle/brain type) alpha(1C) isoform with a 16-aa initial segment. We addressed this question, and the molecular mechanism of NTI module action, by expressing subunits of Ca(v)1.2 in Xenopus oocytes. NT deletions and chimeras identified aa 1-20 of the long-NT as necessary and sufficient to perform NTI module functions. Coexpression of beta2b subunit reproducibly modulated function and surface expression of alpha1C, despite the presence of measurable amounts of an endogenous Ca(v)beta in Xenopus oocytes. Coexpressed beta2b increased surface expression of alpha1C approximately twofold (as demonstrated by two independent immunohistochemical methods), shifted the G-V curve by approximately 14 mV, and increased P(o,max) 2.8-3.8-fold. Neither the surface expression of the channel without Ca(v)beta nor beta2b-induced increase in surface expression or the shift in G-V curve depended on the presence of the NTI module. In contrast, the increase in P(o,max) was completely absent in the short-NT isoform and in mutants of long-NT alpha1C lacking the NTI module. We conclude that regulation of P(o,max) is a discrete, separable function of Ca(v)beta. In Ca(v)1.2, this action of Ca(v)beta depends on NT of alpha1C and is alpha1C isoform specific.     

Conformational Changes during Nucleotide Selection by Sulfolobus solfataricus DNA Polymerase Dpo4

The mechanism of nucleotide selection by Y-family DNA polymerases has been the subject of intense study, but significant structural contacts and/or conformational changes that relate to polymerase fidelity have been difficult to identify. Here we report on the conformational dynamics of a model Y-family polymerase Dpo4 from Sulfolobus solfataricus. Hydrogen-deuterium exchange in tandem with mass spectrometry was used to monitor changes in Dpo4 structure as a function of time and the presence or absence of specific substrates and ligands. Analysis of the data revealed previously unrecognized structural changes that accompany steps in the catalytic cycle leading up to phosphoryl transfer. For example, the solvent accessibility of the αB-loop-αC region in the finger domain decreased in the presence of all four dNTP insertion events, but the rate of deuterium exchange, an indicator of conformational flexibility, only decreased during an accurate insertion event. Of particular note is a change in the region surrounding the H-helix of the thumb domain. Upon binding DNA and Mg²⁺, the H-helix showed a decrease in solvent accessibility and flexibility that was relaxed only upon addition of dCTP, which forms a Watson-Crick base pair with template dG and not during mispairing events. The current study expands upon a previous report from our group that used a fluorescent probe located near the thumb domain to measure the kinetic properties of Dpo4 conformational changes. We now present a model for nucleotide selection by Dpo4 that arises from a synthesis of both structural and kinetic data.The mechanisms utilized by DNA polymerases to catalyze replication and/or repair of genomic material provide a fascinating example of how an enzyme can select a single substrate from multiple candidates, all with similar structural and chemical properties. DNA polymerases inside the cells of every living organism select from a pool of (four) dNTP substrates to catalyze phosphoryl transfer and then extend from a nascent primer strand opposite a DNA template. It has been emphasized that the endpoint of each nucleotide selection event cannot be determined solely by the thermodynamics of a given base pair (1, 2). Instead, the molecular features intrinsic to DNA polymerases are generally thought to guide the free-energy landscape of phosphoryl transfer toward the selection of “Watson-Crick” pairs (3, 4), at least among the four canonical bases. It is now apparent that individual DNA polymerases use variations upon a general mechanism to determine nucleotide selectivity (4–8). However, the mechanistic differences that define each polymerase class are only now being elucidated in any molecular detail.The Y-family DNA polymerases (pols)³ represent one class of polymerases with distinctive structural and functional features (8, 9). Like most other DNA polymerases, the Y-family pols have three domains: the finger for dNTP selection, the palm for catalysis, and the thumb for double strand DNA contact/orientation, which together form a structure that has been likened to a right hand. The Y-family polymerases also possess a unique domain that has been called the little finger or palm-associated domain. Most of the additional domains beyond these four core domains are involved in a complex web of protein-protein interactions and cellular localization events, although there are clearly exceptions to this rule (e.g. the N-clasp of pol κ and the N-digit of REV1) (10, 11). The Sulfolobus solfataricus DNA polymerase Dpo4 has served as the prototypical Y-family polymerase, because it is especially amenable to structural analysis and it shares many features and properties of other Y-family members. Numerous Dpo4 crystal structures have been reported in the literature (12–18). However, structural alignments of binary and ternary structures have failed to reveal the “open” to “closed” transition observed for some other pols (19). It is clear that, upon binding DNA, the little finger of Dpo4 probably undergoes a dramatic translation/rotation through space (20, 21), but few other conformational changes have been observed. The lack of obvious conformational rearrangements has led to the proposal that the Y-family DNA polymerases possess a “pre-formed” active site (22). However, there is substantial kinetic evidence in support of the view that a non-covalent step or steps occur in the Dpo4 reaction cycle prior to nucleophilic attack upon (what is assumed to be) a deprotonated 3′-hydroxyl group at the primer terminus, at least during formation of Watson-Crick geometry (23, 24). Previous work from our group used tryptophan fluorescence to monitor changes during dNTP insertion and following phosphoryl transfer (25). In the present study we sought to identify structural changes that occur during the Dpo4 reaction cycle by using hydrogen-deuterium exchange in tandem with mass spectrometry (HDX-MS). The method of HDX-MS has been used successfully to probe conformational dynamics of proteins and enzymes (26–29). The temporal and structural resolution provided by HDX-MS can serve as an effective bridge between kinetic analysis, which provides evidence of changes but cannot identify locations, and the inherently static nature of crystal structures. Our results provide new insight into Y-family polymerase catalysis, which may lead to a better understanding of how these enzymes select nucleotide substrates and, ultimately, how they contribute to mutagenesis.     

Resilience of Coral-Associated Bacterial Communities Exposed to Fish Farm Effluent

Background

The coral holobiont includes the coral animal, algal symbionts, and associated microbial community. These microbes help maintain the holobiont homeostasis; thus, sustaining robust mutualistic microbial communities is a fundamental part of long-term coral reef survival. Coastal pollution is one major threat to reefs, and intensive fish farming is a rapidly growing source of this pollution.

Methodology & Principal Findings

We investigated the susceptibility and resilience of the bacterial communities associated with a common reef-building coral, Porites cylindrica, to coastal pollution by performing a clonally replicated transplantation experiment in Bolinao, Philippines adjacent to intensive fish farming. Ten fragments from each of four colonies (total of 40 fragments) were followed for 22 days across five sites: a well-flushed reference site (the original fragment source); two sites with low exposure to milkfish (Chanos chanos) aquaculture effluent; and two sites with high exposure. Elevated levels of dissolved organic carbon (DOC), chlorophyll a, total heterotrophic and autotrophic bacteria abundance, virus like particle (VLP) abundances, and culturable Vibrio abundance characterized the high effluent sites. Based on 16S rRNA clone libraries and denaturing gradient gel electrophoresis (DGGE) analysis, we observed rapid, dramatic changes in the coral-associated bacterial communities within five days of high effluent exposure. The community composition on fragments at these high effluent sites shifted towards known human and coral pathogens (i.e. Arcobacter, Fusobacterium, and Desulfovibrio) without the host corals showing signs of disease. The communities shifted back towards their original composition by day 22 without reduction in effluent levels.

Significance

This study reveals fish farms as a likely source of pathogens with the potential to proliferate on corals and an unexpected short-term resilience of coral-associated bacterial communities to eutrophication pressure. These data highlight a need for improved aquaculture practices that can achieve both sustainable industry goals and long-term coral reef survival.     

Functional shifts in unvegetated,perhumid, recently-deglaciated soils do not correlate with shifts in soil bacterial community composition

Past work in recently deglaciated soils demonstrates that microbial communities undergo shifts prior to plant colonization. To date, most studies have focused on relatively ‘long’ chronosequences with the ability to sample plant-free sites over at least 50 years of development. However, some recently deglaciated soils feature rapid plant colonization and questions remain about the relative rate of change in the microbial community in the unvegetated soils of these chronosequences. Thus, we investigated the forelands of the Mendenhall Glacier near Juneau, AK, USA, where plants rapidly establish. We collected unvegetated samples representing soils that had been ice-free for 0, 1, 4, and 8 years. Total nitrogen (N) ranged from 0.00∼0.14 mg/g soil, soil organic carbon pools ranged from 0.6∼2.3 mg/g soil, and both decreased in concentration between the 0 and 4 yr soils. Biologically available phosphorus (P) and pH underwent similar dynamics. However, both pH and available P increased in the 8 yr soils. Nitrogen fixation was nearly undetectable in the most recently exposed soils, and increased in the 8 yr soils to ∼5 ng N fixed/cm²/h, a trend that was matched by the activity of the soil N-cycling enzymes urease and β-l,4-N-acetyl-glucosa-minidase. 16S rRNA gene clone libraries revealed no significant differences between the 0 and 8 yr soils; however, 8 yr soils featured the presence of cyanobacteria, a division wholly absent from the 0 yr soils. Taken together, our results suggest that microbes are consuming allochtonous organic matter sources in the most recently exposed soils. Once this carbon source is depleted, a competitive advantage may be ceded to microbes not reliant on in situ nutrient sources.