Bacterial community shifts in taxa and diversity in response to localized organic loading in the deep sea

The deep sea is a unique and extreme environment characterized by low concentrations of highly recalcitrant carbon. As a consequence, large organic inputs have potential to cause significant perturbation. To assess the impact of organic enrichment on deep sea microbial communities, we investigated bacterial diversity in sediments underlying two whale falls at 1820 and 2893 m depth in Monterey Canyon, as compared with surrounding reference sediment 10–20 m away. Bacteroidetes, Epsilonproteobacteria and Firmicutes were recovered primarily from whale fall‐associated sediments, while Gammaproteobacteria and Planctomycetes were found primarily within reference sediments. Abundant Deltaproteobacteria were recovered from both sediment types, but the Desulfobacteraceae and Desulfobulbaceae families were observed primarily beneath the whale falls. UniFrac analysis revealed that bacterial communities from the two whale falls (～30 km apart) clustered to the exclusion of corresponding reference sediment communities, suggesting that deposition of whale fall biomass is more influential on deep sea microbial communities than specific seafloor location. The bacterial population at whale‐1820 at 7 months post deposition was less diverse than reference sediments, with Delta‐ and Epsilonproteobacteria and Bacteroidetes making up 89% of the community. At 70 months, bacterial diversity in reference sediments near whale‐2893 had decreased as well. Over this time, there was a convergence of each community's membership at the phyla level, although lower‐taxonomic‐level composition remained distinct. Long‐term impact of organic carbon loading from the whale falls was also evident by elevated total organic carbon and enhanced proteolytic activity for at least 17–70 months. The response of the sedimentary microbial community to large pulses of organic carbon is complex, likely affected by increased animal bioturbation, and may be sustained over time periods that span years to perhaps even decades.     

Molecular Detection of Marine Invertebrate Larvae

The ecological patterns of many invertebrate larvae remain an ongoing mystery, in large part owing to the difficult task of detecting them in the water column. The development of nucleic-acid–based technology has the potential to resolve this issue by direct identification and monitoring of embryonic and larval forms in situ. We report herein on the successful development and application of nucleic-acid–based sandwich hybridization assays that detect barnacles using rRNA-targeted probes with both group-(order Thoracica) and species-(Balanus glandula) specificity. Primary results include the determination of target 18S rRNA sequences and the construction of “capture” probes for detection of larvae using hybridization techniques. In addition, we modified existing protocols for whole cell hybridization of invertebrate larvae as confirmation of the sandwich hybridization results. We used both hybridization techniques successfully in the laboratory on a plankton time series collected over 3 months, as well as a week-long in situ deployment of the technique in Monterey Bay, CA. The adaptability of this technology promises to be further applicable to various organisms and could be used to enhance our understanding of larval presence in the world's oceans.     

Impact of disease-related mitochondrial mutations on tRNA structure and function

Over 150 mutations with documented pathogenicity have been identified within the human mitochondrial genome. More than half of the disease-related mutations are located within tRNA genes, a remarkable trend, given that these sequences comprise only 10% of the genome. The discovery of diseases correlated with mitochondrial tRNA mutations provides the first example of a class of pathologies related to RNA function, and the study of these tRNAs provides an interesting opportunity to explore the relationship between physiology and tRNA function. Investigations of both cellular and molecular effects have provided important insights into the structural and functional defects caused by the mutations. The picture that emerges from varied studies is that the effects of tRNA mutations are probably multifaceted and complex, but can be traced to the destabilization of structural features that destroy the native tRNA fold required for all aspects of function.     

Redundant and Distinct Functions for Dynamin-1 and Dynamin-2 Isoforms

A role for dynamin in clathrin-mediated endocytosis is now well established. However, mammals express three closely related, tissue-specific dynamin isoforms, each with multiple splice variants. Thus, an important question is whether these isoforms and splice variants function in vesicle formation from distinct intracellular organelles. There are conflicting data as to a role for dynamin-2 in vesicle budding from the TGN. To resolve this issue, we compared the effects of overexpression of dominant-negative mutants of dynamin-1 (the neuronal isoform) and dynamin-2 (the ubiquitously expressed isoform) on endocytic and biosynthetic membrane trafficking in HeLa cells and polarized MDCK cells. Both dyn1(K44A) and dyn2(K44A) were potent inhibitors of receptor-mediated endocytosis; however neither mutant directly affected other membrane trafficking events, including transport mediated by four distinct classes of vesicles budding from the TGN. Dyn2(K44A) more potently inhibited receptor-mediated endocytosis than dyn1(K44A) in HeLa cells and at the basolateral surface of MDCK cells. In contrast, dyn1(K44A) more potently inhibited endocytosis at the apical surface of MDCK cells. The two dynamin isoforms have redundant functions in endocytic vesicle formation, but can be targeted to and function differentially at subdomains of the plasma membrane.     

Patterns of intraspecific trait variation along an aridity gradient suggest both drought escape and drought tolerance strategies in an invasive herb

Background and AimsIn water-limited landscapes, some plants build structures that enable them to survive with minimal water (drought resistance). Instead of making structures that allow survival through times of water limitation, annual plants may invoke a drought escape strategy where they complete growth and reproduction when water is available. Drought escape and resistance each require a unique combination of traits and therefore plants are likely to have a suite of trait values that are consistent with a single drought response strategy. In environments where conditions are variable, plants may additionally evolve phenotypically plastic trait responses to water availability. Invasive annual species commonly occur in arid and semi-arid environments and many will be subject to reduced water availability associated with climate change. Assessing intraspecific trait variation across environmental gradients is a valuable tool for understanding how invasive plants establish and persist in arid environments.MethodsIn this study, we used a common garden experiment with two levels of water availability to determine how traits related to carbon assimilation, water use, biomass allocation and flowering phenology vary in California wild radish populations across an aridity gradient.Key ResultsWe found that populations from arid environments have rapid flowering and increased allocation to root biomass, traits associated with both drought escape and tolerance. Early flowering was associated with higher leaf nitrogen concentration and lower leaf mass per area, traits associated with high resource acquisition. While trait values varied across low- and high-water treatments, these shifts were consistent across populations, indicating no differential plasticity across the aridity gradient.ConclusionsWhile previous studies have suggested that drought escape and drought resistance are mutually exclusive drought response strategies, our findings suggest that invasive annuals may employ both strategies to succeed in novel semi-arid environments. As many regions are expected to become more arid in the future, investigations of intraspecific trait variation within low water environments help to inform our understanding of potential evolutionary responses to increased aridity in invasive species.     

Mutations in the β-tubulin binding site for peloruside A confer resistance by targeting a cleft significant in side chain binding

Peloruside A is a microtubule-stabilizing macrolide that binds to β-tubulin at a site distinct from the taxol site. The site was previously identified by H-D exchange mapping and molecular docking as a region close to the outer surface of the microtubule and confined in a cavity surrounded by a continuous loop of protein folded so as to center on Y340. We have isolated a series of peloruside A-resistant lines of the human ovarian carcinoma cell line A2780(1A9) to better characterize this binding site and the consequences of altering residues in it. Four resistant lines (Pel A-D) are described with single-base mutations in class I β-tubulin that result in the following substitutions: R306H, Y340S, N337D and A296S in various combinations. The mutations are localized to peptides previously identified by Hydrogen-Deuterium exchange mapping, and center on a cleft in which the drug side chain appears to dock. The Pel lines are 10–15-fold resistant to peloruside A and show cross resistance to laulimalide but not to any other microtubule stabilizers. They show no cross-sensitivity to any microtubule destabilizers, nor to two drugs with targets unrelated to microtubules. Peloruside A induces G₂/M arrest in the Pel cell lines at concentrations 10–15 times that required in the parental line. The cells show notable changes in morphology compared with the parental line.Key words: Apeloruside A, laulimalide, paclitaxel, drug resistance, mitotic arrest, binding site, β-tubulin     

Prospective genomic characterization of the German enterohemorrhagic Escherichia coli O104:H4 outbreak by rapid next generation sequencing technology

An ongoing outbreak of exceptionally virulent Shiga toxin (Stx)-producing Escherichia coli O104:H4 centered in Germany, has caused over 830 cases of hemolytic uremic syndrome (HUS) and 46 deaths since May 2011. Serotype O104:H4, which has not been detected in animals, has rarely been associated with HUS in the past. To prospectively elucidate the unique characteristics of this strain in the early stages of this outbreak, we applied whole genome sequencing on the Life Technologies Ion Torrent PGM? sequencer and Optical Mapping to characterize one outbreak isolate (LB226692) and a historic O104:H4 HUS isolate from 2001 (01-09591). Reference guided draft assemblies of both strains were completed with the newly introduced PGM? within 62 hours. The HUS-associated strains both carried genes typically found in two types of pathogenic E. coli, enteroaggregative E. coli (EAEC) and enterohemorrhagic E. coli (EHEC). Phylogenetic analyses of 1,144 core E. coli genes indicate that the HUS-causing O104:H4 strains and the previously published sequence of the EAEC strain 55989 show a close relationship but are only distantly related to common EHEC serotypes. Though closely related, the outbreak strain differs from the 2001 strain in plasmid content and fimbrial genes. We propose a model in which EAEC 55989 and EHEC O104:H4 strains evolved from a common EHEC O104:H4 progenitor, and suggest that by stepwise gain and loss of chromosomal and plasmid-encoded virulence factors, a highly pathogenic hybrid of EAEC and EHEC emerged as the current outbreak clone. In conclusion, rapid next-generation technologies facilitated prospective whole genome characterization in the early stages of an outbreak.