Site-specific mutagenesis in Legionella pneumophila by allelic exchange using counterselectable ColE1 vectors

Abstract To study the molecular pathogenesis of infection by Legionella pneumophila , a technique of site-specific mutagenesis by allelic exchange was evaluated. To develop this system, we optimized conjugal DNA transfer by isolating a mutant that functions 1000-fold more efficiently as a recipient than the wild type strain, identified two counter-selectable markers, rpsL and sacB , that function in L. pneumophila , and constructed a counterselectable Co1E1 vector. Allelic exchange of a L. pneumophila chrosomal gene was achieved at a frequency of 10⁻⁵ per transconjugant. The allelic exchange procedure itself did not alter the ability of L. pneumophila to infect macrophages, indicating that the system can be used to study this aspect of virulence.     

Conformations and interactions of histone H2A (F2A2, ALK).

Conformational changes in histone H2A (ALK, F2A2, IIbl) as a function of ionic strength and pH have been followed using high resolution nuclear magnetic resonance (NMR), circular dichroism (CD), and infrared (ir). While change in pH from 3 to 7 (no added salt) causes little structural change, added salt induces the formation of both alpha helix (28 percent maximum) and intermolecular associates in the region of the molecule between 25 and 113. No beta structure was observed at high salt. By the use of different salts it was shown that the structural changes were due largely to nonspecific counterion screening by the added anion. Comparison of observed with simulated NMR spectra has led to the proposal that an ionic strength dependent equilibrium exists between largely unstructured coil molecules and fully structured and aggregated molecules. NMR spectra of H2A obtained in the presence of DNA showed that both the N- and C-terminal regions bind to DNA, i.e., not the portion of the chain that is involved in interhistone interactions.     

Demography of an endangered,long-lived fish: Informing management options in the face of cyclic and stochastic climate variation

June sucker (Chasmistes liorus) is a long-lived, endangered fish endemic to Utah Lake, Utah. For several decades June sucker have failed to recruit sufficient numbers to the adult size classes such that the current wild population consists of a small number of old adults and it continues to decline. Vital rates of June sucker are influenced by climate-driven variation in lake level and inflow from the Provo River. We used population projection matrix modeling to assess effects of cyclic and stochastic environmental variation on population growth trajectories of June sucker in Utah Lake. The stable stage distribution is dominated by stage 1 individuals (93% of the total population) in contrast to the current situation where old age classes are the most abundant. Total population size is highly influenced by the stochastic component of climate variation; whereas, the adult population of June sucker closely tracks the systematic drought cycle. If changes in survival of larvae and juveniles can be coordinated such that positive changes in both parameters can occur somewhat simultaneously, then each parameter would only have to be increased by a factor of about 8.8 to achieve sustainable population growth (compared to a 77-fold increase for each parameter separately). Stochastic climatic variation has relatively little long-term effect on population growth. However, the multidecadal cyclic pattern of lake level and river discharge imposes a similar pattern on population growth rates of the June sucker, such that during some periods, populations decline even when the long-term trend is positive.     

Mode of Action of the Antimicrobial Peptide D4E1 on Aspergillus flavus

International Journal of Peptide Research and Therapeutics - The synthetic, linear peptide, D4E1, demonstrates antimicrobial activity against a broad spectrum of organisms including the toxigenic...     

SSU‐rRNA Gene Sequencing Survey of Benthic Microbial Eukaryotes from Guaymas Basin Hydrothermal Vent

Microbial eukaryotes have important roles in marine food webs, but their diversity and activities in hydrothermal vent ecosystems are poorly characterized. In this study, we analyzed microbial eukaryotic communities associated with bacterial (Beggiatoa) mats in the 2,000 m deep‐sea Guaymas Basin hydrothermal vent system using 18S rRNA gene high‐throughput sequencing of the V4 region. We detected 6,954 distinct Operational Taxonomic Units (OTUs) across various mat systems. Of the sequences that aligned with known protistan phylotypes, most were affiliated with alveolates (especially dinoflagellates and ciliates) and cercozoans. OTU richness and community structure differed among sediment habitats (e.g. different mat types and cold sediments away from mats). Additionally, full‐length 18S rRNA genes amplified and cloned from single cells revealed the identities of some of the most commonly encountered, active ciliates in this hydrothermal vent ecosystem. Observations and experiments were also conducted to demonstrate that ciliates were trophically active and ingesting fluorescent bacteria or Beggiatoa trichomes. Our work suggests that the active and diverse protistan community at the Guaymas Basin hydrothermal vent ecosystem likely consumes substantial amounts of bacterial biomass, and that the different habitats, often defined by distances of just a few 10s of cm, select for particular assemblages and levels of diversity.