Surface and subsurface nitrate flow pathways on a watershed scale

Determining the interaction and impact of surface runoff and subsurface flow processes on the environment has been hindered by our inability to characterize subsurface soil structures on a watershed scale. Ground penetrating radar (GPR) data were collected and evaluated in determining subsurface hydrology at four small watersheds in Beltsville, MD. The watersheds have similar textures, organic matter contents, and yield distributions. Although the surface slope was greater on one of the watersheds, slope alone could not explain why it also had a nitrate runoff flux that was 18 times greater than the other three watersheds. Only with knowledge of the subsurface hydrology could the surface runoff differences be explained. The subsurface hydrology was developed by combining GPR and surface topography in a geographic information system. Discrete subsurface flow pathways were identified and confirmed with color infrared imagery, real-time soil moisture monitoring, and yield monitoring. The discrete subsurface flow patterns were also useful in understanding observed nitrate levels entering the riparian wetland and first order stream. This study demonstrated the impact that subsurface stratigraphy can have on water and nitrate (NO3-N) fluxes exiting agricultural lands, even when soil properties, yield distributions, and climate are similar. Reliable protocols for measuring subsurface fluxes of water and chemicals need to be developed.     

Identification of genes encoding exported Mycobacterium tuberculosis proteins using a Tn552'phoA in vitro transposition system

Secreted and cell envelope-associated proteins are important to both Mycobacterium tuberculosis pathogenesis and the generation of protective immunity to M. tuberculosis. We used an in vitro Tn552'phoA transposition system to identify exported proteins of M. tuberculosis. The system is simple and efficient, and the transposon inserts randomly into target DNA. M. tuberculosis genomic libraries were targeted with Tn552'phoA transposons, and these libraries were screened in M. smegmatis for active PhoA translational fusions. Thirty-two different M. tuberculosis open reading frames were identified; eight contain standard signal peptides, six contain lipoprotein signal peptides, and seventeen contain one or more transmembrane domains. Four of these proteins had not yet been assigned as exported proteins in the M. tuberculosis databases. This collection of exported proteins includes factors that are known to participate in the immune response of M. tuberculosis and proteins with homologies, suggesting a role in pathogenesis. Nine of the proteins appear to be unique to mycobacteria and represent promising candidates for factors that participate in protective immunity and virulence. This technology of creating comprehensive fusion libraries should be applicable to other organisms.     

Distinct and overlapping roles of single-repeat MYB genes in root epidermal patterning

Cell specification in the root epidermis of Arabidopsis generates a position-dependent pattern of root-hair cells and non-hair cells. Here we conduct a comprehensive analysis of the five members of a single-repeat R3 MYB gene family, including CAPRICE (CPC), TRIPTYCHON (TRY), ENHANCER of TRY and CPC 1, 2, and 3 (ETC1, ETC2, and ETC3), and study their role and functional relationship in root epidermal cell specification. Based on genetic and expression analyses, CPC, TRY and ETC1, but not ETC2 or ETC3, promote the hair cell fate by inhibiting non-hair specification. Further, we find that single-repeat MYB activity is required for epidermal patterning throughout root development, beginning during embryogenesis. We also identify a novel regulatory interaction whereby GLABRA2 (GL2) promotes TRY (but not CPC or ETC1) expression in the root epidermis, which generates a second lateral inhibition feedback loop. Gene fusion experiments combining CPC regulatory elements with protein-coding regions of each single-repeat MYB gene suggest that all five proteins are functionally similar, although TRY and ETC2 exhibit distinctions from CPC/ETC1/ETC3. These results provide new insight into the function of these single-repeat MYBs and suggest that divergence of their regulatory sequences is largely responsible for their distinct roles in epidermal cell patterning.     

Value of a newly sequenced bacterial genome

Next-generation sequencing(NGS) technologies have made high-throughput sequencing available to medium- and small-size laboratories, culminating in a tidal wave of genomic information. The quantity of sequenced bacterial genomes has not only brought excitement to the field of genomics but also heightened expectations that NGS would boost antibacterial discovery and vaccine development. Although many possible drug and vaccine targets have been discovered, the success rate of genome-based analysis has remained below expectations. Furthermore, NGS has had consequences for genome quality, resulting in an exponential increase in draft(partial data) genome deposits in public databases. If no further interests are expressed for a particular bacterial genome, it is more likely that the sequencing of its genome will be limited to a draft stage, and the painstaking tasks of completing the sequencing of its genome and annotation will not be undertaken. It is important to know what is lost when we settle for a draft genome and to determine the "scientific value" of a newly sequenced genome. This review addresses the expected impact of newly sequenced genomes on antibacterial discovery and vaccinology. Also, it discusses the factors that could be leading to the increase in the number of draft deposits and the consequent loss of relevant biological information.     

Cellular mechanics of germ band retraction in Drosophila

Germ band retraction involves a dramatic rearrangement of the tissues on the surface of the Drosophila embryo. As germ band retraction commences, one tissue, the germ band, wraps around another, the amnioserosa. Through retraction the two tissues move cohesively as the highly elongated cells of the amnioserosa contract and the germ band moves so it is only on one side of the embryo. To understand the mechanical drivers of this process, we designed a series of laser ablations that suggest a mechanical role for the amnioserosa. First, we find that during mid retraction, segments in the curve of the germ band are under anisotropic tension. The largest tensions are in the direction in which the amnioserosa contracts. Second, ablating one lateral flank of the amnioserosa reduces the observed force anisotropy and leads to retraction failures. The other intact flank of amnioserosa is insufficient to drive retraction, but can support some germ band cell elongation and is thus not a full phenocopy of ush mutants. Another ablation-induced failure in retraction can phenocopy mys mutants, and does so by targeting amnioserosa cells in the same region where the mutant fails to adhere to the germ band. We conclude that the amnioserosa must play a key, but assistive, mechanical role that aids uncurling of the germ band.     

Phosphorothioates in molecular biology

The observation that phosphorothioate analogues of the nucleoside triphosphates are substrates for DNA- and RNA-polymerases has proven a boon for the molecular biologist. As these phosphorothioate-containing polymers are stable to degradation by nucleases and the sulfur atom confers many favourable chemical properties, several applications in molecular biology have been developed, including new methods for site-directed mutagenesis and DNA sequencing.