Characterization of the Mycobiome of the Seagrass, Zostera marina, Reveals Putative Associations With Marine Chytrids.

Seagrasses are globally distributed marine flowering plants that are foundation species in coastal ecosystems. Seagrass beds play essential roles as habitats and hatcheries, in nutrient cycling, and in protecting the coastline from erosion. Although many studies have focused on seagrass ecology, only a limited number have investigated their associated fungi. In terrestrial systems, fungi can have beneficial and detrimental effects on plant fitness. However, not much is known about marine fungi and even less is known about seagrass associated fungi. Here we used culture-independent sequencing of the ribosomal internal transcribed spacer (ITS) region to characterize the taxonomic diversity of fungi associated with the seagrass, Zostera marina. We sampled from two Z. marina beds in Bodega Bay over three time points to investigate fungal diversity within and between plants. Our results indicate that there are many fungal taxa for which a taxonomic assignment cannot be made living on and inside Z. marina leaves, roots and rhizomes and that these plant tissues harbor distinct fungal communities. We also identified differences in the abundances of the orders, Glomerellales, Agaricales and Malasseziales, between seagrass tissues. The most prevalent ITS amplicon sequence variants (ASVs) associated with Z. marina tissues could not initially be confidently assigned to a fungal phylum, but shared significant sequence similarity with Chytridiomycota and Aphelidomycota. To obtain a more definitive taxonomic classification of the most abundant ASV associated with Z. marina leaves, we used PCR with one primer targeting a unique region of this ASV's ITS2 and a second primer targeting fungal 28S rRNA genes to amplify part of the 28S rRNA gene region corresponding to this ASV. Sequencing and phylogenetic analysis of the resulting partial 28S rRNA gene revealed that the organism that this ASV comes from is a member of Novel Clade SW-I in the order Lobulomycetales in the phylum Chytridiomycota. This clade includes known parasites of freshwater diatoms and algae and it is possible this chytrid is directly infecting Z. marina leaf tissues. This work highlights a need for further studies focusing on marine fungi and the potential importance of these understudied communities to the larger seagrass ecosystem

Virtual in situs: Sequencing mRNA from cryo-sliced Drosophila embryos to determine genome-wide spatial patterns of gene expression

Complex spatial and temporal patterns of gene expression underlie embryo differentiation, yet methods do not yet exist for the efficient genome-wide determination of spatial expression patterns during development. In situ imaging of transcripts and proteins is the gold-standard, but it is difficult and time consuming to apply to an entire genome, even when highly automated. Sequencing, in contrast, is fast and genome-wide, but is generally applied to homogenized tissues, thereby discarding spatial information. It is likely that these methods will ultimately converge, and we will be able to sequence RNAs in situ, simultaneously determining their identity and location. As a step along this path, we developed methods to cryosection individual blastoderm stage Drosophila melanogaster embryos along the anterior-posterior axis and sequence the mRNA isolated from each 25 micron slice. The spatial patterns of gene expression we infer closely match patterns previously determined by in situ hybridization and microscopy. We applied this method to generate a genome-wide timecourse of spatial gene expression from shortly after fertilization through gastrulation. We identify numerous genes with spatial patterns that have not yet been described in the several ongoing systematic in situ based projects. This simple experiment demonstrates the potential for combining careful anatomical dissection with high-throughput sequencing to obtain spatially resolved gene expression on a genome-wide scale.Comment: 6 pages, 3 figures, 7 supplemental figures (available on request from [email protected]

Systematic identification of gene families for use as markers for phylogenetic and phylogeny- driven ecological studies of bacteria and archaea and their major subgroups

With the astonishing rate that the genomic and metagenomic sequence data sets are accumulating, there are many reasons to constrain the data analyses. One approach to such constrained analyses is to focus on select subsets of gene families that are particularly well suited for the tasks at hand. Such gene families have generally been referred to as marker genes. We are particularly interested in identifying and using such marker genes for phylogenetic and phylogeny-driven ecological studies of microbes and their communities. We therefore refer to these as PhyEco (for phylogenetic and phylogenetic ecology) markers. The dual use of these PhyEco markers means that we needed to develop and apply a set of somewhat novel criteria for identification of the best candidates for such markers. The criteria we focused on included universality across the taxa of interest, ability to be used to produce robust phylogenetic trees that reflect as much as possible the evolution of the species from which the genes come, and low variation in copy number across taxa. We describe here an automated protocol for identifying potential PhyEco markers from a set of complete genome sequences. The protocol combines rapid searching, clustering and phylogenetic tree building algorithms to generate protein families that meet the criteria listed above. We report here the identification of PhyEco markers for different taxonomic levels including 40 for all bacteria and archaea, 114 for all bacteria, and much more for some of the individual phyla of bacteria. This new list of PhyEco markers should allow much more detailed automated phylogenetic and phylogenetic ecology analyses of these groups than possible previously.Comment: 24 pages, 3 figure

Silicon implantation in GaAs

The electrical properties of room-temperature Si implants in GaAs have been studied. The implantations were done at 300 keV with doses ranging from 1.7×10^13 to 1.7×10^15 cm^–2. The implanted samples were annealed with silicon nitride encapsulants in H2 atmosphere for 30 min at temperatures ranging from 800 to 900°C to electrically activate the implanted ions. Results show that the implanted layers are n type, which implies that the Si ions preferentially go into Ga sites substitutionally. For low-dose implants, high (~90%) electrical activation of the implanted ions is achieved and the depth distribution of the free-electron concentration in the implanted layer roughly follows a Gaussian. However, for high-dose implants, the activation is poor (<15% for a 900 °C anneal) and the electron concentration profile is flat and deeper than the expected range

Social disparities in heart disease risk and survivor bias among autoworkers: an examination based on survival models and g-estimation.

ObjectivesTo examine gender and racial disparities in ischaemic heart disease (IHD) mortality related to metalworking fluid exposures and in the healthy worker survivor effect.MethodsA cohort of white and black men and women autoworkers in the USA was followed from 1941 to 1995 with quantitative exposure to respirable particulate matter from water-based metalworking fluids. Separate analyses used proportional hazards models and g-estimation.ResultsThe HR for IHD among black men was 3.29 (95% CI 1.49 to 7.31) in the highest category of cumulative synthetic fluid exposure. The HR for IHD among white women exposed to soluble fluid reached 2.44 (95% CI 0.96 to 6.22). However, no increased risk was observed among white men until we corrected for the healthy worker survivor effect. Results from g-estimation indicate that if white male cases exposed to soluble or synthetic fluid had been unexposed to that fluid type, then 1.59 and 1.20 years of life would have been saved on average, respectively.ConclusionsWe leveraged the strengths of two different analytic approaches to examine the IHD risks of metalworking fluids. All workers may have the same aetiological risk; however, black and female workers may experience more IHD from water-based metalworking fluid exposure because of a steeper exposure-response or weaker healthy worker survivor effect

Bacterial Community Sequences of Submerged Aquatic Vegetation in the Potomac River.

Here, we report results from PCR and sequencing of bacterial 16S rRNA genes from leaf and root surfaces from nine submerged aquatic vegetation (SAV) samples comprising five species. Samples were from four sites along the Potomac River