Diverse,rare microbial taxa responded to the Deepwater Horizon deep-sea hydrocarbon plume

The Deepwater Horizon (DWH) oil well blowout generated an enormous plume of dispersed hydrocarbons that substantially altered the Gulf of Mexico''s deep-sea microbial community. A significant enrichment of distinct microbial populations was observed, yet, little is known about the abundance and richness of specific microbial ecotypes involved in gas, oil and dispersant biodegradation in the wake of oil spills. Here, we document a previously unrecognized diversity of closely related taxa affiliating with Cycloclasticus, Colwellia and Oceanospirillaceae and describe their spatio-temporal distribution in the Gulf''s deepwater, in close proximity to the discharge site and at increasing distance from it, before, during and after the discharge. A highly sensitive, computational method (oligotyping) applied to a data set generated from 454-tag pyrosequencing of bacterial 16S ribosomal RNA gene V4–V6 regions, enabled the detection of population dynamics at the sub-operational taxonomic unit level (0.2% sequence similarity). The biogeochemical signature of the deep-sea samples was assessed via total cell counts, concentrations of short-chain alkanes (C₁–C₅), nutrients, (colored) dissolved organic and inorganic carbon, as well as methane oxidation rates. Statistical analysis elucidated environmental factors that shaped ecologically relevant dynamics of oligotypes, which likely represent distinct ecotypes. Major hydrocarbon degraders, adapted to the slow-diffusive natural hydrocarbon seepage in the Gulf of Mexico, appeared unable to cope with the conditions encountered during the DWH spill or were outcompeted. In contrast, diverse, rare taxa increased rapidly in abundance, underscoring the importance of specialized sub-populations and potential ecotypes during massive deep-sea oil discharges and perhaps other large-scale perturbations.     

Whole Genome Sequence Analysis Using JSpecies Tool Establishes Clonal Relationships between Listeria monocytogenes Strains from Epidemiologically Unrelated Listeriosis Outbreaks

In an effort to build a comprehensive genomic approach to food safety challenges, the FDA has implemented a whole genome sequencing effort, GenomeTrakr, which involves the sequencing and analysis of genomes of foodborne pathogens. As a part of this effort, we routinely sequence whole genomes of Listeria monocytogenes (Lm) isolates associated with human listeriosis outbreaks, as well as those isolated through other sources. To rapidly establish genetic relatedness of these genomes, we evaluated tetranucleotide frequency analysis via the JSpecies program to provide a cursory analysis of strain relatedness. The JSpecies tetranucleotide (tetra) analysis plots standardized (z-score) tetramer word frequencies of two strains against each other and uses linear regression analysis to determine similarity (r²). This tool was able to validate the close relationships between outbreak related strains from four different outbreaks. Included in this study was the analysis of Lm strains isolated during the recent caramel apple outbreak and stone fruit incident in 2014. We identified that many of the isolates from these two outbreaks shared a common 4b variant (4bV) serotype, also designated as IVb-v1, using a qPCR protocol developed in our laboratory. The 4bV serotype is characterized by the presence of a 6.3 Kb DNA segment normally found in serotype 1/2a, 3a, 1/2c and 3c strains but not in serotype 4b or 1/2b strains. We decided to compare these strains at a genomic level using the JSpecies Tetra tool. Specifically, we compared several 4bV and 4b isolates and identified a high level of similarity between the stone fruit and apple 4bV strains, but not the 4b strains co-identified in the caramel apple outbreak or other 4b or 4bV strains in our collection. This finding was further substantiated by a SNP-based analysis. Additionally, we were able to identify close relatedness between isolates from clinical cases from 1993–1994 and a single case from 2011 as well as links between two isolates from over 30 years ago. The identification of these potential links shows that JSpecies Tetra analysis can be a useful tool in rapidly assessing genetic relatedness of Lm isolates during outbreak investigations and for comparing historical isolates. Our analyses led to the identification of a highly related clonal group involved in two separate outbreaks, stone fruit and caramel apple, and suggests the possibility of a new genotype that may be better adapted for certain foods and/or environment.     

Glycine 384 is required for presenilin-1 function and is conserved in bacterial polytopic aspartyl proteases

Endoproteolysis of beta-amyloid precursor protein (betaAPP) and Notch requires conserved aspartate residues in presenilins 1 and 2 (PS1 and PS2). Although PS1 and PS2 have therefore been proposed to be aspartyl proteases, no homology to other aspartyl proteases has been found. Here we identify homology between the presenilin active site and polytopic aspartyl proteases of bacterial origin, thus supporting the hypothesis that presenilins are novel aspartyl proteases.     

Analysis of plastid DNA-like sequences within the nuclear genomes of higher plants

A wide-ranging examination of plastid (pt)DNA sequence homologies within higher plant nuclear genomes (promiscuous DNA) was undertaken. Digestion with methylation-sensitive restriction enzymes and Southern analysis was used to distinguish plastid and nuclear DNA in order to assess the extent of variability of promiscuous sequences within and between plant species. Some species, such as Gossypium hirsutum (cotton), Nicotiana tabacum (tobacco), and Chenopodium quinoa, showed homogenity of these sequences, while intraspecific sequence variation was observed among different cultivars of Pisum sativum (pea), Hordeum vulgare (barley), and Triticum aestivum (wheat). Hypervariability of plastid sequence homologies was identified in the nuclear genomes of Spinacea oleracea (spinach) and Beta vulgaris (beet), in which individual plants were shown to possess a unique spectrum of nuclear sequences with ptDNA homology. This hypervariability apparently extended to somatic variation in B. vulgaris. No sequences with ptDNA homology were identified by this method in the nuclear genome of Arabidopsis thaliana.      

Not so colourful after all: eggshell pigments constrain avian eggshell colour space

Birds'' eggshells are renowned for their striking colours and varied patterns. Although often considered exceptionally diverse, we report that avian eggshell coloration, sampled here across the full phylogenetic diversity of birds, occupies only 0.08–0.10% of the avian perceivable colour space. The concentrations of the two known tetrapyrrole eggshell pigments (protoporphyrin and biliverdin) are generally poor predictors of colour, both intra- and interspecifically. Here, we show that the constrained diversity of eggshell coloration can be accurately predicted by colour mixing models based on the relative contribution of both pigments and we demonstrate that the models'' predictions can be improved by accounting for the reflectance of the eggshell''s calcium carbonate matrix. The establishment of these proximate links between pigmentation and colour will enable future tests of hypotheses on the functions of perceived avian eggshell colours that depend on eggshell chemistry. More generally, colour mixing models are not limited to avian eggshell colours but apply to any natural colour. Our approach illustrates how modelling can aid the understanding of constraints on phenotypic diversity.     

Phylogeny of Greya (Lepidoptera: Prodoxidae), based on nucleotide sequence variation in mitochondrial cytochrome oxidase I and II: congruence with morphological data

The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.      

Comparison of micrometer‐ and scanning electron microscope‐based measurements of avian eggshell thickness

ABSTRACT The study of avian eggshell structure, including composition, pigmentation, thickness, and strength, has important ecological and economic implications. Previous investigators have used a variety of techniques to derive either direct measures or indirect estimates of eggshell thickness. Assessing the repeatability and method agreement of different techniques is necessary to permit comparison of eggshell thickness values from different studies on various genetic stocks, populations, and species. We recorded and analyzed measurements of eggshell thickness using two methods, micrometers and scanning electron microscopy (SEM), for several Palaeognathae and Neognathae taxa, including nonpasserines and passerines. Applying a tolerance‐interval approach, we found that repeatability of measurements for eggs with thinner shells (<300 μm, all Neognathae taxa) was worse than for eggs with thicker shells (Palaeognathae taxa), but was still statistically and biologically reasonable given that the relative magnitude of intramethod agreements was <11%. Our results support previous predictions that measurements made using a micrometer are comparable to those made using SEM. This finding is particularly important given the relative ease and cost efficiency of the micrometer method. Importantly, these new analyses can be used to validate the use of published data from previous studies of micrometer‐based eggshell thickness for both intra‐ and interspecific comparisons.     

Ionizing radiation induces delayed hyperrecombination in Mammalian cells

Exposure to ionizing radiation can result in delayed effects that can be detected in the progeny of an irradiated cell multiple generations after the initial exposure. These effects are described under the rubric of radiation-induced genomic instability and encompass multiple genotoxic endpoints. We have developed a green fluorescence protein (GFP)-based assay and demonstrated that ionizing radiation induces genomic instability in human RKO-derived cells and in human hamster hybrid GM10115 cells, manifested as increased homologous recombination (HR). Up to 10% of cells cultured after irradiation produce mixed GFP(+/-) colonies indicative of delayed HR or, in the case of RKO-derived cells, mutation and deletion. Consistent with prior studies, delayed chromosomal instability correlated with delayed reproductive cell death. In contrast, cells displaying delayed HR showed no evidence of delayed reproductive cell death, and there was no correlation between delayed chromosomal instability and delayed HR, indicating that these forms of genome instability arise by distinct mechanisms. Because delayed hyperrecombination can be induced at doses of ionizing radiation that are not associated with significantly reduced cell viability, these data may have important implications for assessment of radiation risk and understanding the mechanisms of radiation carcinogenesis.