More Than Meets the Eye: Associations of Vaginal Bacteria with Gram Stain Morphotypes Using Molecular Phylogenetic Analysis

Bacterial vaginosis (BV) is a highly prevalent condition associated with adverse health outcomes. Gram stain analysis of vaginal fluid is the standard for confirming the diagnosis of BV, wherein abundances of key bacterial morphotypes are assessed. These Lactobacillus, Gardnerella, Bacteroides, and Mobiluncus morphotypes were originally linked to particular bacterial species through cultivation studies, but no studies have systematically investigated associations between uncultivated bacteria detected by molecular methods and Gram stain findings. In this study, 16S-rRNA PCR/pyrosequencing was used to examine associations between vaginal bacteria and bacterial morphotypes in 220 women with and without BV. Species-specific quantitative PCR (qPCR) and fluorescence in Situ hybridization (FISH) methods were used to document concentrations of two bacteria with curved rod morphologies: Mobiluncus and the fastidious BV-associated bacterium-1 (BVAB1). Rank abundance of vaginal bacteria in samples with evidence of curved gram-negative rods showed that BVAB1 was dominant (26.1%), while Mobiluncus was rare (0.2% of sequence reads). BVAB1 sequence reads were associated with Mobiluncus morphotypes (p<0.001). Among women with curved rods, mean concentration of BVAB1 DNA was 2 log units greater than Mobiluncus (p<0.001) using species-specific quantitative PCR. FISH analyses revealed that mean number of BVAB1 cells was 2 log units greater than Mobiluncus cells in women with highest Nugent score (p<0.001). Prevotella and Porphyromonas spp. were significantly associated with the “Bacteroides morphotype,” whereas Bacteroides species were rare. Gram-negative rods designated Mobiluncus morphotypes on Gram stain are more likely BVAB1. These findings provide a clearer picture of the bacteria associated with morphotypes on vaginal Gram stain.     

Spectral quality of monochromatic LED affects photosynthetic acclimation to high-intensity sunlight of Chrysanthemum and Spathiphyllum

Vertical farming using light-emitting diode offers potential for the early production phase (few weeks) of young ornamental plants. However, once transferred to the greenhouse, the photosynthetic acclimation of these young plants might depend on this initial light regime. To obtain insight about this acclimatization, Chrysanthemum (sun species) and Spathiphyllum (shade species) were preconditioned in growth chambers for 4 weeks under four light qualities: blue (B), red (R), red/blue (RB, 60% R) and white (W) at 100 μmol m⁻² s⁻¹. Monochromatic light (R and B) limited leaf development of both species, which resulted in a lower leaf mass per area when compared to multispectral light (RB for Chrysanthemum, RB and W for Spathiphyllum). R-developed leaves had a lower photosynthetic efficiency in both species. After the light quality pretreatment, plants were transferred to the greenhouse with high-intensity natural light conditions. On the first day of transfer, R and B preconditioned leaves of both species had an inhibited photosynthesis. After 1 week in natural light condition, rapid light curve parameters of Chrysanthemum leaves that developed under B acclimated to sunlight had a similar level than RB-developed leaves unlike R-leaves. Spathiphyllum leaves showed a decrease in maximum electron transport rate and this was most pronounced for the R pretreatment. After 1 month, R-preconditioned Chrysanthemum had the lowest dry mass, while no effects on the dry weight of Spathiphyllum with respect to the pretreatments were observed. Light quality during preconditioning affected the leaf ability to acclimate to natural high light intensities in greenhouse environment.     

A major locus controls a biologically active pheromone component in Heliconius melpomene

Understanding the production, response, and genetics of signals used in mate choice can inform our understanding of the evolution of both intraspecific mate choice and reproductive isolation. Sex pheromones are important for courtship and mate choice in many insects, but we know relatively little of their role in butterflies. The butterfly Heliconius melpomene uses a complex blend of wing androconial compounds during courtship. Electroantennography in H. melpomene and its close relative Heliconius cydno showed that responses to androconial extracts were not species specific. Females of both species responded equally strongly to extracts of both species, suggesting conservation of peripheral nervous system elements across the two species. Individual blend components provoked little to no response, with the exception of octadecanal, a major component of the H. melpomene blend. Supplementing octadecanal on the wings of octadecanal-rich H. melpomene males led to an increase in the time until mating, demonstrating the bioactivity of octadecanal in Heliconius. Using quantitative trait locus (QTL) mapping, we identified a single locus on chromosome 20 responsible for 41% of the parental species’ difference in octadecanal production. This QTL does not overlap with any of the major wing color or mate choice loci, nor does it overlap with known regions of elevated or reduced F_ST. A set of 16 candidate fatty acid biosynthesis genes lies underneath the QTL. Pheromones in Heliconius carry information relevant for mate choice and are under simple genetic control, suggesting they could be important during speciation.     

Polymerase manager protein UmuD directly regulates Escherichia coli DNA polymerase III α binding to ssDNA

Replication by Escherichia coli DNA polymerase III is disrupted on encountering DNA damage. Consequently, specialized Y-family DNA polymerases are used to bypass DNA damage. The protein UmuD is extensively involved in modulating cellular responses to DNA damage and may play a role in DNA polymerase exchange for damage tolerance. In the absence of DNA, UmuD interacts with the α subunit of DNA polymerase III at two distinct binding sites, one of which is adjacent to the single-stranded DNA-binding site of α. Here, we use single molecule DNA stretching experiments to demonstrate that UmuD specifically inhibits binding of α to ssDNA. We predict using molecular modeling that UmuD residues D91 and G92 are involved in this interaction and demonstrate that mutation of these residues disrupts the interaction. Our results suggest that competition between UmuD and ssDNA for α binding is a new mechanism for polymerase exchange.     

Noninvasive diffusive optical imaging of the auditory response to birdsong in the zebra finch

Songbirds communicate by learned vocalizations with concomitant changes in neurophysiological and genomic activities in discrete parts of the brain. Here, we tested a novel implementation of diffusive optical imaging (also known as diffuse optical imaging, DOI) for monitoring brain physiology associated with vocal signal perception. DOI noninvasively measures brain activity using red and near-infrared light delivered through optic fibers (optodes) resting on the scalp. DOI does not harm subjects, so it raises the possibility of repeatedly measuring brain activity and the effects of accumulated experience in the same subject over an entire life span, all while leaving tissue intact for further study. We developed a custom-made apparatus for interfacing optodes to the zebra finch (Taeniopygia guttata) head using 3D modeling software and rapid prototyping technology, and applied it to record responses to presentations of birdsong in isoflurane-anesthetized zebra finches. We discovered a subtle but significant difference between the hemoglobin spectra of zebra finches and mammals which has a major impact in how hemodynamic responses are interpreted in the zebra finch. Our measured responses to birdsong playback were robust, highly repeatable, and readily observed in single trials. Responses were complex in shape and closely paralleled responses described in mammals. They were localized to the caudal medial portion of the brain, consistent with response localization from prior gene expression, electrophysiological, and functional magnetic resonance imaging studies. These results define an approach for collecting neurophysiological data from songbirds that should be applicable to diverse species and adaptable for studies in awake behaving animals.     

Automatic drought stress detection in grapevines without using conventional threshold values

Aims

Because the water status of grapevines strongly affects the quality of the grapes and resulting wine, automated and early drought stress detection is important. Plant measurements are very promising for detecting drought stress, but strongly depend on microclimatic changes. Therefore, conventional stress detection methods require threshold values which define when plants start sensing drought stress. There is however no unique method to define these values. In this study, we propose two techniques that overcome this limitation.

Methods

Two statistical methods were used to automatically distinguish between drought and microclimate effects, based on a short preceding full-irrigated period to extract plant behaviour under normal conditions: Unfold Principal Component Analysis (UPCA) and Functional Unfold Principal Component Analysis (FUPCA). Both techniques aimed at detecting when measured sap flow rate or stem diameter variations in grapevine deviated from their normal behaviour due to drought stress.

Results

The models based on sap flow rate had some difficulties to detect stress on days with low atmospheric demands, while those based on stem diameter variations did not show this limitation, but ceased detecting stress when the stem diameter levelled off after a period of severe shrinkage. Nevertheless, stress was successfully detected with both approaches days before visible symptoms appeared.

Conclusions

UPCA and FUPCA based on plant indicators are therefore very promising for early stress detection.     

Interplant volatile signaling in willows: revisiting the original talking trees

The importance of interplant volatile signaling in plant–herbivore interactions has been a contentious issue for the past 30 years. We revisit willows as the system in which evidence for interplant signaling was originally found, but then questioned. We established three well-replicated experiments with two willow species (Salix exigua and Salix lemmonii) to address whether the receipt of an interplant signal from a neighboring willow reduces herbivore damage. Additionally we tested whether this signal is volatile in nature, and whether plants signal better to themselves than they do to other individuals. In all three experiments, we found evidence that cues from a damaged neighbor reduce subsequent herbivory experienced by willows. In one experiment, we showed that bagging of clipped tissue, which prevents the exchange of volatile signals, removed the effect of neighbor wounding. This was consistent with results from the other two experiments, in which clipping potted neighbors connected only through airborne volatile cues reduced damage of receivers. In one year, we found evidence that the perception of volatile signals from genetically identical clones was more effective at reducing foliar damage to a neighbor than signals from a genetically different individual. However, this trend was not significant in the following year. In three well-replicated experiments, we found strong evidence for the importance of interplant volatile cues in mediating herbivore interactions with willows.     

Semaphorin 3A Binds to the Perineuronal Nets via Chondroitin Sulfate Type E Motifs in Rodent Brains

Chondroitin sulfate (CS) and the CS-rich extracellular matrix structures called perineuronal nets (PNNs) restrict plasticity and regeneration in the CNS. Plasticity is enhanced by chondroitinase ABC treatment that removes CS from its core protein in the chondroitin sulfate proteoglycans or by preventing the formation of PNNs, suggesting that chondroitin sulfate proteoglycans in the PNNs control plasticity. Recently, we have shown that semaphorin3A (Sema3A), a repulsive axon guidance molecule, localizes to the PNNs and is removed by chondroitinase ABC treatment (Vo, T., Carulli, D., Ehlert, E. M., Kwok, J. C., Dick, G., Mecollari, V., Moloney, E. B., Neufeld, G., de Winter, F., Fawcett, J. W., and Verhaagen, J. (2013) Mol. Cell. Neurosci. 56C, 186–200). Sema3A is therefore a candidate for a PNN effector in controlling plasticity. Here, we characterize the interaction of Sema3A with CS of the PNNs. Recombinant Sema3A interacts with CS type E (CS-E), and this interaction is involved in the binding of Sema3A to rat brain-derived PNN glycosaminoglycans, as demonstrated by the use of CS-E blocking antibody GD3G7. In addition, we investigate the release of endogenous Sema3A from rat brain by biochemical and enzymatic extractions. Our results confirm the interaction of Sema3A with CS-E containing glycosaminoglycans in the dense extracellular matrix of rat brain. We also demonstrate that the combination of Sema3A and PNN GAGs is a potent inhibitor of axon growth, and this inhibition is reduced by the CS-E blocking antibody. In conclusion, Sema3A binding to CS-E in the PNNs may be a mechanism whereby PNNs restrict growth and plasticity and may represent a possible point of intervention to facilitate neuronal plasticity.