Seed germination and plant fitness response of a narrowly endemic,rare winter annual to spatial heterogeneity in microenvironment

Environmental heterogeneity affects distributions of plant species, although the effects of fine-scale heterogeneity on plant demographies are not widely studied. Diminutive winter annuals, especially rare taxa, can be sensitive to spatial variation in microenvironment as a consequence of their small stature above and belowground. To address whether spatial environmental heterogeneity affects demography, germination and fitness of Chorizanthe orcuttiana, an endangered winter annual distributed in distinct easterly and westerly microhabitats within an exceedingly narrow niche in California, we performed multiyear observational and empirical studies. We manipulated after-ripening environment, soil moisture and ambient light at both aspects, and profiled microclimate, soil physicochemistry and soil microbiomes at all sites. We show that easterly aspects host larger plants in larger populations, and have lower air temperatures combined with higher soil moisture in comparison to the west-facing sites. Yet, soil physicochemistry and microbiomes were similar across all sites. Manipulations of after-ripening conditions showed that seeds exposed to low humidity (17%) during dormancy and sown at easterly aspects exhibited the highest germination percentages, whereas seeds incubated in situ and subsequently sown at westerly aspects yielded the lowest germination. Simultaneous manipulations of soil moisture and light showed that at both aspects higher moisture combined with shade led to higher germination, whereas ambient soil moisture combined with shade yielded the lowest germination. Altogether, our studies show that the diminutive, rare winter annual C. orcuttiana exhibits higher germination and plant fitness under cooler soil conditions with higher soil moisture while preferring drier environments during after-ripening.     

Conservative Treatment for Postprostatectomy Incontinence

Postprostatectomy incontinence (PPI) is a bothersome complication of radical prostatectomy. Although most men recover from PPI, some men continue to have persistent urinary incontinence. The initial management of persistent PPI usually consists of conservative measures such as pelvic floor muscle exercises. Surgical treatments are usually not entertained for men with urinary incontinence until conservative treatments have failed. This article discusses risk factors for PPI and various options for its treatment, including biofeedback and pharmacotherapy.Key words: Postprostatectomy incontinence, Radical prostatectomy, Pelvic floor muscle training, Pelvic floor stimulationPostprostatectomy incontinence (PPI) is a bothersome complication of radical prostatectomy (RP). Although most men recover from PPI, some men continue to have persistent urinary incontinence and roughly half of these men seek treatment.¹ It is important to understand the natural history of postprostatectomy urinary dysfunction prior to initiating treatment. Generally, the initial management of persistent PPI consists of conservative measures such as pelvic floor muscle exercises.     

The Oxytricha trifallax Macronuclear Genome: A Complex Eukaryotic Genome with 16,000 Tiny Chromosomes

The macronuclear genome of the ciliate Oxytricha trifallax displays an extreme and unique eukaryotic genome architecture with extensive genomic variation. During sexual genome development, the expressed, somatic macronuclear genome is whittled down to the genic portion of a small fraction (∼5%) of its precursor “silent” germline micronuclear genome by a process of “unscrambling” and fragmentation. The tiny macronuclear “nanochromosomes” typically encode single, protein-coding genes (a small portion, 10%, encode 2–8 genes), have minimal noncoding regions, and are differentially amplified to an average of ∼2,000 copies. We report the high-quality genome assembly of ∼16,000 complete nanochromosomes (∼50 Mb haploid genome size) that vary from 469 bp to 66 kb long (mean ∼3.2 kb) and encode ∼18,500 genes. Alternative DNA fragmentation processes ∼10% of the nanochromosomes into multiple isoforms that usually encode complete genes. Nucleotide diversity in the macronucleus is very high (SNP heterozygosity is ∼4.0%), suggesting that Oxytricha trifallax may have one of the largest known effective population sizes of eukaryotes. Comparison to other ciliates with nonscrambled genomes and long macronuclear chromosomes (on the order of 100 kb) suggests several candidate proteins that could be involved in genome rearrangement, including domesticated MULE and IS1595-like DDE transposases. The assembly of the highly fragmented Oxytricha macronuclear genome is the first completed genome with such an unusual architecture. This genome sequence provides tantalizing glimpses into novel molecular biology and evolution. For example, Oxytricha maintains tens of millions of telomeres per cell and has also evolved an intriguing expansion of telomere end-binding proteins. In conjunction with the micronuclear genome in progress, the O. trifallax macronuclear genome will provide an invaluable resource for investigating programmed genome rearrangements, complementing studies of rearrangements arising during evolution and disease.     

The AgI/II Family Adhesin AspA Is Required for Respiratory Infection by Streptococcus pyogenes

Streptococcus pyogenes (GAS) is a human pathogen that causes pharyngitis and invasive diseases such as toxic shock syndrome and sepsis. The upper respiratory tract is the primary reservoir from which GAS can infect new hosts and cause disease. The factors involved in colonisation are incompletely known however. Previous evidence in oral streptococci has shown that the AgI/II family proteins are involved. We hypothesized that the AspA member of this family might be involved in GAS colonization. We describe a novel mouse model of GAS colonization of the nasopharynx and lower respiratory tract to elucidate these interactions. We used two clinical M serotypes expressing AspA, and their aspA gene deletant isogenic mutants in experiments using adherence assays to respiratory epithelium, macrophage phagocytosis and neutrophil killing assays and in vivo models of respiratory tract colonisation and infection. We demonstrated the requirement for AspA in colonization of the respiratory tract. AspA mutants were cleared from the respiratory tract and were deficient in adherence to epithelial cells, and susceptible to phagocytosis. Expression of AspA in the surrogate host Lactococcus lactis protected bacteria from phagocytosis. Our results suggest that AspA has an essential role in respiratory infection, and may function as a novel anti-phagocytic factor.     

Histone Deacetylase Inhibitor Upregulates Peroxisomal Fatty Acid Oxidation and Inhibits Apoptotic Cell Death in Abcd1-Deficient Glial Cells

In X-ALD, mutation/deletion of ALD gene (ABCD1) and the resultant very long chain fatty acid (VLCFA) derangement has dramatically opposing effects in astrocytes and oligodendrocytes. While loss of Abcd1 in astrocytes produces a robust inflammatory response, the oligodendrocytes undergo cell death leading to demyelination in X-linked adrenoleukodystrophy (X-ALD). The mechanisms of these distinct pathways in the two cell types are not well understood. Here, we investigated the effects of Abcd1-knockdown and the subsequent alteration in VLCFA metabolism in human U87 astrocytes and rat B12 oligodendrocytes. Loss of Abcd1 inhibited peroxisomal β-oxidation activity and increased expression of VLCFA synthesizing enzymes, elongase of very long chain fatty acids (ELOVLs) (1 and 3) in both cell types. However, higher induction of ELOVL''s in Abcd1-deficient B12 oligodendrocytes than astrocytes suggests that ELOVL pathway may play a prominent role in oligodendrocytes in X-ALD. While astrocytes are able to maintain the cellular homeostasis of anti-apoptotic proteins, Abcd1-deletion in B12 oligodendrocytes downregulated the anti-apototic (Bcl-2 and Bcl-xL) and cell survival (phospho-Erk1/2) proteins, and upregulated the pro-apoptotic proteins (Bad, Bim, Bax and Bid) leading to cell loss. These observations provide insights into different cellular signaling mechanisms in response to Abcd1-deletion in two different cell types of CNS. The apoptotic responses were accompanied by activation of caspase-3 and caspase-9 suggesting the involvement of mitochondrial-caspase-9-dependent mechanism in Abcd1-deficient oligodendrocytes. Treatment with histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA) corrected the VLCFA derangement both in vitro and in vivo, and inhibited the oligodendrocytes loss. These observations provide a proof-of principle that HDAC inhibitor SAHA may have a therapeutic potential for X-ALD.     

Crystal Structure of Chlorite Dismutase, a Detoxifying Enzyme Producing Molecular Oxygen

Chlorite dismutase (Cld) is a key enzyme of perchlorate and chlorate respiration. This heme-based protein reduces the toxic compound chlorite into the innocuous chloride anion in a very efficient way while producing molecular oxygen. A sequence comparison between Cld homologues shows a highly conserved family. The crystal structure of Azospira oryzae strain GR-1 Cld is reported to 2.1 Å resolution. The structure reveals a hexameric organization of the Cld, while each monomer exhibits a ferredoxin-like fold. The six subunits are organized in a ring structure with a maximal diameter of 9 nm and an inner diameter of 2 nm. The heme active-site pocket is solvent accessible both from the inside and the outside of the ring. Moreover, a second anion binding site that could accommodate the assumed reaction intermediate ClO‾ for further transformation has been identified near the active site.The environment of the heme cofactor was investigated with electron paramagnetic resonance spectroscopy. Apart from the high-spin ferric signal of the five-coordinate resting-state enzyme, two low-spin signals were found corresponding to six-coordinate species. The current crystal structure confirms and complements a recently proposed catalytic mechanism that proceeds via a ferryl species and a ClO‾ anion. Our structural data exclude cooperativity between the iron centers.     

A 2.6 A structure of a serpin polymer and implications for conformational disease.

The function of the serpins as proteinase inhibitors depends on their ability to insert the cleaved reactive centre loop as the fourth strand in the main A beta-sheet of the molecule upon proteolytic attack at the reactive centre, P1-P1'. This mechanism is vulnerable to mutations which result in inappropriate intra- or intermolecular loop insertion in the absence of cleavage. Intermolecular loop insertion is known as serpin polymerisation and results in a variety of diseases, most notably liver cirrhosis resulting from mutations of the prototypical serpin alpha1-antitrypsin. We present here the 2.6 A structure of a polymer of alpha1-antitrypsin cleaved six residues N-terminal to the reactive centre, P7-P6 (Phe352-Leu353). After self insertion of P14 to P7, intermolecular linkage is affected by insertion of the P6-P3 residues of one molecule into the partially occupied beta-sheet A of another. This results in an infinite, linear polymer which propagates in the crystal along a 2-fold screw axis. These findings provide a framework for understanding the uncleaved alpha1-antitrypsin polymer and fibrillar and amyloid deposition of proteins seen in other conformational diseases, with the ordered array of polymers in the crystal resulting from slow accretion of the cleaved serpin over the period of a year.     

A rare temperate terrestrial orchid selects similar Tulasnella taxa in ex situ and in situ environments

Mycorrhizal symbiosis in orchids is unique in that fungal presence is considered a requirement for germination as well as for further development. Additionally, orchid fungal associations can exhibit high specificity in nature. Yet, an important ecological question remains unanswered: ‘With which orchid mycorrhizal fungi (OMF) do un-inoculated orchid seedlings form symbiosis when cultured ex situ?’ Simultaneously, it is asserted that orchid conservation efforts involving ex situ plant culture should exclusively utilize natural symbionts of the respective orchid taxa. We present a first comparison of OMF communities within the roots of asymbiotically cultured plants of the rare orchid Platanthera chapmanii grown ex situ (ES), and those occurring naturally in situ (IS). Nuclear ribosomal internal transcribed spacer (nrITS) barcoding region was used to identify peloton forming OMF from roots collected between 2012 and 2014 from both growing environments. Our 114 sequences clustered into 11 operational taxonomic units (OTUs) belonging to four closely related clades of the fungal family Tulasnellaceae. Shannon–Wiener (H) and Simpson diversity (D) indices were similar (p = 0.81 for both) for ES and IS OMF communities. Beta diversity comparisons also showed similarity between ES and IS treatments based on weighted (p = 0.10) and unweighted (p = 0.20) Bray–Curtis dissimilarity matrices. Bayesian and Maximum Likelihood (ML) phylograms clustered ES and IS derived fungal OTUs into the same clades. Our data suggest that P. chapmanii: (1) forms symbiosis with taxonomically similar fungi in ex situ culture and in its native soil, and (2) exhibits a narrow phylogenetic breadth of mycorrhizal fungal OTUs within the Tulasnellaceae.     

Enhanced transformation of polycyclic aromatic hydrocarbons using a combined Fenton's reagent,microbial treatment and surfactants

The potential for using Fenton's reagent (H₂O₂ + Fe²⁺) as an advanced oxidation pretreatment process to enhance microbial transformation of two model polycyclic aromatic hydrocarbons, anthracene and benzo[a]pyrene, in an aqueous system was evaluated. Fenton's reagent at a concentration of 0.5% H₂O₂ and 10 mM Fe²⁺ (molar ratio, 15:1) was most effective in transforming anthracene at pH 4. Application of non-ionic surfactants during Fenton's pre-treatment was found to be more effective in the transformation of both anthracene and benzo[a]pyrene. The extent of removal of substrates by a combined Fenton's–biotreatment was 2–4 times higher than with Fenton's treatment or biotreatment alone. In a chemical–biological treatment train, 48 h of Fenton's pre-treatment in the presence of a non-ionic surfactant, followed by 7 days of biological treatment resulted in 80–85% removal of PAHs (100 ppm). Electronic Publication