The Physiological and Ecological Characterisation of the First Cultivated Species of the Candidate Division OP10

Bacteria are one of the three taxonomic domains and play a major role in the biological processes on Earth, yet their taxonomy and constituent species are poorly understood. OP10 is a candidate division within the domain Bacteria that up until recently has no cultivated representative. Until the recent isolation of the cultivated OP10 strains T49 and P488 from the Taupo Volcanic Zone in New Zealand (Stott et al. 2008), little was known about this putative phylum despite the fact that its presence was detected in various environmental surveys. This study aims to (i) characterise the physiology of one of the first isolated OP10 species, strain T49; and (ii) investigate and expand upon known OP10 diversity by using targeted oligonucleotide primers. Optimal growth temperature and pH of T49 was determined by cultivation in liquid medium. T49 substrate utilisation was investigated by culturing T49 with various substrates, with a focus on its sugar utilisation profiles. T49 was also characterised for its cell morphology through transmission and scanning electron microscopy as well as phase contrast microscopy. The total fatty acid profile of T49 was characterised through gas chromatography and mass spectroscopy. OP10 diversity was investigated at several geothermal sites using OP10-specific primers designed in the course of this study. The primers target conserved regions of the 16S rRNA gene sequence specific to clades within OP10. The primers were designed in silico by obtaining publically available OP10 16S rRNA gene sequences (through the Greengenes and SILVA databases) as references and dividing the candidate division into distinct clades through phylogenetic analysis with ARB. Primers were selected for specificity and coverage of the reference sequences within each of the clades. The performance of the primers were investigated by applying the primers against environmental DNA samples where OP10 sequences were previously identified, as well as various negative controls. The results were validated by sequencing PCR products generated to identify positive OP10 hits and false positive non-OP10 hits. This study found that T49 had a rod-shape morphology with the dimensions of 0.5-0.7 m by 2.5-3.0 m, more than half (67.4%) of the total fatty acids of the bacteria consisted of stearic acid (18:0 ) and palmitic acid (16:0), and GC content of 54.6%. Furthermore, T49 was found to have a growth temperature range of less than 50 degrees C to 73 degrees C and optimum at 68 degrees C, growth pH range of 4.9 to 5.8 and optimum at pH 5.3. Substrate utilisation experiments showed that strain T49 grew optimally using mono- and di- saccharides such as arabinose, mannose, ribose, galactose, and maltose, as well as amorphous polysaccharides including starch, glycogen, and dextrin. The OP10-specific primers successfully identified the presence of OP10 in environmental samples from Waikite, Waipahihi, and Mount Ngauruhoe , confirming previous unpublished 16S rRNA gene sequence surveys. Furthermore, the OP10-specific primers also identified previously undetected genetic diversity in Tikitere and Mount Ngauruhoe, demonstrating the advantage of the primers over traditional universal primers with their high specificity for OP10 DNA sequences which enabled more sensitive surveys of OP10 in the environment. The characterisation of the first OP10 species and the development of OP10-specific primers enable further investigation into this major taxonomic group within the Bacteria domain. T49 and its related strains may play important role within their environments, which can now be investigated based on these known physiologies. The effectiveness of the OP10-specific primers experimented may be use to detect presence of OP10 species in the environment at a higher sensitivity and selectiveness than traditional primers. The results from this research have expanded our knowledge on this previously unknown phylum. By increasing our knowledge of the OP10 candidate division, we also increase our understanding of the global bacterial diversity and this may help bring about insights into biology and global climate processes as well generating practical solutions in these fields. Many of these initial observations of T49 merit further investigation. These include: The production of pigments and lipids, two distinct cytosolic structures, and interesting growth behaviours such as quorum sensing and biofilm formation. OP10-specific primers developed in this study can be improved upon through further validations with environmental DNA representing clades that were not tested during this study due to unavailability. The improved primers can be developed as a rapid diagnostic tool to detect OP10 in the environment for isolation efforts. The primers may also act as fluorescent probes to identify OP10 in microbial consortia such as biofilms while preserving the structure of the microenvironments

Speech Dereverberation Based on Integrated Deep and Ensemble Learning Algorithm

Reverberation, which is generally caused by sound reflections from walls, ceilings, and floors, can result in severe performance degradation of acoustic applications. Due to a complicated combination of attenuation and time-delay effects, the reverberation property is difficult to characterize, and it remains a challenging task to effectively retrieve the anechoic speech signals from reverberation ones. In the present study, we proposed a novel integrated deep and ensemble learning algorithm (IDEA) for speech dereverberation. The IDEA consists of offline and online phases. In the offline phase, we train multiple dereverberation models, each aiming to precisely dereverb speech signals in a particular acoustic environment; then a unified fusion function is estimated that aims to integrate the information of multiple dereverberation models. In the online phase, an input utterance is first processed by each of the dereverberation models. The outputs of all models are integrated accordingly to generate the final anechoic signal. We evaluated the IDEA on designed acoustic environments, including both matched and mismatched conditions of the training and testing data. Experimental results confirm that the proposed IDEA outperforms single deep-neural-network-based dereverberation model with the same model architecture and training data

The First Insights into the Phylogeny, Genomics, and Ecology of the Novel Bacterial Phylum Armatimonadetes

Currently, a large proportion of novel microbial evolutionary lineages is poorly understood due to limited coverage of representative species. These “candidate” lineages represent significant gaps in our understanding of microbial function and ecology. This study focused on Chthonomonas calidirosea, the earliest isolated species within Armatimonadetes, the most recently-recognised bacterial phylum. The overall aim of this research was to start to understand the ecology and phylogeny of Armatimonadetes, and provide a foundation for future research into the phylum, with the benefit of narrowing the current knowledge gaps on microbial diversity. This was achieved by integrating multiple data types (phylogenetics, genomics, and community profiling metagenomics). The initial stage of this research aimed to address and clarify conflicts in reported phylogeny of the phylum Armatimonadetes. This study generated a comprehensive reference phylogenetic tree of 16S rRNA genes for the phylum, so that the phylogenetic position of newly-identified phylotypes can be reliably associated across studies. Multiple robust statistical methods were used to arrive at a consensus on the partitioning of classes and neighbouring phyla. The process also helped to identify and exclude candidate phyla previously misattributed to Armatimonadetes, thus better defining the phylum for future studies. The deeply-branching phylogenetic relationship of Armatimonadetes with other bacterial phyla was resolved by the sequencing of C. calidirosea T49T genome and analysing concatenated amino acid sequences of conserved genes against homologs in other prokaryotic genomes. The phylogenomic analysis showed Chloroflexi to be the most closely related formal phylum to Armatimonadetes. This publication was the first analysis of a genome from the phylum Armatimonadetes, and provided evolutionarily- and genetically-distinct insights to the overall knowledge of microbial genetic diversity. Analysis of the genome showed a metabolism geared towards non-cellulosic carbohydrates as the carbon and energy source, which coincides with previous culture-based physiological experiments (Lee et al., 2011). Genetic mechanisms behind leucine auxotrophy and narrow pH growth range were also identified. These observations supported the theory that C. calidirosea T49T occupies the niche of a scavenger of diverse species of carbohydrates within geothermal environments, in association with cellulolytic community members. In addition, the genome exhibited an unusual disorganisation of functionally-related genes typically found in conserved operons. The relatively high abundance of sigma factors (relative to genome size) in strain T49T may play an important role in gene regulation and coordination of metabolic pathways to compensate for the scattering of operons. Overall, this research built upon the previous physiological characterisation C. calidirosea T49T (Lee et al., 2011), resulting in a more in-depth and integrated analysis of the bacterium through both phenotypic and genotypic information. Finally, to investigate the genome dynamics of the species (particularly in genome organisation and adaptation to various environments as a scavenger), and to provide ecological and evolutionary context beyond the single genome analysed, the genomes of three additional C. calidirosea isolates cultured from diverse locations across the Taupō Volcanic Zone were extracted, sequenced, and compared to T49T. The genomes exhibited higher within-species conservation than other thermophilic species such as Thermus thermophilus (Henne et al., 2004; Jiang et al., 2013; Oshima & Ariga, 1975) and Sulfolobus islandicus (Reno et al., 2009) isolated from similar geographical distance. No genomic rearrangements were identified between C. calidirosea isolates. The majority of variation was limited to single nucleotide polymorphisms, with a limited number of horizontally-transferred genes and differentially-present fast-evolving genes, such as restriction modification system. The phylogeny and carbohydrate utilisation profiles of the isolates correlated with the geographical relationship between the sample sites rather than with other factors, such as soil geochemistry or microbial communities of the sites. The correlation between geography and phylogeny, low abundance of C. calidirosea at all sample sites (ranging from 0.006 % to 0.3 %), and the high genomic conservation indicated rapid aeolian dispersal and localised extinction as the most probable causes of homogeneity between the populations. The findings contribute to a better understanding of the genome dynamics and ecology of C. calidirosea, as well as the dispersal possibilities of free-living bacteria between distinct and discrete habitats. These studies addressed the overarching aim to investigate the ecology and phylogeny of Armatimonadetes through the research outlined above. This body of work contributed greatly to our understanding of Armatimonadetes phylogeny, both by clarifying its internal taxonomy and its position relative to neighbouring clades. Furthermore, it contributed to understanding of Armatimonadetes ecology by richly describing the ecological niche, genome, and lifestyle of C. calidirosea. Not only does this work greatly increase our understanding of the newest of the 30 prokaryotic phyla (Euzéby, 2011, Retrieved in December 2014), it also provides a rich foundation for future study

Existence theorems for a crystal surface model involving the p-Laplace operator

The manufacturing of crystal films lies at the heart of modern nanotechnology. How to accurately predict the motion of a crystal surface is of fundamental importance. Many continuum models have been developed for this purpose, including a number of PDE models, which are often obtained as the continuum limit of a family of kinetic Monte Carlo models of crystal surface relaxation that includes both the solid-on-solid and discrete Gaussian models. In this paper we offer an analytical perspective into some of these models. To be specific, we study the existence of a weak solution to the boundary value problem for the equation - \Delta e^{-\mbox{div}\left(|\nabla u|^{p-2}\nabla u\right)}+au=f, where $p>1, a>0$ are given numbers and $f$ is a given function. This problem is derived from a crystal surface model proposed by J.L.~Marzuola and J.~Weare (2013 Physical Review, E 88, 032403). The mathematical challenge is due to the fact that the principal term in our equation is an exponential function of a p-Laplacian. Existence of a suitably-defined weak solution is established under the assumptions that $p\in(1,2], \ N\leq 4$, and $f\in W^{1,p}$. Our investigations reveal that the key to our existence assertion is how to control the set where -\mbox{div}\left(|\nabla u|^{p-2}\nabla u\right) is $\pm\infty$