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At least 199 records · Page 11

Carnobacterium pleistocenium sp. nov., a novel psychrotolerant, facultative anaerobe isolated from permafrost of the Fox Tunnel in Alaska

A novel, psychrotolerant, facultative anaerobe, strain FTR1T, was isolated from Pleistocene ice from the permafrost tunnel in Fox, Alaska. Gram-positive, motile, rod-shaped cells were observed with sizes 0.6-0.7 x 0.9-1.5 microm. Growth occurred within the pH range 6.5-9.5 with optimum growth at pH 7.3-7.5. The temperature range for growth of the novel isolate was 0-28 degrees C and optimum growth occurred at 24 degrees C. The novel isolate does not require NaCl; growth was observed between 0 and 5 % NaCl with optimum growth at 0.5 % (w/v). The novel isolate was a catalase-negative chemoorganoheterotroph that used as substrates sugars and some products of proteolysis. The metabolic end products were acetate, ethanol and CO2. Strain FTR1T was sensitive to ampicillin, tetracycline, chloramphenicol, rifampicin, kanamycin and gentamicin. 16S rRNA gene sequence analysis showed 99.8 % similarity between strain FTR1T and Carnobacterium alterfunditum, but DNA-DNA hybridization between them demonstrated 39+/-1.5 % relatedness. On the basis of genotypic and phenotypic characteristics, it is proposed that strain FTR1T (=ATCC BAA-754T=JCM 12174T=CIP 108033T) be assigned to the novel species Carnobacterium pleistocenium sp. nov.

Gram-Positive Asporogenous Rods/classification/gen↗

Integration of Russian Tier-1 Grid Center with High Performance Computers at NRC-KI for LHC experiments and beyond HENP

The LHC experiments are preparing for the precision measurements and further discoveries that will be made possible by higher LHC energies from April 2015 (LHC Run2). The need for simulation, data processing and analysis would overwhelm the expected capacity of grid infrastructure computing facilities deployed by the Worldwide LHC Computing Grid (WLCG). To meet this challenge the integration of the opportunistic resources into LHC computing model is highly important. The Tier-1 facility at Kurchatov Institute (NRC-KI) in Moscow is a part of WLCG and it will process, simulate and store up to 10% of total data obtained from ALICE, ATLAS and LHCb experiments. In addition Kurchatov Institute has supercomputers with peak performance 0.12 PFLOPS. The delegation of even a fraction of supercomputing resources to the LHC Computing will notably increase total capacity. In 2014 the development a portal combining a Tier-1 and a supercomputer in Kurchatov Institute was started to provide common interfaces and storage. The portal will be used not only for HENP experiments, but also by other data- and compute-intensive sciences like biology with genome sequencing analysis; astrophysics with cosmic rays analysis, antimatter and dark matter search, etc.

97 MATHEMATICS AND COMPUTING↗

AHTR Mechanical, Structural, and Neutronic Preconceptual Design

This report provides an overview of the mechanical, structural, and neutronic aspects of the Advanced High Temperature Reactor (AHTR) design concept. The AHTR is a design concept for a large output Fluoride salt cooled High-temperature Reactor (FHR) that is being developed to enable evaluation of the technology hurdles remaining to be overcome prior to FHRs becoming an option for commercial reactor deployment. This report documents the incremental AHTR design maturation performed over the past year and is focused on advancing the design concept to a level of a functional, self-consistent system. The reactor concept development remains at a preconceptual level of maturity. While the overall appearance of an AHTR design is anticipated to be similar to the current concept, optimized dimensions will differ from those presented here. The AHTR employs plate type coated particle fuel assemblies with rapid, off-line refueling. Neutronic analysis of the core has confirmed the viability of a 6-month two-batch cycle with 9 wt. % enriched uranium fuel. Refueling is intended to be performed automatically under visual guidance using dedicated robotic manipulators. The report includes a preconceptual design of the manipulators, the fuel transfer system, and the used fuel storage system. The present design intent is for used fuel to be stored inside of containment for at least six months and then transferred to local dry wells for intermediate term, on-site storage. The mechanical and structural concept development effort has included an emphasis on transportation and constructability to minimize construction costs and schedule. The design intent is that all components be factory fabricated into rail transportable modules that are assembled into subsystems at an on-site workshop prior to being lifted into position using a heavy-lift crane in an open-top style construction. While detailed accident identification and response sequence analysis has yet to be performed, the design concept incorporates fully passive responses to all identified design basis or non-very-low frequency beyond design basis accidents as well as multiple levels of radioactive material containment. Key building design elements include (1) below grade siting to minimize vulnerability to aircraft impact, (2) multiple natural circulation decay heat rejection chimneys, (3) seismic base isolation, and (4) decay heat powered back-up electricity generation.

01 COAL, LIGNITE, AND PEAT↗

Analysis of Salmonella enterica Isolated from a Mixed-Use Watershed in Georgia, USA: Antimicrobial Resistance, Serotype Diversity, and Genetic Relatedness to Human Isolates

As the cases of Salmonella enterica infections associated with contaminated water are increasing, this study was conducted to address the role of surface water as a reservoir of S. enterica serotypes. We sampled rivers and streams (n = 688) over a 3-year period (2015 to 2017) in a mixed-use watershed in Georgia, USA, and 70.2% of the total stream samples tested positive for Salmonella. A total of 1,190 isolates were recovered and characterized by serotyping, antimicrobial susceptibility testing, and pulsed-field gel electrophoresis (PFGE). A wide range of serotypes was identified, including those commonly associated with humans and animals, with S. enterica serotype Muenchen being predominant (22.7%) and each serotype exhibiting a high degree of strain diversity by PFGE. About half (46.1%) of the isolates had PFGE patterns indistinguishable from those of human clinical isolates in the CDC PulseNet database. A total of 52 isolates (4.4%) were resistant to antimicrobials, out of which 43 isolates were multidrug resistant (MDR; resistance to two or more classes of antimicrobials). These 52 resistant Salmonella isolates were screened for the presence of antimicrobial resistance genes, plasmid replicons, and class 1 integrons, out of which four representative MDR isolates were selected for whole-genome sequencing analysis. The results showed that 28 MDR isolates resistant to 10 antimicrobials had blacmy-2 on an A/C plasmid. Persistent contamination of surface water with a high diversity of Salmonella strains, some of which are drug resistant and genetically indistinguishable from human isolates, supports a role of environmental surface water as a reservoir for and transmission route of this pathogen.

59 BASIC BIOLOGICAL SCIENCES↗

Global Genomic Epidemiology of Salmonella enterica Serovar Typhimurium DT104

It has been 30 years since the initial emergence and subsequent rapid global spread of multidrug-resistant Salmonella enterica serovar Typhimurium DT104 (MDR DT104). Nonetheless, its origin and transmission route have never been revealed. In this paper, we used whole-genome sequencing (WGS) and temporally structured sequence analysis within a Bayesian framework to reconstruct temporal and spatial phylogenetic trees and estimate the rates of mutation and divergence times of 315 S. Typhimurium DT104 isolates sampled from 1969 to 2012 from 21 countries on six continents. DT104 was estimated to have emerged initially as antimicrobial susceptible in ~1948 (95% credible interval [CI], 1934 to 1962) and later became MDR DT104 in ~1972 (95% CI, 1972 to 1988) through horizontal transfer of the 13-kb Salmonella genomic island 1 (SGI1) MDR region into susceptible strains already containing SGI1. This was followed by multiple transmission events, initially from central Europe and later between several European countries. An independent transmission to the United States and another to Japan occurred, and from there MDR DT104 was probably transmitted to Taiwan and Canada. An independent acquisition of resistance genes took place in Thailand in ~1975 (95% CI, 1975 to 1990). In Denmark, WGS analysis provided evidence for transmission of the organism between herds of animals. Interestingly, the demographic history of Danish MDR DT104 provided evidence for the success of the program to eradicate Salmonella from pig herds in Denmark from 1996 to 2000. Finally, the results from this study refute several hypotheses on the evolution of DT104 and suggest that WGS may be useful in monitoring emerging clones and devising strategies for prevention of Salmonella infections.

59 BASIC BIOLOGICAL SCIENCES↗

The oncogenic role of meiosis-specific Aurora kinase C in mitotic cells

Highlights: • AURKC is expressed in meiotic cells and some cancers. • AURKC expression increases cell migration and transformation of human mitotic cells. • Deletion of AURKC in U2OS cells decreases cell migration and soft agar growth. • Transcriptome analysis reveals that AURKC alters the extracellular matrix. Aberrant expression of meiosis-specific genes in cancer has recently emerged as a driver of some cancer formation. Aurora kinase C (AURKC) is a member of the Aurora kinase family of proteins known to regulate chromosome segregation during cell divisions. AURKC is normally expressed in meiotic cells; however, elevated levels of AURKC mRNA and protein are frequently measured in cancer cells. To understand the function of AURKC in cancer cells, expression was induced in noncancerous, human retina pigmented epithelial cells. While AURKC expression did not alter cell proliferation over 72 h, it did increase cell migration and anchorage independent growth in soft agar suggesting an oncogenic role in mitotically dividing cells. To evaluate AURKC as a potential therapeutic target, a frameshift mutation in the gene was introduced in U2OS osteosarcoma cells using CRISPR-Cas9 technology resulting in a premature stop codon. Cancer cells lacking AURKC displayed no change in cell proliferation over 72 h but did migrate less and formed fewer colonies in soft agar. Whole transcriptome sequencing analysis uncovered over 400 differentially expressed genes in U2OS cells with and without AURKC. GO analysis revealed alterations in proteinaceous extracellular matrix genes including COL1A1. These data indicate that therapeutics targeting AURKC could decrease cancer cell metastasis and disease progression. Because AURKC is transcriptionally silenced in normal mitotic cells, its disruption could specifically target cancer cells limiting the toxic side effects associated with current therapeutics.

60 APPLIED LIFE SCIENCES↗

Differential expression of members of the annexin multigene family in Arabidopsis

Although in most plant species no more than two annexin genes have been reported to date, seven annexin homologs have been identified in Arabidopsis, Annexin Arabidopsis 1-7 (AnnAt1--AnnAt7). This establishes that annexins can be a diverse, multigene protein family in a single plant species. Here we compare and analyze these seven annexin gene sequences and present the in situ RNA localization patterns of two of these genes, AnnAt1 and AnnAt2, during different stages of Arabidopsis development. Sequence analysis of AnnAt1--AnnAt7 reveals that they contain the characteristic four structural repeats including the more highly conserved 17-amino acid endonexin fold region found in vertebrate annexins. Alignment comparisons show that there are differences within the repeat regions that may have functional importance. To assess the relative level of expression in various tissues, reverse transcription-PCR was carried out using gene-specific primers for each of the Arabidopsis annexin genes. In addition, northern blot analysis using gene-specific probes indicates differences in AnnAt1 and AnnAt2 expression levels in different tissues. AnnAt1 is expressed in all tissues examined and is most abundant in stems, whereas AnnAt2 is expressed mainly in root tissue and to a lesser extent in stems and flowers. In situ RNA localization demonstrates that these two annexin genes display developmentally regulated tissue-specific and cell-specific expression patterns. These patterns are both distinct and overlapping. The developmental expression patterns for both annexins provide further support for the hypothesis that annexins are involved in the Golgi-mediated secretion of polysaccharides.

Non-NASA Center↗

Origin and early evolution of photosynthesis

Photosynthesis was well-established on the earth at least 3.5 thousand million years ago, and it is widely believed that these ancient organisms had similar metabolic capabilities to modern cyanobacteria. This requires that development of two photosystems and the oxygen evolution capability occurred very early in the earth's history, and that a presumed phase of evolution involving non-oxygen evolving photosynthetic organisms took place even earlier. The evolutionary relationships of the reaction center complexes found in all the classes of currently existing organisms have been analyzed using sequence analysis and biophysical measurements. The results indicate that all reaction centers fall into two basic groups, those with pheophytin and a pair of quinones as early acceptors, and those with iron sulfur clusters as early acceptors. No simple linear branching evolutionary scheme can account for the distribution patterns of reaction centers in existing photosynthetic organisms, and lateral transfer of genetic information is considered as a likely possibility. Possible scenarios for the development of primitive reaction centers into the heterodimeric protein structures found in existing reaction centers and for the development of organisms with two linked photosystems are presented.

NASA Program Exobiology↗

Genomic dissection of anthracnose resistance response in sorghum [ Sorghum bicolor (L.) Moench]

Sorghum [ Sorghum bicolor (L.) Moench] is the fifth most important grain crop behind maize, wheat, rice, and barley. Today, it is of interest as a source of fermentable sugars for the production of renewable fuels and chemicals, and as a source of biomass for co-firing. The productivity and profitability of sorghum are limited by several biotic constraints, most notably anthracnose caused by the fungal pathogen Colletotrichum sublineolum . The most cost-effective and environmentally benign strategy to control anthracnose is through the incorporation of resistance genes. Over the last three years, our research efforts have been directed to identify new sources of resistance in temperate adapted and tropical germplasm, and to delimited genomic regions associated with the observe anthracnose resistant response. Three biparental mapping populations derived from the resistant lines SC112-14, QL3 and IS18760 were evaluated for anthracnose resistance response in Texas, Georgia, Florida and Puerto Rico. In parallel, three high density recombination maps were constructed and used to identify resistant loci. Anthracnose resistant response in line SC112-14 is controlled by a major locus on chromosome 5. Segregation analysis of 1,500 progenies delimited the resistance locus on chromosome 5 to a 23-kb region harboring three candidate genes, including Sobic.005G17230 identified by GWAS of the sorghum association panel (SAP). The latter gene belongs to a family of genes encoding F-box proteins indicating that this resistance response involved in signaling cascades and transcriptional reprograming, rather than recognition of pathotype-associated molecular patterns. In contrast, anthracnose resistant response in lines QL3 and IS18760 is controlled by multiple small-effect genes. Greenhouse evaluation of a representative subset of the three mapping populations against nine pathotypes found that lines susceptible in the field could be resistant to a single pathotype in the greenhouse. Thus, the activation of a resistance response system by a single pathotype could not provide a broader resistance response against multiple pathotypes. The screening of 1,801 sweet sorghum accessions from the National Plant Germplasm System identified 654 accessions with Brix value larger than 10, which in turn was used to select a subset of 233 accessions for evaluation of anthracnose resistant response. Even though most of the accessions were not completely infected by anthracnose, 28 accessions were completely resistant against pathotypes from Texas, Georgia, Florida and Puerto Rico. Genotyping-by-sequencing analysis of this subset identified 157,843 single nucleotide polymorphisms. Population structure analysis of the subset based on a subset of 2,345 unlinked SNPs found that the genetic diversity could be divided into four populations. The genetic relatedness among accessions within populations suggests most of the resistant germplasm may contain few different resistance sources. These resistance sources present in sweet sorghum germplasm could expedite the development of new resistant sweet sorghum cultivars and hybrids by avoiding time-consuming introgression breeding approaches with non-sweet sorghums serving as donor of the resistance alleles.

59 BASIC BIOLOGICAL SCIENCES↗

Genomic dissection of anthracnose resistance response in sorghum [Sorghum bicolor (L.) Moench]

Sorghum [Sorghum bicolor (L.) Moench] is the fifth most important grain crop behind maize, wheat, rice, and barley. Today, it is of interest as a source of fermentable sugars for the production of renewable fuels and chemicals, and as a source of biomass for co-firing. The productivity and profitability of sorghum are limited by several biotic constraints, most notably anthracnose caused by the fungal pathogen Colletotrichum sublineolum. The most cost-effective and environmentally benign strategy to control anthracnose is through the incorporation of resistance genes. Over the last three years, our research efforts have been directed to identify new sources of resistance in temperate adapted and tropical germplasm, and to delimited genomic regions associated with the observe anthracnose resistant response. Three biparental mapping populations derived from the resistant lines SC112-14, QL3 and IS18760 were evaluated for anthracnose resistance response in Texas, Georgia, Florida and Puerto Rico. In parallel, three high density recombination maps were constructed and used to identify resistant loci. Anthracnose resistant response in line SC112-14 is controlled by a major locus on chromosome 5. Segregation analysis of 1,500 progenies delimited the resistance locus on chromosome 5 to a 23-kb region harboring three candidate genes, including Sobic.005G17230 identified by GWAS of the sorghum association panel (SAP). The latter gene belongs to a family of genes encoding F-box proteins indicating that this resistance response involved in signaling cascades and transcriptional reprograming, rather than recognition of pathotype-associated molecular patterns. In contrast, anthracnose resistant response in lines QL3 and IS18760 is controlled by multiple small-effect genes. Greenhouse evaluation of a representative subset of the three mapping populations against nine pathotypes found that lines susceptible in the field could be resistant to a single pathotype in the greenhouse. Thus, the activation of a resistance response system by a single pathotype could not provide a broader resistance response against multiple pathotypes. The screening of 1,801 sweet sorghum accessions from the National Plant Germplasm System identified 654 accessions with Brix value larger than 10, which in turn was used to select a subset of 233 accessions for evaluation of anthracnose resistant response. Even though most of the accessions were not completely infected by anthracnose, 28 accessions were completely resistant against pathotypes from Texas, Georgia, Florida and Puerto Rico. Genotyping-by-sequencing analysis of this subset identified 157,843 single nucleotide polymorphisms. Population structure analysis of the subset based on a subset of 2,345 unlinked SNPs found that the genetic diversity could be divided into four populations. The genetic relatedness among accessions within populations suggests most of the resistant germplasm may contain few different resistance sources. These resistance sources present in sweet sorghum germplasm could expedite the development of new resistant sweet sorghum cultivars and hybrids by avoiding time-consuming introgression breeding approaches with non-sweet sorghums serving as donor of the resistance alleles.

59 BASIC BIOLOGICAL SCIENCES↗

Genomic dissection of anthracnose resistance response in sorghum [ Sorghum bicolor (L.) Moench]

Sorghum [Sorghum bicolor (L.) Moench] is the fifth most important grain crop behind maize, wheat, rice, and barley. Today, it is of interest as a source of fermentable sugars for the production of renewable fuels and chemicals, and as a source of biomass for co-firing. The productivity and profitability of sorghum are limited by several biotic constraints, most notably anthracnose caused by the fungal pathogen Colletotrichum sublineolum. The most cost-effective and environmentally benign strategy to control anthracnose is through the incorporation of resistance genes. Over the last three years, our research efforts have been directed to identify new sources of resistance in temperate adapted and tropical germplasm, and to delimited genomic regions associated with the observe anthracnose resistant response. Three biparental mapping populations derived from the resistant lines SC112-14, QL3 and IS18760 were evaluated for anthracnose resistance response in Texas, Georgia, Florida and Puerto Rico. In parallel, three high density recombination maps were constructed and used to identify resistant loci. Anthracnose resistant response in line SC112-14 is controlled by a major locus on chromosome 5. Segregation analysis of 1,500 progenies delimited the resistance locus on chromosome 5 to a 23-kb region harboring three candidate genes, including Sobic.005G17230 identified by GWAS of the sorghum association panel (SAP). The latter gene belongs to a family of genes encoding F-box proteins indicating that this resistance response involved in signaling cascades and transcriptional reprograming, rather than recognition of pathotype-associated molecular patterns. In contrast, anthracnose resistant response in lines QL3 and IS18760 is controlled by multiple small-effect genes. Greenhouse evaluation of a representative subset of the three mapping populations against nine pathotypes found that lines susceptible in the field could be resistant to a single pathotype in the greenhouse. Thus, the activation of a resistance response system by a single pathotype could not provide a broader resistance response against multiple pathotypes. The screening of 1,801 sweet sorghum accessions from the National Plant Germplasm System identified 654 accessions with Brix value larger than 10, which in turn was used to select a subset of 233 accessions for evaluation of anthracnose resistant response. Even though most of the accessions were not completely infected by anthracnose, 28 accessions were completely resistant against pathotypes from Texas, Georgia, Florida and Puerto Rico. Genotyping-by-sequencing analysis of this subset identified 157,843 single nucleotide polymorphisms. Population structure analysis of the subset based on a subset of 2,345 unlinked SNPs found that the genetic diversity could be divided into four populations. The genetic relatedness among accessions within populations suggests most of the resistant germplasm may contain few different resistance sources. These resistance sources present in sweet sorghum germplasm could expedite the development of new resistant sweet sorghum cultivars and hybrids by avoiding time-consuming introgression breeding approaches with non-sweet sorghums serving as donor of the resistance alleles.

09 BIOMASS FUELS↗

Universal Human Papillomavirus Typing Assay: Whole-Genome Sequencing following Target Enrichment

We designed a universal human papillomavirus (HPV) typing assay based on target enrichment and whole-genome sequencing (eWGS). The RNA bait included 23,941 probes targeting 191 HPV types and 12 probes targeting beta-globin as a control. We used the Agilent SureSelect XT2 protocol for library preparation, Illumina HiSeq 2500 for sequencing, and CLC Genomics Workbench for sequence analysis. Mapping stringency for type assignment was determined based on 8 (6 HPV-positive and 2 HPV-negative) control samples. Using the optimal mapping conditions, types were assigned to 24 blinded samples. eWGS results were 100% concordant with Linear Array (LA) genotyping results for 9 plasmid samples and fully or partially concordant for 9 of the 15 cervical-vaginal samples, with 95.83% overall type-specific concordance for LA genotyping. eWGS identified 7 HPV types not included in the LA genotyping. Since this method does not involve degenerate primers targeting HPV genomic regions, PCR bias in genotype detection is minimized. With further refinements aimed at reducing cost and increasing throughput, this first application of eWGS for universal HPV typing could be a useful method to elucidate HPV epidemiology.

59 BASIC BIOLOGICAL SCIENCES↗

Exome sequencing of hepatocellular carcinomas identifies new mutational signatures and potential therapeutic targets

Our genomic analyses promise to improve tumor characterization to optimize personalized treatment for patients with hepatocellular carcinoma (HCC). Exome sequencing analysis of 243 liver tumors identified mutational signatures associated with specific risk factors, mainly combined alcohol and tobacco consumption and exposure to aflatoxin B1. We identified 161 putative driver genes associated with 11 recurrently altered pathways. Associations of mutations defined 3 groups of genes related to risk factors and centered on CTNNB1 (alcohol), TP53 (hepatitis B virus, HBV) and AXIN1. These analyses according to tumor stage progression identified TERT promoter mutation as an early event, whereasFGF3, FGF4, FGF19 or CCND1 amplification and TP53 and CDKN2A alterations appeared at more advanced stages in aggressive tumors. In 28% of the tumors, we identified genetic alterations potentially targetable by US Food and Drug Administration (FDA)–approved drugs. Finally, we identified risk factor–specific mutational signatures and defined the extensive landscape of altered genes and pathways in HCC, which will be useful to design clinical trials for targeted therapy.

59 BASIC BIOLOGICAL SCIENCES↗

Formation of less-known structurally complex ζ{sub b} and orthorhombic quasicrystalline approximant ε{sub n} on solidification of selected Al–Pd–Cr alloys

The evolution of phases was investigated on cooling of Al{sub 71}Pd{sub 24}Cr{sub 5}, Al{sub 73}Pd{sub 20}Cr{sub 7}, and Al{sub 73}Pd{sub 23}Cr{sub 4} alloys from 1350 °C down to ambient temperature with the rate of 10 °C·min{sup −1}. To perform the investigation, differential thermal analysis, synchrotron X-ray powder diffraction, and scanning electron microscopy coupled with energy-dispersive X-ray spectroscopy were used. In all the investigated alloys structurally complex phases ε{sub n} (ε{sub 6} + ε{sub 28}) and ζ{sub b}, as well as the δ-phase were identified. Based on the results of differential thermal analysis, sequences of phase transformations were determined. The Al{sub 71}Pd{sub 24}Cr{sub 5} alloy started to solidify at 1031.4 °C through β. Primary dendrites of ζ{sub b} were observed in Al{sub 73}Pd{sub 20}Cr{sub 7} and Al{sub 73}Pd{sub 23}Cr{sub 4} alloys. In the second step of solidification δ and/or ζ{sub b} were formed. The peritectic reaction, liquid + ζ{sub b} + δ → ε{sub n} + ζ{sub b} + δ, leading to the formation of the quasicrystalline approximant ε{sub n} (ε{sub 6} + ε{sub 28}) took place in the final step of solidification at approximately 792 °C. - Highlights: • Structurally complex ε{sub n} (ε{sub 6} + ε{sub 28}), ζ{sub b} and δ-phases were identified. • The Al{sub 71}Pd{sub 24}Cr{sub 5} alloy started to solidify at 1031.4 °C through the primary β phase. • The Al{sub 73}Pd{sub 20}Cr{sub 7} and Al{sub 73}Pd{sub 23}Cr{sub 4} alloys solidified in the same way. • The quasicrystalline approximant ε{sub n} (ε{sub 6} + ε{sub 28}) was formed at approximately 792 °C.

36 MATERIALS SCIENCE↗

Whole-Genome Sequencing and Bioinformatic Analysis of Environmental, Agricultural, and Human Campylobacter jejuni Isolates From East Tennessee

As a leading cause of bacterial-derived gastroenteritis worldwide, Campylobacter jejuni has a significant impact on human health in both the developed and developing worlds. Despite its prevalence as a human pathogen, the source of these infections remains poorly understood due to the mutation frequency of the organism and past limitations of whole genome analysis. Recent advances in both whole genome sequencing and computational methods have allowed for the high-resolution analysis of intraspecies diversity, leading multiple groups to postulate that these approaches may be used to identify the sources of Campylobacter jejuni infection. To address this hypothesis, our group conducted a regionally and temporally restricted sampling of agricultural and environmental Campylobacter sources and compared isolated C. jejuni genomes to those that caused human infections in the same region during the same time period. Through a network analysis comparing genomes from various sources, we found that human C. jejuni isolates clustered with those isolated from cattle and chickens, indicating these as potential sources of human infection in the region.

59 BASIC BIOLOGICAL SCIENCES↗

Chromosomal mutations in Escherichia coli that improve tolerance to nonvolatile side–products from dilute acid treatment of sugarcane bagasse

Lignocellulosic biomass provides attractive non-food carbohydrates for production of ethanol and dilute acid pretreatment is a biomass-independent process for access to these carbohydrates. However, this pretreatment also releases volatile and nonvolatile inhibitors of fermenting microorganisms. To identify unique gene products contributing to sensitivity/tolerance to non- volatile inhibitors, ethanologenic Escherichia coli strain LY180 was adapted for growth in vacuum-treated sugarcane bagasse acid hydrolysate (VBHz) lacking furfural and other volatile inhibitors. A mutant, strain AQ15, obtained after about 500 generations of growth in VBHz, grew and fermented the sugars in a medium with 50% VBHz. Comparative genome sequence analysis of strains AQ15 and LY180 revealed 95 mutations in strain AQ15. Six of these mutations were also found in strain SL112, an independent inhibitor-tolerant derivative of strain LY180. Among these six mutations, null mutations in mdh and bacA were identified as contributing factors to VBHz tolerance in strain AQ15, based on genetic and physiological analysis. Deletion of either gene in strain LY180 increased tolerance to VBHz from about 30% to 50% (v/v). Here, considering the location and physiological role of the two enzymes in the cell, it is likely that the two enzymes contribute to VBHz sensitivity of ethanologenic E. coli by different mechanisms.

59 BASIC BIOLOGICAL SCIENCES↗

Transcriptome and phenotyping analyses support a role for chloroplast sigma factor 2 in red‐light‐dependent regulation of growth, stress, and photosynthesis

Abstract Sigma factor ( SIG ) proteins contribute to promoter specificity of the plastid‐encoded RNA polymerase during chloroplast genome transcription. All six members of the SIG family, that is, SIG 1– SIG 6, are nuclear‐encoded proteins targeted to chloroplasts. Sigma factor 2 ( SIG 2) is a phytochrome‐regulated protein important for stoichiometric control of the expression of plastid‐ and nuclear‐encoded genes that impact plastid development and plant growth and development. Among SIG factors, SIG 2 is required not only for transcription of chloroplast genes (i.e., anterograde signaling), but also impacts nuclear‐encoded, photosynthesis‐related, and light signaling‐related genes (i.e., retrograde signaling) in response to plastid functional status. Although SIG 2 is involved in photomorphogenesis in Arabidopsis, the molecular bases for its role in light signaling that impacts photomorphogenesis and aspects of photosynthesis have only recently begun to be investigated. Previously, we reported that SIG 2 is necessary for phytochrome‐mediated photomorphogenesis specifically under red (R) and far‐red light, thereby suggesting a link between phytochromes and nuclear‐encoded SIG 2 in light signaling. To explore transcriptional roles of SIG 2 in R‐dependent growth and development, we performed RNA sequencing analysis to compare gene expression in sig2‐2 mutant and Col‐0 wild‐type seedlings at two developmental stages (1‐ and 7‐day). We identified a subset of misregulated genes involved in growth, hormonal cross talk, stress responses, and photosynthesis. To investigate the functional relevance of these gene expression analyses, we performed several comparative phenotyping tests. In these analyses, strong sig2 mutants showed insensitivity to bioactive GA 3 , high intracellular levels of hydrogen peroxide (H 2 O 2 ) indicative of a stress response, and specific defects in photosynthesis, including elevated levels of cyclic electron flow ( CEF ) and nonphotochemical quenching ( NPQ ). We demonstrated that SIG 2 regulates a broader range of physiological responses at the molecular level than previously reported, with specific roles in red‐light‐mediated photomorphogenesis.

Oh, Sookyung↗

Registration of the sorghum carbon–partitioning nested association mapping (CP–NAM) population

The sorghum [ Sorghum bicolor (L.) Moench] carbon-partitioning nested association mapping (CP_NAM) (Reg. no. MP-4, NSL 542189 MAP) population was developed at Clemson University, SC, using 11 diverse, male founder accessions, each crossed with a recurrent female parent ‘Grassl’. The male parents represent all five major botanical races and the four major agronomic types: cellulosic (5), sweet (3), grain (2) and forage (1). A set of 11 recombinant inbred line (RIL) families CP_NAM01 to CP_NAM011 were maintained, which consisted of 2,484 (F 6 ) individuals. Each RIL family contained a minimum of 193 individuals (CP_NAM01) and a maximum of 287 individuals (CP_NAM06). For the development of this population, the founder lines were judiciously selected from the sorghum Bioenergy Association Panel based on carbon-partitioning phenotypes that make this population an ideal genetic resource for dissecting a wide range of agronomic and compositional traits for basic and applied research. The founder accessions of the CP_NAM were phenotypically characterized for various traits, including agronomic, biomass and related components, and additional compositional components. Each of the 11 F 6 RIL families of the CP_NAM were genotyped using genotyping-by-sequencing analysis, and 144,087 single nucleotide polymorphisms were generated for each individual. Genotypic information along with phenotypic data were used for the characterization of this population and to explore the range of phenotypes that permits the understanding of carbon-partitioning dynamics. This population is a unique resource for researchers to study a wide range of contrasting carbon-partitioning characteristics in sorghum to understand the genetic architecture underlying whole-plant carbon partitioning and allocation.

59 BASIC BIOLOGICAL SCIENCES↗