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Antibiotic resistance genes (ARGs) are emerging environmental contaminants of concern to both human and animal health. Dairy manure is considered reservoir of ARGs. This study is focused on investigating prevalence of ARGs in California dairy farm manure under current common manure management. A total of 33 manure samples were collected from multiple manure treatment conditions: 1) flushed manure (FM), 2) fresh pile (FP), 3) compost pile (CP), 4) primary lagoon (PL), and 5) secondary lagoon (SL). After DNA extraction, all fecal samples were screened by PCR for the presence of 8 ARGs: four sulfonamide ARGs (sulI, sulII, sulIII, sulA), two tetracycline ARGs (tetW, tetO), two macrolide-lincosamide-streptogramin B (MLSB) ARGs (ermB, ermF). Samples were also screened for two mobile genetic elements (MGEs) (intI1, tnpA), which are responsible for dissemination of ARGs. Quantitative PCR was then used to screen all samples for five ARGs (sulII, tetW, ermF, tnpA and intI1). Prevalence of genes varied among sample types, but all genes were detectable in different manure types. Results showed that liquid-solid separation, piling, and lagoon conditions had limited effects on reducing ARGs and MGEs, and the effect was only found significant on tetW (p = 0.01). Besides, network analysis indicated that sulII was associated with tnpA (p Psychrobacter and Pseudomonas as opportunistic human pathogens, were potential ARG/MGE hosts (p

A geographically isolated maize landrace cultivated on nitrogen-depleted fields without synthetic fertilizer in the Sierra Mixe region of Oaxaca, Mexico utilizes nitrogen derived from the atmosphere and develops an extensive network of mucilage-secreting aerial roots that harbors a diazotrophic microbiota. Targeting these diazotrophs, we selected nearly 600 microbes from a collection isolated from these plants and confirmed their ability to incorporate heavy nitrogen (15N2) metabolites in vitro. Sequencing their genomes and conducting comparative bioinformatic analyses showed that these genomes had substantial phylogenetic diversity. We examined each diazotroph genome for the presence of nif genes essential to nitrogen fixation (nifHDKENB) and carbohydrate utilization genes relevant to the mucilage polysaccharide digestion. These analyses identified diazotrophs that possessed canonical nif gene operons, as well as many other operon configurations with concomitant fixation and release of >700 different 15N labeled metabolites. We further demonstrated that many diazotrophs possessed alternative nif gene operons and confirmed their genomic potential to derive chemical energy from mucilage polysaccharide to fuel nitrogen fixation. These results confirm that some diazotrophic bacteria associated with Sierra Mixe maize were capable of incorporating atmospheric nitrogen into their small molecule extracellular metabolites through multiple nif gene configurations while others were able to fix nitrogen without the canonical (nifHDKENB) genes.Data SummaryGenetic resources, including biological materials and nucleic acid sequences, were accessed under an Access and Benefit Sharing (ABS) Agreement between the Sierra Mixe community and the Mars Corporation, and with authorization from the Mexican government. An internationally recognized certificate of compliance has been issued by the Mexican government under the Nagoya Protocol for such activities (ABSCH-IRCC-MX-207343-3). Any party seeking access to the nucleic acid sequences underlying the analysis reported here is subject to the full terms and obligations of the ABS agreement and the authorization from the government of Mexico. Individuals wishing to access nucleic acid sequence data for scientific research activities should contact Mars Incorporated Chief Science Officer at CSO@effem.com.

Sierra Mixe maize is a geographically remote landrace variety grown on nitrogen-deficient fields in Oaxaca, Mexico that meets its nutritional requirements without synthetic fertilizer by associating with free-living diazotrophs comprising the microbiota of its aerial root mucilage. We selected nearly 500 diazotrophic (N2-fixing) bacteria isolated from Sierra Mixe maize mucilage and sequenced their genomes. Comparative genomic analysis demonstrated that isolates represented diverse genera and composed three major diazotrophic groups based on nitrogen fixation gene content. In addition to nitrogen fixation, we examined deamination of 1-amino-1-cyclopropanecarboxylic acid, biosynthesis of indole-3-acetic acid, and phosphate solubilization as alternative mechanisms of direct plant growth promotion (PGP). Genome mining showed that isolates of all diazotrophic groups possessed marker genes for multiple mechanisms of direct plant growth promotion (PGP). Implementing in vitro assays corroborated isolate genotypes by measuring each isolate's potential to confer the targeted PGP traits and revealed phenotypic variation among isolates based on diazotrophic group assignment. Investigating the ability of mucilage diazotrophs to confer PGP by direct inoculation of clonally propagated potato plants in planta led to the identification of 16 bio-stimulant candidates. Conducting nitrogen-stress greenhouse experiments demonstrated that potato inoculation with a synthetic community of bio-stimulant candidates, as well as with its individual components, resulted in PGP phenotypes. We further demonstrated that one diazotrophic isolate conferred PGP to a conventional maize variety under nitrogen-stress in the greenhouse. These results indicate that, while many diazotrophic isolates from Sierra Mixe maize possessed genotypes and in vitro phenotypes for targeted PGP traits, a subset of these organisms promoted the growth of potato and conventional maize, potentially through the use of multiple promotion mechanisms.

Dairy farms generate a considerable amount of manure, which is applied in cropland as fertilizer. While the use of manure as fertilizer reduces the application of chemical fertilizers, the main concern with regards to manure application is microbial pollution. Manure is a reservoir of a broad range of microbial populations, including pathogens, which have potential to cause contamination and pose risks to public and animal health. Despite the widespread use of manure fertilizer, the change in microbial diversity of manure under various treatment processes is still not well-understood. We hypothesize that the microbial population of animal waste changes with manure handling used in a farm environment. Consequential microbial risk caused by animal manure may depend on manure handling. In this study, a reconnaissance effort for sampling dairy manure in California Central Valley followed by 16S rRNA analysis of content and diversity was undertaken to understand the microbiome of manure after various handling processes. The microbial community analysis of manure revealed that the population in liquid manure differs from that in solid manure. For instance, the bacteria of genus Sulfuriomonas were unique in liquid samples, while the bacteria of genus Thermos were observed only in solid samples. Bacteria of genus Clostridium were present in both solid and liquid samples. The population among liquid samples was comparable, as was the population among solid samples. These findings suggest that the mode of manure application (i.e., liquid versus solid) could have a potential impact on the microbiome of cropland receiving manure as fertilizers.

Lipid profiles in wheat leaves and the effects of tan spot on the profiles were quantified by mass spectrometry. Inoculation with Pyrenophora tritici-repentis significantly reduced the amount of leaf lipids, including the major plastidic lipids monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), which together accounted for 89% of the mass spectral signal of detected lipids in wheat leaves. Levels of these lipids in susceptible cultivars dropped much more quickly during infection than those in resistant cultivars. Furthermore, cultivars resistant or susceptible to tan spot displayed different lipid profiles; leaves of resistant cultivars had more MGDG and DGDG than susceptible ones, even in noninoculated plants. Lipid compositional data from leaves of 20 noninoculated winter wheat cultivars were regressed against an index of disease susceptibility and fitted with a linear model. This analysis demonstrated a significant relationship between resistance and levels of plastidic galactolipids and indicated that cultivars with high resistance to tan spot uniformly had more MGDG and DGDG than cultivars with high susceptibility. These findings suggest that lipid composition of wheat leaves may be a determining factor in the resistance response of cultivars to tan spot.

Complex glycans cover the gut epithelial surface to protect the cell from the environment. Invasive pathogens must breach the glycan layer before initiating infection. While glycan degradation is crucial for infection, this process is inadequately understood. Salmonella contains 47 glycosyl hydrolases (GHs) that may degrade the glycan. We hypothesized that keystone genes from the entire GH complement of Salmonella are required to degrade glycans to change infection. This study determined that GHs recognize the terminal monosaccharides (N-acetylneuraminic acid (Neu5Ac), galactose, mannose, and fucose) and significantly (p Salmonella used its two GHs sialidase nanH and amylase malS for internalization by targeting different glycan structures. The host glycans were altered during Salmonella association via the induction of N-glycan biosynthesis pathways leading to modification of host glycans by increasing fucosylation and mannose content, while decreasing sialylation. Gene expression analysis indicated that the host cell responded by regulating more than 50 genes resulting in remodeled glycans in response to Salmonella treatment. This study established the glycan structures on colonic epithelial cells, determined that Salmonella required two keystone GHs for internalization, and left remodeled host glycans as a result of infection. These data indicate that microbial GHs are undiscovered virulence factors.

Lipid biomarker profiles are a powerful tool for assessing soil microbial community structure, but intensive laboratory work and data analysis are needed to construct profiles from phospholipid fatty acids and other common biomarkers. Pyrolysis mass spectrometry (Py-MS) is a alternative method that provides a rapid and sensitive ‘fingerprint’ of soil lipids and may be sufficient to characterize lipids from various sites. The objective of this work was to evaluate the capacity of pyrolysis metastable atom bombardment time-of-flight mass spectrometry (Py-MAB-TOF-MS) to provide replicable analysis of soil lipids, compared to a routine gas chromatography-flame ionization detection (GC-FID) method. Soils were collected from six agricultural fields under soybean, corn and asparagus production. Soil lipids extracted with 1:2 chloroform:methanol solvent were analyzed with Py-MAB-TOF-MS or transesterified into fatty acids and then analyzed by GC-FID. The two methods were complementary, but distinct: lipid fingerprints, generated from Py-MAB-TOF-MS spectra, included extractable soil lipids from microbial, animal and plant origins plus non-living organic matter in the samples, whereas fatty acid profiles generally represented lipids from soil bacteria and fungi. We conclude that the soil lipid fingerprints generated from Py-MAB-TOF-MS present more variability than lipid biomarker profiles from the GC-FID method because they include a broader group of extractable soil lipids. Further work is needed to identify the molecular fragment masses in Py-MAB-TOF-MS spectra that could precisely identify soil lipids of microbial origin.

Plants are associated with a complex microbiota that contributes to nutrient acquisition, plant growth, and plant defense. Nitrogen-fixing microbial associations are efficient and well characterized in legumes but are limited in cereals, including maize. We studied an indigenous landrace of maize grown in nitrogen-depleted soils in the Sierra Mixe region of Oaxaca, Mexico. This landrace is characterized by the extensive development of aerial roots that secrete a carbohydrate-rich mucilage. Analysis of the mucilage microbiota indicated that it was enriched in taxa for which many known species are diazotrophic, was enriched for homologs of genes encoding nitrogenase subunits, and harbored active nitrogenase activity as assessed by acetylene reduction and 15N2 incorporation assays. Field experiments in Sierra Mixe using 15N natural abundance or 15N-enrichment assessments over 5 years indicated that atmospheric nitrogen fixation contributed 29%–82% of the nitrogen nutrition of Sierra Mixe maize., Author summary Nitrogen is an essential nutrient for plants, and for many nonlegume crops, the requirement for nitrogen is primarily met by the use of inorganic fertilizers. These fertilizers are produced from fossil fuel by energy-intensive processes that are estimated to use 1% to 2% of the total global energy supply and produce an equivalent share of greenhouse gases. Because maize (Zea mays L.) is a significant recipient of nitrogen fertilization, a research goal for decades has been to identify or engineer mechanisms for biological fixation of atmospheric nitrogen in association with this crop. We hypothesized that isolated indigenous landraces of maize grown using traditional practices with little or no fertilizer might have evolved strategies to improve plant performance under low-nitrogen nutrient conditions. Here, we show that for one such maize landrace grown in nitrogen-depleted fields near Oaxaca, Mexico, 29%–82% of the plant nitrogen is derived from atmospheric nitrogen. High levels of nitrogen fixation are supported, at least in part, by the abundant production of a sugar-rich mucilage associated with aerial roots that provides a home to a complex nitrogen-fixing microbiome.

Members of the Fusarium genus are important components of many plant–soil systems worldwide and are responsible for many crop diseases. Knowledge of the relative influence of biotic and abiotic factors on this genus is therefore of broad economic and ecological importance. In order to address this issue, we examined Fusarium communities in soils nearby apparently healthy and symptomatic asparagus plants in 50 fields scattered in four agricultural regions of Quebec, Canada. Fusarium community structure and abundance were assessed using genus-specific PCR-denaturing gradient gel electrophoresis and CFU counts, respectively. Multivariate statistical analyses were used to detect community patterns related to spatial, abiotic and biotic factors. Results suggested that Fusarium community structure (i.e. the presence and absence of the different Fusarium sequence variants in the samples) in soil is mainly related to biotic factors (arbuscular mycorrhizal fungi and bacterial community structure), whereas Fusarium abundance is more closely related to abiotic factors (mainly clay, organic matter, NH4, Na and Cu). Some degree of influence of spatial patterns was also observed on both Fusarium community structure and abundance with, for instance, a large regional variation in Fusarium community structure. However, Fusarium community structure was not directly related to the disease status of nearby asparagus plants.

Cool-season turfgrasses may experience heat stress during summer. Hybrid bluegrasses (HBGs), crosses between kentucky bluegrass [KBG (Poa pratensis L.)] and native texas bluegrass (Poa arachnifera Torr.), have improved heat tolerance but the mechanisms of heat tolerance are poorly understood. Our objectives were to quantitatively profile membrane lipid molecular species in three cool-season turfgrasses exposed to optimal (22/15 °C, 14/10 h light/dark) and supra-optimal temperatures (35/25 °C and 40/30 °C, 14/10 h light/dark). Grasses included a low heat-tolerant tall fescue [TF (Festuca arundinacea Schreb. ‘Dynasty’)], a mid-heat–tolerant KBG (‘Apollo’), and a heat-tolerant HBG (‘Thermal Blue’). At high temperature, glycolipid digalactosyldiacylglycerol (DGDG) in HBG was 12% and 16% greater than in KBG and TF, respectively, and the ratio DGDG to monogalactosyldiacylglycerol was 19% and 44% greater in HBG than in KBG and TF, respectively. Greater heat tolerance in HBG and KBG was associated with higher contents of phosphatidylethanolamine and phosphatidylglycerol, and with reduced overall unsaturation compared with TF. Overall, 20 lipid molecular species were present in greater amounts and another 20 species in lesser amounts in HBG and KBG than in TF. Results suggest 40 membrane lipid molecules are potential biomarkers for heat tolerance and that compositional changes in membrane lipids in response to heat contribute to differences in heat tolerance among cool-season grasses.

The extraction and transesterification of soil lipids into fatty acid methyl esters (FAMEs) is a useful technique for studying soil microbial communities. The objective of this study was to find the best solvent mixture to extract soil lipids with a pressurized solvent extractor system. Four solvent mixtures were selected for testing: chloroform:methanol:phosphate buffer (1:2:0.8, v/v/v), chloroform:methanol (1:2, v/v), hexane:2-propanol (3:2, v/v) and acetone. Soils were from agricultural fields and had a wide range of clay, organic matter and microbial biomass contents. Total lipid fatty acid methyl esters (TL-FAMEs) were the extractable soil lipids identified and quantified with gas chromatography and flame ionization detection. Concentrations of TL-FAMEs ranged from 57.3 to 542.2 nmole g(-1) soil (dry weight basis). The highest concentrations of TL-FAMEs were extracted with chloroform:methanol:buffer or chloroform:methanol mixtures than with the hexane:2-propanol or acetone solvents. The concentrations of TL-FAMEs in chemical groups, including saturated, branched, mono- and poly-unsaturated and hydroxy fatty acids were assessed, and biological groups (soil bacteria, mycorrhizal fungi, saprophytic fungi and higher plants) was distinguished. The extraction efficiency for the chemical and biological groups followed the general trend of: chloroform:methanol:buffer> or =chloroform:methanol>hexane:2-propanol=acetone. Discriminant analysis revealed differences in TL-FAME profiles based on the solvent mixture and the soil type. Although solvent mixtures containing chloroform and methanol were the most efficient for extracting lipids from the agricultural soils in this study, soil properties and the lipid groups to be studied should be considered when selecting a solvent mixture. According to our knowledge, this is the first report of soil lipid extraction with hexane:2-propanol or acetone in a pressurized solvent extraction system.

Tree cultivation in the dryland agroecosystems is increasingly advocated as a strategy to protect and reverse soil fertility decline, thus sustaining agricultural production. Woody legumes trees like the Ana tree Faidherbia albida (Del.) Chev. host N2-fixing bacteria as well as arbuscular mycorrhizal fungi (AMF), which may contribute to the soil organic carbon pool and soil fertility. The objective of this work was to determine how trees influenced AMF and soil fertility in the agroecosystems of two rural communities (Palmarin and Fimela) of the Saloum Agricultural Eco-Region of Senegal. Smallholder farmers typically cultivated 3–4 fields ranging in size from 0.5 to 2.0 ha with the major crop being millet (Pennisetum glaucum L.). Soil fertility was low to medium, with about 1% soil organic matter (SOM), 17±2.2 mg Bray-1 P kg−1 and 65±5.8 mg extractable K kg−1 in the fields studied. There were seven times more trees per hectare and greater tree diversity in fields around Palmarin, where the smallholder farmers resided, than in the fields they cultivated in Fimela. Social norms appeared to protect trees inside the residential village, while trees farther away are prone to being cut. The relationships between trees, AMF and soil fertility were examined using exploratory path analysis, a structural equation modeling technique. The path analysis model revealed a direct and significant (P

Carbon and nitrogen (C/N) metabolism and allocation within the plant have important implications for plant-parasite interactions. Many plant parasites manipulate the host by inducing C/N changes that benefit their own survival and growth. Plant resistance can prevent this parasite manipulation. We used the wheat–Hessian fly (Mayetiola destructor) system to analyze C/N changes in plants during compatible and incompatible interactions. The Hessian fly is an insect but shares many features with plant pathogens, being sessile during feeding stages and having avirulence (Avr) genes that match plant resistance genes in gene-for-gene relationships. Many wheat genes involved in C/N metabolism were differentially regulated in plants during compatible and incompatible interactions. In plants during compatible interactions, the content of free carbon-containing compounds decreased 36%, whereas the content of free nitrogen-containing compounds increased 46%. This C/N shift was likely achieved through a coordinated regulation of genes in a number of central metabolic pathways, including glycolysis, the tricarboxylic acid cycle, and amino-acid synthesis. Our data on plants during compatible interactions support recent findings that Hessian fly larvae create nutritive cells at feeding (attack) sites and manipulate host plants to enhance their own survival and growth. In plants during incompatible interactions, most of the metabolic genes examined were not affected or down-regulated.

Cellulose synthase complexes (CSCs) at the plasma membrane (PM) are aligned with cortical microtubules (MTs) and direct the biosynthesis of cellulose. The mechanism of the interaction between CSCs and MTs, and the cellular determinants that control the delivery of CSCs at the PM, are not yet well understood. We identified a unique small molecule, CESA TRAFFICKING INHIBITOR (CESTRIN), which reduces cellulose content and alters the anisotropic growth of Arabidopsis (Arabidopsis thaliana) hypocotyls. We monitored the distribution and mobility of fluorescently labeled cellulose synthases (CESAs) in live Arabidopsis cells under chemical exposure to characterize their subcellular effects. CESTRIN reduces the velocity of PM CSCs and causes their accumulation in the cell cortex. The CSC-associated proteins KORRIGAN1 (KOR1) and POM2/CELLULOSE SYNTHASE INTERACTIVE PROTEIN1 (CSI1) were differentially affected by CESTRIN treatment, indicating different forms of association with the PM CSCs. KOR1 accumulated in bodies similar to CESA; however, POM2/CSI1 dissociated into the cytoplasm. In addition, MT stability was altered without direct inhibition of MT polymerization, suggesting a feedback mechanism caused by cellulose interference. The selectivity of CESTRIN was assessed using a variety of subcellular markers for which no morphological effect was observed. The association of CESAs with vesicles decorated by the trans-Golgi network-localized protein SYNTAXIN OF PLANTS61 (SYP61) was increased under CESTRIN treatment, implicating SYP61 compartments in CESA trafficking. The properties of CESTRIN compared with known CESA inhibitors afford unique avenues to study and understand the mechanism under which PM-associated CSCs are maintained and interact with MTs and to dissect their trafficking routes in etiolated hypocotyls.

Slow rates of plant production and decomposition in ombrotrophic bogs are believed to be partially the result of low nutrient availability. To test the effect of nutrient availability on decomposition, carbon dioxide (CO2) flux dynamics, microbial biomass, and nutrients, we added nitrogen (N) with phosphorus (P) and potassium (K), to prevent limitation of the latter 2 nutrients, over 2 growing seasons to plots at Mer Bleue peatland, Ontario, Canada. After the first growing season, increasing N fertilization (with constant P and K) decreased in vitro CO2 production potential and increased microbial biomass measured with a chloroform fumigation-extraction technique in the upper peat profile, while by the end of the second season, CO2 production potential was increased in response to N plus PK treatment, presumably due to more easily decomposable newly formed plant material. In situ CO2 fluxes measured using chamber-techniques over the second

Lipid profiling is a targeted metabolomics platform that provides a comprehensive analysis of lipid species with high sensitivity. Profiling based on electrospray ionization tandem mass spectrometry (ESI-MS/MS) provides quantitative data and is adaptable to high throughput analyses. Here we report the profiling of 140 apparent molecular species of polar glycerolipids in Arabidopsis leaves, flower stalks, flowers, siliques, roots, and seeds. Considerable differences in lipid species occur among these organs, providing insights into the different lipid metabolic activities in a specific organ. In addition, comparative profiling between wild-type and a knockout mutant pldalpha1 (locus ID: AT3G15730) provides insight into the metabolic function of phospholipase D (PLD) in different organs. PLDalpha1 contributes significantly to phosphatidic acid (PA) levels in roots, seeds, flowers, and flower stalks, but little to basal PA levels in siliques and leaves. In seeds of the pldalpha1 mutant plants, levels of PA, lysophosphatidylcholine, and lysophosphatidylethanolamine were significantly lower than those of wild-type seeds, suggesting a role for PLDalpha1 in membrane lipid degradation in seeds.

The diet of earthworms includes soil organic matter, soil microbes and other microfauna, but the relative contribution of these dietary components to earthworm nutrition is not well known. Analysis of fatty acid (FA) profiles can reveal trophic relationships in soil food webs, leading to a better understanding of the energy and nutrient flows from microbiota to earthworms. The objective of this study was to determine the origin of FAs assimilated by the earthworm Lumbricus terrestris L. We analysed the pattern of FAs in: (i) the bulk soil, (ii) soil in the earthworm gut, (iii) the absorptive tissue of the earthworm gut wall, and (iv) the muscular layers of the earthworm body wall. Multivariate analyses performed on the FA profiles suggest that the microbial community in the earthworm gut differs from that in bulk soil. Diverse bacterial and fungal derived FAs, which earthworms cannot synthesize, were found in the earthworm gut wall and body wall, and in the neutral lipids (storage lipids) of the gut wall. The major compounds isolated were 20:4o6, 20:5o3 and 18:2o6, followed by the monoenoic 18:1o7 and 18:1o9c, and the saturated 18:0. The microbial FA assemblage in the gut wall resembled the gut soil more than the bulk soil, and the body wall of L. terrestris showed the same microbial derived FA pattern as the gut wall, although at reduced concentrations. We propose the existence of a specific microbial community in the earthworm gut that provides FAs to the earthworm. It appears that L. terrestris may derive more of its energy and nutrients from gut specific microbiota than from microbiota already present in the ingested soil, based on the trophic relationships revealed through FA analysis. r 2006 Elsevier Ltd. All rights reserved.

The dynamic equilibrium of an ecosystem is driven by mutual feedback interactions between plants and soil microorganisms. Asparagus exerts a particularly strong influence on its soil environment through abundant production of persistent phenolic acids, which impact selectively soil microorganisms and may be involved in Fusarium crown and root rot (FCRR) of asparagus. In a survey of 50 asparagus plantations of the province of Quebec, we found that FCRR was associated with a profound cultivar-specific, reorganization of the soil microbial community, as revealed by phospholipid fatty acid (PLFA) profiling. According to PLFA indicators, microbial biodiversity as well as bacterial and fungal abundance dropped sharply with the onset of FCRR in fields planted with the cultivar Guelph Millenium. This drop was followed by a similar drop in the arbuscular mycorrhizal population. Biodiversity and microbial population size then increased to finally reach a new equilibrium. Discriminant analysis of PLFA profiles obtained from soil samples also indicated a shift in soil microbial community structure associated with FCRR development in fields planted with the cultivar Jersey Giant. Different soil biological conditions, as indicated by microbial biomass C and N and soil enzyme activities, were associated with different cultivars. Preceding crop, manure application, geographical location and tillage depth also influenced the structure of soil microbial communities in asparagus plantations, as determined by PLFA profiling. If higher FCRR incidence is a consequence of the soil microbial community reorganization, means to reduce FCRR incidence in asparagus plantations may be found among practices such as soil organic fertilization, soil tillage and intercropping strategies that would dilute the negative influence of asparagus on the soil microbial community. Finally, FCRR outbreaks were generally promoted by a previous crop of maize. It seems that maize and asparagus host a F. proliferatum teleomorph (Gibberella fujikoroi) of the same mating type.

Epigenetics has an important role for the success of foodborne pathogen persistence in diverse host niches. Substantial challenges exist in determining DNA methylation to situation-specific phenotypic traits. DNA modification, mediated by restriction-modification systems, functions as an immune response against antagonistic external DNA, and bacteriophage-acquired methyltransferases (MTase) and orphan MTases – those lacking the cognate restriction endonuclease – facilitate evolution of new phenotypes via gene expression modulation via DNA and RNA modifications, including methylation and phosphorothioation. Recent establishment of large-scale genome sequencing projects will result in a significant increase in genome availability that will lead to new demands for data analysis including new predictive bioinformatics approaches that can be verified with traditional scientific rigor. Sequencing technologies that detect modification coupled with mass spectrometry to discover new adducts is a powerful tactic to study bacterial epigenetics, which is poised to make novel and far-reaching discoveries that link biological significance and the bacterial epigenome.

Lens opacity 13 (lop13) is a spontaneous, autosomal recessive mouse mutant that exhibits nuclear cataracts. Histological analysis revealed swollen lens fiber cells and the presence of bladder cells within the lens cortex, as well as morgagnian globules and liquefied material at the lens posterior. At 3 months of age, in addition to cataracts, lop13 mice also develop persistent skin wounds. Linkage analysis assigned the lop13 locus to a 1.1-Mb region on mouse Chr 15, encompassing 19 candidate genes. Sequence analysis identified a C3112T mutation in exon 18 of Sterol Regulatory Element Binding-Transcription Factor 2 (Srebf2) resulting in the R1038C substitution of a highly conserved arginine within the Srebf2 regulatory domain. Srebf2 belongs to a family of membrane-bound basic helix-loop-helix leucine zipper transcription factors that control the expression of genes involved in the biosynthesis and uptake of cholesterol and fatty acids. The lack of complementation observed in Srebf2 ( lop13/GT ) compound heterozygotes carrying the Srebf2 gene trapped allele (Srebf2 ( GT )) provides genetic evidence that the identified C3112T substitution in Srebf2 is responsible for the lop13 phenotype. Gas chromatography analysis identified lower levels of cholesterol in the lop13 brain, liver, and lens when compared to wild-type mice. These findings suggest that lop13 is a hypomorphic mutation in Srebf2. As such, the lop13 mouse presents an invaluable in vivo model for studying the contribution of Srebf2 and cholesterol to maintaining the homeostasis of the lens and skin.

Fusarium graminearum is the principal causative agent of Fusarium head blight (FHB), a devastating disease of wheat and barley. This fungus can also colonize Arabidopsis thaliana. Disease resistance was enhanced in transgenic wheat and Arabidopsis plants that constitutively overexpress the NONEXPRESSOR OF PATHOGENESIS-RELATED GENES 1 (NPR1) gene, which regulates salicylic acid (SA) signaling and modulates the activation of jasmonic acid (JA)-dependent defenses. Here, we provide several lines of evidence that reveal an important role for SA and JA signaling in Arabidopsis defense against F. graminearum. SA level was elevated in fungus-inoculated leaves, and SA application and biologically activated systemic acquired resistance enhanced resistance. Furthermore, the disruption of SA accumulation and signaling in the sid2 mutant and NahG transgenic plant, and the npr1 and wrky18 mutants, respectively, resulted in heightened susceptibility to this fungus in leaves and inflorescence. JA signaling was activated in parallel with SA signaling in the fungus-challenged plants. However, the hyperresistance of the JA pathway mutants opr3, coi1, and jar1 indicates that this pathway contributes to susceptibility. Genetic and biochemical experiments indicate that the JA pathway promotes disease by attenuating the activation of SA signaling in fungus-inoculated plants. However, the hypersusceptibility of the jar1 npr1 double mutant compared with the npr1 mutant suggests that JAR1 also contributes to defense, signifying a dichotomous role of JA and a JAR1-dependent mechanism in this interaction.

Exercise has been linked to a reduced cancer risk in animal models. However, the underlying mechanisms are unclear. This study assessed the effect of exercise with dietary consideration on the phospholipid profile in 12-O-tetradecanoylphorbol-13-acetate (TPA)–induced mouse skin tissues. CD-1 mice were randomly assigned to one of the three groups: ad libitum–fed sedentary control; ad libitum–fed treadmill exercise at 13.4 m/min for 60 min/d, 5 d/wk (Ex+AL); and treadmill-exercised but pair-fed with the same amount as the control (Ex+PF). After 14 weeks, Ex+PF but not Ex+AL mice showed ∼25% decrease in both body weight and body fat when compared with the controls. Of the total 338 phospholipids determined by electrospray ionization–tandem mass spectrometry, 57 were significantly changed, and 25 species could distinguish effects of exercise and diet treatments in a stepwise discriminant analysis. A 36% to 75% decrease of phosphatidylinositol (PI) levels in Ex+PF mice occurred along with a significant reduction of PI 3-kinase in TPA-induced skin epidermis, as measured by both Western blotting and immunohistochemistry. In addition, ∼2-fold increase of the long-chain polyunsaturated fatty acids, docosahexaenoic and docosapentaenoic acids, in phosphatidylcholines, phosphatidylethanolamines, and lysophosphatidylethanolamines was observed in the Ex+PF group. Microarray analysis indicated that the expression of fatty acid elongase-1 increased. Taken together, these data indicate that exercise with controlled dietary intake, but not exercise alone, significantly reduced body weight and body fat as well as modified the phospholipid profile, which may contribute to cancer prevention by reducing TPA-induced PI 3-kinase and by enhancing ω-3 fatty acid elongation. Cancer Prev Res; 3(4); 466–77

Members of the Fusarium genus are important components of many plant-soil systems worldwide and are responsible for many crop diseases. Knowledge of the relative influence of biotic and abiotic factors on this genus is therefore of broad economic and ecological importance. In order to address this issue, we examined Fusarium communities in soils nearby apparently healthy and symptomatic asparagus plants in 50 fields scattered in four agricultural regions of Québec, Canada. Fusarium community structure and abundance were assessed using genus-specific PCR-denaturing gradient gel electrophoresis and CFU counts, respectively. Multivariate statistical analyses were used to detect community patterns related to spatial, abiotic and biotic factors. Results suggested that Fusarium community structure (i.e. the presence and absence of the different Fusarium sequence variants in the samples) in soil is mainly related to biotic factors (arbuscular mycorrhizal fungi and bacterial community structure), whereas Fusarium abundance is more closely related to abiotic factors (mainly clay, organic matter, NH(4), Na and Cu). Some degree of influence of spatial patterns was also observed on both Fusarium community structure and abundance with, for instance, a large regional variation in Fusarium community structure. However, Fusarium community structure was not directly related to the disease status of nearby asparagus plants.

Hamel, C., Vujanovic, V., Nakano-Hylander, A., Jeannotte, R., and St-Arnaud, M. 2005. Factors associated with Fusarium crown and root rot of asparagus outbreaks in Quebec. Phytopathology 95:867-873. The Fusarium spp. causing Fusarium crown and root rot (FCRR) are ubiquitous and abundant in soils, but in contrast, disease expression is localized and sporadic. Previous studies have related FCRR infection to phenolic acids released by asparagus, to the repression of Mn-reducers in soil, and to various soil physicochemical conditions. Fifty commercial asparagus plantations were surveyed using an exploratory approach in order to pinpoint the ecological conditions associated with FCRR development. Twenty-eight variables were used to describe the soil environments of the asparagus crops as well as the influence of crop management practices used locally. The data set was analyzed both as a whole and parsed by main cultivars (Jersey Giant and Guelph Millenium). Both field conditions and percentage of field area affected by FCRR varied widely between asparagus plantations. Planting depth was positively correlated with percentage of field area affected by FCRR and, hence, deep planting may favor FCRR infection. Plantation age was positively correlated with percentage of field area affected by FCRR, while soil available Mn was inversely correlated. Most importantly, soil Mn availability decreased with increasing plantation age, supporting the hypothesis of an asparagusmediated negative impact on Mn-reducing bacteria and of the involvement of reduced Mn availability in FCRR development. Improving the availability of Mn could provide a solution to the problem of FCRR in asparagus plantations.

In the past dozen years, many new strategies for mass-spectrometry-based analyses of lipids have been developed. Lipidomics has emerged as a comprehensive approach to analysis of lipids from biological systems, and the most-utilized lipidomics methodologies involve electrospray ionization (ESI) sources and triple quadrupole analyzers. While mass spectral techniques for lipid profiling have advanced, challenges in developing uniform data acquisition methods and in handling, storing, and analyzing mass spectral data remain. Investigation of other ionization methods, including matrix-assisted laser desorption/ionization (MALDI) and atmospheric pressure chemical ionization (APCI), has demonstrated that these are useful in specific applications. APCI is particularly amenable to analysis of less polar lipids, and MALDI provides a rapid technology with application for tissue imaging. Time-of-flight secondary ion mass spectrometry (TOF-SIMS) is particularly suited for imaging of tissues and cells.

In the past dozen years, many new strategies for mass-spectrometry-based analyses of lipids have been developed. Lipidomics has emerged as a comprehensive approach to analysis of lipids from biological systems, and the most-utilized lipidomics methodologies involve electrospray ionization (ESI) sources and triple quadrupole analyzers. While mass spectral techniques for lipid profiling have advanced, challenges in developing uniform data acquisition methods and in handling, storing, and analyzing mass spectral data remain. Investigation of other ionization methods, including matrix-assisted laser desorption/ionization (MALDI) and atmospheric pressure chemical ionization (APCI), has demonstrated that these are useful in specific applications. APCI is particularly amenable to analysis of less polar lipids, and MALDI provides a rapid technology with application for tissue imaging. Time-of-flight secondary ion mass spectrometry (TOF-SIMS) is particularly suited for imaging of tissues and cells.

Quantification of phosphorus (P) concentrations in microbial biomass is required to better understand how P immobilization and turnover in soils are controlled by environmental and anthropogenic factors. Soil microbial biomass P (MBP) is generally extracted using the chloroform fumigation-direct extraction procedure and then analysed for P using the ammonium molybdate-ascorbic acid method on a flow injection analysis (FIA) system. Our objective was to determine whether a microscale malachite green method on a microplate system would provide as accurate MBP analysis as the ascorbic acid method on an FIA system. Twelve soils were collected from agricultural fields in southwestern Quebec, fumigated with chloroform and extracted with 0.5 M NaHCO3 (pH 8.5). The dissolved inorganic phosphorus (DIP) concentration in fumigated soils was not affected by the method of analysis, and results from the two systems of analysis were significantly correlated (r =0.998, P

Convertases of the subtilisin/kexin family are responsible for the biological activation of a variety of pro-proteins, pro-hormones, and pro-trophic factors, and thus can modulate various aspects of embryonic development. We investigated the expression of each convertase by Northern hybridization during cell differentiation in vitro, using the mouse embryonal carcinoma cell line P19 as a model. The neuroendocrine convertase PC2 and 7B2, its specific binding protein, are co-induced during neuronal differentiation of P19 cells with retinoic acid, whereas the other convertases are not or follow different patterns of temporal expression. The mature forms of PC2 and 7B2 proteins are detected together by immunoblotting following induction of mRNA expression, indicating that these proteins are processed early during brain development. These results demonstrate that PC2 and 7B2 gene expression and protein processing are in a close temporal association during neuronal differentiation and point to the value of the P19 cell model to study the significance and the regulation of this relationship in mammalian brain development.

To identify innovative ways for better utilizing flushed dairy manure wastewater, we have assessed the effect of dairy manure and supplementation with synthetic medium on the growth of Chlorella vulgaris. A series of experiments were carried out to study the impacts of pretreatment of dairy wastewater and the benefits of supplementing dairy manure wastewater with synthetic medium on C. vulgaris growth increment and the ultrastructure (chloroplast, starch, lipid, and cell wall) of C. vulgaris cells. Results showed that the biomass production of C. vulgaris in dairy wastewater can be enhanced by pretreatment and using supplementation with synthetic media. A recipe combining pretreated dairy wastewater (40%) and synthetic medium (60%) exhibited an improved growth of C. vulgaris. The effects of dairy wastewater on the ultrastructure of C. vulgaris cells were distinct compared to that of cells grown in synthetic medium. The C. vulgaris growth in both synthetic medium and manure wastewater without supplementing synthetic medium was lower than the growth in dairy manure supplemented with synthetic medium. We anticipate that the results of this study will help in deriving an enhanced method of coupling nutrient-rich dairy manure wastewater for biofuel production.