Fatty acid biomarker analysis to characterize soil microbial communities in soybean agroecosystems with Sclerotinia stem rot disease

Soybean (Glycine max (L.) Merr.) is one the major crops produced worldwide. However, soybean is susceptible to many diseases. Sclerotinia stem rot (SSR) disease caused by Sclerotinia sclerotiorum (Lib.) de Bary is considered one of the most important fungal diseases of soybean. It can be controlled by chemicals (e.g. fungicides), by breeding cultivars with disease resistance and by cultural control (e.g. increasing the width between rows, reducing plant populations). A promising and complementary method of controlling SSR disease in the field is the application of biological control agents. Biological control agents introduced in a soil environment will interact with other soil food web organisms, as do the pathogenic organism and infected plants, which may change the genetic and functional diversity in soil microbial communities. Profiling these changes may lead to an improved understanding of the interactions between these players (biological control agents, pathogens, soil biota and plants) in the biological control phenomenom, permiting us to exploit naturally-occurring ecological relationships and develop more sustainable approaches to control soybean diseases. Fatty acid biomarkers analysis was used to profile microbial communities in soils. Two laboratory studies were conducted to evaluate the methods used for extraction and profiling the fatty acid biomarkers from soil samples with a range of soil properties (clay content, organic matter content), The first study investigated the best solvent mixture for recovering fatty acid biomarkers from soil using an automated pressurized solvent extraction (PSE) system. Solvent mixtures containing chloroform and methanol were more efficient at extracting fatty acids from agricultural soils than hexane:2-propanol and acetone. The second study presented an exploratory pyrolysis-mass spectrometry technique to rapidly fingerprint soil lipids extracted from different agroecosystems. Pyrolysis-mass spectrometry discriminated among soils and crop production systems in the same way as the fatty acid profiling. I also report on the efficicacy of biological control agents to control Sclerotinia stem rot disease in soybean. A two-year study was conducted in soybean fields under conventional or no tillage to determine whether Trichoderma virens (SoilGard(TM)) and arbuscular mycorrhizal fungi (a mixture of Glomus intraradices and G. mosseae ), used alone or in combination, could reduce sclerotinia stem rot (SSR) disease incidence. Generally, SSR disease indicators, as well as the soybean yield, were not affected significantly by the biological control treatments. I then studied whether changes in microbial community composition were related to the inoculation of the biological control agents and the disease incidence in soybean fields. Inoculation of biological control agents changes the expression of many soil fatty acids during both years of the trial. Also, in the plots with severely diseased plants, fatty acids biomarkers of gram positive and actinomycetes bacteria were significantly greater than in plots with healthy plants. I conclude that further improvement in laboratory techniques and procedures will permit researchers to efficiently extract and characterize soil lipids, providing new insight into soil organic matter dynamics and soil microbial ecology. Further study will be needed to verify the efficacy and optimize the application method, dose and timing of biocontrol agents to provide protection against SSR disease in soybean fields

Salmonella Degrades the Host Glycocalyx Leading to Altered Infection and Glycan Remodeling.

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 < 0.05) alter infection. During infection, 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

Virulence factors of Actinobacillus pleuropneumoniae involved in colonization, persistence and induction of lesions in its porcine host

Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia. The virulence factors of this microorganism involved in colonization and the induction of lung lesions have been thoroughly studied and some have been well characterized. A. pleuropneumoniae binds preferentially to cells of the lower respiratory tract in a process involving different adhesins and probably biofilm formation. Apx toxins and lipopolysaccharides exert pathogenic effects on several host cells, resulting in typical lung lesions. Lysis of host cells is essential for the bacterium to obtain nutrients from the environment and A. pleuropneumoniae has developed several uptake mechanisms for these nutrients. In addition to persistence in lung lesions, colonization of the upper respiratory tract – and of the tonsils in particular – may also be important for long-term persistent asymptomatic infection. Information on virulence factors involved in tonsillar and nasal cavity colonization and persistence is scarce, but it can be speculated that similar features as demonstrated for the lung may play a role

Lipid profiles in wheat cultivars resistant and susceptible to tan spot and the effect of disease on the profiles

Citation: Kim, D., . . . & Bockus, W. (2012). Lipid Profiles in Wheat Cultivars Resistant and Susceptible to Tan Spot and the Effect of Disease on the Profiles. Phytopathology, 103(1), 74-80. https://doi.org/10.1094/PHYTO-05-12-0099-RLipid 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

A spontaneous mutation in Srebf2 leads to cataracts and persistent skin wounds in the lens opacity 13 (lop13) mouse.

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

Chlorella vulgaris production enhancement with supplementation of synthetic medium in dairy manure wastewater.

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

Quantification of antibiotic resistance genes and mobile genetic in dairy manure.

BackgroundAntibiotic resistance genes (ARGs) are considered to be emerging environmental contaminants of concern potentially posing risks to human and animal health, and this research studied the prevalence of antimicrobial resistance in dairy manure.MethodsThis study is focused on investigating prevalence of ARGs in California dairy farm manure under current common different 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 eight 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).ResultsPrevalence 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 < 0.05), and Psychrobacter and Pseudomonas as opportunistic human pathogens, were potential ARG/MGE hosts (p < 0.05). This research indicated current different manure management practices in California dairy farms has limited effects on reducing ARGs and MGEs. Improvement of different manure management in dairy farms is thus important to mitigate dissemination of ARGs into the environment

Diazotrophic bacteria from maize exhibit multifaceted plant growth promotion traits in multiple hosts.

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

Diazotrophic bacteria from maize exhibit multifaceted plant growth promotion traits in multiple hosts.

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

Diazotrophic bacteria from maize exhibit multifaceted plant growth promotion traits in multiple hosts.

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