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Wang K.,Samuel Roberts Noble Foundation | Wang K.,Agroecology Research Services Center | Senthil-Kumar M.,Samuel Roberts Noble Foundation | Ryu C.-M.,Samuel Roberts Noble Foundation | And 3 more authors.
Plant Physiology | Year: 2012

Bacterial pathogens colonize a host plant by growing between the cells by utilizing the nutrients present in apoplastic space. While successful pathogens manipulate the plant cell membrane to retrieve more nutrients from the cell, the counteracting plant defense mechanism against nonhost pathogens to restrict the nutrient efflux into the apoplast is not clear. To identify the genes involved in nonhost resistance against bacterial pathogens, we developed a virus-induced gene-silencing-based fastforward genetics screen in Nicotiana benthamiana. Silencing of N. benthamiana SQUALENE SYNTHASE, a key gene in phytosterol biosynthesis, not only compromised nonhost resistance to few pathovars of Pseudomonas syringae and Xanthomonas campestris, but also enhanced the growth of the host pathogen P. syringae pv tabaci by increasing nutrient efflux into the apoplast. An Arabidopsis (Arabidopsis thaliana) sterol methyltransferase mutant (sterol methyltransferase2) involved in sterol biosynthesis also compromised plant innate immunity against bacterial pathogens. The Arabidopsis cytochrome P450 CYP710A1, which encodes C22-sterol desaturase that converts b-sitosterol to stigmasterol, was dramatically induced upon inoculation with nonhost pathogens. An Arabidopsis Atcyp710A1 null mutant compromised both nonhost and basal resistance while overexpressors of AtCYP710A1 enhanced resistance to host pathogens. Our data implicate the involvement of sterols in plant innate immunity against bacterial infections by regulating nutrient efflux into the apoplast. © 2012 American Society of Plant Biologists. All Rights Reserved.


Rojas C.M.,Samuel Roberts Noble Foundation | Senthil-Kumar M.,Samuel Roberts Noble Foundation | Wang K.,Samuel Roberts Noble Foundation | Wang K.,Agroecology Research Services Center | And 4 more authors.
Plant Cell | Year: 2012

In contrast to gene-for-gene disease resistance, nonhost resistance governs defense responses to a broad range of potential pathogen species. To identify specific genes involved in the signal transduction cascade associated with nonhost disease resistance, we used a virus-induced gene-silencing screen in Nicotiana benthamiana, and identified the peroxisomal enzyme glycolate oxidase (GOX) as an essential component of nonhost resistance. GOX-silenced N. benthamiana and Arabidopsis thaliana GOX T-DNA insertion mutants are compromised for nonhost resistance. Moreover, Arabidopsis gox mutants have lower H2O2 accumulation, reduced callose deposition, and reduced electrolyte leakage upon inoculation with hypersensitive response-causing nonhost pathogens. Arabidopsis gox mutants were not affected in NADPH oxidase activity, and silencing of a gene encoding NADPH oxidase (Respiratory burst oxidase homolog) in the gox mutants did not further increase susceptibility to nonhost pathogens, suggesting that GOX functions independently from NADPH oxidase. In the two gox mutants examined (haox2 and gox3), the expression of several defense-related genes upon nonhost pathogen inoculation was decreased compared with wild-type plants. Here we show that GOX is an alternative source for the production of H 2O 2 during both gene-for-gene and nonhost resistance responses. © 2012 American Society of Plant Biologists.


Senthil-Kumar M.,Samuel Roberts Noble Foundation | Wang K.,Agroecology Research Services Center | Mysore K.S.,Samuel Roberts Noble Foundation
Plant Signaling and Behavior | Year: 2013

Stigmasterol and sitosterol, important sterols present in plants, are known to influence permeability and fluidity characteristics of the plasma membrane and other organellar membranes. We had previously demonstrated that the Arabidopsis Atcyp710A1 gene, which catalyzes conversion of sitosterol into stigmasterol, plays a role in plasma membrane permeability, thus influencing leakage of cellular nutrients and ions into apoplast. In this study, we investigated the role of this gene in imparting various abiotic stress tolerances in Arabidopsis. By analyzing Atcyp710a1 mutant and AtCYP710A1 overexpressor lines, we found that the AtCYP710A1 gene plays a role in imparting low and high temperature tolerance. © 2013 Landes Bioscience.


Johnston-Monje D.,University of Guelph | Johnston-Monje D.,Symbiota | Lundberg D.S.,Max Planck Institute for Developmental Biology | Lazarovits G.,Agroecology Research Services Center | And 2 more authors.
Plant and Soil | Year: 2016

Background and aims: To assess the impacts of soil microbes and plant genotype on the composition of maize associated bacterial communities. Methods: Two genotypes of Brazilian maize were planted indoors on sterile sand, a deep underground subsoil, and a nutrient-rich topsoil from the Amazon jungle (terra preta). DNA was extracted from rhizospheres, phyllospheres, and surface sterilized roots for 16S rDNA fingerprinting and next generation sequencing. Results: Neither plant genotype nor soil type appeared to influence bacterial diversity in phyllospheres or endospheres. Rhizospheres showed strikingly similar 16S rDNA ordination of both fingerprinting and sequencing data, with soil type driving grouping patterns and genotype having a significant impact only on sterile sand. Rhizospheres grown in non-sterile soils contained greater bacterial diversity than sterile-sand grown ones, however the dominant OTUs (species of Proteobacteria and Bacteroidetes) were found in all rhizospheres suggesting seeds as a common source of inoculum. Rhizospheres of the commercial hybrid appeared to contain less bacterial diversity than the landrace. Conclusions: Maize rhizospheres receive diverse bacteria from soil, are influenced by the genotype or treatment of the seed, and are dominated by species of Proteobacteria, Actinobacteria, Bacteroidetes, and Firmicutes. As many dominant 16S rDNA sequences were observed in rhizospheres grown in both sterile and non-sterile substrate, we conclude that the most common bacterial cells in juvenile maize rhizospheres are seed transmitted. © 2016 The Author(s)


Borrego-Benjumea A.,Agriculture and Agri Food Canada | Borrego-Benjumea A.,CSIC - Institute for Sustainable Agriculture | Basallote-Ureba M.J.,IFAPA Centro Las Torres Tomejil | Abbasi P.A.,Agriculture and Agri Food Canada | And 3 more authors.
Annals of Applied Biology | Year: 2014

Organic soil amendments play important roles in the reduction of plant diseases caused by soil-borne plant pathogens. This study examined the combined effects of concentrations of organic amendments, temperature and period of incubation in soil on the management of Fusarium wilt of tomato caused by Fusarium oxysporum f. sp. lycopersici (Fol). In an experiment with substrate mixture, Fol reduction was higher when the soils were incubated at 35°C than at 30°C. Disease severity was proportionally reduced as the volume of amendment added increased. Furthermore, disease was significantly lower in substrates incubated for 30-days at both temperatures, as compared to substrates incubated for only 15-days. The most effective control was achieved with pelletised poultry manure (PPM). In experiments with natural sandy soil, the effects of amendments on Fol populations, measured by real-time quantitative PCR with TaqMan probes, were significant. The highest decreases in Fol DNA resulted when the soil was amended with 2% PPM and incubated at 35°C. The reductions in DNA concentrations was most likely related to the accumulations of high concentrations of NH3 (27.3-mM) in soils treated with 2% PPM and incubated at room temperature (RT; 23-±-2°C), or at 35°C. Severity of plants grown in soils incubated at RT decreased by over 40%, and more than 73% when incubated at 35°C, regardless of the rate of PPM. The results indicate that the management with PPM, when combined with heating or solarisation, is an effective control measure against Fusarium wilt of tomato. © 2014 Association of Applied Biologists.


Johnston-Monje D.,University of Guelph | Johnston-Monje D.,Agroecology Research Services Center | Mousa W.K.,University of Guelph | Mousa W.K.,Mansoura University | And 2 more authors.
BMC Plant Biology | Year: 2014

Background: Endophytes are microbes that live within plants such as maize (corn, Zea mays L.) without causing disease. It is generally assumed that most endophytes originate from soil. If this is true, then as humans collected, domesticated, bred and migrated maize globally from its native Mexico, they moved the species away from its native population of endophyte donors. The migration of maize persists today, as breeders collect wild and exotic seed (as sources of diverse alleles) from sites of high genetic diversity in Mexico for breeding programs on distant soils. When transported to new lands, it is unclear whether maize permits only selective colonization of microbes from the Mexican soils on which it co-evolved, tolerates shifts in soil-derived endophytes, or prevents colonization of soil-based microbes in favour of seed-transmitted microbes. To test these hypotheses, non-sterilized seeds of three types of maize (pre-domesticated-Mexican, ancient-Mexican, modern-temperate) were planted side-by-side on indigenous Mexican soil, Canadian temperate soil or sterilized sand. The impact of these soil swaps on founder bacterial endophyte communities was tested using 16S-rDNA profiling, culturing and microbial trait phenotyping. Results: Multivariate analysis showed that bacterial 16S-rDNA TRFLP profiles from young, surface-sterilized maize plants were more similar when the same host genotype was grown on the different soils than when different maize genotypes were grown on the same soil. There appeared to be two reasons for this result. First, the largest fraction of bacterial 16S-signals from soil-grown plants was shared with parental seeds and/or plants grown on sterilized sand, suggesting significant inheritance of candidate endophytes. The in vitro activities of soil-derived candidate endophytes could be provided by bacteria that were isolated from sterile sand grown plants. Second, many non-inherited 16S-signals from sibling plants grown on geographically-distant soils were shared with one another, suggesting maize can select microbes with similar TRFLP peak sizes from diverse soils. Wild, pre-domesticated maize did not possess more unique 16S-signals when grown on its native Mexican soil than on Canadian soil, pointing against long-term co-evolutionary selection. The modern hybrid did not reject more soil-derived 16S-signals than did ancestral maize, pointing against such rejection as a mechanism that contributes to yield stability across environments. A minor fraction of 16S-signals was uniquely associated with any one soil. Conclusion: Within the limits of TRFLP profiling, the candidate bacterial endophyte populations of pre-domesticated, ancient and modern maize are partially buffered against the effects of geographic migration --- from a Mexican soil associated with ancestral maize, to a Canadian soil associated with modern hybrid agriculture. These results have implications for understanding the effects of domestication, migration, ex situ seed conservation and modern breeding, on the microbiome of one of the world's most important food crops. © 2014 Johnston-Monje et al.; licensee BioMed Central Ltd.


Turnbull A.L.,University of Calgary | Turnbull A.L.,Agroecology Research Services Center | Kim W.,University of Calgary | Kim W.,University of Oxford | And 2 more authors.
Canadian Journal of Microbiology | Year: 2012

The sdiA gene encodes for a LuxR-type transcription factor, which is active when bound to N-acyl homoserine lactones (AHLs). Because Salmonella enterica serovar Typhimurium does not produce AHLs, SdiA senses signals produced by other organisms. SdiA is not expressed constitutively, and response is limited to conditions in which elevated expression occurs, but little is known about the regulation of sdiA expression. Here we map the sdiA promoter and define several regulators that directly or indirectly act on the promoter. The major activator of sdiA expression is cAMP-receptor protein (CRP), and we define the CRP operator in the sdiA promoter using promoter and crp mutants. LeuO activates sdiA expression to a lesser extent than does CRP. We demonstrate that LeuO directly binds the sdiA promoter and the Rcs phosphorelay represses sdiA expression. In this study, NhaR, IlvY, and Fur affected sdiA expression indirectly and weakly. Expression in late-stationary phase depended on RpoS. AHL-dependent expression of the SdiA-regulated gene rck correlated to the observed sdiA transcriptional changes in regulator mutants. The data demonstrate that regulation of sdiA involves integration of multiple environmental and metabolic signals.


Turnbull A.L.,Agroecology Research Services Center | Liu Y.,Agroecology Research Services Center | Lazarovits G.,Agroecology Research Services Center
American Journal of Potato Research | Year: 2012

The objective of this study was to isolate and characterize genera of bacteria that had been identified as being the most common residents in the rhizosphere of potato using cpn60 pyrosequencing analysis. Using various semi-selective media targeted to specific genera of interest, 200 isolates of bacteria were collected from rhizosphere soil and the rhizoplane of potatoes grown in soils obtained from a potato farm in Prince Edward Island and Ontario. The procedures employed were successful in selecting out representative bacteria suggested by pyrosequencing to be common in the potato rhizoplane. Results of 16S rRNA sequencing showed that 44 % of the isolates represented new species. All isolates were tested for biological and biochemical activities including phosphate solubilization, phytohormone metabolism, nitrogen fixation, antibiosis, exoenzyme production, and production of acyl-homoserine lactones. Massilia spp. and Chryseobacterium spp. showed the strongest exoenzyme activities. A greater proportion of Agrobacterium tumefaciens rhizosphere strains produced acyl-homoserine lactones compared to rhizoplane strains. Pseudomonas spp. and Lysobacter capsici had the greatest antagonistic activity on laboratory media towards six potato pathogens, and also significantly decreased disease in plants grown in pathogen-infested soil. Four isolates significantly increased growth of potato nodal explants in tissue culture. By using preliminary results derived from next generation sequencing technology and a targeted cultural technique, we were able to gain a better understanding of the biological activities of the most abundant bacterial species in the rhizosphere and rhizoplane of a cultivated crop. © 2012 Potato Association of America.

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