Proteobacteria

Event Date: 
Wednesday, March 25, 2015 - 18:15 - 18:30
Institution: 
University of Florida / UWS
Title: 

Candidatus Liberibacter asiaticus encodes a functional salicylic acid hydroxylase which degrades SA and contributes to the suppression of plant defence

Abstract: 

Salicylate (SA) is a plant hormone and plays important roles in plant defence. SA is synthesized in the chloroplast and transmitted in the phloem. SA hydroxylase is a flavoprotein monooxygenase with the enzyme activity of degradation of SA and is a proximal component of the naphthalene degradation pathway in many bacteria. Candidatus Liberibacter asiaticus, the causal agent of the most devastating citrus disease, is phloem limited and encodes a SA hydroxylase. In this study, we have shown that the SA hydroxylase is functional in degrading SA and its analogs. Ca. L. asiaticus infected plants have reduced PR gene (PR1, PR2, and PR5) expression and SA accumulation in Duncan grapefruit and Valencia sweet orange in response to subsequent inoculation with Xanthomonas citri subsp. citri (Xac) Aw, which is nonpathogenic on both citrus varieties. Ca. L. asiaticus also increased citrus susceptibility to subsequent infection by X. citri. The bacterial populations of XacA and XacAw in grapefruit were significantly higher in Ca. L. asiaticus infected plants compared to healthy control. Our data suggest that Ca. L. asiaticus encodes a functional salicylic acid hydroxylase which degrades SA and contributes to the suppression of plant defence. To counteract this virulence mechanism of Ca. L. asiaticus, foliar spray of SA analogs 2, 6-Dichloroisonicotinic acid (INA) and 2,1,3-Benzothiadiazole (BTH) and SA producing bacterial isolates was conducted to control HLB in large scale field trials. Both INA and BTH in combination with selected bacterial strains slowed down the increase of Ca. L. asiaticus titers in planta and HLB disease severity compared to negative control. SA hydroxylase seems to be an ideal target to develop small molecule inhibitors since no human homolog is present and it is not essential for bacterial growth, hence, the possibility of resistance development is minimized.      Salicylate (SA) is a plant hormone and plays important roles in plant defence. SA is synthesized in the chloroplast and transmitted in the phloem. SA hydroxylase is a flavoprotein monooxygenase with the enzyme activity of degradation of SA and is a proximal component of the naphthalene degradation pathway in many bacteria. Candidatus Liberibacter asiaticus, the causal agent of the most devastating citrus disease, is phloem limited and encodes a SA hydroxylase. In this study, we have shown that the SA hydroxylase is functional in degrading SA and its analogs. Ca. L. asiaticus infected plants have reduced PR gene (PR1, PR2, and PR5) expression and SA accumulation in Duncan grapefruit and Valencia sweet orange in response to subsequent inoculation with Xanthomonas citri subsp. citri (Xac) Aw, which is nonpathogenic on both citrus varieties. Ca. L. asiaticus also increased citrus susceptibility to subsequent infection by X. citri. The bacterial populations of XacA and XacAw in grapefruit were significantly higher in Ca. L. asiaticus infected plants compared to healthy control. Our data suggest that Ca. L. asiaticus encodes a functional salicylic acid hydroxylase which degrades SA and contributes to the suppression of plant defence. To counteract this virulence mechanism of Ca. L. asiaticus, foliar spray of SA analogs 2, 6-Dichloroisonicotinic acid (INA) and 2,1,3-Benzothiadiazole (BTH) and SA producing bacterial isolates was conducted to control HLB in large scale field trials. Both INA and BTH in combination with selected bacterial strains slowed down the increase of Ca. L. asiaticus titers in planta and HLB disease severity compared to negative control. SA hydroxylase seems to be an ideal target to develop small molecule inhibitors since no human homolog is present and it is not essential for bacterial growth, hence, the possibility of resistance development is minimized.      

Event Date: 
Wednesday, March 26, 2014 - 18:00 - 18:15
Institution: 
UNSW
Title: 

Insights into the stress response of the biomining bacterium Acidithiobacillus ferrooxidans using gene expression and proteomic analysis

Abstract: 

Bioleaching is a simple and effective process used for metal extraction from low grade ores and mineral concentrates using microorganisms. The extraction of some metals such as copper from low grade ore is becoming necessary because of gradual depletion of high grade ore. The traditional methods used for extraction of copper are either Pyrometallurgy or Hydrometallurgy. However both the methods are not environmental friendly. There are many techniques proposed to extract metals but these are not practically suitable, as these requires a very high energy input as well as most of them creates environmental pollution problem, that also rises the cost of environmental protection throughout the world. Therefore, bioleaching is recognizable as the most environmentally friendly method of separating metals since it requires less energy and it reduces the amount of greenhouse gasses released to the atmosphere. Bioleaching is also a fairly simple process that does not require a lot of expertise to operate or complicated machinery.
The most commonly used bacterium in bioleaching is Acidithiobacillus ferrooxidans (former Thiobacillus ferrooxidans) and this is due to its capacity to oxidize metal sulphides. A. ferroxidans is a chemolithotrophic bacterium capable of utilizing ferrous iron or reduced sulphur compounds as the sole source of energy for its growth. It thrives optimally around pH 2.0 and 30ºC. During Bioleaching process, A. ferrooxidans is often subject to changes in the ideal growth pH and temperature, and to nutrients starvation. These changes can affect the bacterial physiology and as a consequence, the efficiency of bioleaching. Then, the stress response of this bacterium subject to heat stress and phosphate starvation has been investigated using different approaches, namely, gene expression and proteomic analysis, Fourier transform infrared spectroscopy (FT-IR), as well as morphological analysis by scanning electron microscopy (SEM).
The results showed that under the tested stress conditions A. ferrooxidans cells suffer elongation, a common stress response in bacteria. Alterations in carbohydrates, phospholipids and phosphoproteins were detected by FT-IR. By proteomic analyses (2-DE and tandem mass spectrometry), many differentially expressed protein spots were visualized and identified as proteins belonging to 11 different functional categories. Indeed, the up-regulated proteins were mainly from the protein fate category. Real time quantitative PCR was employed to analyze changes in the expression patterns of heat shock genes, as well as many other genes encoding proteins related to several functional categories in A. ferrooxidans. Cells were submitted to long-term growth and to heat shock, both at 40°C. The results evidenced that heat shock affected the expression levels of most genes while long-term growth at 40°C caused minimal changes in gene expression patterns – with exception of some iron transport related genes, which were strongly down-regulated. Further bioinformatic analysis indicated a putative transcriptional regulation, by the σ32 factor, in most heat-affected genes. These results evidence that A. ferrooxidans has an efficient range of stress-responses, which explains its ability for biotechnological purposes.
 

Event Date: 
Wednesday, March 27, 2013 - 18:15 - 18:30
Institution: 
Hawkesbury Institute for the Environment, University of Western Sydney
Title: 

Effect of Huanglongbing on the structure and functional diversity of microbial communities associated with citrus.

Abstract: 

Plant-microbe interactions lie at the heart of plant performance and ecology. It has been postulated that disruption of multi-trophic interactions in a stable ecosystem under the influence of invading phytopathogens will cause community reorganization and changes in the local feedback interactions. However, there is a paucity of knowledge on the extent to which such community shifts may occur, on the dynamics of changes and on the putative effects regarding the functioning of ecosystems. We have used Citrus-‘Candidatus Liberibacter asiaticus’ [Las, causal agent of devastating Huanglongbing (HLB) disease] as a host pathogen model to characterize the structure, function and interactions of plant-associated microbial communities. We applied a suit of metagenomic techniques to provide detailed census of citrus associated microbiomes. Our results confirmed that Las is the sole causal agent of HLB in Florida and revealed that HLB significantly re-structures the composition of native microbial community present either in leaf, roots and rhizosphere of citrus. Functional microarray (Geochip) and shotgun metagenomic sequencing showed that HLB has severe effects on various functional guilds of bacteria involved in key ecological processes including nitrogen cycling and carbon fixation. Overall, the metagenomic studies provided evidence that change in plant physiology mediated by Las infection could elicit shifts in the composition and functional potential of plant associated microbial communities. In the long term, these fluctuations might have important implications for the productivity and sustainability of citrus producing agro-ecosystems.

JAMS REPORT
Tom Jeffries
 
January JAMS got the new year off to a good start with a solid turnout and some stimulating talks. First up was Olivier Laczka who took us in to the technical realm of biosensors. Olivier’s work has focused on developing cost-effective tools for the rapid identification of micro-organisms relevant to industry and has led to several Patents.

Event Date: 
Tuesday, July 24, 2012 - 18:00 - 18:15
Institution: 
University of NSW
Title: 

Bioremediation of Mixed Chlorinated Solvents by Combining Two Biogeochemical Processes

Abstract: 

Chloroethenes are a class of chlorinated solvents which cause extensive soil and groundwater contamination worldwide. They can be detoxified by anaerobic dehalogenating bacteria, in the process of reductive dechlorination.  However, chloroethenes are often found mixed with chloromethanes, a class of solvents which inhibit the enzymatic detoxification of chloroethenes by dehalogenating strains.  Iron sulfides are powerful chemical reductants for the dechlorination of chloromethanes, and can be generated through the metabolism of iron- and sulfate-reducing bacteria. In this study, a sulfate reducing bacterium was used to produce iron sulfide in the presence of moderate levels of tetrachloroethene and carbon tetrachloride to examine the ability of a sulfate reducing organism to drive reduction of a chloromethane in the presence of chloroethene.

 

Cultures of the sulfate-reducer Desulfovibrio vulgaris were established in the presence of 100 µM each of tetrachloroethene and carbon tetrachloride. Growth, sulfide formation and chlorinated solvents and their dechlorinated products were monitored. The effects of amorphous iron oxide and cyanocobalamin on the fate of chlorinated solvents compared with unamended control cultures were investigated. 

Following growth and sulfide formation, carbon tetrachloride was dechlorinated mostly to carbon disulfide while tetrachloroethene was dechlorinated to trichloroethene and acetylene.  Dechlorination rates were enhanced both by the presence of iron and cyanocobalamin separately, and significantly increased when both were present.

This study illustrates the potential to use sulfate reducing bacteria in zones of mixed chlorinated solvent groundwater pollution in order to produce iron sulfide minerals. Their cyanocobalamin-catalyzed action on chloromethanes, coupled with that of dehalogenating strains on chloroethenes is a promising strategy for the bioremediation of such contaminated areas."

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