scholarly journals In Vitro Evaluation of Five Antimicrobial Peptides against the Plant Pathogen Erwinia amylovora

Biomolecules ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 554
Author(s):  
Rafael J. Mendes ◽  
Laura Regalado ◽  
João P. Luz ◽  
Natália Tassi ◽  
Cátia Teixeira ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Antimicrobial Peptides ◽  
Erwinia Amylovora ◽  
Growth Curves ◽  
Fruit Trees ◽  
Membrane Permeabilization ◽  
Control Measures ◽  
Minimal Bactericidal Concentration ◽  
Pome Fruit

Fire blight is a major pome fruit trees disease that is caused by the quarantine phytopathogenic Erwinia amylovora, leading to major losses, namely, in pear and apple productions. Nevertheless, no effective sustainable control treatments and measures have yet been disclosed. In that regard, antimicrobial peptides (AMPs) have been proposed as an alternative biomolecule against pathogens but some of those AMPs have yet to be tested against E. amylovora. In this study, the potential of five AMPs (RW-BP100, CA-M, 3.1, D4E1, and Dhvar-5) together with BP100, were assessed to control E. amylovora. Antibiograms, minimal inhibitory, and bactericidal concentrations (minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC), growth and IC50 were determined and membrane permeabilization capacity was evaluated by flow cytometry analysis and colony-forming units (CFUs) plate counting. For the tested AMPs, the higher inhibitory and bactericidal capacity was observed for RW-BP100 and CA-M (5 and 5–8 µM, respectively for both MIC and MBC), whilst for IC50 RW-BP100 presented higher efficiency (2.8 to 3.5 µM). Growth curves for the first concentrations bellow MIC showed that these AMPs delayed E. amylovora growth. Flow cytometry disclosed faster membrane permeabilization for CA-M. These results highlight the potential of RW-BP100 and CA-M AMPs as sustainable control measures against E. amylovora.

scholarly journals Comparative Study Regarding in Vitro Infections with Erwinia Amylovora and Pseudomonas Syringae Pv. Syringae on Pomaceae Species

2015 ◽  
Vol 48 (1) ◽  
pp. 97-106
Author(s):  
Irina Paraschiva Chiriac ◽  
Fl. D. Lipşa ◽  
E. Ulea
Keyword(s):  
Pseudomonas Syringae ◽  
Erwinia Amylovora ◽  
Plant Protection ◽  
Fruit Trees ◽  
Pome Fruit ◽  
Occurrence Time ◽  
Major Objective ◽  
Great Economic Importance ◽  
In Vitro Inoculation

Abstract The knowledge about pome fruit trees bacterioses and their evolution in orchards is a major objective for plant protection. Erwinia amylovora and Pseudomonas syringae pv. syringae cause on attacked organs of Pomaceae species similar dieback symptoms in vegetative and flowering shoots of quince, pear and apple in spring. Both bacteria can produce disastrous diseases in orchards and are therefore of great economic importance. Biological materials represented by vegetative shoots, leaves and fruits of Pyrus spp., Malus spp., Cydonia spp. were used after isolation of different E. amylovora and Ps. syringae pv. syringae strains for in vitro infections. Results presented in this study established that for in vitro inoculation of Pomaceae species similar symptoms in case of leaves and, respectively different symptoms for vegetative shoots and fruits occurred. The occurrence time was the only difference, because Ps. syringae pv. syringae spread faster than E. amylovora. The vegetative shoots inoculated with E. amylovora, in comparison to Ps. syringae pv. syringae, were more damaged and for both bacteria the highest values for attack degree were recorded in case of pear species, followed by quince and apples, respectively.

scholarly journals CRISPR genotyping as complementary tool for epidemiological surveillance of Erwinia amylovora outbreaks

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250280
Author(s):  
Rafael J. Mendes ◽  
João Pedro Luz ◽  
Conceição Santos ◽  
Fernando Tavares
Keyword(s):  
Fire Blight ◽  
Erwinia Amylovora ◽  
Blot Hybridization ◽  
Fruit Trees ◽  
Epidemiological Surveillance ◽  
Economic Losses ◽  
Pome Fruit ◽  
Dot Blot ◽  
Virulence Markers ◽  
Clonal Population Structure

Fire blight is a destructive plant disease caused by Erwinia amylovora affecting pome fruit trees, and responsible for large yield declines, long phytosanitary confinements, and high economic losses. In Portugal, the first major fire blight outbreaks occurred in 2010 and 2011, and although later considered eradicated, the emergence of other outbreaks in recent years stressed the need to characterize the E. amylovora populations associated with these outbreaks. In this regard, CRISPR genotyping, assessment of three virulence markers, and semi-quantitative virulence bioassays, were carried out to determine the genotype, and assess the virulence of thirty-six E. amylovora isolates associated with outbreaks occurring between 2010 and 2017 and affecting apple and pear orchards located in the country central-west, known as the main producing region of pome fruits in Portugal. The data gathered reveal that 35 E. amylovora isolates belong to one of the widely-distributed CRISPR genotypes (5-24-38 / D-a-α) regardless the host species, year and region. Ea 680 was the single isolate revealing a new CRISPR genotype due to a novel CR2 spacer located closer to the leader sequence and therefore thought to be recently acquired. Regarding pathogenicity, although dot-blot hybridization assays showed the presence of key virulence factors, namely hrpL (T3SS), hrpN (T3E) and amsG from the amylovoran biosynthesis operon in all E. amylovora isolates studied, pathogenicity bioassays on immature pear slices allowed to distinguish four virulence levels, with most of the isolates revealing an intermediate to severe virulence phenotype. Regardless the clonal population structure of the E. amylovora associated to the outbreaks occurring in Portugal between 2010 and 2017, the different virulence phenotypes, suggests that E. amylovora may have been introduced at different instances into the country. This is the first study regarding E. amylovora in Portugal, and it discloses a novel CRISPR genotype for this bacterium.

scholarly journals Activation of metabolic and stress responses during subtoxic expression of the type I toxin hok in Erwinia amylovora

2021 ◽  
Author(s):  
Jingyu Peng ◽  
Lindsay R. Triplett ◽  
George Sundin
Keyword(s):  
Cell Death ◽  
Stress Responses ◽  
Erwinia Amylovora ◽  
Physiological State ◽  
Fruit Trees ◽  
Toxin Gene ◽  
Type I ◽  
Pome Fruit ◽  
Membrane Rupture ◽  
High Level

Abstract Background: Toxin-antitoxin (TA) systems, abundant in prokaryotes, are composed of a toxin gene and its cognate antitoxin. Several toxins are implied to affect the physiological state and stress tolerance of bacteria in a population. We previously identified a chromosomally encoded hok-sok type I TA system in Erwinia amylovora, the causative agent of fire blight disease on pome fruit trees. A high-level induction of the hok gene was lethal to E. amylovora cells through unknown mechanisms. The molecular targets or regulatory roles of Hok were unknown.Results: Here, we examined the physiological and transcriptomic changes of Erwinia amylovora cells expressing hok at subtoxic levels that were confirmed to confer no cell death, and at toxic levels that resulted in killing of cells. In both conditions, hok caused membrane rupture and collapse of the proton motive force in a subpopulation of E. amylovora cells. We demonstrated that induction of hok resulted in upregulation of ATP biosynthesis genes, and caused leakage of ATP from cells only at toxic levels. We showed that overexpression of the phage shock protein gene pspA largely reversed the cell death phenotype caused by high levels of hok induction. We also showed that induction of hok at a subtoxic level rendered a greater proportion of stationary phase E. amylovora cells tolerant to the antibiotic streptomycin. Conclusions: We characterized the molecular mechanism of toxicity by high-level of hok induction and demonstrated that low-level expression of hok primes the stress responses of E. amylovora against further membrane and antibiotic stressors.

scholarly journals The Role of luxS in the Fire Blight Pathogen Erwinia amylovora Is Limited to Metabolism and Does Not Involve Quorum Sensing

2007 ◽  
Vol 20 (10) ◽  
pp. 1284-1297 ◽  
Author(s):  
Fabio Rezzonico ◽  
Brion Duffy
Keyword(s):  
Gene Expression ◽  
Quorum Sensing ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Mutational Analysis ◽  
Pome Fruit ◽  
Autoinducer 2 ◽  
Metabolic Genes ◽  
Metabolic Role

Erwinia amylovora is a gram-negative phytopathogen that causes fire blight of pome fruit and related members of the family Rosaceae. We sequenced the putative autoinducer-2 (AI-2) synthase gene luxS from E. amylovora. Diversity analysis indicated that this gene is extremely conserved among E. amylovora strains. Quorum sensing mediated by LuxS has been implicated in coordinated gene expression, growth, and virulence in other enterobacteria; however, our evidence suggests this is not the function in E. amylovora. Mutational analysis pointed to a role in colonization of apple blossoms, the primary infection court for fire blight, although little if any role in virulence on apple shoots and pear fruit was observed. Expression of key virulence genes hrpL and dspA/E was reduced in mutants of two E. amylovora strains. Stronger effects on gene expression were observed for metabolic genes involved in the activated methyl cycle with mutants having greater levels of expression. No quorum-sensing effect was observed in co-culture experiments with wild-type and mutant strains either in vitro or in apple blossoms. Known receptors essential for AI-2 quorum sensing, the LuxPQ sensor kinase or the Lsr ABC-transporter, are absent in E. amylovora, further suggesting a primarily metabolic role for luxS in this bacterium.

scholarly journals Activation of metabolic and stress responses during subtoxic expression of the type I toxin hok in Erwinia amylovora

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Jingyu Peng ◽  
Lindsay R. Triplett ◽  
George W. Sundin
Keyword(s):  
Cell Death ◽  
Stress Responses ◽  
Erwinia Amylovora ◽  
Physiological State ◽  
Fruit Trees ◽  
Toxin Gene ◽  
Type I ◽  
Pome Fruit ◽  
Membrane Rupture ◽  
High Level

Abstract Background Toxin-antitoxin (TA) systems, abundant in prokaryotes, are composed of a toxin gene and its cognate antitoxin. Several toxins are implied to affect the physiological state and stress tolerance of bacteria in a population. We previously identified a chromosomally encoded hok-sok type I TA system in Erwinia amylovora, the causative agent of fire blight disease on pome fruit trees. A high-level induction of the hok gene was lethal to E. amylovora cells through unknown mechanisms. The molecular targets or regulatory roles of Hok were unknown. Results Here, we examined the physiological and transcriptomic changes of Erwinia amylovora cells expressing hok at subtoxic levels that were confirmed to confer no cell death, and at toxic levels that resulted in killing of cells. In both conditions, hok caused membrane rupture and collapse of the proton motive force in a subpopulation of E. amylovora cells. We demonstrated that induction of hok resulted in upregulation of ATP biosynthesis genes, and caused leakage of ATP from cells only at toxic levels. We showed that overexpression of the phage shock protein gene pspA largely reversed the cell death phenotype caused by high levels of hok induction. We also showed that induction of hok at a subtoxic level rendered a greater proportion of stationary phase E. amylovora cells tolerant to the antibiotic streptomycin. Conclusions We characterized the molecular mechanism of toxicity by high-level of hok induction and demonstrated that low-level expression of hok primes the stress responses of E. amylovora against further membrane and antibiotic stressors.

scholarly journals The Incessant Battle Against Fire Blight in Pears: 30 Years of Challenges and Successes in Managing the Disease in Israel

Plant Disease ◽  
2015 ◽  
Vol 99 (8) ◽  
pp. 1048-1058 ◽  
Author(s):  
Dani Shtienberg ◽  
Shulamit Manulis-Sasson ◽  
Miriam Zilberstaine ◽  
Dov Oppenheim ◽  
Hagai Shwartz
Keyword(s):  
Crucial Role ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Fruit Trees ◽  
Future Perspectives ◽  
Full Understanding ◽  
Pome Fruit ◽  
Local Experience

Fire blight, caused by the bacterium Erwinia amylovora, is the most destructive disease of pears and other pome fruit trees worldwide. The disease was first detected in Israel in 1985, and in the 30 years since, the intensity of fire blight epidemics has varied markedly. During this time, there were two national pandemics: the first between 1994 and 1996 and the second in 2010. In both cases, it was feared that the Israeli pear industry would not recover. National efforts were devoted to combat the problem and after both pandemics the industry survived. In this paper we indicate some unique characteristics that play a crucial role in the epidemiology of the disease under Israeli conditions. We then describe the continual struggle of the Israeli pear industry with fire blight over the last 30 years, elaborating on the two national pandemics and the efforts devoted to cope with them. Finally, we summarize the conclusions derived from our local experience and present our future perspectives regarding fire blight management. The take-home message of the Israeli fire blight story is that the battle against this hazardous disease is neverending. In some years, growers are able to adequately suppress the disease; in others, the pathogen overcomes management efforts and severe outbreaks occur. The latter could be minimized if growers have a full understanding of the management protocols suitable for the conditions and applied them rigorously.

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