scholarly journals Establishment of Bacterial Antagonists of Erwinia amylovora on Pear and Apple Blossoms as Influenced by Inoculum Preparation

1998 ◽  
Vol 88 (6) ◽  
pp. 506-513 ◽  
Author(s):  
V. O. Stockwell ◽  
K. B. Johnson ◽  
J. E. Loper
Keyword(s):  
Fire Blight ◽  
Erwinia Amylovora ◽  
Culture Media ◽  
Field Trials ◽  
Bacterial Cells ◽  
Fluorescent Microspheres ◽  
Bacterial Antagonists ◽  
Spray Inoculation ◽  
Population Sizes ◽  
Inoculum Preparation

The influence of inoculum preparation on the establishment of bacterial antagonists that suppress fire blight and Erwinia amylovora on blossoms was evaluated. Aqueous suspensions of Pseudomonas fluorescens A506, E. herbicola C9-1R, or E. amylovora 153N were prepared from cells harvested from the surface of an agar medium or from cells that were lyophilized after culture under similar conditions. Bacterial suspensions (1 × 108 CFU/ml) were sprayed on pear and apple trees at 50% bloom near midday. The incidence of recovery (proportion of blossoms containing detectable populations) and the population sizes of the bacteria on individual blossoms with detectable populations were followed over a period of several days. Fluorescent microspheres (1 μm in diameter) were added to sprays at a concentration of 1 × 107 microspheres per ml to mark blossoms that were open during application of bacteria. After dilution-plating, the stigmas and styles of each blossom were examined for the presence of microspheres with an epifluorescence microscope. In three of five trials, bacteria applied as suspensions of lyophilized cells were recovered from a greater proportion of blossoms than bacterial cells harvested directly from culture media. Every blossom harvested within 6 days after spraying had microspheres present on the surfaces of the styles and stigmas; thus, lack of establishment of detectable populations, rather than escape of blossoms from spray inoculation, accounted for the differences in proportion of blossoms colonized by the different preparations of bacteria. The use of lyophilized cells in field trials decreased variability in the establishment of bacteria on blossoms.

scholarly journals Control of Fire Blight by Pseudomonas fluorescens A506 and Pantoea vagans C9-1 Applied as Single Strains and Mixed Inocula

2010 ◽  
Vol 100 (12) ◽  
pp. 1330-1339 ◽  
Author(s):  
V. O. Stockwell ◽  
K. B. Johnson ◽  
D. Sugar ◽  
J. E. Loper
Keyword(s):  
Biological Control ◽  
Pseudomonas Fluorescens ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Field Trials ◽  
Disease Suppression ◽  
Biological Control Agents ◽  
Population Sizes ◽  
Mixed Inocula ◽  
Pantoea Vagans

The biological control agents Pseudomonas fluorescens A506 and Pantoea vagans C9-1 were evaluated individually and in combination for the suppression of fire blight of pear or apple in 10 field trials inoculated with the pathogen Erwinia amylovora. The formulation of pathogen inoculum applied to blossoms influenced establishment of the pathogen and the efficacy of biological control. Pantoea vagans C9-1 suppressed fire blight in all five trials in which the pathogen was applied as lyophilized cells but in none of the trials in which the pathogen was applied as freshly harvested cells. In contrast, Pseudomonas fluorescens A506 reduced disease significantly in only one trial. A mixture of the two strains also suppressed fire blight, but the magnitude of disease suppression over all field trials (averaging 32%) was less than that attained by C9-1 alone (42%). The two biological control agents did not antagonize one another on blossom surfaces, and application of the mixture of A506 and C9-1 to blossoms resulted in a greater proportion of flowers having detectable populations of at least one bacterial antagonist than the application of individual strains. Therefore, the mixture of A506 and C9-1 provided less disease control than expected based upon the epiphytic population sizes of the antagonists on blossom surfaces. We speculate that the biocontrol mixture was less effective than anticipated due to incompatibility between the mechanisms by which A506 and C9-1 suppress disease.

scholarly journals Implications of Pathogenesis by Erwinia amylovora on Rosaceous Stigmas to Biological Control of Fire Blight

2009 ◽  
Vol 99 (2) ◽  
pp. 128-138 ◽  
Author(s):  
K. B. Johnson ◽  
T. L. Sawyer ◽  
V. O. Stockwell ◽  
T. N. Temple
Keyword(s):  
Biological Control ◽  
Population Size ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Fluorescent Protein ◽  
Virulent Strain ◽  
Avirulent Strain ◽  
Bacterial Antagonists ◽  
Population Sizes ◽  
Green Fluorescent

As a prerequisite to infection of flowers, Erwinia amylovora grows epiphytically on stigmas, which provide a conducive habitat for bacterial growth. Stigmas also support growth of several other bacterial genera, which allows for biological control of fire blight; although, in practice, it is very difficult to exclude E. amylovora completely from this habitat. We investigated the dynamics of growth suppression of E. amylovora by comparing the ability of virulent and avirulent strains of E. amylovora to compete with each other on stigmas of pear, apple, and blackberry, and to compete with a co-inoculated mixture of effective bacterial antagonists. When strains were inoculated individually, virulent E. amylovora strain Ea153N attained the highest population size on stigmas, with population sizes that were approximately double those of an avirulent hrpL mutant of Ea153 or the bacterial antagonists. In competition experiments, growth of the avirulent derivative was suppressed by the antagonist mixture to a greater extent than the virulent strain. Unexpectedly, the virulent strain enhanced the population size of the antagonist mixture. Similarly, a small dose of virulent Ea153N added to inoculum of an avirulent hrpL mutant of Ea153 significantly increased the population size of the avirulent strain. A pathogenesis-gene reporter strain, Ea153 dspE::gfp, was applied to flowers and a subset of the population expressed the green fluorescent protein while growing epiphytically on stigmas of apple. These results are consistent with the hypothesis that virulent E. amylovora modifies the epiphytic habitat presented by the stigma through a pathogenesis-related process, which increases host resources available to itself and, coincidentally, to nonpathogenic competitors. Over nine orchard trials, avirulent Ea153 hrpL significantly suppressed the incidence of fire blight four times compared with six for the antagonist mixture. The degree of biological control achievable with an avirulent strain of E. amylovora likely is limited by its inability to utilize the stigmatic habitat to the same degree as a virulent strain.

New potential bacterial antagonists for the biocontrol of fire blight disease (Erwinia amylovora) in Morocco

2018 ◽  
Vol 117 ◽  
pp. 7-15 ◽  
Author(s):  
Smail Ait Bahadou ◽  
Abderrahmane Ouijja ◽  
Abdelkarim Karfach ◽  
Abdessalem Tahiri ◽  
Rachid Lahlali
Keyword(s):  
Fire Blight ◽  
Erwinia Amylovora ◽  
Bacterial Antagonists ◽  
Blight Disease

scholarly journals Epiphytic Bacteria and Yeasts on Apple Blossoms and Their Potential as Antagonists of Erwinia amylovora

2009 ◽  
Vol 99 (5) ◽  
pp. 571-581 ◽  
Author(s):  
P. Lawrence Pusey ◽  
Virginia O. Stockwell ◽  
Mark Mazzola
Keyword(s):  
Relative Humidity ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Acid Methyl Ester ◽  
Washington State ◽  
Microbial Populations ◽  
Acid Methyl ◽  
Population Sizes ◽  
Average Relative Humidity ◽  
Cryptococcus Spp

Apple blossoms were sampled for indigenous epiphytic populations of culturable microorganisms during different stages of bloom at two locations in central Washington State and one site in Corvallis, OR. Frequencies and population sizes of bacteria on stigmas of apple were lower in Washington than at Corvallis, where average relative humidity was higher and possibly favored greater colonization; however, bacteria at Corvallis were mainly pseudomonads, whereas those in Washington were diverse, composed of several genera. In Washington, yeast as well as bacteria were isolated from both stigmatic and hypanthial surfaces. Sampled blossoms were processed immediately to assess microbial populations, or after a 24-h incubation at 28°C and high relative humidity, which broadened the range of detectable taxa evaluated as potential antagonists. Identifications were based on fatty acid methyl ester profiles and rDNA sequence analyses. Yeasts or yeastlike organisms were detected at frequencies similar to or greater than bacteria, particularly in hypanthia. When microbial isolates were tested for their capacity to suppress Erwinia amylovora on stigmas of detached crab apple flowers, many were ineffective. The best antagonists were the bacteria Pantoea agglomerans and Pseudomonas spp. and a few yeasts identified as Cryptococcus spp. Further evaluation of these taxa on flowers could lead to the discovery of additional biocontrol agents for fire blight.

scholarly journals Rates of Epiphytic Growth of Erwinia amylovora on Flowers Common in the Landscape

Plant Disease ◽  
2006 ◽  
Vol 90 (10) ◽  
pp. 1331-1336 ◽  
Author(s):  
K. B. Johnson ◽  
T. L. Sawyer ◽  
T. N. Temple
Keyword(s):  
Growth Rates ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Prunus Avium ◽  
Relative Growth ◽  
Relative Growth Rates ◽  
Scotch Broom ◽  
Rubus Armeniacus ◽  
Population Sizes ◽  
Pollinating Insects

We evaluated epiphytic growth of the fire blight bacterium, Erwinia amylovora, on flowers of plant species common to landscapes where pears and apples are grown. The plants were from genera regarded as important nectar and pollen sources for pollinating insects: Acer, Amelanchier, Brassica, Cytisus, Populus, Prunus, Rubus, Salix, Taraxacum, Trifolium, and Symphoricarpos. Floral bouquets were inoculated with E. amylovora and incubated in growth chambers at 15°C for 96 h. Regardless of their susceptibility to fire blight, all species from the rose family except Prunus domestica (European plum) supported epiphytic populations of E. amylovora that exceeded 1 × 106 CFU/flower with relative growth rates for the populations that averaged 7% per hour. Nonrosaceous plants were generally poor supporters of epiphytic growth of the fire blight pathogen with relative growth rates averaging <4% per hour. In two seasons of field inoculations, the rosaceous non-disease-host plants, Prunus avium (sweet cherry) and Rubus armeniacus (Himalayan blackberry), yielded mean population sizes of E. amylovora that exceeded 1 × 106 CFU/flower; in contrast, at 8 days after inoculation, mean population sizes of the pathogen were in the range of 5 × 103 to 5 × 104 CFU/flower on Cytisus scoparius (Scotch broom) and <1 × 102 CFU on Acer macrophylum (big leaf maple). Because vectors of E. amylovora, principally bees, visit many kinds of flowers in landscape areas between pear and apple orchards, flowers of rosaceous, non-disease-host species could serve as potential sites of inoculum increase during their periods of bloom.

scholarly journals Fire blight (Erwinia amylovora) susceptibility of old Hungarian apple cultivars

2005 ◽  
Vol 11 (3) ◽  
Author(s):  
M. Tóth ◽  
K. Honty ◽  
M. Hevesi
Keyword(s):  
Powdery Mildew ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Latent Infection ◽  
Bacterial Cells ◽  
Potted Plants ◽  
Study Evaluation ◽  
Resistant Sources ◽  
Apple Cultivars ◽  
One Year

The aim of the Hungarian apple breeding program started in the Department of Fruit Science was to find resistant apple cultivars against major diseases (scab, powdery mildew, fire blight). The outbreak of fire blight (Erwinia amylovora) in 1996 motivated us to search new resistant sources principally from old traditional apple cultivars. First of all, cultivars have been gathered since 1997 from Carpathia (Visk) and evaluated between 2001 and 2003. In this recent study evaluation of resistance of old Hungarian genotypes to fire blight collected from the English National Fruit Collection (Brogdale, Faversham) is presented. 13 old Hungarian apple cultivars in 2002, and 38 genotypes in 2005 have been evaluated. We used `Idared' and 'Jonathan M41' as susceptible controls and 'Liberty' and 'Remo' as resistant ones. Shoots of two-year-old potted plants were inoculated with a mixture of virulent E. amylovora isolates (Ea2, Ea60, Ea67) at a concentration of 5 x 108 cells/ml. Resistance of apple cultivars was evaluated weekly, four times after inoculation by disease severity of symptoms. Numbers of bacterial colonies in 1 cm length shoot were determined in the fourth week after infection. 8 cultivars in 2002 and 9 cultivars in 2005 displayed notable resistance to fire blight based on one-year data. Based on the coincident data of both years, out of the cultivars collected also from Carpathia ‘Pónyik', and `Sikulai' were found to be resistant and gene sources additional old Hungarian valuable apple genotypes could be selected: `Szabadkai szercsika' and `Tordai piros !davit'. The cultivar `Szemes alma' originated from Visk has proved to be recurrently resistant. The number of bacterial cells in shoots of the investigated cultivars correlated with the severity of symptoms. With this method, which was introduced by us earlier, we can screen cultivars displaying weak visible symptoms, which cannot be proposed as a source of resistance because of their latent infection.

scholarly journals Antibiosis Contributes to Biological Control of Fire Blight by Pantoea agglomerans Strain Eh252 in Orchards

2002 ◽  
Vol 92 (11) ◽  
pp. 1202-1209 ◽  
Author(s):  
V. O. Stockwell ◽  
K. B. Johnson ◽  
D. Sugar ◽  
J. E. Loper
Keyword(s):  
Biological Control ◽  
Fire Blight ◽  
Field Trials ◽  
Pantoea Agglomerans ◽  
Disease Suppression ◽  
Habitat Modification ◽  
Bacterial Disease ◽  
Apple Trees ◽  
Freeze Dried ◽  
Population Sizes

Fire blight, caused by Erwinia amylovora, is the most serious bacterial disease of pear and apple trees. Biological control with strains of Pantoea agglomerans (syn. Erwinia herbicola) may provide an effective disease management strategy for fire blight. Most strains of P. agglomerans evaluated for suppression of fire blight produce compounds that inhibit the growth of E. amylovora in culture. The role of these inhibitory compounds in fire blight suppression in orchard environments has not been studied. In seven field trials in Oregon, we compared the population dynamics and disease suppression with P. agglomerans Eh252, a strain that produces a single antibiotic, with its near-isogenic antibiotic-deficient derivative, strain 10:12. Water or suspensions of Eh252 or 10:12 (1 × 108 CFU/ml) were applied at 30 and 70% bloom to pear or apple trees. Aqueous suspensions of freeze-dried cells of E. amylovora (3 × 105 CFU/ml) were applied at full bloom. Additional trees were treated with streptomycin or oxytetracycline at 30 and 70% bloom and in some experiments, 1 day after application of the pathogen. Population sizes of Eh252 or 10:12 on pear blossoms were estimated by spreading dilutions of blossom washes on culture media. Average population sizes of Eh252 and 10:12 on blossoms ranged from 105 to 107 CFU, and in five of six trials, the relative area under the population curve of Eh252 was not significantly different than that of its derivative 10:12. Both Eh252 and 10:12 reduced the growth of the pathogen on blossoms compared with inoculated water-treated controls. Eh252 significantly decreased the incidence of fire blight in six of seven field trials compared with the incidence on water-treated trees, and 10:12 similarly reduced the incidence of fire blight in four of seven trials. In three of seven field trials, trees treated with Eh252 had a significantly lower incidence of fire blight compared with trees treated 3 with 10:12. Overall,3 Eh252 reduced the incidence of fire blight by 55 ± 8%, 10:12 by 30 ± 6%, streptomycin by 75 ± 4%, and oxytetracycline by 16 ± 14%. The effectiveness of strain 10:12 compared with water treatment indicates that other mechanisms (e.g., competitive exclusion or habitat modification) also contribute to disease suppression by P. agglomerans. The increased suppression of fire blight by the parental strain Eh252 compared with the antibiotic-deficient mutant 10:12 indicates that antibiosis is an important mechanism of biological control of fire blight.

scholarly journals Duplex Real-Time Polymerase Chain Reaction Reveals Competition Between Erwinia amylovora and E. pyrifoliae on Pear Blossoms

2008 ◽  
Vol 98 (6) ◽  
pp. 673-679 ◽  
Author(s):  
Susan M. Lehman ◽  
Won-Sik Kim ◽  
Alan J. Castle ◽  
Antonet M. Svircev
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Erwinia Amylovora ◽  
Single Species ◽  
Bacterial Cells ◽  
Chain Reaction ◽  
Causative Agents ◽  
Population Sizes ◽  
Polymerase Chain ◽  
Erwinia Species

Erwinia amylovora and E. pyrifoliae are the causative agents of fire blight and Asian pear blight, respectively. The pathogens are closely related, with overlapping host ranges. Data are unavailable on the current distribution of E. pyrifoliae and on the interaction between the two species when they are present together on the same host. In this study, a duplex real-time polymerase chain reaction (PCR) protocol was developed to monitor the population dynamics of E. amylovora and E. pyrifoliae on the surface of Bartlett pear blossoms. Bacterial cells washed from blossoms were used directly as the PCR template without DNA extraction. Primers and a probe based on the E. amylovora levansucrase gene detected all E. amylovora strains. All E. pyrifoliae strains, including the Japanese Erwinia strains previously described as E. amylovora, were detected with a primer and probe combination based on the E. pyrifoliae hrpW gene. Disease development and severity were not significantly different in blossoms inoculated with individual Erwinia species or with a mixture of the two species. However, E. amylovora grew to greater population sizes than did E. pyrifoliae in both single species inoculations and in mixtures, suggesting that E. amylovora has a greater competitive fitness on Bartlett pear blossoms than E. pyrifoliae.

scholarly journals Field trials with non-bactericide products to control fire blight in apple orchards

2004 ◽  
Vol 10 (2) ◽  
Author(s):  
T. Bubán ◽  
P. Sallai ◽  
J. Deme
Keyword(s):  
Active Ingredient ◽  
Fire Blight ◽  
Erwinia Amylovora ◽  
Field Trials ◽  
The Other ◽  
European Countries ◽  
Nutrient Elements ◽  
Chemical Agents ◽  
Shoot Blight ◽  
In Fire

Recently, novel strategies and chemical agents for prophylactic protection against the bacterial (Erwinia amylovora) disease fire blight are being sought. Resistance-inducing compounds, such as prohexadione-Ca represent promising alternatives. Prohexadione-Ca is the active ingredient of the bioregulator Regalis, currently being introduced in several European countries and overseas. Another product used in this study was Biomit Plussz, a leaf fertiliser providing harmonic supply of nutrient elements, the complete supply is assumed to improve the tolerance against diseases. Treatments' effects of both of these products were compared to the effectiveness of treatments with antibiotics repeated twice, three or four times a season. In the years of 2001 and 2002, the effectiveness of both Regalis and Biomit Plussz in reduction of incidence of shoot blight was similar, or proved to be superior to the check treatments consisting of repeated sprayings of antibiotics. Last year (2003) treatments of streptomycin resulted — although within the same magnitude — in a somewhat better control of shoot blight than sprayings with the other compounds. As regards severity of blossom blight, inconsistent results were recorded concerning both Regalis and Biomit Plussz. In general, prohexadione-Ca is less efficient for controlling flower infection by E. amylovora as compared to shoot infections, since successful prophylactic treatments are difficult to carry out early in the season. The highest effectiveness in fire blight management can, therefore, be achieved by using prohexadione-Ca (as preventive protection) in combination with streptomycin or other suitable antibiotics (as curative protection).

scholarly journals Biological Control Agents for Fire Blight of Apple Compared Under Conditions Limiting Natural Dispersal

Plant Disease ◽  
2002 ◽  
Vol 86 (6) ◽  
pp. 639-644 ◽  
Author(s):  
P. L. Pusey
Keyword(s):  
Fire Blight ◽  
Erwinia Amylovora ◽  
Disease Incidence ◽  
Field Trials ◽  
Microbial Populations ◽  
Biological Control Agents ◽  
Two Factors ◽  
Statistical Separation ◽  
Natural Dispersal ◽  
Potential Use

The efficacy of Pantoea agglomerans strain E325 for control of fire blight of apple was determined in comparative field trials involving other bacterial antagonists. Concurrently, the importance of the natural dispersal of bacteria as a complicating factor was assessed. Tests were performed under two sets of conditions, those that allowed for the dispersal of bacteria via honeybee activity and those that minimized it through the use of translucent polyethylene enclosures around single trees. The enclosures also raised daytime temperatures and allowed for controlled wetting, two factors important to the development of blossom blight. Singleantagonist treatments with strain E325, Pseudomonas fluorescens strain A506, and P. agglomerans strain C9-1 were applied to open blossoms on each of 10 enclosed trees and 10 nonenclosed trees. During bloom, suspensions of antagonists (108 CFU/ml) were applied twice with a brush, and a suspension of Erwinia amylovora (107 CFU/ml) was subsequently applied once using the same method. Two days after inoculation with the pathogen, trees were misted to simulate precipitation. Flower-to-flower spread of antagonistic bacteria was less frequent on trees surrounded by plastic enclosures than on nonenclosed trees. The range and statistical separation of means for population size of E. amylovora and disease incidence among treatments were greater for enclosed trees than for nonenclosed trees. Based on these results, the natural spread of antagonists being compared may mask differences in their efficacy as biocontrol agents. Such distortions and resulting misinterpretations could be lessened by separating treatments widely in large orchard blocks and by monitoring microbial populations. Strain E325 from fresh or lyophilized cultures was consistently more effective than standard antagonists in suppressing E. amylovora and reducing disease incidence. To fully assess its potential use for fire blight, larger-scale trials under various conditions will be necessary.

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