Dataset Information

Antibiotic production by Erwinia herbicola Eh1087: its role in inhibition of Erwinia amylovora and partial characterization of antibiotic biosynthesis genes.

ABSTRACT: Mutants of Erwinia herbicola Eh1087 (Ant-), which did not produce antibiotic activity against Erwinia amylovora, the fire blight pathogen, were selected after TnphoA mutagenesis. In immature pear fruit Ant- mutants grew at the same rate as wild-type strain Eh1087 but did not suppress development of the disease caused by E. amylovora. These results indicated that antibiosis plays an important role in the suppression of disease by strain Eh1087. All of the Ant- mutations obtained were located in a 2.2-kb region on a 200-kb indigenous plasmid. Sequence analysis of the mutated DNA region resulted in identification of six open reading frames, designated ORF1 through ORF6, four of which were essential to antibiotic expression. One gene was identified as a gene which encodes a translocase protein which is probably involved in antibiotic secretion. A sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of plasmid proteins produced in Escherichia coli minicells confirmed the presence of proteins whose sizes corresponded to the sizes of the predicted open reading frame products.

SUBMITTER: Kearns LP

PROVIDER: S-EPMC106239 | biostudies-literature | 1998 May

REPOSITORIES: biostudies-literature

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Antibiotic production by Erwinia herbicola Eh1087: its role in inhibition of Erwinia amylovora and partial characterization of antibiotic biosynthesis genes.

Kearns L P LP Mahanty H K HK

Applied and environmental microbiology 19980501 5

Mutants of Erwinia herbicola Eh1087 (Ant-), which did not produce antibiotic activity against Erwinia amylovora, the fire blight pathogen, were selected after TnphoA mutagenesis. In immature pear fruit Ant- mutants grew at the same rate as wild-type strain Eh1087 but did not suppress development of the disease caused by E. amylovora. These results indicated that antibiosis plays an important role in the suppression of disease by strain Eh1087. All of the Ant- mutations obtained were located in a ...[more]

PMID: 9572960

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Project description:Cyclic di-GMP (c-di-GMP) is a ubiquitous bacterial second messenger molecule that is an important virulence regulator in the plant pathogen Erwinia amylovora Intracellular levels of c-di-GMP are modulated by diguanylate cyclase (DGC) enzymes that synthesize c-di-GMP and by phosphodiesterase (PDE) enzymes that degrade c-di-GMP. The regulatory role of the PDE enzymes in E. amylovora has not been determined. Using a combination of single, double, and triple deletion mutants, we determined the effects of each of the four putative PDE-encoding genes (pdeA, pdeB, pdeC, and edcA) in E. amylovora on cellular processes related to virulence. Our results indicate that pdeA and pdeC are the two phosphodiesterases most active in virulence regulation in E. amylovora Ea1189. The deletion of pdeC resulted in a measurably significant increase in the intracellular pool of c-di-GMP, and the highest intracellular concentrations of c-di-GMP were observed in the Ea1189 ?pdeAC and Ea1189 ?pdeABC mutants. The regulation of virulence traits due to the deletion of the pde genes showed two patterns. A stronger regulatory effect was observed on amylovoran production and biofilm formation, where both Ea1189 ?pdeA and Ea1189 ?pdeC mutants exhibited significant increases in these two phenotypes in vitro In contrast, the deletion of two or more pde genes was required to affect motility and virulence phenotypes. Our results indicate a functional redundancy among the pde genes in E. amylovora for certain traits and indicate that the intracellular degradation of c-di-GMP is mainly regulated by pdeA and pdeC, but they also suggest a role for pdeB in regulating motility and virulence.IMPORTANCE Precise control of the expression of virulence genes is essential for successful infection of apple hosts by the fire blight pathogen, Erwinia amylovora The presence and buildup of a signaling molecule called cyclic di-GMP enables the expression and function of some virulence determinants in E. amylovora, such as amylovoran production and biofilm formation. However, other determinants, such as those for motility and the type III secretion system, are expressed and functional when cyclic di-GMP is absent. Here, we report studies of enzymes called phosphodiesterases, which function in the degradation of cyclic di-GMP. We show the importance of these enzymes in virulence gene regulation and the ability of E. amylovora to cause plant disease.

Project description:Erwinia amylovora is a quarantine phytopathogenic bacterium that is the causal agent of fire blight, a destructive disease responsible for killing millions of fruit-bearing plants worldwide, including apple, pear, quince, and raspberry. Efficient and sustainable control strategies for this serious bacterial disease are still lacking, and traditional methods are limited to the use of antibiotics and some basic agricultural practices. This study aimed to contribute to the development of a sustainable control strategy through the identification, characterization, and application of bacteriophages (phages) able to control fire blight on pears. Phages isolated from wastewater collected in the Apulia region (southern Italy) were characterized and evaluated as antibacterial agents to treat experimental fire blight caused by E. amylovora. Transmission electron microscopy (TEM) conducted on purified phages (named EP-IT22 for Erwinia phage IT22) showed particles with icosahedral heads of ca. 90 ± 5 nm in length and long contractile tails of 100 ± 10 nm, typical of the Myoviridae family. Whole genome sequencing (WGS), assembly, and analysis of the phage DNA generated a single contig of 174.346 bp representing a complete circular genome composed of 310 open reading frames (ORFs). EP-IT22 was found to be 98.48% identical to the Straboviridae Erwinia phage Cronus (EPC) (GenBank Acc. n° NC_055743) at the nucleotide level. EP-IT22 was found to be resistant to high temperatures (up to 60 °C) and pH values between 4 and 11, and was able to accomplish a complete lytic cycle within one hour. Furthermore, the viability-qPCR and turbidity assays showed that EP-IT22 (MOI = 1) lysed 94% of E. amylovora cells in 20 h. The antibacterial activity of EP-IT22 in planta was evaluated in E. amylovora-inoculated pear plants that remained asymptomatic 40 days post inoculation, similarly to those treated with streptomycin sulphate. This is the first description of the morphological, biological, and molecular features of EP-IT22, highlighting its promising potential for biocontrol of E. amylovora against fire blight disease.

Dataset Information

Antibiotic production by Erwinia herbicola Eh1087: its role in inhibition of Erwinia amylovora and partial characterization of antibiotic biosynthesis genes.

Publications

Antibiotic production by Erwinia herbicola Eh1087: its role in inhibition of Erwinia amylovora and partial characterization of antibiotic biosynthesis genes.

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