RESUMO
Treating plant bacterial diseases is notoriously difficult because of the lack of available antimicrobials. Pseudomonas syringae pathovar syringae (Pss) is a major pathogen of cherry (Prunus avium) causing bacterial canker of the stem, leaf and fruit, impacting productivity and leading to a loss of trees. In an attempt to find a treatment for this disease, naturally occurring bacteriophage (phage) that specifically target Pss is being investigated as a biocontrol strategy. However, before using them as a biocontrol treatment, it is important to both understand their efficacy in reducing the bacterial population and determine if the bacterial pathogens can evolve resistance to evade phage infection. To investigate this, killing curve assays of five MR phages targeting Pss showed that phage resistance rapidly emerges in vitro, even when using a cocktail of the five phages together. To gain insight to the changes occurring, Pss colonies were collected three times during a 66-h killing curve assay and separately, Pss and phage were also coevolved over 10 generations, enabling the measurement of genomic and fitness changes in bacterial populations. Pss evolved resistance to phages through modifications in lipopolysaccharide (LPS) synthesis pathways. Bacterial fitness (growth) and virulence were affected in only a few mutants. Deletion of LPS-associated genes suggested that LPS was the main target receptor for all five MR phages. Later generations of coevolved phages from the coevolution experiment were more potent at reducing the bacterial density and when used with wild-type phages could reduce the emergence of phage-resistant mutants. This study shows that understanding the genetic mechanisms of bacterial pathogen resistance to phages is important for helping to design a more effective approach to kill the bacteria while minimizing the opportunity for phage resistance to manifest.
Assuntos
Doenças das Plantas , Pseudomonas syringae , Pseudomonas syringae/virologia , Pseudomonas syringae/genética , Doenças das Plantas/microbiologia , Fagos de Pseudomonas/genética , Fagos de Pseudomonas/fisiologia , Bacteriófagos/genética , Bacteriófagos/fisiologiaRESUMO
The genus Rhodococcus contains several species with agricultural, biotechnological and ecological importance. Within this genus, many phyllosphere, rhizosphere and endosphere strains are plant growth promoting bacteria, whereas strains designated as R. fascians are plant pathogens. In this study, we isolated 47 Rhodococcus strains from a range of herbaceous and woody plant species. Phylogenetic analysis based on 16S rDNA, gyrB and alkB genes was used to compare our strains with type strains of Rhodococcus. For most of our strains, sequence similarity of the 16S rDNA, gyrB and alkB regions to type strains ranged from 98-100â%. Results of the concatenated gene sequence comparisons identified 18 strains of R. fascians and three strains of R. kroppenstedtii. The remaining strains were unclassified, and may represent novel species of Rhodococcus. Phylogenetic analysis based on gyrB sequences provided a more precise classification of our strains to species level than 16S rDNA sequences, whereas analysis of alkB sequences was unable to identify strains with orange-coloured colonies to species level.
Assuntos
Filogenia , Plantas/microbiologia , Rhodococcus/classificação , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Genes Bacterianos , Pigmentação , RNA Ribossômico 16S/genética , Rhodococcus/isolamento & purificação , Análise de Sequência de DNA , TunísiaRESUMO
The draft genome sequences of plant-associated Rhodococcus spp. from Tunisia are reported here. Two Rhodococcus fascians strains were obtained from almond rootstocks, and one Rhodococcus kroppenstedtii strain was obtained from a pistachio tree. The fourth Rhodococcus sp. strain was isolated from an ornamental plant.
