Serratia Plymuthica Strain Research Articles

Complete genome sequence of the rapeseed plant-growth promoting Serratia plymuthica strain AS9.

Serratia plymuthica are plant-associated, plant beneficial species belonging to the family Enterobacteriaceae. The members of the genus Serratia are ubiquitous in nature and their life style varies from endophytic to free-living. S. plymuthica AS9 is of special interest for its ability to inhibit fungal pathogens of rapeseed and to promote plant growth. The genome of S. plymuthica AS9 comprises a 5,442,880 bp long circular chromosome that consists of 4,952 protein-coding genes, 87 tRNA genes and 7 rRNA operons. This genome is part of the project entitled “Genomics of four rapeseed plant growth promoting bacteria with antagonistic effect on plant pathogens” awarded through the 2010 DOE-JGI Community Sequencing Program (CSP2010).

Screening bacterial species for antagonistic activities against the Sclerotinia sclerotiorum (Lib.) De Bary causal agent of cucumber white mold disease

In this study, 23 bacteria strains that belong to 19 bacterial species were tested against Sclerotinia sclerotiorum (Lib.) De Bary. In vivo and in vitro testing of bacterial strains showed that Serratia plymuthica strains IK-150 and IK-139, Burkholderia cepacia strain IK-16, Pseudomonas flourocens s train IK-3, Pseudomonas putida strain IK-1, Paenibacillus macerans strain IK-36, Pantoea agglomerans strain IK-147 and Burkholderia pyronici a strain IK-145 totally inhibited mycelial growth and caused loss of viability of sclerotia. The other tested bacterial strains also reduced mycelial growth of the fungus development. These results indicate that bacterial species used in this study could be used in the control of S. sclerotiorum . Key words : Sclerotinia sclerotiorum, white mold, biological control, bacteria.

Characterisation of two quorum sensing systems in the endophytic Serratia plymuthica strain G3: differential control of motility and biofilm formation according to life-style.

BackgroundN-acylhomoserine lactone (AHL)-based quorum sensing (QS) systems have been described in many plant-associated Gram-negative bacteria to control certain beneficial phenotypic traits, such as production of biocontrol factors and plant growth promotion. However, the role of AHL-mediated signalling in the endophytic strains of plant-associated Serratia is still poorly understood. An endophytic Serratia sp. G3 with biocontrol potential and high levels of AHL signal production was isolated from the stems of wheat and the role of QS in this isolate was determined.ResultsStrain G3 classified as Serratia plymuthica based on 16S rRNA was subjected to phylogenetic analysis. Using primers to conserved sequences of luxIR homologues from the Serratia genus, splIR and spsIR from the chromosome of strain G3 were cloned and sequenced. AHL profiles from strain G3 and Escherichia coli DH5α expressing splI or spsI from recombinant plasmids were identified by liquid chromatography-tandem mass spectrometry. This revealed that the most abundant AHL signals produced by SplI in E. coli were N-3-oxo-hexanoylhomoserine lactone (3-oxo-C6-HSL), N-3-oxo-heptanoylhomoserine lactone (3-oxo-C7-HSL), N-3-hydroxy-hexanoylhomoserine lactone (3-hydroxy-C6-HSL), N-hexanoylhomoserine lactone (C6-HSL), and N-heptanoyl homoserine lactone (C7-HSL); whereas SpsI was primarily responsible for the synthesis of N-butyrylhomoserine lactone (C4-HSL) and N-pentanoylhomoserine lactone (C5-HSL). Furthermore, a quorum quenching analysis by heterologous expression of the Bacillus A24 AiiA lactonase in strain G3 enabled the identification of the AHL-regulated biocontrol-related traits. Depletion of AHLs with this lactonase resulted in altered adhesion and biofilm formation using a microtiter plate assay and flow cells coupled with confocal laser scanning microscopy respectively. This was different from the closely related S. plymuthica strains HRO-C48 and RVH1, where biofilm formation for both strains is AHL-independent. In addition, QS in G3 positively regulated antifungal activity, production of exoenzymes, but negatively regulated production of indol-3-acetic acid (IAA), which is in agreement with previous reports in strain HRO-C48. However, in contrast to HRO-C48, swimming motility was not controlled by AHL-mediated QS.ConclusionsThis is the first report of the characterisation of two AHL-based quorum sensing systems in the same isolate of the genus Serratia. Our results show that the QS network is involved in the global regulation of biocontrol-related traits in the endophytic strain G3. However, although free-living and endophytic S. plymuthica share some conservation on QS phenotypic regulation, the control of motility and biofilm formation seems to be strain-specific and possible linked to the life-style of this organism.

Induction of systemic resistance, root colonisation and biocontrol activities of the rhizospheric strain of Serratia plymuthica are dependent on N-acyl homoserine lactones

Quorum sensing regulation, mediated by N-acyl homoserine lactone signals, produced by strain Serratia plymuthica HRO-C48 isolated from the rhizosphere of oilseed rape, was found to be responsible for this strain’s ability to produce the broad spectrum antibiotic pyrrolnitrin. In this study, we have shown that some other biocontrol-related traits of strain HRO-C48, such as protection of cucumbers against Pythium apahnidermatum damping-off disease, induced systemic resistance to Botrytis cinerea grey mold in bean and tomato plants, and that colonisation of the rhizosphere also depends on AHL signalling. The results prove that quorum sensing regulation may be generally involved in interactions between plant-associated bacteria, fungal pathogens and host plants.

Phytase activity and its regulation in a rhizospheric strain of Serratia plymuthica

Serratia plymuthica strain IC1270 isolated from the rhizosphere, possessing antagonistic activity towards a wide range of plant-pathogenic fungi, is able to hydrolyze phytate. Phytase activity was found intracellularly, while no activity was detected in the culture liquid. Optimum activity was found at pH 4-5; it completely disappeared at pH > 7.0 and 2.5. Phytase production was practically absent in the exponential phase and reached a maximum in the late stationary phase. Mutations of genes grrA and grrS, encoding GacA/GacS-like 2-component global regulatory system, or in gene rpoS encoding the sigma factor RpoS subunit of RNA polymerase, led to a deficiency in phytase production. Introduction into mutants of the respective wild-type genes cloned into the wide-range plasmid pJFF224-NX under the control of the bacteriophage T4 gene 32 promoter complemented this deficiency. This is the first report implicating the GacA/GacS global regulators and RpoS factor in phytase production in bacteria.

An effective biocontrol bioformulation against Phytophthora blight of pepper using growth mixtures of combined chitinolytic bacteria under different field conditions

Phytophthora blight of pepper caused by Phytophthora capsici has devastating consequences when combined with other pathogens, including Rhizoctonia solani, Fusarium oxysporum, and Fusarium solani. In order to develop a field-effective biocontrol strategy against Phytophthora blight of pepper, three chitinolytic bacteria, Serratiaplymuthica strain C-1, strongly antagonistic to P. capsici, Chromobacterium sp. strain C-61, strongly antagonistic to R. solani, and Lysobacter enzymogenes strain C-3, antagonistic to R. solani and Fusarium spp., were selected. In pot studies, application of cultures combining the three bacterial strains effectively suppressed Phytophthora blight more than application of any single bacterial strain. Bioformulations developed from growth of the strains in a simple medium containing chitin under large batch conditions resulted in effective control in field applications. Efficacy of the bioformulated product depended on both the dose and timing of application. Although the undiluted product suppressed Phytophthora blight under all field conditions, a 10-fold diluted product was effective in solar-sterilized greenhouses and in fields with crop rotation. These results suggest that the developed product could be a new effective system to control Phytophthora blight disease in pepper.

Mineralization of phenol by a Serratia plymuthica strain GC isolated from sludge sample

Abstract A bacterial strain isolated from a contaminated soil degraded phenol as the sole source of carbon and energy. It was identified as Serratia plymuthica with the help of its biochemical, physiological and morphological characteristics. The bacterium was able to tolerate and degrade phenol up to a concentration of 1050 mg l−1. High concentration of phenol in minimal medium had inhibitory effect on bacteria. The bacterium was able to degrade other aromatic compounds like benzoic acid, ortho-, meta-, para-cresol, protocatechuate, catechol and tryptophan. Phenol was degraded through ortho-pathway. Crude cell extract showed presence of ring cleavage enzyme catechol 1,2-dioxygenase. Specific activity of catechol 1,2-dioxygenase was 0.134 μmole min−1 mg−1 protein. Effect of phenol as inhibitory substrate show that higher the concentration of phenol in the medium more is the negative effect on growth of bacteria and subsequent biodegradation. This bacterium grew well in pH range of 5 to 9 and temperature range of 30 to 40 °C. Presence of other carbon source like glucose in the phenol minimal medium affects negatively the rate of degradation of phenol but increases the specific growth rate of the bacteria.

Characterization of a luxI/ luxR-type quorum sensing system and N-acyl-homoserine lactone-dependent regulation of exo-enzyme and antibacterial component production in Serratia plymuthica RVH1

Quorum sensing by means of N-acyl- l-homoserine lactones (AHLs) is widespread in Gram-negative bacteria, where diverse AHLs influence a wide variety of functions, even in a single genus such as Serratia. Here we report the identification and characterization of the quorum sensing system of Serratia plymuthica strain RVH1. This strain isolated from a raw vegetable processing line produces at least three AHLs which were identified as N-butanoyl- (C4-HSL), N-hexanoyl- (C6-HSL) and N-(3-oxo-hexanoyl)-homoserine lactone (3-oxo-C6-HSL). The identified LuxI homolog SplI synthesizes 3-oxo-C6-HSL, and influences the production of C4-HSL and C6-HSL, as splI gene inactivation resulted in loss of 3-oxo-C6-HSL production and smaller amounts of C4-HSL and C6-HSL produced. SplI-dependent quorum sensing controls 2,3-butanediol fermentation (previously reported) and the production of an extracellular chitinase, nuclease, protease and antibacterial compound. The identity of the latter is not yet elucidated, but appears to be different from the known antibacterial compounds produced by Serratia strains. SplR, the homolog of the LuxR regulator, appears to act as a repressor of synthesis of extracellular enzymes and antibacterial compound and to autorepress its own expression, probably by binding to a 21 bp lux box sequence.

Control of green and blue mould on orange fruit by Serratia plymuthica strains IC14 and IC1270 and putative modes of action

Two biological control agents of Serratia plymuthica, strains IC1270 and IC14 applied separately and in combination were evaluated for suppressing Penicillium digitatum (green mould) or Penicillium italicum (blue mould) on orange. These bacteria were effective and controlled both pathogens at 1 × 10 8 cells/mL. Disease suppression was increased when both bacterial strains were combined. Possible modes of action studied in this work were antibiosis, chitinolytic activity and competition for nutrients. Two mutants of strain IC1270, one deficient in chitinolytic activity and the second deficient in pyrrolnitrin production were obtained by the gene replacement technique. On orange fruit, mutant IC1270-C7 deficient in chitinase production and mutant IC1270-P1 deficient in pyrrolnitrin production showed a similar efficiency against P. digitatum to the parental strain. However, in vitro IC1270-P1 lost its antifungal activity and no inhibition zone was observed when it was tested against P. digitatum or P. italicum. In similar experiments, a chitinase-deficient mutant of strain IC14 was as effective as the parental strain IC14, suggesting no evidence for a possible role of chitinases in controlling green mould caused by P. digitatum. Interactions between strains IC1270 or IC14 and P. digitatum were studied in tissue culture plates with diluted orange peel extract as the nutrient source. Strain IC1270 decreased germination of P. digitatum conidia when it was physically separated from the pathogen by a membrane filter, which permits nutrient and metabolite interchange, while strain IC14 did not affect germination. Significant inhibition of conidial germination of P. digitatum was achieved, however, when the pathogen and IC14 were in physical contact. Competition for nutrients appears to be the main mode of action of strain IC1270, while a direct cell-to-cell interaction between IC14 and the pathogen is needed for antagonism.

Field performance of a weed-suppressing Serratia plymuthica strain applied with conventional spraying equipment

In previous glasshouse experiments, the soilbacterium Serratia plymuthica, strainA153, showed strong growth-suppressingactivities against a range of broad-leavedweeds after foliar spraying. In field tests ofthis strain in spring wheat, spring barley andpotatoes, variable effects were achieved on arange of weeds including Chenopodiumalbum, Stellaria media, Polygonumconvolvulus and Galeopsis speciosa. Atone site, good suppression of C. albumwas observed when the strain was applied in atank mix with another bacterial isolate or withreduced doses of a herbicide. Effects on weedsappeared to be independent of the applicationvolume (1000, 600, 500 l ha−1), but weedswere in some cases more strongly suppressed athigher bacterial doses. Barley yields weresomewhat reduced by the bacterial application,but wheat yields were less affected. AlthoughS. plymuthica suppressed certain weedswhen applied in the field in a simple aqueousformulation and with conventional sprayingequipment, the level of weed suppression wasunsatisfactory from a practical standpoint.

In vitro and In vivo Activities of a Biocontrol Agent, Serratia plymuthica A2l-4, Against Phytophthora capsici

In vitro and in vivo activities of a biocontrol agent, Serratia plymuthica strain A2l-4, was evaluated for the control of Phytophthora blight of pepper, Strain A2l-4 inhibited mycelial growth, germination of zoosporangia and cystospores, and formation of zoospore and zoosporangia of Phytophthora capsici in vitro. In the pot experiment, incidence of Phytophthora blight of pepper in non-treated control was 100% at 14 days after inoculation, while no disease was observed in the plot treated with S. plymuthica A2l-4. In the greenhouse test, infection rate of pepper in the non-treated plots was 74.5%, while it was only 12.6％ in the plots treated with A2l-4. Results indicate that S. plymuthica A2l-4 is a potential biocontrol agent for Phytophthora blight of pepper.

Suppression of Sclerotinia sclerotiorum apothecial formation by the soil bacterium Serratia plymuthica: identification of a chlorinated macrolide as one of the causal agents

A selection of soil bacteria was screened for their ability to interfere with carpogenic germination of Sclerotinia sclerotiorum. Nine out of 300 bacterial isolates were found to significantly suppress apothecial formation. One of these isolates, identified as a strain of Serratia plymuthica, was highly effective in inducing complete suppression of apothecial formation at high concentrations, and also strongly inhibited the germination of ascospores as well as hyphal growth of S. sclerotiorum. A bioassay-guided purification procedure starting with the cell-free supernatant of the bacterial culture led to the identification of a chlorinated macrolide as an active compound able to induce the observed inhibition. Spectroscopic data showed the compound to be identical to haterumalide A. The data presented show the ability of this compound to inhibit apothecial formation and ascospore germination. Other possible mechanisms involved in inhibition of apothecial formation and mycelial and hyphal growth of S. sclerotiorum by the same isolate are discussed. The relevance of our observations to natural systems will be the subject of further research.

Biological Control of Fungal Strawberry Diseases by Serratia plymuthica HRO-C48.

To develop a biological control product for commercial strawberry production, the chitinolytic rhizobacterium Serratia plymuthica strain HRO-C48 was evaluated for plant growth promotion of strawberries and biological control of the fungal pathogens Verticillium dahliae and Phytophthora cactorum. In phytochamber experiments, treatment with S. plymuthica HRO-C48 resulted in a statistically significant enhancement of plant growth dependent on the concentration of the bacterium that was applied. In greenhouse trials, bacterial treatment reduced the percentage of Verticillium wilt (18.5%) and Phytophthora root rot (33.4%). In three consecutive vegetation periods, field trials were carried out in soil naturally infested by both soilborne pathogens on commercial strawberry farms located in various regions of Germany. Dipping plants in a suspension of S. plymuthica prior to planting reduced Verticillium wilt compared with the nontreated control by 0 to 37.7%, with an average of 24.2%, whereas the increase of yield ranged from 156 to 394%, with an average of 296%. Bacterial treatment reduced Phy-tophthora root rot by 1.3 to 17.9%, with an average of 9.6%, and increased strawberry yield by 60% compared with the nontreated control. Under field conditions, strain HRO-C48 survived at approximately log10 3 to 7 CFU/g of root in the strawberry rhizosphere at 14 months after root application. Although results of the field trials were influenced by pathogen inoculum density, cropping history of the field site, and weather conditions, S. plymuthica HRO-C48 successfully controlled wilt and root rot of strawberry.

Identification of environmental Serratia plymuthica strains with the new combo panels type 1S.

Automated systems are required when numerous samples need to be processed, offering both high through put and test of a multiple simultaneously. This study was performed to compare the MicroScan WalkAway automated identification system in conjunction with the new MicroScan Combo Neg Panels Type 1S with conventional biochemical methods for identifying ten environmental Serratia plymuthica strains. High correlation between both methods were observed for all the 21 tests evaluated, and the MicroScan system was found capable of correctly identifying all S. plymuthica strains tested. In all tests, the percentage of correlation was 100%, except in raffinose test (91%).

Bacterial-Mediated Induced Resistance in Cucumber: Beneficial Effect of the Endophytic Bacterium Serratia plymuthica on the Protection Against Infection by Pythium ultimum

ABSTRACT The potential of the endophytic bacterium Serratia plymuthica strain R1GC4 in stimulating defense reactions in cucumber (Cucumis sativus) seedlings inoculated with the soilborne pathogen Pythium ultimum was explored at the cellular level. Bacterial treatment prior to Pythium inoculation resulted in less seedling disease development as compared with that in nontreated control plants, in which typical root symptoms were visible by 3 days after inoculation with the pathogen. Histological investigations of root samples revealed striking differences in the extent of plant defense reactions between bacterized and nonbacterized plants. These observations were further confirmed at the ultrastructural level with the demonstration that restriction of fungal colonization to the outermost root tissues of bacterized seedlings correlated with the deposition of enlarged callose-enriched wall appositions at sites of potential pathogen penetration and the accumulation of an osmiophilic material in the colonized areas. Hyphae of the pathogen, surrounded by this electron-opaque material, exhibited considerable changes including cytoplasm disorganization and, in many cases, loss of the protoplasm. However, labeling with the beta-1,4-exoglucanase resulted in a regular labeling of Pythium cell walls, even at a time when these walls were entirely coated by the osmiophilic material. This material was also found to infiltrate into the invading hyphae to form either an internal coating of the cell wall or a network of polymorphic droplets in the area previously occupied by the cytoplasm. Cytochemical investigations revealed that callose, pectin, and cellulose appeared in the wall appositions. In addition, glucosides, lipids, and phenolics were detected in the electron-dense aggregates forming the core of most wall appositions. Finally, galactose residues were among the minor polysaccharidic compounds detected in the wall appositions. Evidence is provided in this study showing that treatment with S. plymuthica sensitizes susceptible cucumber plants to react more rapidly and more efficiently to Pythium attack through the formation of physical and chemical barriers at sites of potential fungal entry.

Evaluation of plant growth-promoting rhizobacteria for biological control of pythium root rot of cucumbers grown in rockwool and effects on yield

Three strains ofPseudomonas fluorescens (63-49, 63-28, and 15), one strain ofPseudomonas corrugata (13) and one strain ofSerratia plymuthica (R1GC4) were tested on rockwool-grown cucumbers for their ability to reduce Pythium root-rot caused byPythium aphanidermatum. These strains were previously selected for biocontrol ability from collections of >4000 bacteria. Strains 63-49 and 63-28 were tested on cucumber plants grown in rockwool in two replicatedPythium-inoculated trials conducted in British Columbia (B.C). Another inoculated, replicated trial was conducted in Quebec with all five strains. Cucumber yields (fruit number and weight) were measured over a ten-week harvest period. Strain 63-49 caused an early promotion of plant growth and increased cucumber yields at early harvests. No measurable effect ofPythium inoculation on disease development was observed in the Quebec trial, due to unfavourable cool weather. However, 63-49 significantly increased the total number of cucumbers (12%) and cucumber weight (18%), compared to the non-treated control. Strains 13, 15 and R1GC4 slightly increased the cumulative cucumber yields, but strain 63-28 had no effect. In the B.C. trial, inoculation withP. aphanidermatum reduced the number and weight of cucumbers by 27%. Treatments ofPythium-inoculated cucumbers with 63-49 significantly increased fruit number and weight by 18%, compared to thePythium-inoculated control. Strain 63-28 increased the cumulative number of cucumbers over time, compared to thePythium-inoculated control, but the increase was less than with 63-49. The use ofPseudomonas spp. in rockwool-grown cucumbers can increase yields, both in the presence and absence of Pythium root rot, and with variable seasonal conditions and disease pressures.

Biocontrol of post‐harvest fungal diseases on Dutch white cabbage by Pseudomonas and Serratia antagonists in storage trials

Post‐harvest rotting of Dutch white cabbage was reduced by post‐harvest treatment with Pseudomonas fluorescens strains CL42, CL66, CL82, Serratia plymuthica strain CL43 or Serratia liquefaciens CL80, in three storage trials carried out in an experimental cold store at Manchester University, Manchester, UK and in a commercial cold store at L.W. van Geest Farms Ltd, Spalding, UK. The amount of surface area covered by fungal growth was assessed at 6‐week intervals during storage and the trimming losses were determined after 8 to 10 months. Only strains CL80 and CL82 were found to reduce fungal spoilage significantly in all three trials, and control by strain CL82 was similar to that achieved by post‐harvest treatment with fungicides. In the commercial cold store, CL42 showed better results than any of the other bacterial strains.

Structural and serological characterisation of an O-specific polysaccharide from Serratia plymuthica

The surface polysaccharides of a strain of Serratia plymuthica were characterised and shown to consist of a linear, acidic galactoglucomannan as well as a major and a minor neutral galactan. Immunoblotting results demonstrated cross-reactions between this strain and others with similar galactans (S. marcescens O16 and O20, Klebsiella O1, and Pasteurella haemolytica T4 and T10).

Biocontrol of Botrytis cinerea and Alternaria brassicicola on Dutch white cabbage by bacterial antagonists at cold‐store temperatures

Bacterial antagonists against Botrytis cinerea were isolated from different Brassica spp. and identified. All isolates showing in vitro antagonism at 4°C were shown to be either fluorescent pseudomonads or Serratia spp. In vitro antagonism against B. cinerea and Alternaria brassicicola was found to depend on the temperature and concentration of nutrients in the medium.Bacterial strains which showed in vitro antagonism were tested for in vivo antagonism at 4°C against B. cinerea and A. brassicicola using a leaf disc bioassay. Pseudomonas fluorescens isolates CL42, CL66, CL82 and Serratia plymuthica strain CL43 showed inhibition of Botrytis growth on leaf discs; P. fluorescens isolate CL74 and all Serratia liquefaciens isolates exhibited intermediate control. All other fluorescent pseudomonad isolates showed poor control or caused rotting of the cabbage tissue.

Isolation of CB-25-I, an antifungal antibiotic, from Serratia plymuthica.

A new antifungal antibiotic, CB-25-I, was isolated from the culture broth of a strain of Serratia plymuthica. The antibiotic, a water-soluble dipeptide, is structurally related to Sch 37137 and A 19009, both produced by strains of Actinomycetales. The antibiotic exhibits inhibitory activity against Candida albicans in YNB medium (a synthetic medium), but the activity is significantly reduced in Sabouroud dextrose medium.