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  • 1
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The plant-growth promoting rhizobacterium Azospirillum lipoferum strain 4B generates in vitro a stable phase variant designated 4VI at frequencies of 10−4 to 10−3 per cell per generation. Variant 4VI displays pleitropic modifications, such as the loss of swimming motility and the inability to assimilate certain sugars compared to the wild type. The mechanism underlying phase variation is unknown. To determine whether RecA-mediated processes are involved in phase variation, the recA gene of A. lipoferum 4B was cloned and sequenced and a recA mutant (termed 4BrecA) was constructed by allelic exchange. Strain 4BrecA showed increased sensitivity to UV and MMS compared with 4B and impaired recombinase activity. The ability to generate variants in vitro was not altered; the variants from 4BrecA exhibited all morphological and biochemical features characteristic of the variant generated by strain 4B. However, the frequency of variants generated by 4BrecA was increased by up to 10-fold. So, in contrast with many studies showing the abolition or a large reduction of the frequency of phase variation in recA mutants, this study describes an enhancement of phase variation in the absence of a functional recA.
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  • 2
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology ecology 18 (1995), S. 0 
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract: The role of plasmids in the saprophytic growth of Rhizobium is mostly unknown. Plasmid-cured and complemented derivatives of R. leguminosarum bv. trifolii strain W14-2 were used to investigate the role of plasmids in the growth of this strain in sterile soil incubated under favorable moisture and temperature conditions. Strain W14-2 contains four plasmids (a,b,c,d). Absence of single plasmids in plasmid-cured derivatives generally did not reduce growth in soil when compared to the wild-type but absence of plasmid a delayed growth. Derivatives were unable to grow in soil when only plasmids a or d were present in cells. When only plasmids b or c were present, growth was delayed and the final population in 7 days was approximately 10% of the wild-type population. When the wild-type was co-inoculated at equal population into soil with derivatives lacking plasmids a, c, or d, the population of the wild-type at 7 days incubation was approximately 10 times larger than those of the derivatives. Elimination of only plasmid b did not reduce the ability of the strain to grow in soil when competing with the wild-type. Plasmids were involved in saprophytic growth of strain W14-2 in soil and may be important to the ecology of Rhizobium.
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  • 3
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Calystegines are tropane alkaloids produced by the roots of a few plant species. A bioassay was developed to identify roots with a microbial rhizosphere community capable of calystegine degradation (i.e. MCD roots). In a field survey, the proportion of MCD roots of Zea mays (calystegine-negative) varied from 20 to 80%. In field experiments, the proportions of MCD roots of Z. mays and Calystegia sepium (calystegine-positive) grown in a particular plot were similar to each other but varied with time and, overall, were higher than those of Z. mays roots from adjacent plots free of C. sepium. In autoclaved soil, no root of C. sepium or Z. mays plants propagated as seeds was MCD, indicating that calystegine-degrading microorganisms were not seed-borne. However, MCD roots were found as early as 1 day after planting of rhizomes of C. sepium in autoclaved soil or planting of axenic seedlings of either plant in natural soil microcosms. In total, microorganisms capable of degrading calystegines were harboured not only in the rhizosphere of the calystegine-producing plant but also in that of the calystegine-negative plant and probably in bulk soil.
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  • 4
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: A spontaneous rifampicin-resistant mutant of the biocontrol agent Pseudomonas fluorescens CHA0 was released as soil inoculant in large outdoor lysimeters and its ability to colonise the roots of winter wheat, spring wheat (grown after Phacelia) and maize at the later stages of plant development was investigated by colony counts. The inoculant (i.e. CHA0-Rif) colonised the rhizosphere and the interior of the roots of both wheat varieties but CFUs at ripening were about 2 log (g root)−1 or lower. In contrast, the roots of maize were colonised poorly by the pseudomonad at flowering, but the latter was found at 3 or more log CFU (g root)−1 on and inside the roots in late ripening stage. Furthermore, CHA0-Rif was recovered at more than 5 log CFU (g root)−1 from the interior of several maize root samples. Whereas most cells of CHA0-Rif in soil were small and did not respond to Kogure's viability test, the pseudomonad was present as viable, unusually large (7 mm long) rods inside maize roots. In a microcosm experiment performed with similar sandy-loam soil, the CFUs of maize root-associated CHA0-Rif were higher where the shoots of the plant had been cut off, confirming that older and/or decaying maize roots represent a favourable niche for the inoculant. Overall, the results indicate that Pseudomonas inoculants have the potential to colonise the roots of certain crops (e.g. maize but not wheat for strain CHA0-Rif) at later stages of plant development.
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  • 5
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract: The ability to utilise additional siderophores may increase the ecological fitness of biocontrol inoculants of Pseudomonas in the rhizosphere. Plasmid pCUP2 carries a copy of the gene pbuA coding for the membrane receptor of ferric pseudobactin M114. Pseudomonas sp. B24Rif containing pCUP2 can utilise ferric pseudobactin of P. fluorescens M114 in addition to its own siderophore. A larger fraction of the culturable resident fluorescent pseudomonads in the rhizosphere of sugarbeet grown in a low-iron sandy loam soil could supply siderophore-complexed iron to B24Rif(pCUP2) rather than to B24Rif. However, B24Rif and B24Rif(pCUP2) were found at similar population levels in the rhizosphere for 21 days after their inoculation on seeds. A total of 25 of 43 isolates of resident fluorescent Pseudomonas unable to cross-feed iron to B24Rif could cross-feed B24Rif(pCUP2) and they were subdivided into seven different strains by arbitrary-primed PCR fingerprinting. The siderophores produced by 11 of them were typed by HPLC and they were similar to pseudobactin M114. However, the ability to utilise ferric pseudobactin M114 did not improve the ecological fitness of B24Rif in the rhizosphere of sugarbeet although a larger fraction of the culturable resident fluorescent pseudomonads could supply pseudobactin M114-complexed iron to B24Rif(pCUP2) than to B24Rif.
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  • 6
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Little is known on the behavior of soil-inoculated biocontrol pseudomonads once they are transported to deeper soil layers and/or groundwater levels after a heavy rain. This issue was investigated in inoculated microcosms containing lysimeter effluent water, and experimental conditions mimicking a worse-case scenario for potential bacterial dissemination were chosen. First, the survival of the polyketide-producing biocontrol strain Pseudomonas fluorescens CHA0-Rif was studied for 175 days at two inoculation levels in unamended and nutrient-amended lysimeter effluent water, and its impact on numbers of resident culturable bacteria was determined. Cell numbers of CHA0-Rif declined to 3–4 log cells ml−1 (at high inoculum level) or reached the detection limit or below (at low inoculum level) by day 175, without generating significant numbers of non-culturable cells. At high inoculum level, strain CHA0-Rif resulted durably (from day 50 to 175) in higher numbers of the total resident culturable bacteria when compared with the uninoculated control. This effect, which did not take place at low inoculum level or when nutrients had been added, contrasts with the transient ecological impact of the strain on rhizosphere bacterial populations in previous studies. Neither 2,4-diacetylphloroglucinol nor pyoluteorin were found in the water using HPLC, and inoculation with CHA0-Rif had no effect on the percentages of the total culturable aerobic bacteria sensitive to either antimicrobial polyketide on day 20. Second, the impact of CHA0-Rif on numbers of resident culturable bacteria was compared with that of CHA0-Rif(pME3424). Plasmid pME3424 carries an extra copy of the strain's rpoD gene (encoding sigma factor σ70). CHA0-Rif(pME3424) disappeared within 50 days in the water, but had the same impact as CHA0-Rif on the total number of resident culturable bacteria. This suggests that the impact of CHA0-Rif took place at the early stages of the experiment and was probably linked to the release of nutrients by introduced cells during inoculant decline.
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  • 7
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: In biocontrol fluorescent pseudomonads, phlD encodes a polyketide synthase required for the synthesis of the antifungal compound 2,4-diacetylphloroglucinol (Phl). Here, PCR-restriction fragment length polymorphism analysis was used to compare phlD alleles in 77 dicot-associated pseudomonads originating from various countries worldwide and 10 counterparts from a monocotyledonous host (wheat). The 16 restriction patterns obtained were mostly unrelated to geographic location or dicot host. Cluster analysis distinguished eight phlD clusters at a similarity level of 0.63. One cluster grouped 18 pseudomonads that produced also the antifungal polyketide pyoluteorin but could not assimilate D-galactose, D-galactonate lactone, D-sorbitol, L-arabinose, D-saccharate or D-xylose. These 18 pseudomonads, along with the eight pseudomonads from a second phlD cluster, were the only isolates that failed to deaminase 1-aminocyclopropane-1-carboxylate (ACC), a rare root growth promotion trait. Overall, assessment of phlD polymorphism, ACC deaminase activity and catabolic profiles pointed to a cosmopolitan distribution of Phl-producing biocontrol fluorescent pseudomonads of worldwide origin associated with dicotyledonous crop plants.
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  • 8
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Certain soils from Morens, Switzerland, are naturally suppressive to Thielaviopsis basicola-mediated black root rot of tobacco, and fluorescent pseudomonads are involved in this suppressiveness. Here, we compared two conducive, one moderately suppressive and one suppressive soil from Morens. Disease levels on tobacco after heavy T. basicola inoculation varied from 29% to 85% for the two conducive soils, 10% to 78% for the moderately suppressive soil and 11% to 42% for the suppressive soil, depending on time of the year. In the absence of T. basicola inoculation, disease levels were between 0% and 40% and varied also in time. Fluorescent pseudomonads were isolated from the rhizosphere and roots of tobacco subjected to T. basicola inoculation and characterized for production of the biocontrol metabolites 2,4-diacetylphloroglucinol (Phl) and HCN. No difference in population size was found between the suppressive and the conducive soils for total, Phl+ and HCN+ fluorescent pseudomonads colonizing the rhizosphere or roots of tobacco. Yet, the percentage of Phl+ isolates was significantly higher (30–32% vs. 6–11%) in the rhizosphere and roots for plants grown in the suppressive soil compared with the moderately suppressive and conducive soils. Different restriction profiles for phlD, one of the Phl biosynthetic genes, were often found when analyzing Phl+ isolates colonizing the same plant. Most phlD alleles were recovered from both suppressive and conducive soils, except one allele found only in root isolates from the suppressive soil.
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  • 9
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Erwinia carotovora subspecies atroseptica is the agent of soft rot of potato and causes important crop damage in Europe. Synthetic 2,4-diacetylphloroglucinol (DAPG) inhibited the growth of E. carotovora subsp. atroseptica under in vitro conditions and Pseudomonas fluorescens F113, which produces DAPG, was studied for biocontrol of E. carotovora subsp. atroseptica. Wild-type F113 (or the spontaneous rifampicin-resistant mutant F113Rif) inhibited growth of E. carotovora subsp. atroseptica on solid medium, displayed bactericidal activity towards the pathogen in liquid medium, and prevented Erwinia-mediated rotting of wounded potato tuber under in vitro conditions. F113Rif reduced the population size of E. carotovora subsp. atroseptica in soil and on potato tuber dice in competition experiments carried out with unplanted soil and soil planted with diced potato tubers, respectively. Co-inoculation of potato tuber seeds with F113Rif and E. carotovora subsp. atroseptica reduced Erwinia contamination of the seed tubers compared with single inoculation with the pathogen. F113G22 is a Tn5::lacZY-induced DAPG-negative biosynthetic derivative of F113 and showed no antibiosis towards E. carotovora subsp. atroseptica in vitro. In contrast to F113Rif, F113G22 did not inhibit Erwinia-mediated rotting of wounded potato tuber in vitro, did not influence survival of E. carotovora subsp. atroseptica in unplanted soil or soil planted with potato tuber dice and did not reduce Erwinia contamination of potato seed tubers. F113G22(pCU203) is a complemented derivative with restored DAPG-producing ability. F113G22(pCU203) had similar effects against E. carotovora subsp. atroseptica as F113 (or F113Rif) under in vitro conditions and in soil microcosms. The results indicate that P. fluorescens F113 is a promising biocontrol agent against the potato soft rot agent E. carotovora subsp. atroseptica and suggest that the pseudomonad's ability to produce DAPG is a key factor in its inhibition of the pathogen.
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  • 10
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The physical organization of phytobeneficial genes was investigated in the plant growth-promoting rhizobacterium Azospirillum lipoferum 4VI by hybridization screening of a bacterial artificial chromosome (BAC) library. Pulsed-field gel electrophoresis gave an estimated 5.7-Mb genome size for strain 4VI and a coverage level of 9 for the BAC library. The phytobeneficial genes nifH (associative nitrogen fixation) and ipdC (synthesis of the phytohormone indoleacetic acid) are chromosomal, but no BAC clone containing both genes was found, pointing to the absence of any genetic island containing nifH and ipdC. A 11.8-kb fragment containing nifH was analyzed. Neighboring genes implicated in nitrogen fixation (nifH, draT, draG) or not (arsC, yafJ and acpD) were organized as in A. brasilense. In contrast, the region located downstream of acpD contained four housekeeping genes (i.e. genes encoding DapF-, MiaB- and FtsY-like proteins, as well as gene amn) and differed totally from the one found in A. brasilense.
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