Root Surface Colonization Research Articles

Proteomic Analysis of a Global Regulator GacS Sensor Kinase in the Rhizobacterium, Pseudomonas chlororaphis O6

The GacS/GacA system in the root colonizer Pseudomonas chlororaphis O6 is a key regulator of many traits relevant to the biocontrol function of this bacterium. Proteomic analysis revealed 12 proteins were down-regulated in a gacS mutant of P. chlororaphis O6. These GacS-regulated proteins functioned in combating oxidative stress, cell signaling, biosynthesis of secondary metabolism, and secretion. The extent of regulation was shown by real-time RT-PCR to vary between the genes. Mutants of P. chlororaphis O6 were generated in two GacS-regulated genes, trpE, encoding a protein involved in tryptophan synthesis, and prnA, required for conversion of tryptophan to the antimicrobial compound, pyrrolitrin. Failure of the trpE mutant to induce systemic resistance in tobacco against a foliar pathogen causing soft rot, Pectobacterium carotovorum SCCI, correlated with reduced colonization of root surfaces implying an inadequate supply of tryptophan to support growth. Although colonization was not affected by mutation in the prnA gene, induction of systemic resistance was reduced, suggesting that pyrrolnitrin was an activator of plant resistance as well as an antifungal agent. Study of mutants in the other GacS-regulated proteins will indicate further the features required for biocontrol-activity in this rhizobacterium.

The global regulator GacS regulates biofilm formation in Pseudomonas chlororaphis O6 differently with carbon source

An aggressive root colonizer, Pseudomonas chlororaphis O6 produces various secondary metabolites that impact plant health. The sensor kinase GacS is a key regulator of the expression of biocontrol-related traits. Biofilm formation is one such trait because of its role in root surface colonization. This paper focuses on the effects of carbon source on biofilm formation. In comparison with the wild type, a gacS mutant formed biofilms at a reduced level with sucrose as the major carbon source but at much higher level with mannitol in the defined medium. Biofilm formation by the gacS mutant occurred without phenazine production and in the absence of normal levels of acyl homoserine lactones, which promote biofilms with other pseudomonads. Colonization of tomato roots was similar for the wild type and gacS mutant, showing that any differences in biofilm formation in the rhizosphere were not of consequence under the tested conditions. The reduced ability of the gacS mutant to induce systemic resistance against tomato leaf mold and tomato gray mold was consistent with a lack of production of effectors, such as phenazines. These results demonstrated plasticity in biofilm formation and root colonization in the rhizosphere by a beneficial pseudomonad.

Influence of Bacillus spp. strains on seedling growth and physiological parameters of sorghum under moisture stress conditions

Microorganisms isolated from stressed ecosystem may prove as ideal candidates for development of bio-inoculants for stressed agricultural production systems. In the present study, moisture stress tolerant rhizobacteria were isolated from the rhizosphere of sorghum, pigeonpea, and cowpea grown under semiarid conditions in India. Four isolates KB122, KB129, KB133, and KB142 from sorghum rhizosphere exhibited plant growth promoting traits and tolerance to salinity, high temperature, and moisture stress. These isolates were identified as Bacillus spp. by 16S rDNA sequence analysis. The strains were evaluated for growth promotion of sorghum seedlings under two different moisture stress conditions (set-I, continuous 50% soil water holding capacity (WHC) throughout the experiment and set-II, 75% soil WHC for 27 days followed by no irrigation for 5 days) under greenhouse conditions. Plate count and scanning electron microscope studies indicated successful root surface colonization by inoculated bacteria. Plants inoculated with Bacillus spp. strains showed better growth in terms of shoot length and root biomass with dark greenish leaves due to high chlorophyll content while un-inoculated plants showed rolling of the leaves, stunted appearance, and wilting under both stress conditions. Inoculation also improved leaf relative water content and soil moisture content. However, variation in proline and sugar content in the different treatments under two stress conditions indicated differential effect of microbial treatments on plant physiological parameters under stress conditions.

Early root surface colonization by human periodontal ligament fibroblasts following treatment with different biomaterials

Objectives. The present in-vitro study examined the effects of different biomaterials on early root surface colonization by human periodontal ligament (PDL) fibroblasts using confocal-laser-scanning-microscopy (CLSM). Materials and methods. Fifteen periodontally-diseased teeth were extracted, treated with scaling/root planing and longitudinally cut to obtain 30 root fragments. Fragments were treated either with 24% EDTA following application of enamel matrix derivative (EMD), 24% EDTA or EMD only, nanocrystalline hydroxyapatite (NHA) paste or oily calcium hydroxide suspension (OCHS) for 1 h each. The analogue untreated root specimens served as controls. Root fragments were incubated with human PDL fibroblasts and cellular proliferation and morphology were evaluated after 1, 3, 5 and 8 days using CLSM-visualization and image recognition software. Results. The rate of cellular proliferation was different among treatment modalities examined (p = 0.019). Except treatment with NHA paste all treatment modalities improved cellular proliferation on root surfaces at all different points of time compared with the control specimens. A significant difference between treatment modalities was observed between EMD and NHA paste (p = 0.008). No synergistic effect could be demonstrated comparing root surface conditioning with 24% EDTA and EMD application compared to 24% EDTA or EMD application only. Conclusion. The present results suggest that initial root surface colonization by PDL fibroblasts may be enhanced by root surface conditioning with 24% EDTA and application of EMD, application of 24% EDTA or EMD alone and OCHS. The addition of 24% EDTA for root surface conditioning prior to EMD application provided no synergistic effects in terms of early root surface colonization by PDL fibroblasts.

Amendment of degraded desert soil with wastewater debris containing immobilized Chlorella sorokiniana and Azospirillum brasilense significantly modifies soil bacterial community structure, diversity, and richness

The main goal of this study was to expand our knowledge of what happens to the soil bacterial community in an eroded desert soil when improvement of soil fertility is derived from the application of debris of tertiary wastewater treatment containing immobilized microalgae Chlorella sor- okiniana and the plant growth-promoting bacterium (PGPB) Azospirillum brasilense. We hypothesized that an "im- proved" non-agricultural desert soil will exhibit substantial changes in the structure of the bacterial community in a relatively short time after amendment. To assess the effect of the amendments, microalgae and PGPB alone or com- bined, on the structure of the rhizosphere bacterial commu- nity, changes in species richness and bacterial diversity over time were based on sequence differences in the 16S rRNA gene, performed with PCR-denaturing gradient gel electro- phoresis (DGGE) and then analyzed by similarity test and non-metric multidimensional scaling analysis. Root surface colonization and persistence in the rhizosphere of A. brasi- lense was monitored by fluorescent in situ hybridization and sequencing of DGGE bands. Application of waste debris significantly changed the rhizosphere bacterial population structure, whether comparisons were made over time, be- tween inoculated and non-inoculated soil, and among dif- ferent inoculated microorganisms. Species richness and diversity increased when the waste debris contained the microalgae-bacteria association and also over time. Even as its secondary role as an inoculant after wastewater treat- ment, A. brasilense colonized the root surface profusely and persisted within the rhizosphere bacterial community. This study demonstrated that small organic amendment to desert soil significantly changed soil bacterial community com- pared to the original soil and also 2 months after amend- ments were added.

The Effect of Tillage System and Rimsulfuron Application on Weed Flora, Arbuscular Mycorrhizal (AM) Root Colonization and Yield of Maize (Zea mays L.)

Field experiments were conducted to determine the effects of tillage system and rimsulfuron application on weed flora and growth of maize (Zea mays L. Mitic F1) at a site with no history of pesticide use for the last 5 years. A randomized complete block design was employed with three replicates per treatment (conventional tillage without rimsulfuron application (control), no-tillage with rimsulfuron application (NT+Rim) and conventional tillage with rimsulfuron application (CT+Rim)). The lowest leaf area index, dry weight and yield of maize were recorded in NT+Rim plots. In addition, the highest soil bulk density was determined in NT+Rim plots. Moreover, the rimsulfuron application resulted in significant reduction in the root growth and arbuscular mycorrhizal (AM) colonization. The lowest root biomass, root length density, root surface and AM root colonization was found in CT+Rim plots. The lowest weed number and biomass was also observed at CT+Rim plots. Sustainability yield index (SYI) shown that the maize crop is more stable under conventional tillage compared with no-tillage. Our results indicated that root growth was reduced significant by rimsulfuron application but statistically significant lower maize yield was obtained in no-tillage system.

A combination of cellular automata and agent-based models for simulating the root surface colonization by bacteria

Models of root colonization by bacteria facilitate conceptual and practical understanding of fundamental processes that influence plant health, mineral nutrition, and stress tolerance. In this study, we explored the use of cellular automata and agent-based models to simulate the primary colonization of roots by bacteria as determined by selected parameters related to bacterial growth (nutrient uptake, fitness, reproduction and starvation) and environmental constraints (space, nutrient depletion, and pH). The results were then compared with observations from experiments examining the colonization of plant roots by a GFP – strain of Escherichia coli. The latter experiments were conducted with plants grown in agar medium containing calcium phosphate that allowed visualization of bacterial distribution (aggregates and abundance) and phosphate solubilization at root microsites. The numerical models revealed outcomes for diverse numerical scenarios, which agreed with the in vivo data and provided a basic framework for describing bacterial colonization of plant roots. Further efforts will be required to evaluate factors affecting the competence and ecology of bacterial communities at rhizosphere microsites, but offer promise for the development of precise predictive models with practical applications for agriculture.

The insect‐pathogenic fungus Metarhizium robertsii (Clavicipitaceae) is also an endophyte that stimulates plant root development

The soil-inhabiting insect-pathogenic fungus Metarhizium robertsii also colonizes plant roots endophytically, thus showing potential as a plant symbiont. Metarhizium robertsii is not randomly distributed in soils but preferentially associates with the plant rhizosphere when applied in agricultural settings. Root surface and endophytic colonization of switchgrass (Panicum virgatum) and haricot beans (Phaseolus vulgaris) by M. robertsii were examined after inoculation with fungal conidia. We used light and confocal microscopy to ascertain the plant endophytic association with GFP-expressing M. robertsii. Root lengths, root hair density, and lateral roots emerged were also observed. Initially, M. robertsii conidia adhered to, germinated on, and colonized roots. Furthermore, plant roots treated with Metarhizium grew faster and the density of plant root hairs increased when compared with control plants. The onset of plant root hair proliferation was initiated before germination of M. robertsii on the root (within 1-2 d). Plants inoculated with M. robertsii ΔMAD2 (plant adhesin gene) took significantly longer to show root hair proliferation than the wild type. Cell free extracts of M. robertsii did not stimulate root hair proliferation. Longer-term (60 d) associations showed that M. robertsii endophytically colonized cortical cells within bean roots. Metarhizium appeared as a mycelial aggregate within root cortical cells as well as between the intercellular spaces with no apparent damage to the plant. These results suggest that M. robertsii is not only rhizosphere competent but also displays a beneficial endophytic association with plant roots that results in the proliferation of root hairs.

Exopolysaccharide Production Is Required for Biofilm Formation and Plant Colonization by the Nitrogen-Fixing Endophyte Gluconacetobacter diazotrophicus

The genome of the endophytic diazotrophic bacterial species Gluconacetobacter diazotrophicus PAL5 (PAL5) revealed the presence of a gum gene cluster. In this study, the gumD gene homologue, which is predicted to be responsible for the first step in exopolysaccharide (EPS) production, was insertionally inactivated and the resultant mutant (MGD) was functionally studied. The mutant MGD presented normal growth and nitrogen (N(2)) fixation levels but did not produce EPS when grown on different carbon sources. MGD presented altered colony morphology on soft agar plates (0.3% agar) and was defective in biofilm formation on glass wool. Most interestingly, MGD was defective in rice root surface attachment and in root surface and endophytic colonization. Genetic complementation reverted all mutant phenotypes. Also, the addition of EPS purified from culture supernatants of the wild-type strain PAL5 to the mutant MGD was effective in partially restoring wild-type biofilm formation and plant colonization. These data provide strong evidence that the PAL5 gumD gene is involved in EPS biosynthesis and that EPS biosynthesis is required for biofilm formation and plant colonization. To our knowledge, this is the first report of a role of EPS in the endophytic colonization of graminaceous plants by a nitrogen-fixing bacterium.

Root Colonization and ISR-mediated Anthracnose Disease Control in Cucumber by Strain Enterobacter asburiae B1

Here, we show that an endophytic bacterial strain, Enterobacter asburiae B1 exhibits the ability to elicit ISR in cucumber, tobacco and Arabidopsis thaliana. This indicates that strain B1 has a widespread ability to elicit ISR on various host plants. In this study, E. asburiae strain B1 did not show antifungal activity against tested major fungal pathogens, Colletotrichum orbiculare, Botrytis cinerea, Phytophthora capsici, Rhizoctonia solani, and Fusarium oxysporum. Moreover, the siderophore production by E. asburiae strain B1 was observed under in vitro condition. In greenhouse experiments, the root treatment of strain B1 significantly reduced disease severity of cucumber anthracnose caused by fungal pathogen C. orbiculare compared to nontreated control plants. By root treatment of strain B1 more than 50% disease control against anthracnose on cucumber was observed in all greenhouse experiments. Simultaneously, under the greenhouse condition, the soil drench of strain B1 and a chemical inducer benzothiadiazole (BTH) to tobacco plants induced GUS activity which is linked with activation of PR promoter gene. Furthermore, in Arabidopsis thaliana plants the soil drench of strain B1 induced the defense gene expression of PR1 and PDF1.2 related to salicylic acid and jasmonic acid/ethylene signaling pathways, respectively. In this study, for the main focus on root colonization by strain B1 associated with defense responses, bacterial cells of strain B1 was tagged with the gfp gene encoding the green fluorescent protein in order to determine the colonization pattern of strain B1 in cucumber. The gfp-tagged B1 cells were found on root surface and internal colonization in root, stem, and leaf. In addition to this, the scanning electron microscopy observation showed that E. asburiae strain B1 was able to colonized cucumber root surface.

Colonization process of olive tissues by Verticillium dahliae and its in planta interaction with the biocontrol root endophyte Pseudomonas fluorescens PICF7.

SummaryThe colonization process of Olea europaea by the defoliating pathotype of Verticillium dahliae, and the in planta interaction with the endophytic, biocontrol strain Pseudomonas fluorescens PICF7 were determined. Differential fluorescent protein tagging was used for the simultaneous visualization of P. fluorescens PICF7 and V. dahliae in olive tissues. Olive plants were bacterized with PICF7 and then transferred to V. dahliae‐infested soil. Monitoring olive colonization events by V. dahliae and its interaction with PICF7 was conducted using a non‐gnotobiotic system, confocal laser scanner microscopy and tissue vibratoming sections. A yellow fluorescently tagged V. dahliae derivative (VDAT‐36I) was obtained by Agrobacterium tumefaciens‐mediated transformation. Isolate VDAT‐36I quickly colonized olive root surface, successfully invaded root cortex and vascular tissues via macro‐ and micro‐breakages, and progressed to the aerial parts of the plant through xylem vessel cells. Strain PICF7 used root hairs as preferred penetration site, and once established on/in root tissues, hindered pathogen colonization. For the first time using this approach, the entire colonization process of a woody plant by V. dahliae is reported. Early and localized root surface and root endophytic colonization by P. fluorescens PICF7 is needed to impair full progress of verticillium wilt epidemics in olive.

Landscape- and age-based factors affecting fungal colonization of conifer seedling roots at the alpine tree line

Survival of seedlings is a demographic bottleneck for tree populations in alpine tree-line ecotones (ATE) and may play a pivotal role in determining the elevation of timberlines and tree lines. We evaluated (1) fungal colonization of root surfaces of natural seedlings of Picea engelmannii Parry ex Engelm. and Abies lasiocarpa (Hook.) Nutt. in an ATE, (2) the relative importance of soil versus microclimate variation to differences in colonization in the ATE, determined using a reciprocal soil transplant experiment, and (3) ecophysiological responses of seedlings to natural and experimental variation in colonization. Many root systems lacked ectomycorrhizae but had extensive quantities of white mycelium attached to root surfaces. This mycelium was composed of a group of species referred to as the "alpine white complex" (AWC), and included Mortierella parvispora Linneman, Trichoderma viride Pers.:Fr., and a basidiomycete. Nearly all seedlings that survived beyond the first year of growth were colonized by AWC, whereas <10% of emergent seedlings had AWC where they commonly do not survive, in microsites distant from trees. No relationships of AWC and microclimate, photosynthesis, or water relations of seedlings were detected, and low colonization levels likely resulted from scarcity of AWC in ATE soils. Determining if and how AWC or emergent conifer seedlings benefit from their symbiosis in ATEs will require further inquiry.

Microbial colonization in the seagrass Posidonia spp. roots

The pattern of colonization by microorganisms on root surfaces from three species of seagrass belonging to the genus Posidonia was assessed. Microbial abundance on roots was measured by two electronic microscope techniques. Trends in microbial colonization between species and root order were defined. In addition, eutrophication status of the sampling sites and physiological status of Posidonia oceanica (L.) Delile roots have been taken into account. Our results show high microbial abundance in the Mediterranean species P. oceanica, in comparison with the low rates of colonization found in the Australian species P. australis Hook f. and P. sinuosa Cambridge et Kuo. Microbial density tended to decrease as root order increased, and living roots always showed higher microbial abundance than dead ones. Colonization of P. oceanica roots at the three sites with different environmental status follows different trends according to root order. It is suggested that root age influences the rate of microbial colonization of seagrass roots and that colonization of root surface by microorganisms is associated with organic exudates from the roots rather than with decaying root tissues.

Effect of enamel matrix derivative on human periodontal fibroblasts: proliferation, morphology and root surface colonization. An in vitro study.

Several clinical trials have shown the effectiveness of Emdogain(R) (EMD) in promoting tissue regeneration, even though the underlining biological mechanism is still poorly known. The aim of the present study was to verify the effect of EMD on the proliferation of human periodontal ligament (PDL) fibroblasts and on their colonization and differentiation following contact with the root surface of extracted teeth in vitro. Fibroblasts from PDL were seeded on Petri dishes and cell growth was evaluated by cell counting in the presence and absence of EMD, after 1, 3 and 8 d of culture. A significant effect of EMD upon cellular proliferation at d 3 and 8 was detected. When PDL cells were grown for 12 d with EMD on etched human root surface, a change in cell morphology was observed. Scanning electron microscopy revealed that cells grown on root EMD-treated surface present a body with a flattened surface closely adherent to the substrate and an outer smooth surface rounded in shape. From the flattened surface some thin and elongated cellular processes connecting with the substrate were also observable. PDL cells grown on EMD-treated surface showed lack of alkaline phosphatase activity, as some authors noticed in cementoblasts in vitro. In conclusion, our data indicate that EMD enhances human PDL fibroblast proliferation. Furthermore, the cells in the presence of EMD show morphological changes that make them more similar to cementoblasts than to fibroblasts, suggesting a process of cellular differentiation that could play an important role in periodontal tissue repair.

Root-surface colonization of black mangrove seedlings by Azospirillum halopraeferens and Azospirillum brasilense in seawater

Inoculation of axenic black mangrove seedlings in seawater for 8 days with either the terrestrial halotolerant plant growth-promoting bacterium Azospirillum halopraeferens or with Azospirillum brasilense produced heavy colonization of the root surface. The colonization pattern was different for the two strains. A. halopraeferens yielded mainly single cells embedded in a thick sheath, whereas A. brasilense produced primarily microaggregates. A. brasilense cells were anchored to the root surfaces and to themselves by a network of fibrillar material. Both bacterial strains survived in seawater (approximately 104 colony forming units per ml) for more than 30 days, for 70 days in saline water (A. brasilense) and colonized mangrove roots at a very high population density. A. halopraeferens was a better root surface colonizer, whereas the A. brasilense population was greater in the entire root. This work is the initial stage of studies designed to assess the feasibility of using terrestrial plant growth-promoting bacteria for the inoculation of marine plants.

Root-surface colonization of black mangrove seedlings by Azospirillum halopraeferens and Azospirillum brasilense in seawater

Inoculation of axenic black mangrove seedlings in seawater for 8 days with either the terrestrial halotolerant plant growth-promoting bacterium Azospirillum halopraeferens or with Azospirillum brasilense produced heavy colonization of the root surface. The colonization pattern was different for the two strains. A. halopraeferens yielded mainly single cells embedded in a thick sheath, whereas A. brasilense produced primarily microaggregates. A. brasilense cells were anchored to the root surfaces and to themselves by a network of fibrillar material. Both bacterial strains survived in seawater (approximately 10 4 colony forming units per ml) for more than 30 days, for 70 days in saline water ( A. brasilense) and colonized mangrove roots at a very high population density. A. halopraeferens was a better root surface colonizer, whereas the A. brasilense population was greater in the entire root. This work is the initial stage of studies designed to assess the feasibility of using terrestrial plant growth-promoting bacteria for the inoculation of marine plants.

A transcriptional regulator of the LuxR-UhpA family, FlcA, controls flocculation and wheat root surface colonization by Azospirillum brasilense Sp7.

Genetic complementation of a spontaneous mutant, impaired in flocculation, Congo red binding, and colonization of root surface, led to the identification of a new regulatory gene in Azospirillum brasilense Sp7, designated flcA. The deduced amino acid sequence of flcA shared high similarity with a family of transcriptional activators of the LuxR-UphA family. The most significant match was with the AgmR protein, an activator for glycerol metabolism in Pseudomonas aeruginosa. Derivatives of Sp7 resulting from site-directed Tn5 mutagenesis in the flcA coding sequence were constructed by marker exchange. Characterization of the resulting mutant strains showed that flcA controls the production of capsular polysaccharides, the flocculation process in culture, and the colonization of the root surface of wheat. This study provides new information on the genetic control of the mechanism of plant root colonization by Azospirillum.

Inoculum density, temperature, and genotype effects on in vitro growth promotion and epiphytic and endophytic colonization of tomato (Lycopersicon esculentumL.) seedlings inoculated with a pseudomonad bacterium

The effects of inoculum density (0, 4.6 × 107, 4.2 × 108, and 8.8 × 108 cfu∙mL−1), temperature (10, 20, and 30 °C), and plant genotype (cultivars Celebrity, Blazer, Scotia, and Mountain Delight) on bacterial colonization and plant growth promotion were investigated in a gnotobiotic system. An in vitro dual culture of tomato (Lycopersicon esculentum L.) plantlets and a Pseudomonas sp., strain PsJN, were used. Epiphytic (external) and endophytic (internal) bacterial populations were determined to evaluate plantlet colonization. Shoot and root biomass of bacterized plantlets was significantly higher (p ≤ 0.05) than that of nonbacterized controls. Growth promotion was best with inoculum densities of 3 × 108– 7 × 108 cfu∙mL−1at 20 °C, particularly in the early maturing cultivars Blazer and Scotia. Lower inoculum densities were required to maximize root growth (approximately 1 × 108 cfu∙mL−1) than shoot growth (approximately 3 × 108 cfu∙mL−1). Shoot surface populations did not vary with inoculum density or temperature, but the bacterium colonized the shoot exterior of cultivars Celebrity, Mountain Delight, and Scotia better than cultivar Blazer. The root surface populations increased linearly with increasing inoculum density (within a range of 107–108 cfu∙mL−1), decreased with increasing temperatures (from 10 to 30 °C), and were higher for the main season cultivar Celebrity than for cultivars Blazer, Scotia, and Mountain Delight. Populations of shoot endophytes did not vary with initial inoculum density or genotype but were affected by temperature; the highest colonization was at 10 °C. The number of root endophytes was also highest at 10 °C at the inoculum density of approximately 4 × 108 cfu∙mL−1and did not vary with genotypes. The experiments clearly indicate that there was no relationship between root surface colonization and plant growth promotion. However, the range of inoculum levels (3 × 108– 7 × 108 cfu∙mL−1) that promoted colonization of the inner root tissues (endophytic) also best promoted plant growth. A possible biostimulation threshold within the tissues of the inoculated plants under conditions favourable to the growth of tomato is proposed.Key words: Pseudomonas sp., tomato, colonization, growth promotion.

Root surface colonization of non-cereal crop plants by pleomorphic Azospirillum brasilense Cd

Summary: Root surface colonization byAzospirillum brasilenseCd of tomato, pepper and cotton plants under normal growth conditions and soybean plants under normal and water-stress conditions was monitored by scanning electron microscopy and bacterial counts.A. brasilenseCd was capable of efficiently colonizing the elongation and root-hair zones of all four plant species tested. In these zones, the bacteria mainly colonized the root surface (tomato, soybean), root-hairs (pepper), or both (cotton), by single cells (tomato, soybean), micro-aggregates (pepper), or a combination of the two (cotton). All inoculated plants demonstrated (i) larger amounts of mucigel-like substance on the root surface than non-inoculated plants and (ii) fibrillar material which anchored the bacterial cells to the root surface and established connections between cells within bacterial aggregates. On non-water-stressed soybean plants, mostA. brasilenseCd cells in the rhizosphere occurred as vibroid forms whereas those on water-stressed plants (wilting) were cyst-like. A lower rhizosphere bacterial population was observed on water-stressed plants. When water-stress conditions were eliminated, the bacterial cells reverted to the vibroid form and a concomitant increase in the bacterial population was observed. It is suggested that cyst-like formation is a natural response forA. brasilenseCd in the rhizosphere of water-stressed plants.

A root tissue culture system to study winter wheat-rhizobacteria interactions

A tissue culture procedure was developed for growing winter wheat roots. This system was used to study bacterial-root interactions and root colonization by Pseudomonas cepacia strains R55 and R85, Azospirillum brasilense ATCC 29729 and Azotobacter chroococcum ATCC 9043. Axenic root tissue cultures were inoculated with bacteria and incubated at 25° C on a rotary shaker (150 rpm) for up to 3 weeks. At various intervals, root morphology and root hair development, bacterial colonization of root surfaces and nitrogenase activity were determined. As determined by plate counting techniques, bacterial attachment to the roots varied from 7.5 × 104 to 3.2 × 107 colony-forming units cm−1. Scanning electron microscopy of inoculated roots revealed that some rhizobacteria (e.g., P. cepacia R85) significantly enhanced root hair development, and others (e.g., P. cepacia R55) specifically colonized root exudation sites. Nitrogenase activity of roots inoculated with either A. brasilense ATCC 29729 or A. chroococcum ATCC 9043 was stimulated if the inorganic nitrogen sources in the medium were replaced with 300 μg glutamine ml−1. We propose that the use of root tissue culture systems will facilitate studies on plant root-microbe interactions.