CURRENT MICROBIOLOGY Vol. 55 (2007), pp. 314–322 DOI: 10.


Current Microbiology
An International Journal
ª Springer Science+Business Media, LLC 2007

Biocontrol and PGPR Features in Native Strains Isolated from Saline Soils of Argentina
Analía Príncipe, Florencia Alvarez, Marina G. Castro, Lucía Zachi, Sonia E. Fischer, Gladys B. Mori, Edgardo JofrØ
Facultad de Ciencias Exactas, Físico-Químicas y Naturales, Universidad Nacional de Río Cuarto, Ruta 36-Km 601-5800, Río Cuarto-Córdoba, Argentina Received: 27 December 2006 / Accepted: 21 February 2007

Abstract. A bacterial collection of approximately one thousand native strains, isolated from saline soils of Cordoba province (Argentina), was established. From this collection, a screening to identify those strains showing plant growth promotion and biocontrol activities, as well as salt tolerance, was performed. Eight native strains tolerant to 1 M NaCl and displaying plant growth promotion and/or biocontrol features were selected for further characterization. Strains MEP2 18, MRP2 26, MEP2 11a, MEP3 1, and MEP3 3b significantly increased the growth of maize seedlings under normal and saline conditions, whereas isolates ARP2 3, AEP1 5, and ARP2 6 were able to increase the root dry weight of agropyre under saline conditions. On the other hand, strains MEP2 18 and ARP2 3 showed antagonistic activity against phytopathogenic fungi belonging to Sclerotinia and Fusarium genus. Antifungal activity was found in cell-free supernatants, and it was heat and protease resistant. Strains MEP218 and ARP23 were identified as Bacillus sp. and strains MEP211a and MEP33b as Ochrobactrum sp. according to the sequence analysis of 16S rRNA gene.

Salinity is one of the most important abiotic problems in agriculture, turning agronomically useful lands into unproductive areas. In the southeast of Cordoba province (Argentina) there are 1.5 · 106 ha with hydrohalomorphic characteristics due to deficient superficial and subterranean drainage and the ascent of the freatic layer [7]. Salinity may directly or indirectly inhibit cell division and the elongation growth state; therefore, leaves and stems of the affected plants appear stunted. On the other hand, plants suffering saline stress present alterations in their homeostasis, mainly because of a reduction in the osmotic potential and an inadequate ionic distribution, which causes a nutritional imbalance [26, 41]. In addition to the use of traditional breeding and transgenic plants, the utilization of plant growthpromoting rhizobacteria (PGPR) is useful in developing strategies to facilitate plant growth in saline soils [31]. PGPR mechanisms can be classified into direct or indirect growth promotion [18]. Direct promotion occurs

Correspondence to: Edgardo JofrØ; email:

when compounds synthesized by bacteria, such as fixed nitrogen, soluble phosphate, phytohormones, and iron that has been sequestered by bacterial siderophores, are provided to the plant. Many PGPR also produce the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase that metabolizes ACC, the precursor of ethylene. Indirect promotion or biocontrol occurs when PGPR reduces or prevents the deleterious effects of phytopathogenic organisms. This may be achieved by the induction of systemic resistance (ISR) and/or by the synthesis of antimicrobial compounds [29, 31]. Furthermore, Ryu et al. [38] demonstrated that volatile compounds (butanediol, acetoin) released by members of the genus Bacillus can stimulate plant growth and promote the ISR response. Among biotic stress, fungi are considered important plant pathogens, particularly members of the genera Sclerotinia and Fusarium, are able to infect a wide range of plants including several crop, vegetable, ornamental, and fruit species. Field crops reported as main hosts for Sclerotinia include peanut, soybean, sunflower, bean, lettuce, alfalfa, tomato, and peppermint, causing dis-

the concentrated supernatants were fractionated (1. [47]. soil (third letter plus sub index) and isolate number. Fusarium verticillioides RC 2000.01% (wt/vol) for 15 s. Universidad Nacional de Río Cuarto). Azospirillum brasilense Cd kindly supplied by EMPRAPA (Brazil). proteinase k. 11. soil type. PGPR Tests. eases such as blight in peanut. ethanol. and an intended result is sometimes difficult to obtain. Seedlings were aseptically . Then. Approximately 70 isolates were randomly selected from the pool of bacteria tolerant to 1 M NaCl and were phenotypically characterized by Gram stain (Britania Laboratories). Sclerotium rolfsii. Growth Promotion of Maize and Agropyro under Salinity Conditions. In this sense.5 mL) and dried in a rotary vacuum evaporator. Cellulase synthesis: The ability to produce cellulase was measured on plates containing minimal medium with 2% (wt/vol) 1-carboxymethylcellulose as carbon source according to Hankin and Anagnostakis [21]. roots were taken out from these suspensions. Salt-tolerant strains Growth in Nitrogen-Free Medium Fresh colonies grown in NA 500 mM NaCl were inoculated into vials containing 5 mL of semisolid Nfb (nitrogen free medium) or LGI media with and without 2. Azospirillum brasilense Cd was used as positive control [2]. as described before. and all Fusarium species that infect cereals can produce one or more mycotoxins [12]. For this reason. plant species. and rot in sunflower capitulum and in soybean stem. soaked in Cl2Hg 0. and roots were suspended in sterile physiological solution and vortexed to remove firmly adhered soil (rhizosphere soil). Materials and Methods Isolation of Native Strains from Saline Soils. An aliquot of 100 lL of each concentrated supernatant was spotted onto a paper disk on PDA plates to test the antifungal activity.5 g L)1 NH4Cl as nitrogen source. in particular different soil types [37]. and Bacillus megaterium NRRL B-939 (kindly supplied by Universidad Nacional de La Plata) were used as indicator strains.A. and incubated for 10 days at 28°C. at 29° 30Õ latitude south and in the south at 35°. An effective biological control strain isolated from one region may not perform in the same way in other soil and climatic conditions [8. 25]. The indol acetic acid (IAA) and/or analogues production was tested using SalkowskyÕs reagent [5]. Production of antimicrobial substances: The capacity to inhibit the growth of other bacteria was determined in several of the isolates according to Parret et al. Fusarium solani. On the other hand. Soils of this region used in this study are udorthentic haplustolls [23]. rhizosphere or endorhizosphere (second letter). and washed seven times with sterile distilled water. Pseudomonas fluorescens (ATCC 11253). 100 lL of the treated antifungal solution was spotted onto a paper disk on PDA plates to test the antimicrobial activity. surface disinfected by immersion in 95% (vol/vol) ethanol for 3 min. Samples were serially diluted and plated on nutrient agar medium (NA). Synthesis of proteases: protease activity was detected on 3% (wt/ vol) powdered milk-agar plates according to Walsh et al. The roots were then macerated in physiological solution (endorhizosphere).: Native Strains Isolated from Saline Soils 315 were maintained on nutritive agar supplemented with 500 mM NaCl and designated according to the host plant (first letter). and catalase activity. In order to evaluate the stability of cell-free supernatants to temperature. Seeds of Zea mays and Agropyrum elongatum were surface sterilized and germinated as described before. The immediate response to soil inoculation with PGPR varies considerably depending on the bacteria. washed with tap water to remove adhered soil particles. Siderophore production was screened by a plate assay using chrome azurol-S agar plates [40]. Plants were carefully harvested from the pots. and their ability to promote plant growth and/or to inhibit the growth of phytopathogen fungi was studied. 24]. or distilled water (to evaluate the effect of autoclaving and hydrolytic enzyme). Those isolates inhibiting mycelia growth on PDA medium were selected for further characterization [9]. Biocontrol Tests. The plants were incubated under greenhouse conditions (25°C and 12 h light–dark periods) for 15–20 days. native strains adapted to saline soils were isolated. Strains were tested against Sclerotinia sclerotiorum. Cell-free MEP2 18 and ARP2 3 cultures grown for 72 h in potato dextrose liquid medium were obtained by centrifugation at 10. The tolerance of isolates to salinity was tested by growing the strains on NA supplemented with increasing concentrations of NaCl (300–500–700–1000 mM). Fusarium head blight of small grain cereals and ear rot in maize are significant diseases across the world. and organic solvents. Antifungal Activity of Cell-Free Supernatants. Recently. Sclerotinia minor. damping-off in soybean. Príncipe et al. Finally. In this work. [33]. oxidase test (Britania Laboratories). The effect of the inoculation with native strains on the growth of gramineous plants (maize and agropyro) was evaluated. MØxico) and agropyro (Agropyrum elongatum) were surface sterilized [16. The control of these diseases becomes difficult because of the formation of resistance structures called sclerotia [35]. P. The resulting pellets were dissolved in chloroform.000 rpm for 20 min and filtered with 0. inoculum density. and environment conditions. The inoculated bacteria sometimes do not survive in the soil when competing with the betteradapted indigenous microflora [4]. and Fusarium graminearum RC 664 in plate bioassays (fungal strains were kindly supplied by the laboratories of Phytopathology and Mycology. The seeds were germinated and seedlings were aseptically transferred to pots containing saline/arid soils of the three different areas of the south of the Cordoba province. clumps of soil loosely attached to the roots were removed. Fusarium proliferatum RC 479. corrugata NCPPB 2445. our laboratory has attempted to isolate native strains with PGPR ability. Soil is an unpredictable environment. Mineral phosphate solubilization activity was assayed on plates according to Frioni [17].45 lm pore-size filters and then lyophilized and resuspended (to a 10-fold concentration) in distilled water or methanol. one important factor to be considered when screening new isolates is their activity in the range of environments in which they would be expected to be used. Cordoba province (Argentina) is located in the north. Seeds of maize (Zea mays) tolerant to 200 mM NaCl (CIMMYT. we have isolated and characterized Pseudomonas strains able to promote wheat growth under greenhouse conditions [15].

Almost all the selected isolates were catalase-positive. and MEP31 significantly increased the shoot dry weight in normal conditions as compared to the un-inoculated plants (control). plants inoculated and grown in saline conditions also showed an increased shoot dry matter as compared to the un-inoculated plant grown in the same conditions. Biocontrol Tests. Most strains were able to produce siderophore (70%). respectively. and protease production were determined. 3b). Sequences were analyzed using the BLASTN algorithm available in GeneBank (http://www. which largely overcomes the limit to be considered as saline soil (4 mS) [6]. the strains ARP2 3 and MEP2 18 were selected for their ability to inhibit all tested phytopathogenic fungi (Fig.nih. Different treatments such as protease and heat did not show any effect on the biocontrol activity of cellfree supernatants (Fig. Plants were grown under greenhouse conditions (25°C. DQ409215 (Ochrobactrum sp. Sequencing of 16S rRNA gene. On the other hand.05). 3a). 55 (2007) 3 and MEP2 18 displayed the best antifungal activity among the analyzed isolates. A graphic representation of the tree was made using Njplot software [34]. Interestingly. LZ). in both normal and saline conditions (100 mM NaCl). and the highest production was observed at the end of the stationary phase after 5 days of culture (Fig. showed a 99% identity with Ochrobactrum genus while the strains MEP2 18 and ARP2 3 were assigned to the Bacillus genus. with the mean values compared by using the FisherÕs protected least significant differences (LSD) analysis (p £ 0. 12 h light). An about [1]. plants inoculated with these strains and irrigated with 150 mM NaCl accumulated shoot dry weights to approximately the same extent as non-inoculated plants grown in the absence of salt (Fig. chloroform. but there were no effects on the shoot dry weight (Fig. 11a). Gram-positive as well as Gram-negative rods were isolated from rhizosphere and endorhizosphere in all the tested soils. Approximately 1000 bacterial isolates were obtained from saline soils of three different areas of the south of Cordoba province by using maize and agropyro plants as trap host. CURRENT MICROBIOLOGY Vol. The double strand sequencing was performed by custom service of MACROGEN Inc. Some isolates showing PGPR traits were selected for the experiments of maize and agropyroÕs inoculation under greenhouse conditions. and they were positive for all indirect PGPR tests (Table 2). PGPR Test. a screening in culture media containing different NaCl concentration was performed. Nucleotide Sequence Accession Number. MEP3 3b. The sequences obtained in this study were deposited in the GenBank nucleotide sequence database under the accession number DQ343613 (Bacillus sp.05) the root dry weight only under stress conditions (Fig.nlm. In addition. Isolates ARP2 . Isolates MEP2 11a and MEP3 3b. The PCR products were purified and sequenced.ncbi. oxidase-negative.5 kb fragment of 16S rRNA gene of three isolates was amplified by PCR using universal primers fD1 y rD1. 3b).05) increased the root dry weight only in normal conditions. Figures 5 Results Bacterial Isolation. 1). Growth Promotion of Maize and Agropyro by Native Strains Under Salinity Conditions. DQ 343615 (Bacillus sp ARP2 3). In order to know the mechanisms of the fungal inhibition. 2A and 2B). Bioactive compounds were found in the extracellular medium. cellulase. and DQ342340 (Ochrobactrum sp. The strains ARP2 3 and AEP1 5 significantly increased root dry weight of agropyro. using Clustal W program [44]. Extraction of the supernatants with ethanol.316 transferred to pots with sterile sand and perlite (1:1) and watered with nitrogen-free Hoagland nutrient solution [22] or nitrogen-free Hoagland solution containing 100 (for agropyro) or 150 mM NaCl (for maize). whereas 26–28% of isolates were able to solubilize phosphate and synthetize IAA. and showed rod morphology. Some Gram-positive isolates inhibited the growth of phytopathogenic fungi in vitro. 2C and 2D). The construction of a neighbor-joining tree was realized [39]. About 44% of isolates were able to grow in nitrogen-free medium. The sequences of isolates were aligned. Sequences of 16S rDNA. shoot and root dry weight were measured and the data obtained were analyzed by twoway analysis of variance. bacteriocin. Those isolates able to grow in 1 M NaCl were selected for further characterization. inoculated with 1 mL of bacterial suspension (107 cell mL)1). The inoculation of maize with bacterial strains MEP2 11a. In addition to PGPR properties described above. and after 7 days. PGPR tests of 72 isolates are shown in Table 1. The fraction soluble in methanol showed greater activity than the aqueous fraction. Seventy-two of 400 isolates able to grow on the highest NaCl concentration tested were selected because of their phenotypic and PGPR characteristics (Table 1). In order to select those isolates tolerant to salinity in vitro. The sequences obtained were analyzed using the BLAST program. The 16S rRNA gene was amplified by polymerase chain reaction (PCR) using universal primers rD1 and fD1 [48]. The utilized soils showed an electric conductivity higher than 25 mS. with the sequences retrieved from the Genbank database. whereas MEP211a was able to significantly increase (p < 0. Four weeks post-inoculation. MEP33b significantly (p < 0. 4). compared with the control. and methanol resulted in the transfer of antifungal activity into the organic phase.

Plant growing-Promoting rhizobacteria traits of bacterial isolates obtained from different saline soils using maize and agropyro plants as trap-host Isolates from maize MRP1 13 MRP1 15 MRP1 25 MRP1 28 MRP1 29 MRP1 32 MRP1 36 MRP1 13 MRP1 15 MRP1 24 MRP1 26 MRP1 27 MRP1 30 a MRP1 30 b MRP1 31 MRP1 33 MRP1 34 MRP1 35 MEP1 4 MEP1 38 MEP1 18 MEP1 42 MEP1 31 MEP1 12 MEP1 50 MRP2 10 MRP2 13 MRP2 25 a MRP2 17 MRP2 26 MRP2 7 MRP2 15 MRP2 12 MRP2 19 MEP2 18 MEP2 24 MEP2 11 a MEP2 44 MRP3 30 MRP3 20 b MRP3 23 MRP3 21 MRP3 15 MRP3 10 MRP3 6 MRP3 25 MRP3 1 MRP3 3 a MRP3 21 b MEP3 36 MEP3 1 MEP3 3 b MEP3 11 Isolates from agropyro ARP1 13 ARP1 3 IAA and /or analogues ) ) ) ) ) ) ) ) ) ) ) ) ) + ) ) ) ) ) ) ) ) + + + + ) + ) ) ) ) + + + ) ) ) ) + ) ) ) ) ) ) ) ) ) + ) + + IAA and /or analogues ) ) Solubilization of phosphate ) ng ) ) ) ) ) ) ng + ) ) + ng ) ) + ) ) + + ng ) ) ) ) ) ) + + ng + ) ) ) ) + + ) ng ) ) ) ) ) ) ) ) + ) + + ) Solubilization of phosphate ) + Growth in nitrogen free medium + ) + ng ) ) + + ) ng + ) ) ) ) ) ) ) ng + + + ) ) ) ) ng ng ) + ng + + + + ) + ) + ) ng ) ) ng ) ) ) ) ng ) + ) ) Growth in nitrogen free medium + ng Inhibition of phytopathogens nd nd ng ) ) ) ng nd nd ) ) ) ng ) ng ng ) ) nd ) ) ) ) ) nd + + ) ) ) ) ) ) + + ) ) + ) ) ) ) ) ) ) ) ) ) nd ) ) ) Inhibition of phytopathogens + ) Morphology G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G(+) Rod G(+) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G()) Rod G()) Rod G()) Rod G(+) Rod G(+) Rod G()) Rod G()) Rod G(+) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G(+) Rod G()) Rod G()) Rod G()) Rod Morphology G(+) Rod G()) Rod Siderophores nd + ng + + ng ng nd + ng ) + ng ) ng + ng + + + + ) ) ) nd ) + + + + + + + + + + + + + ) + + + + + + + + + ) + + ) Siderophores + + .A. Príncipe et al.: Native Strains Isolated from Saline Soils 317 Table 1.

G Gram stain. Azospirillum spp. we isolated native strains from saline soils of Cordoba province and screened for salt-tolerant bacteria with PGPR abilities. nd not determined. the growth reduction was decreased and an increased dry biomass compared to uninoculated plants was observed. causing a diminished yield. MEP211a. ARP2 6 (agropyro). This phytohormone has been implicated in increasing root growth and length. Acetobacter diazotrophicus. Bacillus sp. Moreover. P2. Taking this into account. ARP23. when plants were inoculated with the isolates MEP211a and MEP3 3b (maize) and ARP2 3. P1. In addition. An important factor to be considered when screening new isolates is their activity in the range of environments in which they would be expected to be used.. there are 1. Continued Isolates from maize AEP1 1 AEP1 4 AEP1 5 AEP1 6 ARP2 2 ARP2 3 ARP2 6 ARP2 32 ARP2 7 ARP2 8 ARP2 9 ARP2 10 ARP2 18 ARP2 2a AEP2 12 AEP2 141 AEP2 125 AEP2 126 AEP2 140 IAA and /or analogues ) ) ) ) + + ) + ) ) + ) + ) ) + ) ) + Solubilization of phosphate ) ) ) ) ) ) ) + + ) ) ) + ) ) ) + ) + CURRENT MICROBIOLOGY Vol. such as Azotobacter chroococcum [27]. MEP211a and MEP3 3b also promote maize growth under normal conditions. and changes in Na+/K+ ratio. which frequently is immobilized and unavailable for plants. 20].318 Table 1. 55 (2007) Morphology G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G()) Rod G()) Rod G()) Rod G()) Rod G()) Rod G(+) Rod G(+) Rod G(+) Rod G(+) Rod G()) Rod Siderophores + ng + + + + + + + ng + + + + + + + + ) Growth in nitrogen free medium + + + ng + + + ) + + + + ng ng + + + + + Inhibition of phytopathogens ) ng + ) + + ) + + ) ) ) ) ) ) + ) + ) A or M agropyro or maize. the increased resistance to environmental stress of plants treated with PGPR has been described.. and MEP218. Some isolates were selected for plant inoculation experiments under normal and saline conditions. which enables the plant to access more nutrients from soil. R or E rhizosphere or endorhizosphere. The isolates . and P. In the southeast of Cordoba province (Argentina). Some mechanisms include reduction of stress ethylene production via the action of ACC deaminase [19. However. Among the IAA producer PGPR. For example. There is evidence that the mode of action of many PGPR is by increasing the availability of nutrients for the plant [18]. the siderophore produced by rhizobacteria may contribute to an increased mobility of Fe in the soil and rhizosphere. For example. and 6 show the phylogenetic position of four isolates within different genus of PGPR. resulting in greater root surface area. Another essential macronutrient is phosphorus. putida [9]. ng nogrowth. making it more available for the plant.5 · 106 ha affected by salinity [7]. We observed greater reduction of growth in maize and agropyro seedlings when they were irrigated with a high concentration of salt. Herbaspirillum seropedicae has been described [10]. IAA is very commonly produced by PGPR. The 16S rRNA sequence was obtained from isolates MEP33b. accumulation of osmoprotectants such as proline in wetland rice seedlings [36]. Isolates in italic were selected for greenhouse experiments. or P3 soil used. Discussion Saline conditions are known to suppress the growth of plants. On the other hand. Some PGPR are able to solubilize phosphate. which could be associated with some PGPR mechanisms determined in these isolates such as IAA and siderophores production and/or phosphate solubilization. it has been demonstrated that Azospirillum brasilense increased the iron absorption and translocation by sorghum [3].

s + + - 319 Antimicrobial substances B + + + + + C + + + + + + + Pr + + + + - S. MEP33b and MEP211a were identified as Ochrobactrum sp. Príncipe et al. g Fusarium graminearum. The comparative sequence analysis of the 16S rRNA sequences showed the location of strains MEP211a and MEP33b within the family Brucellaceae. L) by isolates ARP23 (A. This result clearly showed that the isolates MEP211a and MEP33b belong to the genus Ochrobactrum. was recently proved by the description of strains with complete symbiotic ability in Acacia and Lupinus nodules [32. F).v + + F. P Protease. s Sclerotinia sclerotiorum. H. rhizosphere. E. O.p + + F. Fusarium solani (G. B). water. F. members of the genus Sclerotinia and Fusarium are well-known phytopathogens. and K) and MEP218 (B. are frequently associated with plants. producing significant losses in crop yield. Fusarium graminearum (E.A. L). Ochrobactrum strains occur in different habitats including soil. On the other hand. The diazotrophy of Ochrobactrum. Indirect Plant growing-Promoting rhizobacteria mechanisms of the strains selected for greenhouse experiment Indirect promotion Inhibition of phytopathogens Isolate ARP2 3 AEP1 5 ARP2 6 MEP2 18 MRP2 26 MEP2 11a MEP3 1 MEP3 3b S. G. C Cellulase. v Fusarium verticillioides.: Native Strains Isolated from Saline Soils Table 2. F. Recent reports have described the isolation of Ochrobactrum from plant tissue of deep-water rice (Oriza sativa) [45] as well as from soils and sediments. p Fusarium proliferatum. Given the . Fusarium verticillioides (I. B Bacteriocin. have been described as freeliving a-proteobacteria. D. and humans. F. C. S. D). 1. s Fusarium solani. H). F. Fusarium proliferatum (K. anthropi possess several enzymes including D-stereospecific aminopeptidase [13] and glutathione S-transferase that degrades atrazine as the sole source of carbon [14]. Inhibition of Sclerotinia sclerotiorum (A. Sclerotinia minor (C. Moreover. s + + + + S. g + + F. J). plant. In vitro growth inhibition of fungal phytopathogens. Isolates proceeding from soils and sediments were associated with the detoxification of xenobiotics. Fig. particularly in the utilization of halobenzoates under denitrifying conditions [42]. and ARP23 and MEP218 as Bacillus sp. Ochrobactrum spp. suspected from earlier studies [30]. animals. 46]. F.m + +/) + F. m Sclerotinia minor. I. The environmental isolates of Ochrobactrum spp. because they have mostly been isolated from the rhizosphere or the rhizoplane [28]. J.

In addition.05. 2) 100 lL. A. the antifungal activity was resistant to high temperature and protease activity. 2) untreated supernatant. Antagonistic activity of cell-free supernatants against Sclerotinia sclerotiorum. isolated from saline soils of Cordoba province. 4. ACKNOWLEDGMENTS S. Moreover. showed strong antagonism in vitro against different species of Sclerotinia and Fusarium. Means with different letters indicate significant difference with p £ 0. 55 (2007) Fig. can play an essential role in helping plants to establish and grow in salinity conditions. with 10 plants per treatment. 2. indicating that biocontrol metabolites were secreted into the culture medium. In this sense. native Bacillus strains described in this study could be employed to minimize the utilization of synthetic fungicides.E. Effect of ARP23. with 10 plants per treatment. Fig. isolates ARP2 3 and MEP2 18 identified as Fig. 2) 100 lL.320 CURRENT MICROBIOLOGY Vol. AEP1 5. Further analysis revealed that the biocontrol activity was observed in cell-free supernatants. members of the genus Bacillus. B. Further studies are being carried out in order to determine the mechanism of alleviation observed in inoculated plants under saline stress conditions as well as to identify the antifungal metabolites produced by Bacillus sp. D. 3. importance of this economic damage. 3) 150 lL. and ARP2 6 on root (A) and shoot (B) dry weight of agropyro grown in normal and saline conditions (0 and 100 mM NaCl). C. Means with different letters indicate significant difference with p £ 0. contributing to the preservation of the environment. 2) untreated LB medium (10-fold concentrated). From these results we conclude that the native strains with PGPR activity. A.E. according to the comparative sequence analysis of their 16S rRNA sequences and representative Bacillus strains. Fischer and E. Values are the means of two replicates € S. Effect of proteinase K on medium LB as control: 1) treated LB medium (10-fold concentrated). MEP3 3b. JofrØ are members of the Scientific Researcher Career-CONICET (National Council of Technological Researchs). Values are the means of two replicates € S. Supernatant of isolate ARP23: 1) 50 lL. . Supernatant of isolate MEP218: 1) 50 lL. It is well known that the genus Bacillus produces bioactive compounds belonging to the cyclic lipopeptides group with high stability attributable to their structure [43].05. Effect of proteinase K on the antifungal activity of supernatant of isolate ARP23: 1) treated supernatant. 3) 100 lL of LB medium (control). Effect of MEP211a. it is interesting to find PGPR able to control these plant diseases. MEP31 on shoot (A) and root (B) dry weight of maize grown in normal and salinity conditions (0 and 150 mM NaCl).

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