Genetic and Functional Diversity among Fluorescent Pseudomonads Isolated from the Rhizosphere of Banana

Fluorescent pseudomonads from banana rhizospheric soil were isolated and screened for the production of enzymes and hormones such as phosphatase, indole-3-acetic acid (IAA), 1-aminocyclopropane-1-carboxylate (ACC) deaminase, protease, and antifungal metabolites. Of 95 isolates, 50 (52%) isolates solubilized tri-calcium phosphate (TCP), 63 (66%) isolates produced plant growth hormone IAA, 10 (11%) isolates exhibited ACC deaminase, and 23 (24%) isolates produced protease. Isolates were screened for antifungal activity toward phytopathogenic fungi. Gene-specific primers have identified the putative antibiotic producing isolates. These putative isolates were grown in the production media and production of antibiotics was confirmed by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). Genotypic analysis by BOX (bacterial repetitive BOX element)-polymerase chain reaction (PCR) resulted into three distinct genomic clusters at a 50% similarity level and 62 distinct BOX profiles. Based on the sequence similarity of 16S rRNA and construction of subsequent phylogenetic tree analysis, isolates were designated as Pseudomonas monteilii, P. plecoglossicida, P. fluorescens, P. fulva, P. mosselii, P. aeruginosa, P. alcaligenes, and P. pseudoalcaligenes. Present study revealed the genetic and functional diversity among isolates of fluorescent pseudomonads associated with rhizospheric soil of banana and also identified P. monteilii as dominant species. The knowledge on genetic and functional diversity of fluorescent pseudomonads associated with banana rhizosphere is useful to understand their ecological role and for their utilization in sustainable agriculture.     

Isolation and characterization of soybean-associated bacteria and their potential for plant growth promotion

Endophytic and epiphytic bacteria were isolated from two soybean cultivars (Foscarin and Cristalina). Significant differences were observed in bacterial population densities in relation to season of isolation, soybean growth phase and the tissues from which the isolates were obtained. The isolates were identified by partial 16S rDNA sequence analysis, with most of the isolates belonging to the Pseudomonaceae, Burkholderiacea and Enterobacteriaceae groups. The potential of the isolates for plant growth promotion was evaluated by screening for indoleacetic acid (IAA) production and mineral phosphate solubilization; 34% of endophytic bacteria produced IAA and 49% were able to solubilize mineral phosphate whereas only 21% of epiphytic bacteria produced IAA although 52% were able to solubilize mineral phosphate. A high frequency of IAA producing isolates occurred in the early ripening Foscarin cultivar whereas a high percentage of phosphate solubilizing isolates were obtained from plants in the initial development stage (V6). We also found that 60% of endophytic and 69% of epiphytic isolates that produced IAA and solubilized mineral phosphate were also able to fix nitrogen in vitro. The soybean-associated bacteria showing characteristics related to plant growth promotion were identified as belonging to the genera Pseudomonas, Ralstonia, Enterobacter, Pantoea and Acinetobacter.     

Rapid identification of marine bioluminescent bacteria by amplified 16S ribosomal RNA gene restriction analysis

To rapidly identify natural isolates of marine bioluminescent bacteria, we developed amplified ribosomal DNA restriction analysis (ARDRA) methods. ARDRA, which is based on the restriction patterns of 16S rRNA gene digested with five enzymes (EcoRI, DdeI, HhaI, HinfI, RsaI), clearly distinguished the 14 species of marine bioluminescent bacteria currently known, which belong to the genera Vibrio, Photobacterium, and Shewanella. When we applied ARDRA to 129 natural isolates from two cruises in Sagami Bay, Japan, 127 were grouped into six ARDRA types with distinctive restriction patterns; these isolates represented the bioluminescent species, P. angustum, P. leiognathi, P. phosphoreum, S. woodyi, V. fischeri, and V. harveyi. The other two isolates showing unexpected ARDRA patterns turned out to have 16S rRNA gene sequences similar to P. leiognathi and P. phosphoreum. Nevertheless, ARDRA provides a simple and fairly robust means for rapid identification of the natural isolates of marine bioluminescent bacteria, and is therefore useful in studying their diversity.     

Genetic diversity and biological control activity of novel species of closely related pseudomonads isolated from wheat field soils in South Australia

Rhizobacteria closely related to two recently described species of pseudomonads, Pseudomonas brassicacearum and Pseudomonas thivervalensis, were isolated from two geographically distinct wheat field soils in South Australia. Isolation was undertaken by either selective plating or immunotrapping utilizing a polyclonal antibody raised against P. brassicacearum. A subset of 42 isolates were characterized by amplified 16S ribosomal DNA restriction analysis (ARDRA), BIOLOG analysis, and gas chromatography-fatty acid methyl ester (GC-FAME) analysis and separated into closely related phenetic groups. More than 75% of isolates tested by ARDRA were found to have >95% similarity to either Pseudomonas corrugata or P. brassicacearum-P. thivervalensis type strains, and all isolates had >90% similarity to either type strain. BIOLOG and GC-FAME clustering showed a >70% match to ARDRA profiles. Strains representing different ARDRA groups were tested in two soil types for biological control activity against the soilborne plant pathogen Gaeumannomyces graminis var. tritici, the causative agent of take-all of wheat and barley. Three isolates out of 11 significantly reduced take-all-induced root lesions on wheat plants grown in a red-brown earth soil. Only one strain, K208, was consistent in reducing disease symptoms in both the acidic red-brown earth and a calcareous sandy loam. Results from this study indicate that P. brassicacearum and P. thivervalensis are present in Australian soils and that a level of genetic diversity exists within these two novel species but that this diversity does not appear to be related to geographic distribution. The result of the glasshouse pot trial suggests that some isolates of these species may have potential as biological control agents for plant disease.     

Molecular typing of Pasteurella pneumotropica isolated from rodents by amplified 16S ribosomal DNA restriction analysis and pulsed-field gel electrophoresis

A total of 52 isolates of Pasteurella pneumotropica obtained from rodents were examined for their genetic heterogeneity. On the basis of DNA restriction analysis, including amplified 16S ribosomal DNA restriction analysis (ARDRA) and pulsed-field gel electrophoresis (PFGE), differences were identified among the isolates. ARDRA typing with Hae III revealed 4 different banding patterns of the P. pneumotropica isolates. Eighty-two percent of the 23 isolates identified as a-1 were derived from mice, whereas all the isolates identified as a-3 were derived from rats. Most of the isolates, which showed hemolytic activity on blood agar, obtained from mice and rats, were identified as a-2 and a-4, respectively. By restriction analysis of genomic DNA, Apa I and Not I digestion differentiated 9 variants and an undiscriminating group. However, no close relation with regard to the phenotypic characteristics was observed among the variants. The isolates identified as a-2 and a-4 could not be distinguished by PFGE analysis. DNA restriction analysis revealed that the genetic diversity of the P. pneumotropica isolates was more complex than the phenotypic characteristics among the species, and that at least the P. pneumotropica isolates were clearly differentiated into 4 groups by ARDRA typing with Hae III.     

Screening and Optimization of Indole-3-Acetic Acid Production and Phosphate Solubilization from Rhizobacteria Aimed at Improving Plant Growth

A total of 216 bacterial strains were isolated from rice rhizospheric soils in Northern Thailand. The bacterial strains were initially tested for solubilization of inorganic phosphate, indole acetic acid (IAA) production, selected strains were then tested for optimized conditions for IAA production and whether these caused stimulatory effects on bean and maize seedling growth. It was found that all strains had solubilized inorganic phosphate (P), but only 18.05% produced IAA. The best IAA producer was identified by biochemical testing and 16S rDNA sequence analysis as Klebsiella SN 1.1. In addition to being the best IAA producer, this strain was a high P-solubilizer and produced the highest amount of IAA (291.97 ± 0.19 ppm) in culture media supplemented with l-tryptophan. The maximum production of IAA was achieved after 9 days of incubation. The culture requirements were optimized for maximum IAA production. The tested of IAA production by selected isolates was studied in a medium with 0, 0.1, 0.2, 0.5, 0.7, and 0.9% (v/v) l-tryptophan. Low levels (12.6 ppm) of IAA production was recorded without tryptophan addition. Production of IAA in Klebsiella SN 1.1 increased with an increase to 0.2% (v/v) tryptophan concentration. The production of IAA was further confirmed by extraction of crude IAA from this isolate and subsequent Thin Layer Chromatography (TLC) analysis. A specific spot from the extracted IAA production was found to correspond with a standard spot of IAA with the same R _f value. The Klebsiella strain SN 1.1 also demonstrated stimulatory effects on bean seedlings in vivo.     

Screening of free-living rhizospheric bacteria for their multiple plant growth promoting activities

Plant growth promoting rhizobacteria (PGPR) are known to influence plant growth by various direct or indirect mechanisms. In search of efficient PGPR strains with multiple activities, a total of 72 bacterial isolates belonging to Azotobacter, fluorescent Pseudomonas, Mesorhizobium and Bacillus were isolated from different rhizospheric soil and plant root nodules in the vicinity of Aligarh. These test isolates were biochemically characterized. These isolates were screened in vitro for their plant growth promoting traits like production of indoleacetic acid (IAA), ammonia (NH(3)), hydrogen cyanide (HCN), siderophore, phosphate solubilization and antifungal activity. More than 80% of the isolates of Azotobacter, fluorescent Pseudomonas and Mesorhizobium ciceri produced IAA, whereas only 20% of Bacillus isolates was IAA producer. Solubilization of phosphate was commonly detected in the isolates of Bacillus (80%) followed by Azotobacter (74.47%), Pseudomonas (55.56%) and Mesorhizobium (16.67%). All test isolates could produce ammonia but none of the isolates hydrolyzed chitin. Siderophore production and antifungal activity of these isolates except Mesorhizobium were exhibited by 10-12.77% isolates. HCN production was more common trait of Pseudomonas (88.89%) and Bacillus (50%). On the basis of multiple plant growth promoting activities, eleven bacterial isolates (seven Azotobacter, three Pseudomonas and one Bacillus) were evaluated for their quantitative IAA production, and broad-spectrum (active against three test fungi) antifungal activity. Almost at all concentration of tryptophan (50-500 microg/ml), IAA production was highest in the Pseudomonas followed by Azotobacter and Bacillus isolates. Azotobacter isolates (AZT(3), AZT(13), AZT(23)), Pseudomonas (Ps(5)) and Bacillus (B(1)) showed broad-spectrum antifungal activity on Muller-Hinton medium against Aspergillus, one or more species of Fusarium and Rhizoctonia bataticola. Further evaluation of the isolates exhibiting multiple plant growth promoting (PGP) traits on soil-plant system is needed to uncover their efficacy as effective PGPR.     

Characterization of halotolerant rhizobia isolated from root nodules of Canavalia rosea from seaside areas

Twelve nodule isolates from Canavalia rosea, an indigenous leguminous halophyte growing in the seaside areas of southern Taiwan, were effective symbionts for the original host and able to grow at NaCl concentrations up to 3-3.5% (w/v). The taxonomy of these isolates was investigated using a polyphasic approach, including phenotypic characteristics, banding patterns of total proteins from sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), genomic fingerprint patterns from random amplified polymorphic DNA (RAPD) analysis, pulsed-field gel electrophoresis (PFGE) analysis, amplified 16S rDNA restriction analysis (ARDRA), 16S rRNA gene sequencing, and nifH gene sequencing. Based on the SDS-PAGE, RAPD, PFGE and ARDRA results, the 12 isolates are highly diverse. The 16S rRNA and nifH gene sequences were determined for isolates with distinct ARDRA patterns and compared with other members of the rhizobial species. We propose these isolates should be classified into the genus Sinorhizobium and distinguished from the current species of this genus.     

Analysis of genomic diversity among photosynthetic stem-nodulating rhizobial strains from northeast Argentina

The genomic diversity among photosynthetic rhizobia from northeast Argentina was assessed. Forty six isolates obtained from naturally occurring stem and root nodules of Aeschynomene rudis plants were analyzed by three molecular typing methods with different levels of taxonomic resolution: repetitive sequence-based PCR (rep-PCR) genomic fingerprinting with BOX and REP primers, amplified 16S rDNA restriction analysis (ARDRA), and 16S-23S rDNA intergenic spacer-restriction fragment length polymorphism (IGS-RFLP) analysis. The in vivo absorption spectra of membranes of strains were similar in the near infrared region with peaks at 870 and 800 nm revealing the presence of light harvesting complex I, bacteriochlorophyll-binding polypeptides (LHI-Bchl complex). After extraction with acetone-methanol the spectra differed in the visible part displaying peaks belonging to canthaxanthin or spirilloxanthin as the main carotenoid complement. The genotypic characterization by rep-PCR revealed a high level of genomic diversity among the isolates and almost all the photosynthetic ones have identical ARDRA patterns and fell into one cluster different from Bradyrhizobium japonicum and Bradyrhizobium elkanii. In the combined analysis of ARDRA and rep-PCR fingerprints, 7 clusters were found including most of the isolates. Five of those contained only photosynthetic isolates; all canthaxanthin-containing strains grouped in one cluster, most of the other photosynthetic isolates were grouped in a second large cluster, while the remaining three clusters contained a few strains. The other two clusters comprising reference strains of B. japonicum and B. elkanii, respectively. The IGS-RFLP analysis produced similar clustering for almost all the strains. The 16S rRNA gene sequence of one representative isolate was determined and the DNA sequence analysis confirmed the position of photosynthetic rhizobia in a distinct phylogenetic group within the Bradyrhizobium rDNA cluster.     

具有ACC脱氨酶活性的麻疯树根际促生菌(PGPR)的分离筛选及系统发育分析

[目的]获得具有产ACC、IAA,铁载体,能固氮或解磷的潜在促生菌株.[方法]通过稀释涂布的方法,从麻疯树根际土壤中分离得到98株细菌,从中选取28株以产l-氨基环丙烷-1-羧酸(ACC)脱氨酶为主要促生指标进行筛选,同时检测了其产吲哚乙酸(IAA)、固氮、解磷及铁载体等促生指标的能力.[结果]结果显示,46％的菌株能产ACC脱氨酶,其含量最高可达到128.308 μmol α-KA/(mg.h),68％的菌株能产生IAA,54％的菌株有固氮的能力,32％的菌株有解磷的能力.少量菌株同时具有产ACC脱氨酶、IAA,固氮,解磷等能力.挑选代表性菌株进行16S rRNA序列分析,这些菌株属于芽孢杆菌属(Bacillus)、节杆菌属(Arthrobacter)、假单胞菌属(Pseudomonas)和产碱杆菌属(Advenella)等8个属,其中多数菌株(50％)属于芽孢杆菌属,系统发育分析表明菌株KLBMP 4817、KLBMP 4821和KLBMP 4824为窄食单胞菌属(Stenotrophomonas)和类芽孢杆菌属(Paenibacillus)的潜在新种.[结论]攀枝花麻疯树根际土壤细菌中含有丰富的遗传多样性,且存在大量的促生菌株.其中,菌株KLBMP 4804产ACC脱氨酶含量最高.菌株KLBMP4820产IAA含量最显著.     

Genotypic and phenotypic diversity of PGPR fluorescent pseudomonads isolated from the rhizosphere of sugarcane (Saccharum officinarum L.)

The genetic diversity of plant growth-promoting rhizobacterial (PGPR) fluorescent pseudomonads associated with the sugarcane (Saccharum officinarum L.) rhizosphere was analyzed. Selected isolates were screened for plant growthpromoting properties including production of indole acetic acid, phosphate solubilization, denitrification ability, and production of antifungal metabolites. Furthermore, 16S rDNA sequence analysis was performed to identify and differentiate these isolates. Based on 16S rDNA sequence similarity, the isolates were designated as Pseudomonas plecoglossicida, P. fluorescens, P. libaniensis, and P. aeruginosa. Differentiation of isolates belonging to the same group was achieved through different genomic DNA fingerprinting techniques, including randomly amplified polymorphic DNA (RAPD), amplified ribosomal DNA restriction analysis (ARDRA), repetitive extragenic palindromic (REP), enterobacterial repetitive intergenic consensus (ERIC), and bacterial repetitive BOX elements (BOX) analyses. The genetic diversity observed among the isolates and rep-PCR-generated fingerprinting patterns revealed that PGPR fluorescent pseudomonads are associated with the rhizosphere of sugarcane and that P. plecoglossicida is a dominant species. The knowledge obtained herein regarding the genetic and functional diversity of fluorescent pseudomonads associated with the sugarcane rhizosphere is useful for understanding their ecological role and potential utilization in sustainable agriculture.     

Diversity analysis of pseudomonas in rice rhizosphere for multifaceted plant growth promotion

This investigation was carried out based on the hypothesis that there may be some pseudomonad strains, which could exist in rhizosphere of plant species contributing multifaceted beneficial activities. For this purpose, 21 pseudomonad isolates from the rhizosphere of rice, cultivated in western parts of Tamil Nadu were screened. All the 21 isolates were authenticated as pseudomonads by a genus-specific PCR screening. The molecular diversity of these isolates was investigated by Amplified Ribosomal DNA Restriction Analysis (ARDRA) and the dendrogram obtained from the analysis revealed that all the 21 isolates clustered into seven groups. Further, these isolates were screened for plant growth promoting activities such as diazotrophy (PCR amplification of nifH gene and acetylene reduction assay), Indole acetic acid (IAA) and siderophore production (spectrometrically), 1-Aminocyclopropane-1-carboxylic acid (ACC) deaminase for ethylene regulation (PCR screening), mineral solubilization (biochemically) and antagonistic potential against soil pathogenic fungi (dual culture assay). Based on the results, two elite Pseudomonas isolates (S9 and O3) were chosen as multi-functional plant growth-promoting rhizobacteria, paving way for potential use as bioinoculants in rice.     

Analysis of the abilities of endophytic bacteria associated with banana tree roots to promote plant growth

A total of 40 endophytic bacterial isolates obtained from banana tree roots were characterized for their biotechnological potential for promoting banana tree growth. All isolates had at least one positive feature. Twenty isolates were likely diazotrophs and formed pellicles in nitrogen-free culture medium, and 67% of these isolates belonged to the genus Bacillus sp. The isolates EB-04, EB-169, EB-64, and EB-144 had N fixation abilities as measured by the Kjeldahl method and by an acetylene reduction activity assay. Among the 40 isolates, 37.5% were capable of solubilizing inorganic phosphate and the isolates EB-47 and EB-64 showed the highest solubilization capacity. The isolate EB-53 (Lysinibacillus sp.) had a high solubilization index, whereas 73% of the isolates had low solubilization indices. The synthesis of indole-3-acetic acid (IAA) in the presence of L-tryptophan was detected in 40% of the isolates. The isolate EB-40 (Bacillus sp.) produced the highest amount of IAA (47.88 μg/ml) in medium supplemented with L-tryptophan and was able to synthesize IAA in the absence of L-tryptophan. The isolates EB-126 (Bacillus subtilis) and EB-47 (Bacillus sp.) were able to simultaneously fix nitrogen, solubilize phosphate and produce IAA in vitro. The results of this study demonstrated that the isolates analyzed here had diverse abilities and all have the potential to be used as growth-promoting microbial inoculants for banana trees.     

Positive control of swarming, rhamnolipid synthesis, and lipase production by the posttranscriptional RsmA/RsmZ system in Pseudomonas aeruginosa PAO1

In Pseudomonas aeruginosa, the small RNA-binding, regulatory protein RsmA is a negative control element in the formation of several extracellular products (e.g., pyocyanin, hydrogen cyanide, PA-IL lectin) as well as in the production of N-acylhomoserine lactone quorum-sensing signal molecules. RsmA was found to control positively the ability to swarm and to produce extracellular rhamnolipids and lipase, i.e., functions contributing to niche colonization by P. aeruginosa. An rsmA null mutant was entirely devoid of swarming but produced detectable amounts of rhamnolipids, suggesting that factors in addition to rhamnolipids influence the swarming ability of P. aeruginosa. A small regulatory RNA, rsmZ, which antagonized the effects of RsmA, was identified in P. aeruginosa. Expression of the rsmZ gene was dependent on both the global regulator GacA and RsmA, increased with cell density, and was subject to negative autoregulation. Overexpression of rsmZ and a null mutation in rsmA resulted in quantitatively similar, negative or positive effects on target genes, in agreement with a model that postulates titration of RsmA protein by RsmZ RNA.     

Exploration and characterization of agriculturally and industrially important haloalkaliphilic bacteria from environmental samples of hypersaline Sambhar lake, India

Screening of bacteria from Sambhar lake, an extreme hypersaline environment of India, led to the isolation of 93 haloalkaliphilic bacteria growing optimally in media with 2?C25?% salt and 6?C12 pH. Based on 16S rRNA gene sequences, 93 isolates were further categorized into 32 groups, with each group representing a different taxa belonging to 3 phyla (Firmicutes, Proteobacteria and Actinobacteria). Majority of the isolates (53.12?%) showed similarity with phylum Firmicutes which was followed by Proteobacteria (40.63?%) and Actinobacteria (6.25?%). The isolates belonging to 32 representative groups were further evaluated for the production of extracellular enzymes viz. amylase, cellulase, protease and xylanase, plant growth promoting attributes and BIOLOG? substrate usage. Among all the isolates, xylanase producing isolates were in maximum (68?%) as compared to protease (56?%), cellulase (40?%), and amylase (37?%) producing strains. Similarly, among plant growth promoting activities, ammonia producing isolates were highest (56?%) when compared to those producing ACC deaminase (53?%), IAA (50?%), hydrogen cyanide (28?%), siderophore (21?%) and solubilizing P (34?%). Isolates showing enzymatic and PGP activities could be further utilized for promoting plant growth in saline affected area.     

Molecular and functional characteristics, growth promoting effect and persistence of selected parent isolates and streptomycin resistant derivatives of rice rhizobacteria

Molecular and functional characteristics of seven azospirilla and five phosphorus solubilizing bacteria (PSB) isolates of rice rhizosphere, growth promotion ability of two efficient strains, Azospirillum amazonense A10 (MTCC4716) and Bacillus megaterium P5 (MTCC4714) and their persistence based on streptomycin resistant derivatives (SRD), were determined. SDS-PAGE and isozyme banding patterns of the isolates were used to arbitrarily group the azospirilla into 4 and PSB into 3 clusters and as markers to ascertain their identity. The azospirilla produced 2.0 to 10.5 ppm of IAA like substances and showed nitrogenase activity of 0.02 to 3.55 nmole C2H4/hr/ml of pure culture. PSB isolates produced 7.8 to 15.0 ppm IAA like substances and 20 to 128 ppm soluble P. Induction of resistance to streptomycin resulted in changes of these properties. Co-inoculation of rice with SRD A10 and SRD P5 and their parental strains in separate treatments enhanced grain yield over control by 31 and 12.4%, respectively. Nitrogenase activity of rice roots under SRD co-inoculated treatment was higher (4.16 nmole C2H4/hr/hill) than that-under parental strains co-inoculated treatment (3.76 nmole C2H4/hr/hill). SDS-PAGE profile and population count of the strains confirmed their establishment in rice rhizosphere and persistence over a year after inoculation.     

Evaluation of diversity of Bacilli from chickpea rhizosphere by 16S ARDRA and assessment of their plant-growth-promoting attributes

A total of eight motile, aerobic, Gram-positive and straight rod-shaped, endospore forming Bacillus spp. were isolated from rhizosphere of chickpea plants collected from different agricultural fields. Phylogeny of isolates was studied by partial sequencing of 16S rDNA and comparative analysis of the sequence data confirmed that the isolates belong to distinct phylogenetic lineage corresponding to Bacillus. Phenotyping clusters correlate with ARDRA pattern and showed resemblance to partial 16S rDNA sequencing. Bacillus spp. BSK5 and Bacillus subtilis BSK17 were the most potent strains for having plant-growth-promoting attributes. These two strains solubilised inorganic phosphate, produced Indole acetic acid, siderophore, Hydrocyanic acid and secreted extracellular chitinase and mild β-1,3-glucanase which antagonised and caused mycelial deformities in two phytopathogens Macrophomina phaseolina and Fusarium oxysporum in dual culture and by culture filtrate.     

Production of N-acyl Homoserine Lactones and Virulence Factors of Waterborne Aeromonas hydrophila

Aeromonads are inhabitants of aquatic ecosystems and are described as being involved in intestinal disturbances and other infections. The purpose of this study was to investigate the production of N-acyl-homoserine lactone (AHL) signal molecules and some virulence factors, including hemolysins, proteases, extracellular nucleases production and cytotoxicity by waterborne Aeromonas hydrophila. A total of 24 strains isolated from fresh-water or diseased fish were used in the study. The majority A.hydrophila strains produce two AHL molecules (21/24), one is N-butanoyl homoserine lactone (BHL), and the other is N-hexanoyl homoserine lactone (HHL) according to thin-layer chromatography analysis. Among the virulence factors tested, more than 83 % of the isolates produced β haemolysin when inoculated on sheep blood agar, only 50 % of the isolates displayed DNase activity, 75 % of the isolates shown proteolytic activity on skimmed milk plate, and cytotoxic activity was detected in 20 of 24 of the isolates. The strains producing AHLs possessed one or more virulence factors. In conclusion, the production of quorum sensing signal molecules is common among the strains that we examined, and there seems to some relationships between quorum sensing signal production and virulence factors in A. hydrophila.     

Genetic Diversity and Biological Control Activity of Novel Species of Closely Related Pseudomonads Isolated from Wheat Field Soils in South Australia

Rhizobacteria closely related to two recently described species of pseudomonads, Pseudomonas brassicacearum and Pseudomonas thivervalensis, were isolated from two geographically distinct wheat field soils in South Australia. Isolation was undertaken by either selective plating or immunotrapping utilizing a polyclonal antibody raised against P. brassicacearum. A subset of 42 isolates were characterized by amplified 16S ribosomal DNA restriction analysis (ARDRA), BIOLOG analysis, and gas chromatography-fatty acid methyl ester (GC-FAME) analysis and separated into closely related phenetic groups. More than 75% of isolates tested by ARDRA were found to have >95% similarity to either Pseudomonas corrugata or P. brassicacearum-P. thivervalensis type strains, and all isolates had >90% similarity to either type strain. BIOLOG and GC-FAME clustering showed a >70% match to ARDRA profiles. Strains representing different ARDRA groups were tested in two soil types for biological control activity against the soilborne plant pathogen Gaeumannomyces graminis var. tritici, the causative agent of take-all of wheat and barley. Three isolates out of 11 significantly reduced take-all-induced root lesions on wheat plants grown in a red-brown earth soil. Only one strain, K208, was consistent in reducing disease symptoms in both the acidic red-brown earth and a calcareous sandy loam. Results from this study indicate that P. brassicacearum and P. thivervalensis are present in Australian soils and that a level of genetic diversity exists within these two novel species but that this diversity does not appear to be related to geographic distribution. The result of the glasshouse pot trial suggests that some isolates of these species may have potential as biological control agents for plant disease.