Bacterial Culture Filtrates Research Articles

Control of grey mould disease on strawberry using the effective agent, Bacillus amyloliquefaciens Y1

ABSTRACT This study was conducted to investigate the effect of Bacillus amyloliquefaciens Y1 on the control of grey mould disease caused by Botrytis cinerea and subsequent growth promotion of strawberry. Strain Y1 strongly inhibited mycelial growth of B. cinerea on different media with the highest inhibition of 61.22% on V8 juice agar medium. The higher the bacterial culture filtrate (BCF) concentration of strain Y1 blended in the PDA medium, the stronger the rate of inhibition on mycelial growth of B. cinerea. Microscopic observation revealed that the hyphal structure of the fungal pathogen was severely affected by the BCF indicating abnormal growth with excessive branching and swelling. Strain Y1 showed the promising ability to suppress grey mould disease under in vivo conditions and significantly lessened disease severity on flowers and fruits leading to an increase in disease control value when compared to fungicide treatment. Moreover, application of strain Y1 resulted in a significant promotion of strawberry plants with higher values in most growth characteristics compared to other treatments. The detection of higher levels of the defence-related enzyme, β-1,3-glucanase in the Y1-treated plants indicated that induction of resistance by this strain was at least partly involved in disease suppression. Even though there were reductions in the overall numbers of the soil bacterial population in the Y1-treated plants, no significant impact on the growth of strawberry was observed. Based on our results, strain Y1 is a promising candidate for environment-friendly disease control strategies.

ABA analogue produced by Bacillus marisflavi modulates the physiological response of Brassica juncea L. under drought stress

The present study aims to understand the molecular mechanism involved in beneficial rhizobacteria mediated alleviation of drought stress in the host plant. Bacillus marisflavi CRDT-EB-1 isolated from the rhizosphere soil was found effective in inducing resistance against drought stress in mustard seedlings. Among the different bacterial derivatives tested, the culture filtrate was found to contain bioactive molecules. Solvent extract of bacterial culture filtrate yielded seven distinct bands/fractions on thin layer chromatography (TLC). The fraction four (F4) with Rf value 0.35–0.40 was significant in reducing the adverse effect of drought stress in host plants. Application of F4 resulted in delayed drooping point and higher drought tolerance index (3.34), induced stomatal closure (9.648 μ), seed germination inhibition (12%), and reduced the GA3 induced α-amylase activity in germinating barley seeds. On TLC, F4 turned colorless to orange color upon the spray of 2,4-dinitrophenylhydrazine reagent indicated the presence of aldehyde group. Supporting to this, the peaks between 9.8 and 10.0 ppm in 1H NMR chromatogram confirmed the presence of the aldehyde group. Upon LC-MS/MS analysis of a crude extract of culture filtrate and F4 revealed the presence of compounds with the molecular mass 250.33 and 266.33. By analyzing these data, the identity of the bioactive compounds was predicted as xanthoxin and xanthoxic acid, which are a well-known precursor of Abscisic acid (ABA) in plants. The present study concludes the capability of ABA analogue (xanthoxin like compounds) production by B. marisflavi CRDT-EB-1 and its involvement in inducing drought stress tolerance in the host plant.

Non-chemical based management of Tylenchulus semipenetrans, a major threat to the citrus industry

There is a vast variety of microbes available in soil performing diverse functions for plants including root colonization for protection against pathogens and nutrient acquisition. In the present study, efficacy of rhizospheric microbial strains and plant extracts were evaluated against citrus nematode for their nematicidal activity. Three different concentrations of plant extracts were evaluated in vitro, results depicts that 10% solution of Tagetes erecta (root) has given 46% the highest percentage mortality at 48hrs followed by Tagetes erecta (leaf) and Datura stramonium killed 41% and 37% nematodes respectively, similarly for 20% of plant extract in distilled water, the highest mortality rate was recorded for T. erecta (root) 68% followed by D. stramonium 47% and Acacia nilotica 44% at 48hrs of application. While using a higher concentration 30% of SS. the percentage mortality elevated multifold i.e. Marigold root has killed maximum 76% nematodes followed by Marigold leaves and Dhatura with mortality rate 69% and 58% respectively at 48 hrs of interval. While Eucalyptus killed lowest no of nematodes. Additionally, five bacterial isolates were evaluated for their nematicidal activity. Results indicated that for bacterial cell suspension, Pseudomonas putida has shown maximum mortality rate at 84% followed by Bacillus subtilis 73% & Pseudomonas geniculata 65% at 48hrs of interval. Similarly, in the 2nd set of experiment bacterial culture filtrates has exhibited promising results, maximum mortality rate was recorded for Bacillus spp. 81% followed by B. subtilis 76%, Ps putida 63%, after 48hrs of incubation. Moreover, an insight investigation of bacterial extracellular metabolites and plant extracts would enable researchers to develop purposeful products and compositions

Role of bacterial antagonists of fungal pathogens, Bacillus thuringiensis KYC and Bacillus velezensis CE 100 in control of root-knot nematode, Meloidogyne incognita and subsequent growth promotion of tomato

ABSTRACT In this study, the biocontrol ability of bacterial antagonists of fungal pathogens, B. thuringiensis KYC and B. velezensis CE 100 was investigated for the inhibition of root-knot nematode, Meloidogyne incognita, and subsequent growth promotion of tomato. Based on the results of dual-culture assay, both Bacillus strains showed antagonistic activities against six different phytopathogenic fungi. The bacterial culture filtrates (BCF) of two Bacillus strains had a strong toxic effect on egg hatching and second-stage juvenile (J2) survival of M. incognita. With increasing concentration of BCF and incubation period, the hatching rate of the nematode egg was significantly inhibited and J2 mortality was remarkably increased. Moreover, the crude enzymes which were extracted from culture supernatants of two Bacillus strains caused structural damage to nematode eggs and J2 at 1, 3 and 5 days after inoculation. The pot experiment revealed that B. thuringiensis KYC and B. velezensis CE 100 significantly promoted tomato shoot growth compared to fertiliser treatment. However, root fresh weight was significantly increased in fertiliser treatment due to numerous masses in roots. The number of egg masses and root gall index in the treatment group with both Bacillus strains were significantly decreased compared to those in fertiliser treatment. Higher populations of B. thuringiensis KYC and B. velezensis CE 100 (4.12 and 6.52 × 106 CFU g−1of soil) were detected in the tomato rhizosphere. These results demonstrated the potential of B. thuringiensis KYC and B. velezensis CE 100 as biocontrol agents to control plant-pathogenic fungi and root-knot nematode and as biofertilisers to promote plant growth.

Antagonistic activity of cowshed Bacillus sp. bacteria against aflatoxigenic and sclerotic Aspergillus flavus

Mycotoxins produced by many food spoilage fungi cause serious damage to human as well as to livestock. Aflatoxins are one such group of mycotoxins produced by Aspergillus flavus in many agricultural and food products including rice. The fungus can virtually grow in any environment and also produces resistive structures such as sclerotia to overcome unfavourable environmental conditions. Hence, the need of the time is to control this toxigenic and sclerotic fungus through an eco-friendly approach. In the current study four biocontrol bacteria belonging to Bacillus species were isolated from Indian cow shed environment and these bacteria could efficiently control not only the mycelia growth of A. flavus but also the germination and growth of sclerotia. Both active bacterial culture broth and cell free culture filtrate could limit the growth of the fungus up to more than 90%. Older culture broth and filtrate lost their inhibition efficiency. Unlike many similar studies the current investigation emphasizes the importance of cowshed environment instead of cow dung and the control of sclerotia instead of fungal spores.

Experimental exposure of Burkholderia pseudomallei crude culture filtrate upregulates PD-1 on T lymphocytes.

Burkholderia pseudomallei is the causative agent for melioidosis. Because of its intracellular nature, the bacterium is capable of replicating within a plethora of eukaryotic cell lines. B. pseudomallei can remain dormant within host cells without symptoms for years, causing recrudescent infections. Here, we investigated the pathogenesis mechanism behind the suppression of T cell responses by B. pseudomallei . Peripheral blood mononuclear cells (1×106 cells/well) isolated by Ficoll Paque (Sigma-Aldrich) density gradient centrifugation were incubated with optimized concentrations of bacterial crude culture filtrate antigens (CFAs) (10 ug ml−1) and heat-killed bacteria [1 : 10 multiplicity of infection (m.o.i.)]. Following incubation, cells were investigated for surface expression of coinhibitory molecules by flow cytometry. We found that B. pseudomallei induced the upregulation of programmed death 1 (PD-1), a molecule responsible for T cell exhaustion, on T cells in vitro following exposure to crude CFAs of B. pseudomallei . This upregulation of PD-1 probably contributes to poor immune surveillance and disease pathogenesis.

Zinc solubilizing plant growth promoting microbes produce zinc nanoparticles

Strains of Pseudomonas, Bacillus, and Azospirillum with plant growth promoting ability were checked for their zinc solubilizing ability at ICAR-CRIDA, Hyderabad, India. Efficient zinc solubilizing microorganisms were evaluated for their ability to produce nano-scale zinc particles. The nanoparticles from the cell-free culture filtrates obtained from these strains were characterized for particle size, Zeta potential and functional groups. Presence of Zn nanoparticles in the bacterial culture filtrate was confirmed by particle distribution and Scanning electron microscope (SEM) analysis. Most properties of nanoparticles are size dependent. Zinc nanoparticles were observed to be spherical in shape and size ranged from 52.0 to 106.0nm. Zeta potential of the Zn nanoparticles was estimated to understand the stability of the particles. The measured zeta potentials varied from -14.5mV to +179.10 mV indicating high stability and dispersion of the zinc nanoparticles. FTIR peaks at different wave numbers depicted the role of functional groups of proteins in the biosynthesis of Zn nanoparticles. These results demonstrate the green synthesis of zinc nanoparticles by the plant growth promoting and zinc solubilizing strains of Azospirillum, Pseudomonas and Bacillus.

In vitro evaluation of some isolated bacteria against the plant parasite nematode Meloidogyne incognita

BackgroundMicrobial cultures are extensively used as environment friendly ways for biological control of parasitic pests, including the root-knot nematodes, alternative to the use of chemical nematicides. The present study was conducted to isolate some lytic rhizobacteria and examine their nematicidal activity against Meloidogyne incognita J2 mortality and egg hatching in in vitro test.ResultsLytic Rhizobacteria were isolated from soil samples adhering to tomato plant roots from different localities at Giza governorate, Egypt. Six bacterial isolates, exhibited high efficacy against root-knot nematodes, were identified based on the analysis of the 16S rRNA gene sequence as Pseudomonas aeruginosa, Paenibacillus polymyxa, Lysinibacillus sphaericus, Bacillus cereus, Bacillus subtilis, and Achromobacter xylosoxidans. These strains showed high production of chitinase, chitosanase, and protease using colloidal chitin and soluble chitosan as carbon sources. Two strains, Paenibacillus polymyxa and Bacillus subtilis, produced the highest levels of chitinase in the media. Except for Pseudomonas aeruginosa, all the bacterial strains produced high levels of chitosanase.ConclusionThe results revealed that high amounts of protease were, however, secreted by Pseudomonas aeruginosa as compared to the other strains. In in vitro tests, all the bacterial culture filtrates potentially displayed nematicidal effect in M. incognita egg hatching and an obvious increase in J2 mortality as compared to control. Paenibacillus polymyxa caused 100% juvenile mortality followed by Bacillus subtilis 97.25%, Bacillus cereus 94%, Achromobacter xylosoxidans 93%, Lysinibacillus sphaericus 92%, and Pseudomonas aeruginosa 84.29% after 48 h of exposure, as compared to control.

A Rapid Isolation Method of Extracellular Proteins Produced by Pseudomonad Strains

Manufacturing of protein/enzymes by biotechnological methods may contribute to develop new protein/enzyme preparations useful in various sectors of industry. Microbial enzymes released in growing medium and difficult to isolate. In this study, a rapid method has been developed for isolating extracellular proteins from cultured nutrient broth. After removing the bacterial cells from cultured broth through 0.4mm filter paper and dissolved proteins recovered by precipitation with 5% trichloroacetic acid (TCA). Sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the isolated crude enzymes from bacterial cell free culture filtrate showed the presence of different bands in the electrophoretic field.

In vitro interaction between Bacillus megaterium strains and Caco-2 cells.

To further our understanding of the virulence potential of Bacillus megaterium strains, cell association and invasion assays were conducted in vitro by infecting human enterocytes (Caco-2 cells) with 53 strains of this bacterium isolated from honey. Two series of experiments were performed: (i) necrosis and cell detachment assays with the supernatants of bacterial culture filtrates from 16-h cultures and (ii) adhesion/invasion assays in which cultured enterocytes incubated with bacteria from 3-h cultures were resuspended in Dulbecco's modified Eagle's medium and chloramphenicol. The detachment of Caco-2 cells was evaluated by staining the cells with crystal violet. Necrosis was assessed by fluorescence microscopy of cells labeled with propidium iodide. Association (adhesion plus invasion) was determined by plate counts and invasion in an aminoglycoside protection assay. The results showed that spent culture supernatants detached and necrotized Caco-2 cells in a strain-dependent manner. Seven out of 53 B. megaterium filtered culture supernatants caused complete cell detachment. Suspensions of these same bacterial strains adhered and invaded enterocytes in 2-h infection experiments. To our knowledge, this is the first report on the interaction between B. megaterium and intestinal epithelial Caco-2 cells.

Unraveling the mode of antifungal action of Bacillus subtilis and Bacillus amyloliquefaciens as potential biocontrol agents against aflatoxigenic Aspergillus parasiticus

Some filamentous fungi are plant pathogens that cause serious economic losses in foodstuffs and agricultural commodities. In the present study, the antifungal effects of culture filtrates of soil strains of Bacillus subtilis and Bacillus amyloliquefaciens were studied against some common field-pathogenic fungi with special reference to the possible mode of actions against Aspergillus parasiticus NRRL2999. The fungal growth and aflatoxin production were determined by microbioassay technique. Changes in membrane ergosterol content, mitochondrial dehydrogenase activity, and chitin and β-1,3-glucan content of the fungal cell wall were evaluated as the different possible targets in the presence of various concentrations of bacterial culture filtrates. According to the results, both bacteria showed strong antifungal activity against the broad spectrum of filamentous fungi. They were able to suppress A. parasiticus growth (up to 92%) and aflatoxin production (up to 100%) and showed good proteolytic activities (up to 10.5 ± 0.4 mm clear zones on gelatin agar). Both antagonistic bacteria decreased ergosterol content of the fungal cell membrane of A. parasiticus (9.0–80%) in a dose-dependent manner. They also affected dehydrogenase activity of fungal cell mitochondria. Interestingly, by using 2500 μl of bacterial culture filtrate, the slight resume of fungal growth and cell wall compositions, referred as the paradoxical effect, was exhibited. Scanning and transmission electron microscopy showed drastic changes in structure and shapeof the treated fungal hyphae and vesicles including folding, wrinkling, cell depletion and vacuolization. Altogether, our results show B. subtilis and B. amyloliquefaciens target different sites in the fungal cells and thus, they could act as biological control agents to combat A. parasiticus growth and subsequent aflatoxin contamination of crops and agricultural commodities.

Isolation and characterization of metabolites from Bacillus licheniformis MH48 with antifungal activity against plant pathogens.

In this study, we isolated Bacillus licheniformis MH48 from rhizosphere soil and demonstrated that this strain shows significant antifungal activity against Rhizoctonia solani, Colletotrichum gloeosporioides, and Phytophthora capsici. Our results showed that a 50% concentration of bacterial cell-free culture filtrate of B.licheniformis MH48 shows strong activity against fungal pathogens. Benzoic acid produced by B.licheniformis MH48 was purified by various chromatographic techniques and identified by nuclear magnetic resonance and gas chromatography-mass spectrometry analysis. Benzoic acid displayed antifungal activity against R.solani and C.gloeosporides with minimum inhibitory concentration of 128μg/mL against mycelial growth. Microscopic examination revealed that benzoic acid (50μg/mL and 100μg/mL) transformed C.gloeosporioides conidial morphology and inhibited conidial germination. In addition, benzoic acid (100μg/mL and 200μg/mL) degraded R.solani mycelia. Therefore, our results demonstrate that B.licheniformis MH48 strain shows potential for utility as a biological agent for the control of various fungal pathogens of plants.

Effective control of root-knot nematode disease with Pseudomonad rhizobacteria filtrate

Root-knot disease poses a worldwide problem to agriculture; hence, several environmentally hazardous chemical nematicides are being used to control it. Rhizobacteria could be one of the important biocontrol agents which can replace these chemicals. This study focused on the effect of bacterial cell-free culture filtrate against Root-knot nematodes. Experiment was conducted to observe the effect of different dilutions of cell-free culture filtrate on Meloidogyne incognita under laboratory and in greenhouse conditions. In laboratory, toxicity effect of filtrate was recorded on second stage juveniles (J2), 100% mortality was recorded in all treatment above the 75% concentration. In greenhouse, all the growth and nematode induced parameters were significantly differed after culture filtrate treatment to the tomato plants. The finding indicates the capability of fluorescent Pseudomonad's culture filtrate as a powerful biocontrol tool for the management of Root-knot nematodes.

Dickeya dadantii pectic enzymes necessary for virulence are also responsible for activation of the Arabidopsis thaliana innate immune system.

Soft-rot diseases of plants attributed to Dickeya dadantii result from lysis of the plant cell wall caused by pectic enzymes released by the bacterial cell by a type II secretion system (T2SS). Arabidopsis thaliana can express several lines of defence against this bacterium. We employed bacterial mutants with defective envelope structures or secreted proteins to examine early plant defence reactions. We focused on the production of AtrbohD-dependent reactive oxygen species (ROS), callose deposition and cell death as indicators of these reactions. We observed a significant reduction in ROS and callose formation with a bacterial mutant in which genes encoding five pectate lyases (Pels) were disrupted. Treatment of plant leaves with bacterial culture filtrates containing Pels resulted in ROS and callose production, and both reactions were dependent on a functional AtrbohD gene. ROS and callose were produced in response to treatment with a cellular fraction of a T2SS-negative mutant grown in a Pels-inducing medium. Finally, ROS and callose were produced in leaves treated with purified Pels that had also been shown to induce the expression of jasmonic acid-dependent defence genes. Pel catalytic activity is required for the induction of ROS accumulation. In contrast, cell death observed in leaves infected with the wild-type strain appeared to be independent of a functional AtrbohD gene. It was also independent of the bacterial production of pectic enzymes and the type III secretion system (T3SS). In conclusion, the work presented here shows that D. dadantii is recognized by the A. thaliana innate immune system through the action of pectic enzymes secreted by bacteria at the site of infection. This recognition leads to AtrbohD-dependent ROS and callose accumulation, but not cell death.

Native Pseudomonas spp. suppressed the root-knot nematode in in vitro and in vivo, and promoted the nodulation and grain yield in the field grown mungbean

The study describes the effects of Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas stutzeri and Pseudomonas striata on root nodulation (Bradyrhizobium sp.), root-knot disease (Meloidogyne incognita), and on the plant growth and yield of field grown mungbean (Vigna radiata). In vitro the Pseudomonas spp. produced hydrogen cyanide (P. fluorescens), ammonia, siderophore and indole acetic acid, and solubilized phosphorus. The bacterial cells and cell free culture filtrate of Pseudomonas spp. suppressed egg hatching and induced mortality to the juveniles of M. incognita with overall greater effects of P. fluorescens and P. striata. The seeds of mungbean cv. T-44 treated with Pseudomonas spp. (1011–12colony formingunits/ml) and Bradyrhizobium (109–10CFU/ml, 5ml/kg seeds) were sown at 45 spots in every field plot (4×2m) with or without the nematode inoculation of 2000juveniles/spot. In the plots without nematodes, grain yield and NPK acquisition of mungbean plants were significantly higher with P. fluorescens or P. striata treatments (P⩽0.05). The Pseudomonas treatments synergized with Bradyrhizobium sp., leading to significant enhancements in the root nodulation, leghemoglobin contents, and the population of rhizobia in the nodule as well as in the soil. The nematode infection caused severe galling on the roots and decreased the yield of mungbean by 21% in the absence of Pseudomonas spp. The nematode infection also suppressed the nodulation, leghemoglobin contents and rhizobia population. However, treatments with Pseudomonas spp. improved the nodulation and other parameters of nematode infected plants. Among the four Pseudomonas spp., P. fluorescens was found more effective against the root-knot nematode, and its application reduced the gall formation, reproduction and soil population of M. incognita (P⩽0.05). The suppressive effect of P. fluorescens on the nematode was relatively less than carbofuran (5g active ingredient/kg seed), but the yield of nematode infected plants with the bacterial treatment was 14% (P⩽0.05) greater than the chemical treatment. Hence, seed treatment with P. fluorescens offers a better substitute of the nematicide in mungbean cultivation.

유해조류번성 주변의 해수와 침전물에서 살조균의 분리

This study mainly focused on isolation of marine algicidal bacteria associated with phytoplankton blooms and characterization of algicidal activity against harmful algae. Harmful algal blooms (HABs) found naturally in surface waters have caused many environmental problems worldwide. In this study, forty bacterial strains that have capability of inhibiting harmful algal growth were isolated from Masan Bay, Jinhae Bay, Dol Island, Jangmok Bay, and the Tongyeong Sea, Republic of Korea. The bacteria were screened furthermore for the characteristics on algicidal activities against Cochlodinium polykrikoides, Chattonella marina, Skeletonema costatum, Heterosigma akashiwo, Heterocapsa triquetra, Prorocentrum minimum, and Scrippsiella trochoidea. As a result, the algicidal bacteria that were screened from double over layer agar and microscopic counts tests belonged to genera Pseudomonas, Vibrio, Bacillus, Pseudoalteromonas, Ruegeria, Joostella, Marinomonas, Stakelama, Porphyrobacter, and Albirhodobacter. One of the most important HAB species is Co. polykrikoides and the strongest algicidal activity against the dinoflagellate was 94.00% after 6 h treatment with 10% bacterial culture filtrate. In this study, Marinomonas sp. M Jin 1-8, Stakelama sp. ZB Yeonmyeong 1-11 & 1-13, Porphyrobacter sp. M Yeonmyeong 2-22, and Albirhodobacter sp. 6-R Jin 6-1 were found to be as new genera of bacteria having anti-algal activity. These results suggest that these bacteria might play an important role in controlling phytoplankton blooms.

Streptococcus mutans Can Modulate Biofilm Formation and Attenuate the Virulence of Candida albicans.

Streptococcus mutans and Candida albicans are found together in the oral biofilms on dental surfaces, but little is known about the ecological interactions between these species. Here, we studied the effects of S. mutans UA159 on the growth and pathogencity of C. albicans. Initially, the effects of S. mutans on the biofilm formation and morphogenesis of C. albicans were tested in vitro. Next, we investigate the influence of S. mutans on pathogenicity of C. albicans using in vivo host models, in which the experimental candidiasis was induced in G. mellonella larvae and analyzed by survival curves, C. albicans count in hemolymph, and quantification of hyphae in the host tissues. In all the tests, we evaluated the direct effects of S. mutans cells, as well as the indirect effects of the subproducts secreted by this microorganism using a bacterial culture filtrate. The in vitro analysis showed that S. mutans cells favored biofilm formation by C. albicans. However, a reduction in biofilm viable cells and inhibition of hyphal growth was observed when C. albicans was in contact with the S. mutans culture filtrate. In the in vivo study, injection of S. mutans cells or S. mutans culture filtrate into G. mellonella larvae infected with C. albicans increased the survival of these animals. Furthermore, a reduction in hyphal formation was observed in larval tissues when C. albicans was associated with S. mutans culture filtrate. These findings suggest that S. mutans can secrete subproducts capable to inhibit the biofilm formation, morphogenesis and pathogenicity of C. albicans, attenuating the experimental candidiasis in G. mellonella model.

Root endophytic bacteria of a 137Cs and Mn accumulator plant, Eleutherococcus sciadophylloides, increase 137Cs and Mn desorption in the soil

We found that root endophytes of 137Cs accumulator plant produce siderophores, resulting in the desorption of 137Cs from the contaminated soil collected at Fukushima, Japan. We selected an endemic Japanese deciduous tree, Eleutherococcus sciadophylloides (Franch. et Sav), that accumulates high concentrations of 137Cs and Mn. Root endophytic bacteria were isolated from E. sciadophylloides and microbial siderophore production was evaluated via chrome azurol S (CAS) Fe and CAS Al assays. Of the 463 strains that we isolated, 107 (23.1%) produced the siderophores. Using eight strains that showed high siderophore production in our assays, we examined desorption of 137Cs, Mn, Fe and Al by the bacterial culture filtrates from 137Cs-contaminated soil after decomposing the soil organic matter using hydrogen peroxide. We found 137Cs and Mn desorption concomitant with Al and Fe desorption, as well as a decrease of pH. We also detected succinic acid, a well-known siderophore, in the bacterial culture filtrates of our two root endophytic bacteria. Our results strongly suggest that the root endophytic bacteria of E. sciadophylloides produce the siderophores that enhance 137Cs and Mn desorption in the rhizosphere, making the resulting 137Cs and Mn ions easier for E. sciadophylloides to absorb from the rhizosphere.

Isolation and Biocontrol Potential of Bacillus amyloliquefaciens Y1 against Fungal Plant Pathogens

This study was performed to investigate thermophilic bacteria from soil having broad antifungal spectrum against Rhizoctonia solani, Colletotrichum gloeosporioides, Phytophthora capsici, Fusarium oxysporum f.sp. lycopersici, and Botrytis cinerea. One isolate selected could resist heat shock of 60°C for one hour, and had broad antifungal activity in dual culture assay against all tested fungal pathogens and was identified as Bacillus amyloliquefaciens Y1 using 16S rRNA gene sequence. Further investigation for antifungal activity of bacterial culture filtrate (BCF) and butanol crude extract (BCE) of various concentrations showed broad spectrum antifungal activity and fungal growth inhibition significantly increased with increasing concentration with highest growth inhibition of 100% against R. solani with 50% BCF and 11 mm of zone of inhibition against R. solani with 4 mg BCE concentration. Treatment of butanol crude extract resulted in deformation, lysis or degradation of C. gloeosporioides and P. capsici hyphae. Furthermore, B. amyloliquefaciens Y1 produced volatile compounds inhibiting growth of R. solani (70%), C. gloeosporioides (65%) and P. capsici (65-70%) when tested in volatile assay. The results from the study suggest that B. amyloliquefaciens Y1 could be a biocontrol candidate to control fungal diseases in crops.

Mechanism of plant growth promotion elicited by Bacillus sp. LKE15 in oriental melon

Development of organic fertilizers using microbial extracts is an emerging method in organic farming. The objective of this study was to identify and characterize the bioactive substances in a plant growth-promoting bacterial culture, intended to enhance the growth of oriental melons (Cucumis melo L.). LKE15 was isolated from oriental melon rhizosphere soil and identified as Bacillus sp. LKE15 through 16S rDNA sequencing of the bacterial genomic DNA and a subsequent phylogenetic analysis. Bacillus sp. LKE15 was found to enhance the shoot length, fresh weight, dry weight, and chlorophyll content of both dwarf rice (gibberellins [GAs]-deficient mutant, waito-C) and normal rice (active GAs biosynthesis, Hwayoungbyeo) cultivars, as compared to their controls. The presence of bioactive metabolites such as GAs (GA4, GA9, and GA34), indole-3-acetic acid, and high concentrations of phosphate in the bacterial culture filtrate (CF) significantly promote host plant growth. The plant growth-stimulating effect of CF was confirmed by the increased shoot length, root length, and plant fresh weight of oriental melon as compared to the controls. Similarly, Bacillus sp. LKE15 inoculation also enhanced plant growth but at a lower rate than CF. The increased synthesis of chlorophyll and individual amino acids and their derivatives, including taurine, aspartic acid, threonine, serine, glycine, alanine, citrulline, isoleucine, leucine, tyrosine, β-alanine, γ-amino-n-butyric acid, ethanolamine, 1-methylhistidine, and hydroxyproline in CF-treated plants reflected a significant increase in the growth of treated plants, as compared to the control. However, magnesium, potassium, and phosphorus contents were significantly enhanced in plants treated with CF. Results of this study suggest that the availability of bioactive metabolites in the CF of Bacillus sp. LKE15 stimulates plant growth, and that treatment with the CF of Bacillus sp. LKE15 would be a useful tool for the organic farming of crop plants.