Effect of electrostatic spray parameters on the viability of two bacterial biocontrol agents and their deposition on blueberry flower stigmas

The fungus Monilinia vaccinii-corymbosi infects blueberry flowers via the stigma-style ovary pathway to cause mummy berry disease. Previous laboratory experiments documented considerable activity of stigma-applied biofungicides containing the bacteria Bacillus subtilis and, to a lesser extent, Pseudomonas fluorescens against flower infection by the pathogen. However, adequate and targeted delivery of the biocontrol agents to the stigmatic surfaces of open flowers in the field has remained problematic. Here we consider the application of the biofungicides Serenade AS (containing B. subtilis QST713) and BlightBan A506 (containing P. fluorescens A506) to blueberry flowers by air-assisted electrostatic spraying. In laboratory experiments with typical field-use rates, viability of B. subtilis and P. fluorescens was unaffected by different levels of induction-charging voltage (0–1.2 kV) and atomizing pressure (138–276 kPa) applied to an electrostatic spray-charging nozzle, showing that the bacteria in both formulations readily survived exposure to the intense electrical fields and near-sonic atomizing air shear encountered during electrostatic spraying. Electrostatically charged application significantly (P<0.0001) increased deposition of B. subtilis on the stigmatic surfaces of detached blueberry flower clusters by a factor of 4.5 compared with conventional hydraulic spraying; a similar comparison showed that population densities of P. fluorescens on the stigma were increased by a factor of 2.9, but this effect was not statistically significant (P=0.1487). For Serenade, the increased coverage and/or retention on the flower stigma, along with the excellent bacterial survival, portend well for electrostatic application for mummy berry disease control in the field.     

Biosynthesis of Amino Acids from Hydrocarbons

Nine strains of bacteria, capable of synthesising amino acids from hydrocarbons, have been identified. Pseudomonas arvilla, Ps. fluorescens and Micrococcus ureae, previously reported as hydrocarbons assimilators, were found capable of synthesising amino acids. Pseudomonas taetrolens, Bacillus cereus var. mycoides, Bacillus subtilis and Bacillus sphaericus also synthesised amino acids from hydrocarbons.     

Biological control of onion leaf blight disease by bulb and foliar application of powder formulation of antagonist mixture

Abstract

Three antagonists: Pseudomonas fluorescens (Pf1), Bacillus subtilis and Trichoderma viride, were tested alone and in combination for suppression of onion leaf blight (Alternaria palandui) disease under glasshouse and field conditions. The average mean of disease reduction was 24.81% for single strains and 42.44% for mixtures. In addition to disease suppression, treatment with a mixture of antagonists promoted plant growth in terms of increased plant height and ultimately bulb yield. Though seed treatment of either single strain or strain mixtures alone could reduce the disease, subsequent application to root, leaves or soil further reduced the disease and enhanced the plant growth. The mixture consisting of Pseudomonas fluorescens Pf1 plus Bacillus subtilis plus Trichoderma viride was the most effective in reducing the disease and in promoting plant growth and bulb yield in greenhouse and field tests.     

Potential of rhizobacteria for prohibition of potato golden cyst nematode activity

Potato golden cyst nematode is an important causal agent of potato disease. Nematode cysts were isolated and identified by phenotypic and genotypic characteristics. Biocontrol activity of 120 isolated bacteria from rhizosphere towards Globodera rostochiensis, were evaluated. Nematode cysts were exposed to root extracts and released juveniles (J2) subjected to cultural filtrate of isolated bacteria and their mortality were recorded after 24 and 48 h. In total, 33 strains showed inhibitory activities which they caused mortality ranged from 61.10% to 78.74%. Five representatives of inhibitory strains were applied on potato cultivar Marfona under greenhouse conditions using tuber coating and soil drenching in a complete randomised block design with three replicates. The active bacterial strains were identified as Bacillus subtilis, Bacillus megaterium, Pseudomonas fluorescens bv. I, P. fluorescens bv. II and P. putida bv. B. In the greenhouse, significant differences were observed among the tested strains, where the highest disease severity reduction was 42.31%.     

Effect of Pseudomonas fluorescens RB4 and Bacillus subtilis 189 on the phytoremediation potential of Catharanthus roseus (L.) in Cu and Pb-contaminated soils

The remediation of heavy metal-contaminated soils has become a critical issue due to toxic effects of these metals on living organisms. The current research was conducted to study the effect of Pseudomonas fluorescens RB4 and Bacillus subtilis 189 on the growth and phytoremediation potential of Catharanthus roseus in Cu- and Pb-contaminated soils. The bacterial strains exhibited significantly higher level of water-extractable Pb and Cu in Pb, Cu, and Cu+Pb-contaminated. The P. fluorescens RB4 inoculated plants, produced 102%, 48%, and 45% higher fresh weight (FW) in soils contaminated with Cu, Pb, and both elements, respectively, as compared to un-inoculated control plants. Similarly, B. subtilis 189 inoculated plants produced 108%, 43%, and 114% more FW in the presence of Cu, Pb, and both elements. The plants co-cultivated with both bacteria exhibited 121%, 102%, and 177% higher FW, in Cu, Pb, and both elements contaminated soils, as compared to respective un-inoculated control. Co-cultivation of P. fluorescens RB4, B. subtilis 189, and P. fluorescens RB4 + B. subtilis 189 resulted in higher accumulation of Cu and Pb in shoots of the C. roseus grown in contaminated soils as compared to un-inoculated control. Bacterial treatments also improved the translocation and metal bioconcentration factors. The growth and phytoextraction capability of C. roseus was improved by inoculation of P. fluorescens RB4 and B. subtilis 189.     

Plant growth promoting bacteria enhance water stress resistance in green gram plants

Plant growth promoting bacterial (PGPB) strains Pseudomonas fluorescens Pf1 and endophytic Bacillus subtilis EPB5, EPB22, EPB 31 were tested for their capacity to induce water stress related proteins and enzymes in green gram (Vigna radiata) plants. Among the different bacteria used, P. fluorescens Pf1 increased the vigour index, fresh weight and dry weight of green gram seedlings in vitro. Quantitative and qualitative analyses of stress-related enzymes indicated the greater activity of catalase and peroxidase in green gram plants bacterized with P. fluorescens Pf1 against water stress when compared to untreated plants. The greater accumulation of proline was recorded in Pf1 treated plants compared to untreated plants. The pot culture study revealed the greater resistance to water stress by green gram plants treated with P. fluorescens Pf1 compared to untreated plants. The greater activity of stress-related enzymes in green gram plants mediated by PGPB could pave the way for developing drought management strategies.     

A suite of recombinant luminescent bacterial strains for the quantification of bioavailable heavy metals and toxicity testing

Background  

Recombinant whole-cell sensors have already proven useful in the assessment of the bioavailability of environmental pollutants like heavy metals and organic compounds. In this work 19 recombinant bacterial strains representing various Gram-positive (Staphylococcus aureus and Bacillus subtilis) and Gram-negative (Escherichia coli, Pseudomonas fluorescens) bacteria were constructed to express the luminescence encoding genes luxCDABE (from Photorhabdus luminescens) as a response to bioavailable heavy metals ("lights-on" metal sensors containing metal-response elements, 13 strains) or in a constitutive manner ("lights-off" constructs, 6 strains).     

Extracellular factors responsible for the adaptation ofPseudomonas fluorescens batch cultures to unfavorable conditions

The culture liquid filtrate of an exponential-phasePseudomonas fluorescens batch culture added to anotherP. fluorescens culture at the moment of inoculation was found (1) to prevent or diminish cell adsorption of the flask walls, (2) to enhance the intensity of cell respiration, (3) to shorten the period of adaptation of LB-grown cells to growth in glucose-containing mineral M9 medium, (4) to stimulate bacterial growth at supraoptimum temperature (36°C) and pH values (4.8 and 9.2), and (5) to decrease the death rate of bacteria at the supraoptimum growth temperature. These results were interpreted as indicating thatP. fluorescens cultures produce two types of regulatory exometabolites similar to those revealed earlier inEscherichia coli andBacillus subtilis cultures: the direct-action adaptogenic factorX ₁ capable of increasing bacterial resistance to unfavorable growth conditions (temperature and pH) and factor of accelerated adaptation to new media. Both factors are presumably low-molecular-weight hydrophilic nonprotein compounds.     

Effects of Rhamnolipids from Pseudomonas aeruginosa DS10-129 on Luminescent Bacteria: Toxicity and Modulation of Cadmium Bioavailability

In this study, the mixture of mono- and di-rhamnolipids produced by Pseudomonas aeruginosa DS10-129 was characterized for its toxicity and modulatory effects on Cd availability to different bacteria. Gram-negative naturally bioluminescent Vibrio fischeri and recombinant bioluminescent Pseudomonas fluorescens, P. aeruginosa, Escherichia coli, and Gram-positive Bacillus subtilis were used as model organisms. Rhamnolipids reduced the bioluminescence of these bacteria in less than a second of exposure even in relatively low concentrations (30-min EC₅₀ 45–167 mg l⁻¹). Toxicity of Cd to Gram-negative bacteria (30-min EC₅₀ values 0.16 mg l⁻¹ for E. coli, 0.96 mg l⁻¹ for P. fluorescens, and 4.4 mg l⁻¹ for V. fischeri) was remarkably (up to 10-fold) reduced in the presence of 50 mg l⁻¹ rhamnolipids. Interestingly, the toxicity of Cd to Gram-positive B. subtilis (30-min EC₅₀ value 0.49 mg l⁻¹) was not affected by rhamnolipids. Rhamnolipids had an effect on desorption of Cd from soil: 40 mg l⁻¹ rhamnolipids increased the water-extracted fraction of Cd twice compared with untreated control. However, this additionally desorbed fraction of Cd remained bound with rhamnolipids and was not available to bacteria. Hence, in carefully chosen concentrations (still effectively complexing heavy metals but not yet toxic to soil bacteria), rhamnolipids could be applied in remediation of polluted areas.     

Potential of biotic inducers on disease severity and variation of Cucumber mosaic cucumovirus in cucumber plants

Six bacterial strains (Bacillus subtilis, Bacillus polymyxa, Bacillus circulans, Pseudomonas putida, Pseudomonas fluorescens 2 and Pseudomonas fluorescens 8) and one fungal isolate (Trichoderma harzianum) were tested for their ability to protect Cucumis sativus L. cv. Beith Alpha against the disease development of Cucumber mosaic cucumovirus (CMV). Seed treatment with individual bacterial and fungal liquid cultures significantly and consistently reduced the disease severity (DS) of infected cucumber plants, after 14 days from CMV inoculation onto cotyledons. All seven biotic inducers reduced the CMV infection at the range of 16.6–39% and 0–46.5% under sterilised and non-sterilised soils, respectively. The effect of treatment with each biotic inducer has a significant difference in the percentage of CMV DS. In sterilised soil, B. circulans has a low percentage of DS (42%), while P. fluorescens 8 has a high DS percentage (62.9%). In non-sterilised soil, the low DS percentage was 30% for T. harzianum, while P. putida had the highest DS percentage (70%). On the other hand, CMV variability on Chenopodium amaranticolor revealed that all biotic treatments differed according to the local lesion number, similarity and morphology.     

Ribsome and messenger specificity in protein synthesis by bacteria

Ribosomes from Escherichia coli and Pseudomonas fluorescens, two Gram-negative bacteria, translated messenger RNA preparations from all bacteria tested (3 Gram-negative species, and 6 Gram-positive species) as well as f2 RNA and T4 early messenger RNA. Ribosomes from Clostridium pasteurianum, Streptococcus faecalis, and Bacillus subtilis, three Gram-positive organisms, did not translate messenger RNA preparations derived from any of the Gram-negative organisms, the f2 RNA, or the T4 early messenger RNA but did translate messenger preparations from all 6 Gram-positive bacterial species tested.     

Management of postharvest disease of mango anthracnose incited by Colletotrichum gleosporioides

Abstract

Among the seven fungal (Gliocladium virens, Trichoderma hamatum, T. harzianum, T. koningii, T. longibrachiatum, T. pseudokoningii and T. viride) and two bacterial (Bacillus subtilis and Pseudomonas fluorescens) antagonists screened against C. gloeosporioides under in vitro conditions, T. harzianum exhibited maximum inhibition followed by Pseudomonas fluorescens at 5 days after incubation. These fungal and bacterial antagonists were selected for application to fruits infected with pathogens. Fruits inoculated with C. gloeosporioides were dipped in spore/cell suspensions of fungal/bacterial antagonists and kept for different durations. The fungal antagonists T. harzianum and P. fluorescens were effective in checking the spread of pathogens on fruits compared with the pathogen-inoculated control.     

The effect of accelerated seed ageing on cucumber germination following seed treatment with fungicides and microbial biocontrol agents for managing gummy stem blight by Didymella bryoniae

This study examined (1) the effect of the accelerated seed ageing on cucumber germination with treatments: Bacillus subtilis QST713 or Pseudomonas fluorescens CA in 1% methylcellulose and fungicides difenoconazole, carboxin or pyraclostrobin in 5% polyvinyl alcohol, and (2) the impact on disease severity of gummy stem blight (GSB) caused by Didymella bryoniae by the seed treatments and foliar spray application of methylcellulose-formulated B. subtilis or P. fluorescens. Difenoconazole, pyraclostrobin and microorganisms suppressed growth of D. bryoniae in a laboratory dual culture; carboxin had no effect on D. bryoniae growth. Germination of fungicide-treated seed was unaffected by accelerated seed ageing. Greenhouse: GSB disease severity with PVA and non-treated seed was 89% and 84%, respectively, whereas, difenoconazole, carboxin and pyraclostrobin, was significantly reduced, 56%, 53% and 40%, respectively. Germination of Bacillus-treated seed was unaffected by accelerated seed ageing, but was significantly reduced with Pseudomonas-treated seed. GSB disease severity with B. subtilis or P. fluorescens-treated seed was inconsistent; however, foliar spray application of B. subtilis significantly reduced GSB. Accelerated seed ageing exposed a significant negative impact on seed germination with P. fluorescens. Based on the accelerated ageing test, P. fluorescens-treated cucumber seed is detrimental to seed survival and therefore, is not a candidate for biocontrol activities for cucumber requiring seed storage.     

Role of polymer complexes in the formation of biofilms by corrosive bacteria on steel surfaces

The composition of exopolymer complexes (EPCs), synthesized by the monocultures Desulfovibrio sp. 10, Bacillus subtilis 36, and Pseudomonas aeruginosa 27 and by microbial associations involved in the corrosion of metal surfaces has been studied. An analysis of the monosaccharide composition of carbohydrate components, as well as the fatty acid composition of the lipid part of EPCs, was carried out by gasliquid chromatography (GLC). It was found that bacteria in biofilms synthesized polymers; this process was dominated by glucose, while the growth of bacteria in a suspension was marked by a high rhamnose content. Hexouronic acids and hexosamine have been revealed as a part of B. subtilis 36 and P. aeruginosa 27 EPCs. Qualitative differences were revealed in the fatty acid composition of exopolymers in biofilms and in a bacterial suspension. It was shown that the transition to a biofilm form of growth led to an increase in the unsaturation degree of fatty acids in the exopolymers of associative cultures. The results can be used to develop methods to control microbial corrosion of metal surfaces.     

Metal Sorption to Pseudomonas fluorescens: Influence of pH,Ionic Strength and Metal Concentrations

Migration behavior of radionuclides should be understood in order to estimate the impact of high-level radioactive waste (HLW) disposal on the environment. Bacteria, one of the major organic sorbents in solid and aquatic environments, can affect the fate of actinides and lanthanides by sorption onto their cell surfaces. In this study, the sorption of the radionuclide Americium (Am(III)) and several metal ions (Eu(III), Cu(II) and Ca(II)) to Pseudomonas fluorescens were measured under various conditions. It was revealed that as pH decreased, the sorption of Eu(III) and Am(III) increased when the metals were at relatively low concentrations but decreased at higher metal concentrations. On the other hand, sorption of Cu(II) followed the opposite trend. In the case of calcium, an increase in calcium ions was observed due to release from the cells. These findings suggest that the sorption mechanisms for low levels of Eu(III) and Am(III) on the cells of Pseudomonas fluorescens are different from those of Cu(II), Ca(II), and high concentrations of Eu(III) (> 10 ^{? 5} M).     

Foam separation of bacteria with a cationic surfactant

An experimental investigation is presented of the foam separation of six species of bacteria: Escherichia coli, Serratia marcescens, Proteus vulgaris, Pseudomonas fluorescens, Bacillus cereus, and Bacillus subtilis var niger. A cationic surfactant, ethylhexadeeyldimethylammonium bromide is used and results are evaluated in terms of total cell count, using a membrane filtration technique. From similar neutral distilled water suspensions of the pure cultures (approximately 10⁷ cells/ml.) and using the same operating conditions, ratios of cell concentrations in the residual suspensions to those in the initial suspensions range from 0.0013 for Bacillus subtilis var niger to 0.25 for Serratia marcescens. The presence of bacteria, compared to pure surfactant solutions, produces lower collapsed foam volumes; the foam volumes have a strong influence on the separations achieved with the various species, with enrichment ratios ranging from 27 to 3088 and residual ratios ranging from 0.001 to 0.247.     

A novel method for measuring the charge distribution of airborne microbes

A novel method for measuring bioaerosol charge distribution was investigated using electrostatic sampling and quantitative polymerase chain reaction (qPCR). Here, Bacillus subtilis var niger and Pseudomonas fluorescens were aerosolized and precipitated into different regions of two 96-well plates placed inside an electrostatic sampler. The concentrations of negatively charged bacteria in the air samples collected were quantified using qPCR, and the relevant bioaerosol charge levels were calculated using an equation developed in this study. Experimental results showed that B. subtilis var niger bioaerosols had wider charge distribution than P. fluorescens. For B. subtilis var niger, 93% of them carried a charge level of 13–42 units, 2% carried a charge level of 8–13 or lower, and 5% carried a charge level of 42–195 or higher. While for P. fluorescens, 99% of P. fluorescens aerosols carried a charge level of 14–34 elementary units. The indicative charge levels of bioaerosols collected at a certain point in the electrostatic sampling line largely depends on the particle size, bacterial species, electrostatic field strength, and the sampling flow rates for a given electrostatic sampler. In general, the trends for the charge levels here were similar to those obtained in previous studies. Use of qPCR technology in this study can distinguish between biological and non-biological particles collected. Thus, this avoided counting the non-biological particles in traditional bioaerosol charge measurements in which an optical particle counter is used. The developed technique here can be also used to analyze the charge levels for other types of bioaerosols.     

Vermicompost-based bioformulation for the management of sugarbeet root rot caused by Sclerotium rolfsii

Vermicompost-based bioformulations of bacterial and fungal biocontrol agents were examined against sugarbeet root rot caused by Sclerotium rolfsii. The result showed that the Pseudomonas fluorescens strain Pf1 in combination with either Trichoderma asperellum strain TTH1 or Bacillus subtilis strain EPCO-16 performed better in reducing disease next to the chemical difenoconazole. Similarly, enhanced yield was observed in the same combination treatments under both pot and field conditions.     

Experimental Measurements of the Adsorption of Bacillus subtilis and Pseudomonas mendocina Onto Fe-Oxyhydroxide-Coated and Uncoated Quartz Grains

In this study, we compared the adsorption of the gram-positive bacterium Bacillus subtilis with adsorption of the gram-negative bacterium Pseudomonas mendocina onto Fe-oxyhydroxide-coated and uncoated quartz grains as a function of pH and bacteria: mineral mass ratio. We studied metabolically-inactive cells in order to focus on the initial bacterial attachment mechanisms. The data show that the presence of Fe-oxyhydroxide-coatings on quartz surfaces significantly enhances the adsorption of bacteria and that in general the extent of adsorption decreases with increasing pH and with decreasing bacteria: mineral mass ratio. B. subtilisadsorbs to a greater extent than does P. mendocina onto the surface of the Fe-coated quartz. The adsorption behavior appears to be controlled by the overall surface charge of both bacterial and mineral surfaces. We model the adsorption data using a semi-empirical chemical equilibrium model that accounts for the site speciation of the adsorbing surfaces. Models describing bacterial adsorption to Fe-oxyhydroxide-coated quartz can account for changes in pH and bacteria: mineral mass ratio using one set of equilibrium constants.     

Phosphonic acid analogue of forfenicine — synthesis, antibacterial activity and degradation byPseudomonas fluorescens

Summary Phosphonic acid analogue of forfenicine, amino (p-formylbenzyl)-phosphonic acid, was synthesized and evaluated as antibacterial agent. As indicated by disc diffusion test this compound was found to inhibit significantly the growth ofBacillus subtilis andStaphylococcus aureus and moderately the growth ofEscherichia coli. Resistance ofPseudomonas fluorescens to the action of the aminophosphonate may result from the ability of the strain to degrade this compound.