Microbial Conversion of Petro-sulfur Compounds

For the purpose of proving possibility of desulfurization from petroleum oil by microbial procedures, first approach was made to obtaining microorganisms capable of converting petro-sulfur compounds. Dibenzothiophene was used in this study as a model of the sulfur compounds in heavy oil.

Six strains of microorganisms were isolated with dibenzothiophene from various soils. These strains produced organic acid compounds containing sulfur from dibenzothiophene. As a result of the taxonomic studies, three of the strains were found to be new species belonging to the Genus Pseudomonas, so the authors proposed to assign the names, Pseudomonas abikonensis nov. sp. to one strain, and Pseudomonas jianii nov. sp. to the other two strains.     

Biodegradation of 4-chlorobenzoic acid by <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> PA01 NC

A bacterial consortium capable of degrading chloroaromatic compounds was isolated from pulp and paper mill effluents by selective enrichment on 4-chlorobenzoic acid as sole source of carbon and energy. The four different bacterial isolates obtained from bacterial consortium were identified as Pseudomonas aeruginosa AY792969 (A), P. aeruginosa PA01 NC (B), Pseudomonas sp. ZZ5 DQ113452 (C) and Pseudomonas sp. AY762360 (D) based on their morphological and biochemical characteristics and by phylogenetic analysis based on 16S rRNA gene sequences. These bacterial isolates were found to be versatile in degrading a variety of chloroaromatic compounds including fluoro- and iodobenzoic acids. P. aeruginosa PA01 NC utilized 4-chlorobenzoic acid at 2 g/l as growth substrate. Biodegradation studies have revealed that this organism degraded 4-chlorobenzoic acid through 4-chlorocatechol which was further metabolized by ortho-cleavage pathway and the dechlorination occurred after the ring-cleavage.     

The Production of α-Ketoglutaric Acid from Salicylic Acid by Pseudomonas sp.†

Metabolites from salicylic acid by microorganisms were investigated. About eighty strains of bacteria which were able to utilize salicylic acid as a sole source of carbon were isolated from soil and other natural sources.

Among these bacteria, several strains produced a large amount of keto acids in the culture fluid during the cultivation. The acid was isolated from the culture fluid of strain K 102 in crystalline form. The crystal was identified as α-ketoglutaric acid by physicochemical methods. From the taxonomical studies, the isolated bacterial strains K 102 and K 362 were assumed to be Pseudomonas sp.     

Arsenic-resistant <Emphasis Type="BoldItalic">Pseudomonas</Emphasis> spp. and <Emphasis Type="BoldItalic">Bacillus</Emphasis> sp. bacterial strains reducing As(V) to As(III), isolated from Alps soils,Italy

Five arsenic-resistant bacterial strains (designated MP1400, MP1400a, MP1400d, APSLA3, and BPSLA3) were isolated from soils collected at the Alps region (Italy), which showed no contamination by arsenic. Phylogenetic analysis of the 16S rRNA gene sequences assigned them to the genera Pseudomonas and Bacillus. Bacillus sp. strain 1400d and Pseudomonas spp. strains APSLA3 and MP1400 showed higher tolerance to As(III), as indicated by minimum inhibitory concentrations of 10 mmol/L. Pseudomonas sp. strain MP1400 exhibited higher tolerance to As(V) (minimum inhibitory concentration of 135 mmol/L). The isolated arsenic-resistant strains were able to reduce As(V) to As(III), especially Pseudomonas sp. strain MP1400 reducing 2 mmol/L of As(V) to As(III) within 24 h. The results suggest that the isolated bacterial strains play a role in the arsenic biogeochemical cycle of arsenic-poor soils in the Alps mount area.     

Plant growth-promoting rhizobacteria modulate root-system architecture in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> through volatile organic compound emission

Extensive communication occurs between plants and microorganisms during different stages of plant development in which signaling molecules from the two partners play an important role. Volatile organic compounds (VOCs) emission by certain plant-growth promoting rhizobacteria (PGPR) has been found to be involved in plant growth. However, little is known about the role of bacterial VOCs in plant developmental processes. In this work, we investigated the effects of inoculation with twelve bacterial strains isolated from the rhizosphere of lemon plants (Citrus aurantifolia) on growth and development of Arabidopsis thaliana seedlings. Several bacterial strains showed a plant growth promoting effect stimulating biomass production, which was related to differential modulation of root-system architecture. The isolates L263, L266, and L272a stimulated primary root growth and lateral root development, while L254, L265a and L265b did not significantly alter primary root growth but strongly promoted lateral root formation. VOC emission analysis by SPME-GC-MS identified aldehydes, ketones and alcohols as the most abundant compounds common to most rhizobacteria. Other VOCs, including 1-octen-3-ol and butyrolactone were strain specific. Characterization of L254, L266 and L272a bacterial isolates by 16S rDNA analysis revealed the identity of these strains as Bacillus cereus, Bacillus simplex and Bacillus sp, respectively. Taken together, our data suggest that rhizospheric bacterial strains can modulate both plant growth promotion and root-system architecture by differential VOC emission.     

Antimicrobial activities of rhizobacterial strains of <Emphasis Type="Italic">Pseudomonas</Emphasis> and <Emphasis Type="Italic">Bacillus strains</Emphasis> isolated from rhizosphere soil of carnation (<Emphasis Type="Italic">Dianthus caryophyllus</Emphasis> cv. Sunrise)

Under the present study, an attempt was made to characterize rhizobacteria i.e. Pseudomonas and Bacillus species isolated from rhizosphere of carnation to evaluate their growth promoting effect on carnation so as to select and develop more efficient indigenous plant growth promoting and disease suppressing bioagents of specific soil type and specific plant type. Maximum strains of Pseudomonas and Bacillus sp. showed significant antimicrobial activities against most of the microorganisms tested. On the basis of in vitro antagonistic activities, the best strains were selected and used in field trial to study the influence of these strains on the growth of carnation. Results have shown marked effect on growth parameters and disease incidence has also been reduced significantly.     

Simultaneous biodegradation of pyridine and quinoline by two mixed bacterial strains

Experiments were conducted to provide data on the effectiveness of bioaugmentation in the removal of pyridine and quinoline from different wastewaters. A pyridine-degrading bacterial strain (Paracoccus sp. BW001) and a quinoline-degrading strain (Pseudomonas sp. BW003) were isolated from the activated sludge of a coking wastewater treatment plant. In this study, a consortium of these two bacterial strains was used as inoculum to simultaneously degrade pyridine and quinoline in three types of wastewaters: sterile synthetic, domestic, and industrial. In addition, variation of the bacterial community structures during degradation was monitored by denaturing gradient gel electrophoresis and amplicon length heterogeneity polymerase chain reaction techniques. The results of our experiments indicate that pyridine and quinoline can be removed efficiently using this inoculum but that the degradation process results in the production of ammonium as a by-product. Also, in the two actual wastewaters investigated, we observed that several autochthonous strains of bacteria in both the domestic and industrial wastewater were tolerant of pyridine and quinoline and grew rapidly.     

Different co-occurring bacteria enhance or decrease the growth of the microalga Nannochloropsis sp. CCAP211/78

Marine photosynthetic microalgae are ubiquitously associated with bacteria in nature. However, the influence of these bacteria on algal cultures in bioreactors is still largely unknown. In this study, eighteen different bacterial strains were isolated from cultures of Nannochloropsis sp. CCAP211/78 in two outdoor pilot-scale tubular photobioreactors. The majority of isolates was affiliated with the classes Alphaproteobacteria and Flavobacteriia. To assess the impact of the eighteen strains on the growth of Nannochloropsis sp. CCAP211/78, 24-well plates coupled with custom-made LED boxes were used to simultaneously compare replicate axenic microalgal cultures with addition of individual bacterial isolates. Co-culturing of Nannochloropsis sp. CCAP211/78 with these strains demonstrated distinct responses, which shows that the technique we developed is an efficient method for screening the influence of harmful/beneficial bacteria. Two of the tested strains, namely a strain of Maritalea porphyrae (DMSP31) and a Labrenzia aggregata strain (YP26), significantly enhanced microalgal growth with a 14% and 12% increase of the chlorophyll concentration, respectively, whereas flavobacterial strain YP206 greatly inhibited the growth of the microalga with 28% reduction of the chlorophyll concentration. Our study suggests that algal production systems represent a ‘natural’ source to isolate and study microorganisms that can either benefit or harm algal cultures.     

Molecular characterization of plant growth promoting rhizobacteria that enhance peroxidase and phenylalanine ammonia-lyase activities in chile (<Emphasis Type="Italic">Capsicum annuum</Emphasis> L.) and tomato (<Emphasis Type="Italic">Lycopersicon esculentum</Emphasis> Mill.)

Pythium and Phytophthora species are associated with damping-off diseases in vegetable nurseries and reduce seedling stand and yield. In this study, bacterial isolates were selected on the basis of in vitro antagonism potential to inhibit mycelial growth of damping-off pathogens along with plant growth properties for field assessment in wet and winter seasons. We demonstrate efficacy of bacterial isolates to protect chile and tomato plants under natural vegetable nursery and artificially created pathogen-infested (Pythium and Phytophthora spp.) nursery conditions. After 21 days of sowing, chile and tomato plants were harvested and analysed for peroxidase and phenylalanine ammonia-lyase activities. Pseudomonas sp. strains FQP PB-3, FQA PB-3 and GRP₃ were most effective in increasing shoot length (P > 0.05%) in both artificial and natural field sites. For example, Pseudomonas sp. FQA PB-3 treatment increased shoot length by 40% in the artificial Pythium 4746 infested nursery site in chile plants in the wet season. The bacterial treatments significantly increased the activity of peroxidase and phenylalanine ammonia-lyase in chile and tomato plant tissues, which are well known as indicators of an active lignification process. Thus, we conclude that treatment with potential bacterial plant growth promoting agents help plants against pathogen invasion by modulating plant peroxidase and phenylalanine ammonia-lyase activities.     

Comparison of the bacterial community and characterization of plant growth-promoting rhizobacteria from different genotypes of <Emphasis Type="Italic">Chrysopogon zizanioides</Emphasis> (L.) Roberty (Vetiver) rhizospheres

Molecular approaches [PCR-denaturing gradient gel electrophoresis (DGGE)] were used to determine whether three different vetiver (Chrysopogon zizanioides) genotypes, commercially used in Brazil and considered economically important over the world, select specific bacterial populations to coexist in their rhizospheres. DGGE profiles revealed that the predominant rhizospheric bacterial community hardly varies regarding the vetiver genotype. Moreover, using traditional cultivation methods, bacterial strains were isolated from the different rhizospheres. Colonies presenting different morphologies (83) were selected for determining their potential for plant growth promotion. More than half of the strains tested (57.8%) were amplified by PCR using nifH-based primers, specific for the enzyme nitrogenase reductase. The production of siderophores was observed in 88% of the strains, while the production of antimicrobial substances was detected in only 14.5% of the isolates when Micrococcus sp. was used as the indicator strain. Production of indole-3-acetic acid and the solubilization of phosphate were observed in 55.4% and 59% of the isolates, respectively. In total, 44 strains (53%) presented at least three characteristics of plant growth promotion and were submitted to amplified ribosomal DNA restriction analysis. Twenty-four genetic groups were formed at 100% similarity and one representative of each group was selected for their identification by partial 16S rRNA gene sequencing. They were affiliated with the genera Acinetobacter, Comamonas, Chryseobacterium, Klebsiella, Enterobacter, Pantoea, Dyella, Burkholderia, or Pseudomonas. These strains can be considered of great importance as possible biofertilizers in vetiver.     

Membrane fatty acids adaptive profile in the simultaneous presence of arsenic and toluene in <Emphasis Type="Italic">Bacillus</Emphasis> sp. ORAs2 and <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. ORAs5 strains

Bacillus sp. ORAs2 and Pseudomonas sp. ORAs5, two arsenic-resistant bacterial strains previously isolated from sediments of the Orbetello Lagoon, Italy, were tested for their adaptation to mixed contaminants on the level of membrane fatty acid composition. The two bacterial strains were characterized by high levels of arsenic resistance, and Pseudomonas sp. ORAs5 was also shown to be solvent-tolerant. The bacterial strains were exposed to mixtures of two toxic compounds: arsenic at fixed concentrations and toluene in variable amounts or, alternatively, toluene at constant values along with arsenic added at variable concentrations. Both strains react to the contaminants by changing the composition of their membrane fatty acids. Bacillus sp. strain ORAs2 showed a correlation between growth rate decreases and fatty acids degree of saturation increases in both cases, although pointedly in the presence of 1, 2, and 3 mM of toluene and different additions of arsenic, counteracting membranes fluidity induced by toxic compounds. In Pseudomonas sp. ORAs5, adaptive changes in membrane composition was observed both in terms of increases in the degree of saturation and in the trans/cis ratio of unsaturated fatty acids in the presence of varying toluene and constant arsenic concentrations, whereas only minor changes occurred with increasing arsenic and constant toluene concentrations. Thus, on the level of membrane composition, Bacillus sp. ORAs2 showed a higher potential for adaptation to the presence of mixed pollutants, suggesting its probable suitability for bioremediation purposes.     

Isolation and characterization of soil strains producing glutaryl-7-aminocephalosporanic acid acylase

A search was undertaken to screen microorganisms that produce an enzyme capable of deacylating glutaryl-7-aminocephalosporanic acid to 7-aminocephalosporanic acid in soil samples. The screening was carried out by preparing enrichment cultures containing glutaryl-7ACA and cephalosporin C as selective carbon sources. A non-β-lactam model compound, glutaryl-p-nitroanilide, was synthesized as a substrate suitable for the rapid screening of microorganisms isolated from the enrichment cultures. Two isolates exhibiting acylase activity, designated BY7.4 and BY8.1, were identified as strains ofPseudomonas species.Pseudomonas BY8.1 showed higher acylase activity toward Gl-7ACA thanPseudomonas BY7.4. Environmental conditions for the optimal acylase activity ofPseudomonas BY8.1 were shown to be pH 9 and 30°C.     

Catabolic degradation of 4-chlorobiphenyl byPseudomonas sp. DJ-12 via consecutive reaction ofmeta-cleavage and hydrolytic dechlorination

Pseudomonas sp. strain DJ-12 is a bacterial isolate capable of degrading 4-chlorobiphenyl (4CBP) as a carbon and energy source. The catabolic degradation of 4CBP by the strain DJ-12 was studied along with the genetic organization of the genes responsible for the crucial steps of the catabolic degradation. The catabolic pathway was characterized as being conducted by consecutive reactions of themeta-cleavage of 4CBP, hydrolytic dechlorination of 4-chlorobenzoate (4CBA), hydroxylation of 4-hydroxybenzoate, andmeta-cleavage of protocatechuate. ThepcbC gene responsible for themeta-cleavage of 4CBP only showed a 30 to 40% homology in its deduced amino acid sequence compared to those of the corresponding genes from other strains. The amino acid sequence of 4CBA-CoA dechlorinase showed an 86% homology with that ofPseudomonas sp. CBS3, yet only a 50% homology with that ofArthrobacter spp. However, thefcb genes for the hydrolytic dechlorination of 4CBA inPseudomonas sp. DJ-12 showed an uniquely different organization from those of CBS3 and other reported strains. Accordingly, these results indicate that strain DJ-12 can degrade 4CBP completely viameta-cleavage and hydrolytic dechlorination using enzymes that are uniquely different in their amino acid sequences from those of other bacterial strains with the same degradation activities.     

Two naphthalene degrading bacteria belonging to the genera Paenibacillus and Pseudomonas isolated from a highly polluted lagoon perform different sensitivities to the organic and heavy metal contaminants

Two bacterial strains were isolated in the presence of naphthalene as the sole carbon and energy source from sediments of the Orbetello Lagoon, Italy, which is highly contaminated with both organic compounds and metals. 16S rRNA gene sequence analysis of the two isolates assigned the strains to the genera Paenibacillus and Pseudomonas. The effect of different contaminants on the growth behaviors of the two strains was investigated. Pseudomonas sp. ORNaP2 showed a higher tolerance to benzene, toluene, and ethylbenzene than Paenibacillus sp. ORNaP1. In addition, the toxicity of heavy metals potentially present as co-pollutants in the investigated site was tested. Here, strain Paenibacillus sp. ORNaP1 showed a higher tolerance towards arsenic, cadmium, and lead, whereas it was far more sensitive towards mercury than strain Pseudomonas sp. ORNaP2. These differences between the Gram-negative Pseudomonas and the Gram-positive Paenibacillus strain can be explained by different general adaptive response systems present in the two bacteria.     

Interactions between plants and beneficial <Emphasis Type="Italic">Pseudomonas</Emphasis> spp.: exploiting bacterial traits for crop protection

Specific strains of fluorescent Pseudomonas spp. inhabit the environment surrounding plant roots and some even the root interior. Introducing such bacterial strains to plant roots can lead to increased plant growth, usually due to suppression of plant pathogenic microorganisms. We review the modes of action and traits of these beneficial Pseudomonas bacteria involved in disease suppression. The complex regulation of biological control traits in relation to the functioning in the root environment is discussed. Understanding the complexity of the interactions is instrumental in the exploitation of beneficial Pseudomonas spp. in controlling plant diseases.     

Molecular and phenotypic characterization of potential plant growth-promoting <Emphasis Type="Italic">Pseudomonas</Emphasis> from rice and maize rhizospheres

In this study, Pseudomonas species were isolated from the rhizospheres of two plant hosts: rice (Oryza sativa cultivar Pathum Thani 1) and maize (Zea mays cultivar DK888). The genotypic diversity of isolates was determined on basis of amplified rDNA restriction analysis (ARDRA). This analysis showed that both plant varieties selected for two distinct populations of Pseudomonas. The actual biocontrol and plant promotion abilities of these strains was confirmed by bioassays on fungal (Verticillum sp., Rhizoctonia solani and Fusarium sp.) and bacterial (Ralstonia solanacearum and Bacillus subtilis) plant pathogens, as well as indole-3-acetic acid (IAA) production and carbon source utilization. There was a significant difference between isolates from rice and maize rhizosphere in terms of biological control against R.  solanacearum and B.  subtilis. Interestingly, none of the pseudomonads isolated from maize rhizosphere showed antagonistic activity against R.  solanacearum. This study indicated that the percentage of pseudomonad isolates obtained from rice rhizosphere which showed the ability to produce fluorescent pigments was almost threefold higher than pseudomonad isolates obtained from maize rhizosphere. Furthermore, the biocontrol assay results indicated that pseudomonad isolated from rice showed a higher ability to control bacterial and fungal root pathogens than pseudomonad isolates obtained from maize. This work clearly identified a number of isolates with potential for use as plant growth-promoting and biocontrol agents on rice and maize.     

Microbial corrosion monitoring by an amperometric microbial biosensor developed using whole cell of Pseudomonas sp.

A microbial biosensor was developed for monitoring microbiologically influenced corrosion (MIC) of metallic materials in industrial systems. The Pseudomonas sp. isolated from corroded metal surface was immobilized on acetylcellulose membrane and its respiratory activity was estimated by measuring oxygen consumption. The microbial biosensor was used for the measurement of sulfuric acid in a batch culture medium contaminated by microorganisms. A linear relationship between the microbial sensor response and the concentration of sulfuric acid was observed. The response time of biosensor was 5 min and was dependent on the immobilized cell loading of Pseudomonas sp., pH, temperature and corrosive environments. The microbial biosensor response was stable, reproducible and specific for sensing of sulfur oxidizing bacterial activity.     

Combined use of Alkane-Degrading and Plant Growth-Promoting Bacteria Enhanced Phytoremediation of Diesel Contaminated soil

Inoculation of plants with pollutant-degrading and plant growth-promoting microorganisms is a simple strategy to enhance phytoremediation activity. The objective of this study was to determine the effect of inoculation of different bacterial strains, possessing alkane-degradation and 1-amino-cyclopropane-1-carboxylic acid (ACC) deaminase activity, on plant growth and phytoremediation activity. Carpet grass (Axonopus affinis) was planted in soil spiked with diesel (1% w/w) for 90 days and inoculated with different bacterial strains, Pseudomonas sp. ITRH25, Pantoea sp. BTRH79 and Burkholderia sp. PsJN, individually and in combination. Generally, bacterial application increased total numbers of culturable hydrocarbon-degrading bacteria in the rhizosphere of carpet grass, plant biomass production, hydrocarbon degradation and reduced genotoxicity. Bacterial strains possessing different beneficial traits affect plant growth and phytoremediation activity in different ways. Maximum bacterial population, plant biomass production and hydrocarbon degradation were achieved when carpet grass was inoculated with a consortium of three strains. Enhanced plant biomass production and hydrocarbon degradation were associated with increased numbers of culturable hydrocarbon-degrading bacteria in the rhizosphere of carpet grass. The present study revealed that the combined use of different bacterial strains, exhibiting different beneficial traits, is a highly effective strategy to improve plant growth and phytoremediation activity.     

Atrazine degradation in saline wastewater by Pseudomonas sp strain ADP

Wastewater from atrazine manufacturing plants contains large amounts of residual atrazine and atrazine synthesis products, which must be removed before disposal. One of the obstacles to biological treatment of these wastewaters is their high salt content, eg, up to 4% NaCl (w/v). To enable biological treatment, bacteria capable of atrazine mineralization must be adapted to high-salinity conditions. A recently isolated atrazine-degrading bacterium, Pseudomonas sp strain ADP, originally isolated from contaminated soils was adapted to biodegradation of atrazine at salt concentrations relevant to atrazine manufacturing wastewater. The adaptation mechanism was based on the ability of the bacterium to produce trehalose as its main osmolyte. Trehalose accumulation was confirmed by natural-abundance ¹H NMR spectral analysis. The bacterium synthesized trehalose de novo in the cells, but could not utilize trehalose added to the growth medium. Interestingly, the bacterium could not produce glycine betaine (a common compatible solute), but addition of 1 mM of glycine betaine to the medium induced salt tolerance. Osmoregulated Pseudomonas sp strain ADP, feeding on citrate decreased the concentration of atrazine in non-sterile authentic wastewater from 25 ppm to below 1 ppm in less than 2 days. The results of our study suggest that salt-adapted Pseudomonas sp strain ADP can be used for atrazine degradation in salt-containing wastewater. Received 26 August 1997/ Accepted in revised form 06 December 1997     

Aerobic Biodegradation of 2,2′-Dithiodibenzoic Acid Produced from Dibenzothiophene Metabolites

Dibenzothiophene is a sulfur heterocycle found in crude oils and coal. The biodegradation of dibenzothiophene through the Kodama pathway by Pseudomonas sp. strain BT1d leads to the formation of three disulfides: 2-oxo-2-(2-thiophenyl)ethanoic acid disulfide, 2-oxo-2-(2-thiophenyl)ethanoic acid-2-benzoic acid disulfide, and 2,2′-dithiodibenzoic acid. When provided as the carbon and sulfur source in liquid medium, 2,2′-dithiodibenzoic acid was degraded by soil enrichment cultures. Two bacterial isolates, designated strains RM1 and RM6, degraded 2,2′-dithiodibenzoic acid when combined in the medium. Isolate RM6 was found to have an absolute requirement for vitamin B₁₂, and it degraded 2,2′-dithiodibenzoic acid in pure culture when the medium was supplemented with this vitamin. Isolate RM6 also degraded 2,2′-dithiodibenzoic acid in medium containing sterilized supernatants from cultures of isolate RM1 grown on glucose or benzoate. Isolate RM6 was identified as a member of the genus Variovorax using the Biolog system and 16S rRNA gene analysis. Although the mechanism of disulfide metabolism could not be determined, benzoic acid was detected as a transient metabolite of 2,2′-dithiodibenzoic acid biodegradation by Variovorax sp. strain RM6. In pure culture, this isolate mineralized 2,2′-dithiodibenzoic acid, releasing 59% of the carbon as carbon dioxide and 88% of the sulfur as sulfate.