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The type VI secretion system (T6SS) is a phage-derived contractile nanomachine primarily involved in interbacterial competition. Its pivotal component, TssA, is indispensable for the assembly of the T6SS sheath structure, the contraction of which propels a payload of effector proteins into neighboring cells. Despite their key function, TssA proteins exhibit unexpected diversity and exist in two major forms, a short form (TssA S ) and a long form (TssA L ). While TssA L proteins interact with a partner, called TagA, to anchor the distal end of the extended sheath, the mechanism for the stabilization of TssA S -containing T6SSs remains unknown. Here we discover a class of structural components that interact with short TssA proteins and contribute to T6SS assembly by stabilizing the polymerizing sheath from the baseplate. We demonstrate that the presence of these components is important for full sheath extension and optimal firing. Moreover, we show that the pairing of each form of TssA with a different class of sheath stabilization proteins results in T6SS apparatuses that either reside in the cell for some time or fire immediately after sheath extension. We propose that this diversity in firing dynamics could contribute to the specialization of the T6SS to suit bacterial lifestyles in diverse environmental niches., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

Pseudomonas is considered as the specific spoilage bacteria in meat and meat products. The purpose of this study was to evaluate the inactivation efficiency and mechanisms of slightly acidic electrolyzed water (SAEW) against Pseudomonas deceptionensis CM2, a strain isolated from spoiling chicken breast. SAEW caused time-dependent inactivation of P. deceptionensis CM2 cells. After exposure to SAEW (pH 5.9, oxidation-reduction potential of 945 mV, and 64 mg/L of available chlorine concentration) for 60 s, the bacterial populations were reduced by 5.14 log reduction from the initial load of 10.2 log 10 CFU/mL. Morphological changes in P. deceptionensis CM2 cells were clearly observed through field emission-scanning electron microscopy as a consequence of SAEW treatment. SAEW treatment also resulted in significant increases in the extracellular proteins and nucleic acids, and the fluorescence intensities of propidium iodide and n-phenyl-1-napthylamine in P. deceptionensis CM2 cells, suggesting the disruption of cytoplasmic and outer membrane integrity. These findings show that SAEW is a promising antimicrobial agent.

For combating multidrug-resistant microorganisms, exploration of natural compounds from plant endophytes increases the chance of finding novel compounds. An efficient bioactive metabolites producing endophytic fungal strain AE1 was isolated from leaves of Azadirachta indica A. Juss. The metabolites were found to be thermostable, non-proteinacious and produced prominent zones of inhibition against numbers of Gram positive and Gram negative bacteria. Based on 28S rDNA (D1/D2) sequence homology the isolate AE1 was identified as Alternaria alternata. Malt extract broth was found effective for the maximum production of bioactive metabolites by the isolate and was subjected for solvent extraction. The Ethyl acetate (EA) fraction of AE1 showed MIC values of 300-400 μg/ml against Gram positive and Gram negative bacteria tested. The cidal mode of action of EA fraction was detected by treating bacterial cultures at mid log phase. Scanning electron microscopic study supported morphological disintegration of bacterial cells. Release of nucleic acid, protein and potassium ions (K+) also suggested lysis of bacterial cells or leakage of cell membrane upon treatment. In addition, reduction of the activity of EMP pathway, TCA cycle and gluconeogenic enzymes in all bacteria suggested the interference of antibacterial principles with central carbohydrate metabolic pathways. Thin layer chromatographic separation followed by GC-MS analysis of EA fraction suggested numbers of antimicrobial compound production by AE1. In addition, DPPH free radical as well as superoxide radical scavenging assay also suggested strong antioxidant potential of AE1 with an IC50 value of 38.0±1.7 μg/ml and 11.38±1.2 μg/ml respectively. On the basis of above facts it can be concluded that the strain AE1 will be a good source of bioactive compounds having medicinal importance., Competing Interests: The authors have declared that no competing interests exist.

Three-dimensional microscopy is increasingly prevalent in biology due to the development of techniques such as multiphoton, spinning disk confocal, and light sheet fluorescence microscopies. These methods enable unprecedented studies of life at the microscale, but bring with them larger and more complex datasets. New image processing techniques are therefore called for to analyze the resulting images in an accurate and efficient manner. Convolutional neural networks are becoming the standard for classification of objects within images due to their accuracy and generalizability compared to traditional techniques. Their application to data derived from 3D imaging, however, is relatively new and has mostly been in areas of magnetic resonance imaging and computer tomography. It remains unclear, for images of discrete cells in variable backgrounds as are commonly encountered in fluorescence microscopy, whether convolutional neural networks provide sufficient performance to warrant their adoption, especially given the challenges of human comprehension of their classification criteria and their requirements of large training datasets. We therefore applied a 3D convolutional neural network to distinguish bacteria and non-bacterial objects in 3D light sheet fluorescence microscopy images of larval zebrafish intestines. We find that the neural network is as accurate as human experts, outperforms random forest and support vector machine classifiers, and generalizes well to a different bacterial species through the use of transfer learning. We also discuss network design considerations, and describe the dependence of accuracy on dataset size and data augmentation. We provide source code, labeled data, and descriptions of our analysis pipeline to facilitate adoption of convolutional neural network analysis for three-dimensional microscopy data., Competing Interests: The authors have declared that no competing interests exist.

We identified that Pseudomonas guguanensis produced macromolecular mono-rhamnolipid (1264.52 Da) upon sensing n-hexadecane/diesel/kerosene from its surroundings. Permutation experiments were done to improve the laboratory-scale mono-rhamnolipid production (ie, a three-fold increase) using RSM validation. Consequently, maximal mono-rhamnolipids production [40-50 mg/L] and emulsification abilities [65-70%] were encountered on day 8 using vegetable oil, peptone + yeast extract. EI 24 values for the rhamnolipids were found to be 78±1.75% at 12.5 mg/ mL. Production and secretion of rhamnolipids were accompanied by aggregation of cells at day 6 as pictured in SEM. Pure monorhamnolipids of P. guguanensis was found to lower the surface tension of water to 32.98±0.3 mN/m than the crude and CFSs of P. aeruginosa indicating efficient activity. Utilization and subsequent removal of hexadecane was 77.2% and the breakdown products were fatty acids [decanoic, hexadecanoic, octadecanoic acids and methyl stearates] as signified in Head-space GC-MS. The breakdown products of hexadecane are also present in the synthesized rhamnolipids suggesting their biosynthetic role. Rapid degradation of hexadecane, diesel and kerosene by this emulsifier combined with non-pathogenic trait of P. guguanensis identifies this organism as a viable option to remove n-alkanes from aquatic environments., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Three bacterial strains capable of degrading phthalates namely Pseudomonas sp. PKDM2, Pseudomonas sp. PKDE1 and Pseudomonas sp. PKDE2 were isolated and characterized for their degradative potential. These strains efficiently degraded 77.4%-84.4% of DMP, 75.0%-75.7% of DEP and 71.7%-74.7% of DEHP, initial amount of each phthalate is 500mgL -1 of each phthalate, after 44h of incubation. GC-MS results reveal the tentative DEHP degradation pathway, where hydrolases mediate the breakdown of DEHP to phthalic acid (PA) via an intermediate MEHP. MEHP hydrolase is a serine hydrolase which is involved in the reduction of the MEHP to PA. The predicted 3D model of MEHP hydrolase from Pseudomonas mosselii was docked with phthalate monoesters (PMEs) such as MEHP, mono-n-hexyl phthalate (MHP), mono-n-butyl phthalate (MBP) and mono-n-ethyl phthalate (MEP), respectively. Docking results show the distance between the carbonyl carbon of respective phthalate monoester and the hydroxyl group of catalytic serine lies in the range of 2.9 to 3.3Å, which is similar to the ES complex of other serine hydrolases. This structural study highlights the interaction and the role of catalytic residues of MEHP hydrolase involved in the biodegradation of PMEs to phthalate., (Copyright © 2017 Elsevier B.V. All rights reserved.)

High quality spectra of Pseudomonas sp. strain ADP in the planktonic and biofilm state were obtained using Raman microspectroscopy. These spectra enabled the identification of key differences between free and biofilm cells in the fingerprint region of Raman spectra in the nucleic acid, carbohydrate, and protein regions. Scanning electron microscopy (SEM) enabled detailed visualization of ADP biofilm with confirmation of associated extracellular matrix structure. Following extraction and Raman analysis of extracellular polymeric substances, Raman spectral differences between free and biofilm cells were largely attributed to the contribution of extracellular matrix components produced in mature biofilms. Raman spectroscopy complemented with SEM proves to be useful in distinguishing physiological properties among cells of the same species. Graphical Abstract Raman spectroscopy complemented with SEM proves to be useful in distinguishing physiological properties among cells of the same species.

Two bacterial strains, 46-1 and 46-2 T , were isolated from garden soil. These strains were observed to be aerobic, Gram-stain negative, rod-shaped, non-spore-forming, motile and catalase and oxidase positive. Phylogenetic analysis based on 16S rRNA gene sequences showed that the two strains shared 100 % sequence similarity with each other and belong to the genus Pseudomonas in the class Gammaproteobacteria. The concatenated 16S rRNA, gyrB, rpoB and rpoD gene sequences further confirmed that the isolates belong to the Pseudomonas koreensis subgroup (SG), with P. koreensis Ps 9-14 T , Pseudomonas moraviensis 1B4 T and Pseudomonas granadensis F-278,770 T as their close relatives (>96 % pairwise similarity). DNA-DNA hybridization with the closely related type strain P. koreensis SG revealed a low level of relatedness (<50 %). A cladogram constructed using whole-cell matrix-assisted laser desorption/ionization time-of-flight (WC-MALDI-TOF) MS analysis showed the isolates formed a completely separate monophyletic group. The isolates were negative for utilization of glycogen, D-psicose, α-keto butyric acid, α-keto valeric acid, succinamic acid and D, L-α-glycerol phosphate. In contrast, all these reactions were positive in P. koreensis JCM 14769 T and P. moraviensis DSM 16007 T . The fatty acid C 17:0 cyclo was detected as one of the major cellular fatty acids (>15 %) in the isolates but it was a minor component (<4 %) in both reference type strains. In contrast, the fatty acid, C 12:0 was not observed in the isolates but was present in both reference strains. Based on differences such as phylogenetic position, low-level DNA-DNA hybridization, WC-MALDI-TOF MS analysis, fluorescence pigmentation, fatty acid profiles, and substrate utilization, we propose that the isolates 46-1 and 46-2 T represent a novel species of the genus Pseudomonas, for which the name Pseudomonas kribbensis sp. nov. is proposed; the type strain is 46-2 T (=KCTC 32541 T  = DSM 100278 T ).

Recently, it has been shown that procyanidins from Fallopia spp. inhibit bacterial denitrification, a phenomenon called biological denitrification inhibition (BDI). However, the mechanisms involved in such a process remain unknown. Here, we investigate the mechanisms of BDI involving procyanidins, using the model strain Pseudomonas brassicacearum NFM 421. The aerobic and anaerobic (denitrification) respiration, cell permeability and cell viability of P. brassicacearum were determined as a function of procyanidin concentration. The effect of procyanidins on the bacterial membrane was observed using transmission electronic microscopy. Bacterial growth, denitrification, NO3- and NO2-reductase activity, and the expression of subunits of NO3- (encoded by the gene narG) and NO2-reductase (encoded by the gene nirS) under NO3 or NO2 were measured with and without procyanidins. Procyanidins inhibited the denitrification process without affecting aerobic respiration at low concentrations. Procyanidins also disturbed cell membranes without affecting cell viability. They specifically inhibited NO3- but not NO2-reductase.Pseudomonas brassicacearum responded to procyanidins by over-expression of the membrane-bound NO3-reductase subunit (encoded by the gene narG). Our results suggest that procyanidins can specifically inhibit membrane-bound NO3-reductase inducing enzymatic conformational changes through membrane disturbance and that P. brassicacearum responds by over-expressing membrane-bound NO3-reductase. Our results lead the way to a better understanding of BDI., (© FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

ZJ0273 (propyl 4-(2-(4, 6-demethoxy pyrimidin-2-yloxy) benzylamino) benzoate) is a novel pyrimidynyloxybenzoic-based herbicide developed in China for oilseed crop. This study was aimed to construct new strains capable of degrading naphthalene and ZJ0273 by protoplast fusion between Amycolatopsis sp. M3-1 and Pseudomonas sp. Nai8. Eight recombinant strains were successfully produced, and the strains could simultaneously utilize ZJ0273 and naphthalene as the sole carbon and energy source, respectively. One of recombinant strains, MN6 with higher degrading efficiency, was chosen for further study. Under the condition of pH 7.0, 30 °C, ZJ0273 and naphthalene degradation percent by the recombinant strain MN6 could reach 65.10% (20 days) and 88.46% (48 h), respectively. According to the identified six metabolites (M1-M6) by LC-MS/MS, biodegradation pathway of ZJ0273 was proposed. ZJ0273 biodegradation catalyzed by the recombinant strain MN6 involved continuous biocatalytic reactions such as de-estering, hydrolysis, acylation, C-N cleavage, de-methyl, and ether cleavage reactions.

A bacterium strain, designated as CMF-2, was isolated from the jellyfish Cyanea capillata and its culture supernatant exhibited a significant antimicrobial activity. The strain CMF-2 was identified as Pseudomonas sp. based on the morphological, biochemical and physiological characteristics as well as 16S rRNA sequence analysis. In this study, an antimicrobial protein, named as CAP-1, was isolated from the culture of CMF-2 through ammonium sulfate precipitation and gel filtration chromatography. According to the result of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), a major band indicated that the antimicrobial protein had a molecular mass of about 15 kDa, and it was identified as a hypothetical protein by MALDI-TOF-MS analysis and Mascot searching. CAP-1 displayed a broad antimicrobial spectrum against the indicator bacteria and fungus, including Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Candida albicans, especially some marine-derived microorganisms such as Vibrio vulnificus, Vibrio alginolyticus, Vibrio parahaemolyticus, Vibrio cholera, and Vibrio anguillarum, but showed little impact on tumor cells and normal human cells. The protein CAP-1 remained a stable antimicrobial activity in a wide range of temperature (20-80°C) and pH (2-10) conditions. These results suggested that CAP-1 might have a specific antimicrobial function not due to cytotoxicity., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Microbial communities are ubiquitous in both natural and artificial environments. However, microbial diversity is usually reduced under strong selection pressures, such as those present in habitats rich in recalcitrant or toxic compounds displaying antimicrobial properties. Caffeine is a natural alkaloid present in coffee, tea and soft drinks with well-known antibacterial properties. Here we present the first systematic analysis of coffee machine-associated bacteria. We sampled the coffee waste reservoir of ten different Nespresso machines and conducted a dynamic monitoring of the colonization process in a new machine. Our results reveal the existence of a varied bacterial community in all the machines sampled, and a rapid colonisation process of the coffee leach. The community developed from a pioneering pool of enterobacteria and other opportunistic taxa to a mature but still highly variable microbiome rich in coffee-adapted bacteria. The bacterial communities described here, for the first time, are potential drivers of biotechnologically relevant processes including decaffeination and bioremediation.

Background: Pseudomonas tolaasii is a problematic pathogen of cultured mushrooms, forming dark brown 'blotches' on mushroom surfaces and causing spoilage during crop growth and post-harvest . Treating P. tolaasii infection is difficult, as other, commensal bacterial species such as Pseudomonas putida are necessary for mushroom growth, so treatments must be relatively specific., Results: We have found that P. tolaasii is susceptible to predation in vitro by the δ-proteobacterium Bdellovibrio bacteriovorus. This effect also occurred in funga, where B. bacteriovorus was administered to post-harvest mushroom caps before and after administration of the P. tolaasii pathogen. A significant, visible improvement in blotch appearance, after incubation, was observed on administration of Bdellovibrio. A significant reduction in viable P. tolaasii cell numbers, recovered from the mushroom tissue, was detected. This was accompanied by a more marked reduction in blotch severity on Bdellovibrio administration. We found that there was in some cases an accompanying overgrowth of presumed-commensal, non-Pseudomonas bacteria on post-harvest mushroom caps after Bdellovibrio-treatment. These bacteria were identified (by 16SrRNA gene sequencing) as Enterobacter species, which were seemingly resistant to predation. We visualised predatory interactions occuring between B. bacteriovorus and P. tolaasii on the post-harvest mushroom cap surface by Scanning Electron Microscopy, seeing predatory invasion of P. tolaasii by B. bacteriovorus in funga. This anti-P. tolaasii effect worked well in post-harvest supermarket mushrooms, thus Bdellovibrio was not affected by any pre-treatment of mushrooms for commercial/consumer purposes., Conclusions: The soil-dwelling B. bacteriovorus HD100 preys upon and kills P. tolaasii, on mushroom surfaces, and could therefore be applied to prevent spoilage in post-harvest situations where mushrooms are stored and packaged for sale.

An enzootic disease characterized by granulomas in internal organs occurred in cage-farmed large yellow croaker, Larimichthys crocea (Richardson), in April and November 2010, in Ningbo, Zhejiang Province. One bacterial strain, named XSDHY-P, was isolated from the diseased fish and identified by biochemical characterization, fatty acid methyl ester (FAME) analysis and multilocus sequence analysis (MLSA). According to the results obtained from the biochemical tests, FAME analysis and phylogenetic analysis derived from 16S ribosomal RNA, gyrB, oprF, oprI, oprL and rpoD gene sequencing, the bacterial isolate, XSDHY-P, was identified as Pseudomonas plecoglossicida. Moreover, lethal dose, 50% trials were carried out to demonstrate the virulence of XSDHY-P in large yellow croaker when administered at 2.13 9 105 colony-forming units per fish. Visceral granulomas were found in the experimentally infected fish as well as in the naturally infected fish, indicating that P. plecoglossicida is another bacterial pathogen that causes granulomatosis in L. crocea.

The detachment of single cells from biofilms is an intrinsic part of this surface-associated mode of bacterial existence. Pseudomonas sp. strain CT07gfp biofilms, cultivated in microfluidic channels under continuous flow conditions, were subjected to a range of liquid shear stresses (9.42 mPa to 320 mPa). The number of detached planktonic cells was quantified from the effluent at 24-h intervals, while average biofilm thickness and biofilm surface area were determined by confocal laser scanning microscopy and image analysis. Biofilm accumulation proceeded at the highest applied shear stress, while similar rates of planktonic cell detachment was maintained for biofilms of the same age subjected to the range of average shear rates. The conventional view of liquid-mediated shear leading to the passive erosion of single cells from the biofilm surface, disregards the active contribution of attached cell metabolism and growth to the observed detachment rates. As a complement to the conventional conceptual biofilm models, the existence of a biofilm surface-associated zone of planktonic cell proliferation is proposed to highlight the need to expand the traditional perception of biofilms as promoting microbial survival, to include the potential of biofilms to contribute to microbial proliferation.

Pseudomonas extremaustralis is a versatile Antarctic bacterium, able to grow under microaerobic and anaerobic conditions and is related to several non-pathogenic Pseudomonads. Here we report on the role of the global anaerobic regulator Anr, in the early steps of P. extremaustralis biofilm development. We found that the anr mutant was reduced in its ability to attach, to form aggregates and to display twitching motility but presented higher swimming motility than the wild type. In addition, microscopy revealed that the wild type biofilm contained more biomass and was thicker, but were less rough than that of the anr mutant. In silico analysis of the P. extremaustralis genome for Anr-like binding sites led to the identification of two biofilm-related genes as potential targets of this regulator. When measured using Quantitative Real Time PCR, we found that the anr mutant expressed lower levels of pilG, which encodes a component of Type IV pili and has been previously implicated in cellular adhesion. Levels of morA, involved in signal transduction and flagella development, were also lower in the mutant. Our data suggest that under low oxygen conditions, such as those encountered in biofilms, Anr differentially regulates aggregation and motility thus affecting the first stages of biofilm formation.

In recent years, improvements in transmission electron microscopy (TEM) techniques and the use of tomography have provided a more accurate view of the complexity of the ultrastructure of prokaryotic cells. Cryoimmobilization of specimens by rapid cooling followed by freeze substitution (FS) and sectioning, freeze fracture (FF) and observation of replica, or cryoelectron microscopy of vitreous sections (CEMOVIS) now allow visualization of biological samples close to their native state, enabling us to refine our knowledge of already known bacterial structures and to discover new ones. Application of these techniques to the new Antarctic cold-adapted bacterium Pseudomonasdeceptionensis M1(T) has demonstrated the existence of a previously undescribed cytoplasmic structure that does not correspond to known bacterial inclusion bodies or membranous formations. This structure, which we term a "stack", was mainly visualized in slow growing cultures of P. deceptionensis M1(T) and can be described as a set of stacked membranous discs usually arranged perpendicularly to the cell membrane, but not continuous with it, and found in variable number in different locations within the cell. Regardless of their position, stacks were mostly observed very close to DNA fibers. Stacks are not exclusive to P. deceptionensis M1(T) and were also visualized in slow-growing cultures of other bacteria. This new structure deserves further study using cryoelectron tomography to refine its configuration and to establish whether its function could be related to chromosome dynamics.

An iopromide (IOPr)-degrading bacterium was isolated from activated sludge of a wastewater treatment plant in Shanghai. Based on its morphology, physiological-biochemical characteristics and a phylogenetic analysis of its 16S rRNA sequence, the bacterium was identified and named as Pseudomonas sp. I-24. The optimum condition for degrading IOPr was at 30°C and pH 7.0. After 5 days, strain I-24 could degrade 30 mg/L IOPr by 99% in a basal salts medium with a 5% (V/V) inoculum and 200 mg/L starch as the primary substrate. When applied to an Anaerobic-Anoxic/Aerobic (A2/O) process, with the coexistence of other bacteria, the strain I-24 got lower (61.3%) IOPr removal, but in two A2/O systems (with and without I-24 inoculation), the CODcr removal were both approximately 95%. The trial dosed with strain I-24 showed better IOPr removal than the un-dosed one. I-24 sustained its abundance in the A2/O system during the experiment., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Variability of properties and antibiotic activity, as well as cells survival of Pseudomonas batumici 17/20--the producer of batumin (antistaphylococcal antibiotic) after long-term storage under vaseline oil layer have been studied. The main culture-morphological and physiological biochemical properties of the mutant strain have been investigated. It has been shown that storage under vaseline oil allows to preserve high level of antibiotic activity: batumin synthesis by the producer was 150 mg/l. Therewith, the survival of cells decreases by two orders during 5 years of storage. The conditions of strain maintenance have been formulated.

We report on the biodegradation of pure hydrocarbons and chemotaxis towards these compounds by an isolated chlorophenol degrader, Pseudomonas strain H. The biochemical and phylogenetic analysis of the 16S rDNA sequence identified Pseudomonas strain H as having 99.56% similarity with P. aeruginosa PA01. This strain was able to degrade n-hexadecane, 1-undecene, 1-nonene, 1-decene, 1-dodecene and kerosene. It grew in the presence of 1-octene, while this hydrocarbons is toxic to other hydrocarbons degraders. Pseudomonas strain H was also chemotactic towards n-hexadecane, kerosene, 1-undecene and 1-dodecene. These results show that this Pseudomonas strain H is an attractive candidate for hydrocarbon-containing wastewater bioremediation in controlled environments. Since the classical standard techniques for detecting chemotaxis are not efficient at low bacterial concentrations, we demonstrate the use of the dynamic speckle laser method, which is simple and inexpensive, to confirm bacterial chemotaxis at low cell concentrations (less than 10(5) colony-forming unit per millilitre (CFU/mL)) when hydrocarbons are the attractants.

Pseudomonas phage ϕKZ and its two close relatives ϕPA3 and 201ϕ2-1 are very large bacteriophages that form a separate branch in phage classification because their genomes are very different from the rest of GenBank sequence data. The contractile tail of ϕKZ is built from at least 32 different proteins, but a definitive structural function is assigned to only one of them-the tail sheath protein. Here, we report the crystal structure of the C-terminal domain of another phiKZ tail protein, gene product 131 (gp131C). We show that gp131 is located at the periphery of the baseplate and possibly associates with fibers that emanate from the baseplate. Gp131C is a seven-bladed β-propeller that has a shape of a skewed toroid. A small but highly conserved and negatively charged patch on the surface of gp131C might be important for substrate binding or for interaction with a different tail protein., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Electrospun hollow polymeric microfibers (microtubes) were evaluated as an encapsulation method for the atrazine degrading bacterium Pseudomonas sp. ADP. Pseudomonas sp. ADP cells were successfully incorporated in a formulation containing a core solution of polyethylene oxide dissolved in water and spun with an outer shell solution made of polycaprolactone and polyethylene glycol dissolved in a chloroform and dimethylformamide. The resulting microtubes, collected as mats, were partially collapsed with a ribbon-like structure. Following encapsulation, the atrazine degradation rate was low (0.03 ± 0.01 mg atrazine/h/g fiber) indicating that the electrospinning process negatively affected cell activity. Atrazine degradation was restored to 0.5 ± 0.1 mg atrazine/h/g fiber by subjecting the microtubes to a period of growth. After 3 and 7 days growth periods, encapsulated cells were able to remove 20.6 ± 3 and 47.6 ± 5.9 mg atrazine/g mat, respectively, in successive batches under non-growth conditions (with no additional electron donor) until atrazine was detected in the medium. The loss of atrazine degrading capacity was regained following an additional cell-growth period.

Seventy-four unrelated clinical isolates of Streptococcus pneumoniae harboring the tet(M) gene were studied. Seven strains with low tetracycline (Tc) MICs (0.25 to 0.5 μg/ml) were found to harbor truncated tet(M) alleles that were inactivated by different frameshift mutations. In contrast, five strains bore deletions in the tet(M) promoter region, among which four displayed increased Tc MICs (16 to 64 μg/ml). The same promoter mutations were detected in Tc-resistant mutants selected in vitro from various susceptible strains. Sequence analysis revealed that these deletions might impede the formation of the transcriptional attenuator located immediately upstream of tet(M). Expression in Enterococcus faecalis of a tet(M) reporter gene transcribed from these promoter mutants conferred a level of Tc resistance similar to that observed in the parental S. pneumoniae strains. These results show that different levels of Tc susceptibility found in clinical isolates of S. pneumoniae can be explained by frameshift mutations within tet(M) and by alterations of the upstream transcriptional attenuator.

Although microorganisms have the potential to reduce metals, products with elementary forms are unusual. In the present study, a strain of Pseudomonas sp. MBR was tested for its ability to reduce metal ions to their elementary forms coupled to biomineralization under aerobic conditions. The Pseudomonas sp. MBR strain was able to reduce metals such as Fe(III), Mn(II), Cu(II), Ni(II), Cd(II), Co(II), Al(III), Se(IV), and Te(IV) as electron acceptors to elementary forms using citrate, lactate, pyruvate, succinate, malate, glucose, or ethanol as electron donors. Growth and reduction during biomineralization occurred within the pH range of 6.0 to 11.0 and temperature range of 4 to 40 °C, with an optimum growth temperature of 28 °C. The resistance of Ni(II) varied from 0.5 to 5 mM. Ni(II) reduction was still observed when nitrate was present in addition to oxygen as a potential electron acceptor. The Ni(II) reduction efficiency was related with the molar ratio of the electron donor to Ni(II). Unlike other dissimilatory metal-reducing bacteria, which oxidizes organic matter with Fe(III) or Mn(IV) as the sole electron acceptor coupled to energy production under facultative anaerobic conditions, this strain used oxygen as an electron acceptor combined with metal reduction. The aerobic metal reduction may relate to a co-metabolic reduction. Transmission electron microscopy images demonstrated that the cells had the ability to accumulate heavy metals, and that the detoxicity mechanism was intracellular metal reduction. These results suggested that the use of Pseudomonas sp. MBR could be promising for toxic heavy metal bioremediation and biological metallurgy.

A Gram-negative, non-mobile, polar single flagellum, rod-shaped bacterium WZBFD3-5A2(T) was isolated from a wheat soil subjected to herbicides for several years. Cells of strain WZBFD3-5A2(T) grow optimally on Luria-Bertani agar medium at 30 °C in the presence of 0-4.0 % (w/v) NaCl and pH 8.0. 16S rRNA gene sequence analysis revealed that strain WZBFD3-5A2(T) belongs to the genus Pseudomonas. Physiological and biochemical tests supported the phylogenetic affiliation. Strain WZBFD3-5A2(T) is closely related to Pseudomonas nitroreducens IAM1439(T), sharing 99.7 % sequence similarity. DNA-DNA hybridization experiments between the two strains showed only moderate reassociation similarity (33.92 ± 1.0 %). The DNA G+C content is 62.0 mol%. The predominant respiratory quinine is Q-9. The major cellular fatty acids present are C(16:0) (28.55 %), C(16:1ω6c) or C(16:1ω7c) (20.94 %), C(18:1ω7c) (17.21 %) and C(18:0) (13.73 %). The isolate is distinguishable from other related members of the genus Pseudomonas on the basis of phenotypic and biochemical characteristics. From the genotypic, chemotaxonomic and phenotypic data, it is evident that strain WZBFD3-5A2(T) represents a novel species of the genus Pseudomonas, for which the name Pseudomonas nitritereducens sp. nov. is proposed. The type strain is WZBFD3-5A2(T) (=CGMCC 1.10702(T) = LMG 25966(T)).

The herbicide propanil has long been used in rice production in southern Brazil. Bacteria isolated from contaminated soils in Massaranduba, Santa Catarina, Brazil, were found to be able to grow in the presence of propanil, using this compound as a carbon source. Thirty strains were identified as Pseudomonas (86.7%), Serratia (10.0%), and Acinetobacter (3.3%), based on phylogenetic analysis of 16S rDNA. Little genetic diversity was found within species, more than 95% homology, suggesting that there is selective pressure to metabolize propanil in the microbial community. Two strains of Pseudomonas (AF7 and AF1) were selected in bioreactor containing chemotactic growth medium, with the highest degradation activity of propanil exhibited by strain AF7, followed by AF1 (60 and 40%, respectively). These strains when encapsulated in alginate exhibited a high survival rate and were able to colonize the rice root surfaces. Inoculation with Pseudomonas strains AF7 and AF1 significantly improved the plant height of rice. Most of the Pseudomonas strains produced indoleacetic acid, soluble mineral phosphate, and fixed nitrogen. These bacterial strains could potentially be used for the bioremediation of propanil-contaminated soils and the promotion of plant growth.

The objective of this study was to study the simultaneous biodegradation of Ni-citrate complexes and removal of Ni from solutions by Pseudomonas alcaliphila. Adding excess citrate to 1:1 Ni-citrate complexes promoted the degradation of the complexes and removal of Ni. The alkaline pH generated by the metabolism of excess citrate caused partial dissociation of citrate from the Ni-citrate complexes, allowing degradation, and the released Ni was removed through bioaccumulation and precipitation. Addition of Fe(3+) enhanced the degradation of Ni-citrate complexes and removal of Ni from solutions. The displacement of Ni from recalcitrant Ni-citrate complexes by Fe(3+) and subsequent biodegradation of the degradable Fe(III)-citrate complex resulted in complete metabolism of citrate. The almost complete removal of Ni (>98%) can be attributed to the combination of coprecipitation with Fe(3+), bioaccumulation and precipitation. P. alcaliphila potentially could be applied in the treatment of effluent containing Ni-citrate complexes., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The ability of Pseudomonas sp. strain DY1 to adsorb Acid Black 172 was studied to determine the kinetics and mechanisms involved in biosorption of the dye. Kinetic data for adsorption fit a pseudo-second-order model. Increased initial dye concentration could significantly enhance the amount of dye adsorbed by heat-treated biomass in which the maximum amount of dye adsorbed was as high as 2.98 mmol/g biomass, whereas it had no significant influence on dye sorption by live biomass. As treated temperature increased, the biomass showed gradual increase of dye sorption ability. Experiments using potentiometric titration and Fourier transform infrared spectroscopy (FTIR) indicated that amine groups (NH2) played a prominent role in biosorption of Acid Black 172. Scanning electron microscopy (SEM), atomic force microscopy (AFM) and transmission electron microscopy (TEM) analysis indicated that heat treatment of the biomass increased the permeability of the cell walls and denatured the intracellular proteins. The results of biosorption experiments by different cell components confirmed that intracellular proteins contributed to the increased biosorption of Acid Black 172 by heat-treated biomass. The data suggest that biomass produced by this strain may have application for removal of metal-complex dyes from wastewater streams generated from the dye products industry., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Removal of detrimental biofilms from surfaces exposed in the marine environment remains a challenge. A strain of Bacillus pumilus was isolated from the surface of titanium coupons immersed in seawater in the vicinity of Madras Atomic Power Station (MAPS) on the East coast of India. The bacterium formed extensive biofilms when compared to species such as Bacillus licheniformis, Pseudomonas aeruginosa PAO1 and Pseudomonas aureofaciens. A commercially available rhamnolipid was assessed for its ability to inhibit adhesion and disrupt pre-formed B. pumilus biofilms. The planktonic growth of B. pumilus cells was inhibited by concentrations >1.6mM. We studied the effect of various concentrations (0.05-100mM) of the rhamnolipid on adhesion of B. pumilus cells to polystyrene microtitre plates, wherein the effectiveness varied from 46 to 99%. Biofilms of B. pumilus were dislodged efficiently at sub-MIC concentrations, suggesting the role of surfactant activity in removing pre-formed biofilms. Scanning electron microscopy (SEM) confirmed the removal of biofilm-matrix components and disruption of biofilms by treatment with the rhamnolipid. The results suggest the possible use of rhamnolipids as efficient anti-adhesive and biofilm-disrupting agents with potential applications in controlling biofilms on surfaces., (Copyright (c) 2010 Elsevier B.V. All rights reserved.)

In this study, we isolated Pseudomonas sp. strain HB01 from a soil sample contaminated with brominated pollutants, and found that it degraded gamma-hexabromocyclododecane (gamma-HBCD). The strain degraded 81% of 1 mM gamma-HBCD within 5 d of culture. Furthermore, it demonstrated biodegradation of structurally related (bromo) alkanes. To the best of our knowledge, this is the first report that outlines the isolation of a bacterial strain that degrades gamma-HBCD.

The endophytic phase of Pseudomonas savastanoi pv. savastanoi in olive stems and the structural and ultrastructural histogenesis of olive knots have been studied. Construction of a stable plasmid vector expressing the green fluorescent protein, in combination with the use of in vitro olive plants, allowed real-time monitoring of P. savastanoi pv. savastanoi infection. The infection process was also examined by bright field and epifluorescence microscopy as well as by scanning and transmission electron microscopy. Hypertrophy of the stem tissue was concomitant with the formation of bacterial aggregates, microcolonies and multilayer biofilms, over the cell surfaces and the interior of plasmolysed cells facing the air-tissue interface of internal opened fissures, and was followed by invasion of the outer layers of the hypertrophied tissue. Pathogenic invasion of the internal lumen of newly formed xylem vessels, which were connected with the stem vascular system, was also observed in late stages of infection. Ultrastructural analysis of knot sections showed the release of outer membrane vesicles from the pathogen surface, a phenomenon not described before for bacterial phytopathogens during host infection. This is the first real-time monitoring of P. savastanoi disease development and the first illustrated description of the ultrastructure of P. savastanoi-induced knots., (© 2009 The Authors. Journal compilation © 2009 Society for Applied Microbiology and Blackwell Publishing Ltd.)

Heavy metal sequestration by a multimetal resistant Pseudomonas strain isolated from a uranium mine was characterized for its potential application in metal bioremediation. 16S rRNA gene analysis revealed phylogenetic relatedness of this isolate to Pseudomonas fluorescens. Metal uptake by this bacterium was monophasic, fast saturating, concentration and pH dependent with maximum loading of 1048 nmol Ni(2+) followed by 845 nmol Co(2+), 828 nmol Cu(2+) and 700 nmol Cd(2+)mg(-1) dry wt. Preferential metal deposition in cell envelope was confirmed by TEM and cell fractionation. FTIR spectroscopy and EDX analysis revealed a major role of carboxyl and phosphoryl groups along with a possible ion exchange mechanism in cation binding. Binary system demonstrated selective metal binding affinity in the order of Cu(2+)>Ni(2+)>Co(2+)>Cd(2+). A comparison with similar metal uptake reports considering live bacteria strongly indicated the superiority of this strain in metal sequestration, which could be useful for developing efficient metal removal system.

The mechanism and chemical nature of uranium and thorium sequestration by a Pseudomonas strain was investigated by transmission electron microscopy, energy dispersive X-ray (EDX) analysis, FTIR spectroscopy and X-ray diffractometry. Atomic force microscopy (AFM) used in the tapping mode elucidated the morphological changes in bacterial cells following uranium and thorium binding. Transmission electron microscopy revealed intracellular sequestration of uranium and thorium throughout the cell cytoplasm with electron dense microprecipitations of accumulated metals. Energy dispersive X-ray analysis confirmed the cellular deposition of uranium and thorium. EDX and elemental analysis of sorption solution indicated the binding of uranium and thorium by the bacterial biomass via displacement of cellular potassium and calcium. The strong involvement of cellular phosphate, carboxyl and amide groups in radionuclide binding was ascertained by FTIR spectroscopy. X-ray powder diffraction (XRD) analyses confirmed cellular sequestration of crystalline uranium and thorium phosphates. Overall results indicate that a combined ion-exchange-complexation-microprecipitation mechanism could be involved in uranium and thorium sequestration by this bacterium. Atomic force microscopy and topography analysis revealed an undamaged cell surface with an increase in cell length, width and height following radionuclide accumulation. The arithmetic average roughness (R(a)) and root mean square (RMS) roughness (R(q)) values indicated an increase in surface roughness following uranium and thorium sequestration.

The effect of inorganic nutrients (sulfate, phosphate, and ammonium chloride) on the aerobic biodegradation of benzene, toluene, and xylene (BTX) by Pseudomonas spp. was studied in the laboratory using a glass sand tank. The increase of nutrient levels resulted in enhanced bacterial growth and BTX degradation. Sulfate and phosphate serve as key electron acceptors in the microbiological processes degrading BTX. The observed bacterial morphological changes during BTX degradation reveal that the filamentous bacteria were the dominant species at low temperatures about 20 degrees C. The spherical and rod-shaped cells became dominant at higher temperatures ranging from 25 degrees C to 28 degrees C. When the BTX mixture was allowed to be biodegraded for longer incubation periods of 21-42 h at high phosphate concentrations, large amounts of rod-shaped cells were clustered. The morphological adaptation appears to be controlled by the temperature and nutrient levels in the sandy medium where Pseudomonas spp. thrives.

Pseudomonas guezennei biovar. tikehau was isolated from a microbial mat on the atoll of Tikehau in French Polynesia, and is able to synthesize medium chain length poly-beta-hydroxyalkanaote copolymers when grown on coprah oil. A two-step cultivation process was used and the biosynthesis of PHAs was followed along 52h by regular culture sampling. The polyester was purified from freeze-dried cells and analysed by nuclear magnetic resonance (NMR), Fourier transform infra red (FTIR), and gas chromatography mass spectrometries. The copolyester produced by P. guezennei biovar. tikehau from coprah oil mainly consisted of saturated monomers, i.e. 3-hydroxyoctanoate (3HO) and 3-hydroxydecanoate (3HD), and the monomeric composition of the polyester did not change during the fermentation process. However, yield of PHAs production varied from 4% of the cellular dry weight (CDW) to 63% obtained after 36h. Scan electron microscopy was used to study the morphology and organization of PHAs granules within the cells and revealed the presence of several granules occupying almost the entire cell volume.

Cystlike resting cells (CRC) of non-spore-forming gram-negative bacteria of the genus Pseudomonas, P. aurantiaca and P. fluorescens, were obtained and characterized for the first time; their physiological and morphological diversity was demonstrated. The following properties were common for all the revealed types of CRC as dormant forms: (1) long-term (up to 6 months or longer) maintenance of viability in the absence of culture growth and cell respiration; (2) absence of an experimentally detectable level of metabolism; (3) higher resistance to damage and autolysis under the action of provoking factors than in metabolically active vegetative cells; and (4) specific features of ultrastructural organization absent in vegetative cells: thickened and lamellar envelopes, clumpy structure of the cytoplasm, and condensed DNA in nucleoid. The differences in various types of CRC concern the thickness and lamellar structure of cell envelopes, as well as the presence and thickness of the capsular layer. In particular, forms ultrastructurally similar to typical bacterial cysts were revealed in pseudomonad populations growing on soil agar. Physiological diversity was revealed in different levels of viability preservation and thermal resistance in various types of CRC and depended on the conditions of their formation. The optimal conditions and procedures for obtaining P. aurantiaca and P. fluorescens CRC that retain the ability to form colonies on standard nutrient media are as follows: (1) a twofold decrease of nitrogen content in the growth medium; (2) an increased level of anabiosis autoinducer (C12-AHB, 10(-4) M) in stationary cultures; (3) transfer of the cells from stationary cultures to a starvation medium with silica; (4) cultivation in soil extract; and (5) development of cultures on soil agar. The CRC from the cultures grown in soil extract or starvation medium with silica proved to be resistant to heat treatment (60 degrees C, 5 min). In the CRC formed in nitrogen-limited media, the degree of heat resistance increased at longer incubation (1.5 to 6 months). CRCs on soil agar surface were resistant to desiccation. The ultrastructure of the morphologically varied types of P. aurantiaca CRC formed under simulated natural conditions is described for the first time. The data on the intraspecies diversity of pseudomonad dormant forms contribute to the concept of plasticity of the life style and adaptive reactions that ensure survival of these bacteria in unfavorable environmental conditions.

Objective: To screen antagonistic bacteria from rhizosphere of tobacco against Phytophthora parasitica var. nicotianae (Breda de Hann) Tucker and to analyze its antagonistic activity., Methods: The antagonistic bacteria were screened by dual culture with P. parasitica var. nicotianae (Breda de Hann) Tucker on low iron (2 micromol/L FeCl3) Sucrose-L-Asparagine (SA) plate. The chrome azurol S (CAS) assay was used to detect the siderophore production and its affinity to ferric ion. The strain was identified by using morphological, biochemical and physiological characteristics, 16S rRNA sequence homology, phylogenetics and specific species molecular analysis. The siderophore was isolated by column chromatography on Amberlite XAD-2 and analyzed by spectrophotometer. Antagonistic activity of the siderophore was confirmed by incubation of P. parasitica var. nicotianae (Breda de Hann) Tucker in liquid SA medium with different concentration of siderophore and FeCl3., Results: We obtained an antagonistic bacterium G-229-21T against P. parasitica var. nicotianae (Breda de Hann) Tucker. The strain produced high-affinity siderophore under low iron conditions and was identified as Pseudomonas mediterranea. This strain produced carboxylate-type siderophore against P. parasitica var. nicotianae (Breda de Hann) Tucker. The suppression ratio was up to 92.3% under low iron conditions (0.16 micromol/L-10 micromol/L FeCl3). However, it was only 2.0% under Fe-replete conditions (100 micromol/L FeCl3)., Conclusion: P. mediterranea G-229-21T could produce high-affinity carboxylate-type siderophore against P. parasitica var. nicotianae (Breda de Hann) Tucker under low iron conditions.

An acclimatized mixed microbial culture, predominantly Pseudomonas sp., was enriched from a sewage treatment plant, and its potential to simultaneously degrade mixtures of phenol and m-cresol was investigated during its growth in batch shake flasks. A 2(2) full factorial design with the two substrates at two different levels and different initial concentration ranges (low and high), was employed to carry out the biodegradation experiments. The substrates phenol and m-cresol were completely utilized within 21 h when present at low concentrations of 100 mg/L for each, and at high concentration of 600 mg/L for each, a maximum time of 187 h was observed for their removal. The biodegradation results also showed that the presence of phenol in low concentration range (100-300 mg/L) did not inhibit m-cresol biodegradation. Whereas the presence of m-cresol inhibited phenol biodegradation by the culture. Moreover, irrespective of the concentrations used, phenol was degraded preferentially and earlier than m-cresol. A sum kinetics model was used to describe the variation in the substrate specific degradation rates, which gave a high coefficient of determination value (R2 > 0.98) at the low concentration range of the substrates. From the estimated interaction parameter values obtained from this model, the inhibitory effect of phenol on m-cresol degradation by the culture was found to be more pronounced compared to that of m-cresol on phenol. This study showed a good potential of the indigenous mixed culture in degrading mixed substrate of phenolics.

GacS and GacA proteins form a two component signal transduction system in bacteria. Here, Tn5 transposon gacS and gacA (Gac) mutants of Pseudomonas sp. KL28, an alkylphenol degrader, were isolated by selecting for smooth colonies of strain KL28. The mutants exhibited reduced ability to migrate on a solid surface. This surface motility does not require the action of flagella unlike the well-studied swarming motility of other Pseudomonas sp. The Gac mutants also showed reduced levels of biofilm and pellicle formation in liquid culture. In addition, compared to the wild type KL28 strain, these mutants were more resistant to high concentrations of m-cresol but were more sensitive to H2O2, which are characteristics that they share with an rpoS mutant. These results indicate that the Gac regulatory cascade in strain KL28 positively controls wrinkling morphology, biofilm formation, surface translocation and H2O2 resistance, which are important traits for its capacity to survive in particular niches.

A soil bacterium SW4, capable of degrading the sulfonylurea herbicide ethametsulfuron-methyl (ESM), was isolated from the bottom soil of a herbicide factory. Based on physiological characteristics, biochemical tests and phylogenetic analysis of the 16S rRNA gene sequence, the strain was identified as a Pseudomonas sp. The total degradation of ESM in the medium containing glucose was up to 84.6% after 6 days of inoculation with SW4 strain. The inoculation of strain SW4 to soil treated with ESM resulted in a higher degradation rate than in noninoculated soil regardless of the soil sterilized or nonsterilized. Five metabolites of ESM degradation were analyzed by liquid chromatography/mass spectrometry. Based on the identified products, strain SW4 seemed to degrade ESM after two separate and different pathways: one leads to the cleavage of the sulfonylurea bridge, whereas the other to the dealkylation and opening of the triazine ring of ESM.

The adhesion of microbial cells to metal surfaces in aqueous media is an important phenomenon in both the natural environment and engineering systems. The adhesion of two anaerobic sulfate-reducing bacteria (Desulfovibrio desulfuricans and a local marine isolate) and an aerobe (Pseudomonas sp.) to four polished metal surfaces (i.e., stainless steel 316, mild steel, aluminum, and copper) was examined using a force spectroscopy technique with an atomic force microscope (AFM). Using a modified bacterial tip, the attraction and repulsion forces (in the nano-Newton range) between the bacterial cell and the metal surface in aqueous media were quantified. Results show that the bacterial adhesion force to aluminum is the highest among the metals investigated, whereas the one to copper is the lowest. The bacterial adhesion forces to metals are influenced by both the electrostatic force and metal surface hydrophobicity. It is also found that the physiological properties of the bacterium, namely the bacterial surface charges and hydrophobicity, also have influence on the bacteria-metal interaction. The adhesion to the metals by Pseudomonas sp. and D. desulfuricans was greater than by the marine SRB isolate. The cell-cell interactions show that there are strong electrostatic repulsion forces between bacterial cells. Cell probe atomic force microscopy has provided some useful insight into the interactions of bacterial cells with the metal surfaces.

A new Pseudomonas strain, designated GP72, was isolated from green pepper rhizosphere and identified as a member of species Pseudomonas chlororaphis based on morphology; conventional biochemical and physiologic tests; Biolog GN system (Biolog Inc., Hayward, CA); and 16S rDNA sequence analysis. The secondary metabolites produced by this strain have shown broad-spectrum antifungal activity against various phytopathogens of agricultural importance in vitro. Two main antifungal substances produced by this strain proved to be phenazine-1-carboxylic acid and 2-hydroxyphenazine with further purification and structure elucidation based on ultraviolet-absorbent spectrum scanning, atmospheric pressure chemical ionization-mass spectrometry (APCI-MS) spectrum, and (1)H,(13)C nuclear magnetic resonance spectrums. Strain GP72 could produce quorum-sensing signaling molecules of N-butanoyl-L-homoserine lactone and N-hexanoyl-L-homoserine lactone, which were found to accumulate with different quantities in King's medium B and pigment producing medium, respectively.

Constructed ammonium oxidizing biofilms (CAOB) and constructed nitrite oxidizing biofilms (CNOB) were characterized during the bioremediation of a wastewater effluent. The maximum ammonium removal rate and removal efficiency in CAOB was 322 mg N-NH4+ m(-3) d(-1) and 96%, respectively, while in CNOB a maximum removal rate of 255 mg N-NH4+ m(-3) d(-1) and a removal efficiency of 76% was achieved. Both constructed biofilms on low-density polyester Dacron support achieved removal efficiencies higher than that of the concentrations normally present in reactors without constructed biofilms (P < 0.05). Nitrifying bacteria from the constructed biofilms cultures were typed by sequencing 16S rRNA genes that had been amplified by PCR from genomic DNA. Analysis of enrichment biofilms has therefore provided evidence of high removal of ammonium and the presence of Nitrosomonas eutropha, N. halophila and N. europaea in CAOB, while in CNOB Nitrobacter hamburgensis, N. winogradskyi and N. alkalicus were identified according to 16S rRNA gene sequences comparison. The biofilm reactors were nitrifying over the whole experimental period (15 days), showing a definite advantage of constructed biofilms for enhancing a high biomass concentration as evidenced by environmental electron microscopic analysis (ESEM). Our research demonstrates that low-density polyester Dacron can be effectively used for the construction of nitrifying biofilms obtaining high removal efficiencies of nitrogen in a relatively short time from municipal effluents from wastewater treatment plants. CAOB and CNOB are potentially promissory for the treatment of industrial wastewaters that otherwise requires very large and expensive reactors for efficient bioremediation of effluents.

Pseudomonas viridiflava is a common pathogen of Arabidopsis thaliana in wild populations, yet very little is known about mechanisms of resistance and virulence in this interaction. We examined the induced defense response of A. thaliana to several strains of P. viridiflava collected from this host by quantifying the expression of PR-1 and LOX2/PDF1.2, which serve as markers for induction of the salicylic and jasmonic acid (JA) pathways, respectively. Growth of these strains then was assessed on Col-0, the fad3/7/8 and coil-1 mutants deficient in JA- and ethylene (ET)-induced defense responses, and the sid2-1 mutant deficient in salicylic acid-induced defense responses. All strains of P. viridiflava induced high expression of LOX2 and PDF1.2 on Col-0. In contrast, PR-1 expression was delayed and reduced relative to PDF1.2 expression. Additionally, three of four P. viridiflava strains were more virulent on fad3/7/8 relative to Col-0, whereas all strains were more virulent on coil-1 relative to Col-0, indicating that P. viridiflava generally may be suppressed by JA/ET-mediated defense responses. In contrast, no increase in the growth of P. viridiflava strains was observed in the sid2-1 mutant relative to Col-0. Parallel experiments were performed with the closely related P. syringae pv. tomato for comparative purposes. In addition, we assessed the role of pectate lyase and the alternative sigma factor HrpL in P. viridiflava virulence on A. thaliana and found that pectate lyase activity is correlated with virulence, whereas the removal of pectate lyase or HrpL significantly reduced virulence.

A horizontal-flow anaerobic immobilized biomass reactor (HAIB) containing denitrifying biomass was evaluated with respect to its ability to remove, separately and in a short operating period (30 days), organic matter, nitrate, and the hydrocarbons benzene (41.4 mg L-1), toluene (27.8 mg L-1), ethylbenzene (31.1 mg L-1), o-xylene (28.5 mg L-1), m-xylene (28.4 mg L-1) and p-xylene (32.1 mg L-1). The purified culture, which was grown in the presence of the specific hydrocarbon, was used as the source of cells to be immobilized in the polyurethane foam. After 30 days of operation, the foam was removed and a new immobilized biomass was grown in the presence of another hydrocarbon. The average hydrocarbon removal efficiency attained was 97%. The organic matter, especially ethanol, was removed with an average efficiency of 83% at a mean influent concentration of 1185.0 mg L-1. A concomitant removal of 97% of nitrate was observed for a mean influent concentration of 423.4 mg L-1. The independent removal of each hydrocarbon demonstrated that these contaminants can be biodegraded separately, without the need for a compound to be the primary substrate for the degradation of another. This study proposes the application of the system for treatment of areas contaminated with these compounds, with substitution and formation of a biofilm in a 30-day period.

The flocculating activity of a novel bioflocculant MMF1 produced by multiple-microorganism consortia MM1 was investigated. MM1 was composed of strain BAFRT4 identified as Staphylococcus sp. and strain CYGS1 identified as Pseudomonas sp. The flocculating activity of MMF1 isolated from the screening medium was 82.9%, which is remarkably higher than that of the bioflocculant produced by either of the strains under the same condition. Brewery wastewater was also used as the carbon source for MM1, and the cost-effective production medium for MM1 mainly comprised 1.0 L brewery water (chemical oxygen demand (COD) 5000 mg/L), 0.5 g/L urea, 0.5 g/L yeast extract, and 0.2 g/L (NH4)2SO4. The optimal conditions for the production of MMF1 was inoculum size 2%, initial pH 6.0, cultivating temperature 30 degrees C, and shaking speed 160 r/min, under which the flocculating activity of the MMF1 reached 96.8%. Fifteen grams of purified bioflocculant could be recovered from 1.0 L of fermentation broth. MMF1 was identified as a macromolecular substance containing both protein and polysaccharide. It showed good flocculating performance in treating indigotin printing and dyeing wastewater, and the maximal removal efficiencies of COD and chroma were 79.2% and 86.5%, respectively.

Truncation and acylation were combined to investigate the broad-spectrum bactericidal and hemolytic peptide S4(1-15). Substitution of up to seven residues with dodecanoic acid (C(12)) gradually led to specific antipseudomonal activity: out of 40 bacterial strains tested in vitro, C(12)-S4(8-15) displayed similar minimal inhibitory concentrations (MICs) as S4(1-15) against Pseudomonas aeruginosa sp. (identical MIC(90)) but was practically inactive against most other bacteria or erythrocytes. Surface plasmon resonance and isothermal titration calorimetry experiments revealed the binding properties of S4(1-15) to be consistent with its nonselective activities, while discriminative activities of C(12)-S4(8-15) correlated with high binding affinity to a membrane containing pseudomonal lipopolysaccharides and with lower affinities to membranes containing nonpseudomonal lipopolysaccharides or cholesterol. Various mechanistic studies failed to detect significant differences in secondary structure, bactericidal kinetics, or ability to perturb the cytoplasmic membrane, pointing to a similar mode of action.

The susceptibility of planktonic and biofilm cells of Pseudomonas marginalis toward four commonly used biocides at different temperatures (15 and 30 degrees C) and biofilm growth times (24 and 48 h) was assessed. Using the MBEC biofilm device, biofilm production in maple sap was shown to be highly reproducible for each set of conditions tested. Biofilm formation was influenced by growth temperature and time. A temperature of 15 degrees C and incubation time of 24 h yielded fewer CFU per peg and showed fewer adhered cells and typical biofilm structures, based on scanning electron microscopy observations as compared with other conditions. Minimal biofilm eradication concentration values for P. marginalis were significantly greater (P. < 0.001) than were MBCs for planktonic cells and for every biocide tested, with the exception of minimal biofilm eradication concentration values for peracetic acid at 15 degrees C and 24 h. Sodium hypochlorite and peracetic acid sanitizers were able to eliminate P. marginalis biofilms at lower concentrations as compared with hydrogen peroxide- and quaternary ammonium-based sanitizers (P < 0.001). According to the results obtained, sodium hypochlorite and peracetic acid sanitizers would be more appropriate for maple sap collection system sanitation.

Lanthanum biosorption by a Pseudomonas sp. was characterized in terms of equilibrium metal loading, model fitting, kinetics, effect of solution pH, lanthanum-bacteria interaction mechanism and recovery of sorbed metal. Lanthanum sorption by the bacterium was rapid and optimum at pH 5.0 with equilibrium metal loading as high as 950 mg g(-1) biomass dry wt. Scatchard model and potentiometric titration suggested the presence of at least two types of metal-binding sites, corresponding to a strong and a weak binding affinity. The chemical nature of metal-microbe interaction has been elucidated employing FTIR spectroscopy, energy dispersive X-ray analysis (EDX) and X-ray diffraction analysis (XRD). FTIR spectroscopy and XRD analysis revealed strong involvement of cellular carboxyl and phosphate groups in lanthanum binding by the bacterial biomass. EDX and the elemental analysis of the sorption solution ascertained the binding of lanthanum with the bacterial biomass via displacement of cellular potassium and calcium. Transmission electron microscopy exhibited La accumulation throughout the bacterial cell with some granular deposits in cell periphery and in cytoplasm. XRD confirmed the presence of LaPO4 crystals onto the bacterial biomass after La accumulation for a long period. A combined ion-exchange-complexation-microprecipitation mechanism could be involved in lanthanum accumulation by the biomass. Almost 98% of biomass-bound La could be recovered using CaCO3 as the desorbing agent.