Cultures Of Pseudomonas Research Articles

Elucidating the interactions and phytotoxicity of zinc oxide nanoparticles with agriculturally beneficial bacteria and selected crop plants.

There is a growing interest in the use of bioinoculants to assist mineral fertilizers in improving crop production and yield. Azotobacter and Pseudomonas are two agriculturally relevant strains of bacteria which have been established as efficient bioinoculants. An experiment involving addition of graded concentrations of zinc oxide (ZnO) nanoparticles was undertaken using log phase cultures of Azotobacter and Pseudomonas. Growth kinetics revealed a clear trend of gradual decrease with Pseudomonas; however, Azotobacter exhibited a twofold enhancement in growth with increase in the concentration of ZnO concentration. Scanning electron microscopy (SEM), supported by energy-dispersive X-ray (EDX) analyses, illustrated the significant effect of ZnO nanoparticles on Azotobacter by the enhancement in the abundance of globular biofilm-like structures and the intracellular presence of ZnO, with the increase in its concentration. It can be surmised that extracellular mucilage production in Azotobacter may be providing a barrier to the nanoparticles. Further experiments with Azotobacter by inoculation of wheat and tomato seeds with ZnO nanoparticles alone or bacteria grown on ZnO-infused growth medium revealed interesting results. Vigour index of wheat seeds reduced by 40-50% in the presence of different concentrations of ZnO nanoparticles alone, which was alleviated by 15-20%, when ZnO and Azotobacter were present together. However, a drastic 50-60% decrease in vigour indices of tomato seeds was recorded, irrespective of Azotobacter inoculation.

ЧУТЛИВІСТЬ ЗБУДНИКІВ БАКТЕРІАЛЬНИХ ХВОРОБ ПТИЦІ ДО АНТИБАКТЕРІАЛЬНИХ ПРЕПАРАТІВ

The using of antibiotics and antimicrobials drugs without control may leads to the development of numerous complications and resistance of microorganisms to antibiotics. The using of antibiotics and antimicrobials drugs should are controlled on poultry farms. That is why the monitoring and determination of sensitivity of bacterial diseases agents to antimicrobial drugs are very important. Results of salmonellas’ and kolibakterias’ monitoring in poultry’s farms of Ukraine are introduced in the article. Researches were conducted at the Department of veterinary sanitary examination, microbiology, zoohygiene and safety and quality of livestock products of Sumy NAU. Sampling for microbiological studies was conducted from the hatchery and from pathological material and premises where poultry of different age groups was held. The spread of the disease, morbidity, mortality, mortality rate, age characteristics, economic loss what diseases cause to the poultry farms were counted. Identification of Salmonella and Escherichia was conducted by ELISA with using of RIDASCREEN® and LOCATE® test systems, according to methodical recommendations of RIDASCREEN® and LOCATE® test systems using. The results were read visually or after addition of storageco with ELISA–photometer (reader) at 450 nm. Sensitivity to antibiotics was determined by disco – diffusion method in agar. Microbiological monitoring of a number of poultry farms in Ukraine has shown that agents of bacterial diseases’ are widely spread. Between the isolated microflora largest number were accounted for Salmonella (54.1%) and the Escherichia (30.8 per cent). The rest (15,1%) were isolated cultures of Proteus, Pseudomonas, Klebsiella, Salmonella, Campylobacteria, Enterobacteria, and Clostridia Citrobacter. This indicates that systematic control over the availability of the causative agents of bacterial infections in all critical points of production of poultry products is very necessary. Among isolates that were isolated from ill poultry and poultry objects, differences in their sensitivity to antimicrobial agents from active substances that officially have registered in our country were discovered. Bactericidal activity of relatively isolated cultures was showed by colistin, ftorfenicol, zeftiocur, TimTil 250, doxicyclin, enroxil and sarafloxacin.

Detection and Molecular Identification of Persistent Water Vessel Colonizing Bacteria in a Table Water Factory in Nigeria

Aims: To establish organisms persistent in a table water production facility and to determine the points they entered in order to eliminate any risk of having pathogens in a water production system. Study Design: Analytic observational studies Place and Duration of Study: Bottling facility in Nigeria and University of Nottingham, United Kingdom. Study was between August 2011 and Sept 2013. Methodology: Sample control points were subjected to counts of bacteria, yeasts, and mold using membrane filtration to trace the source of increased bacteria counts in a table water production factory. Organisms were identified by 16S rRNA sequencing and biofilm formation was assessed Original Research Article Nwaiwu and Nwachukwu; BMRJ, 13(5): 1-12, 2016; Article no.BMRJ.24378 2 with micro titer dish biofilm formation analysis. Results: Total bacteria and Pseudomonas Spp. count were highest in the carbon filter and 5% (v/v) chlorine used for disinfection was found to be effective against planktonic cells of 18 hour cultures of Pseudomonas Spp. isolated from the carbon filter tank. No yeasts and mold were detected and after a sand blasting exercise to clean affected tanks, total bacteria counts on tryptone glucose extract medium decreased to less than factory allowable limits of 25 cfu per 100 ml of water with no further growth of Pseudomonas Spp. on centrimide medium. Other bacteria that emerged after the sand blasting exercise were regarded as persistent but had less biofilm forming ability (p=.02) when compared with Pseudomonas aeruginosa PAO1. The persistent bacteria identified after sequencing were Aeromonas hydrophila and Serratia proteamaculans. Conclusion: Apart from Pseudomonas Spp., other bacteria can persist in the water tanks of a water bottling facility and routine checks may fail to detect an underlying problem until it becomes obvious. Prompt corrective action ensures that public safety is not compromised. This was the first time Aeromonas hydrophila and Serratia proteamaculans were identified from the water tanks in the bottling facility.

A novel application of Paracoccus pantotrophus for the decolorization of melanoidins from distillery effluent under static conditions

Melanoidin is the hazardous byproduct formed during the production of ethanol in distilleries. In the present study, a highly effective melanoidin decolorizing bacterial isolate, SAG1, was isolated from the effluent enriched soil of a distillery. This strain, identified as Paracoccus pantotrophus, was highly efficient to decolorize melanoidins up to 81.2 ± 2.43% in the presence of glucose and NH4NO3. The effects of autoclaved as well as living cells and inoculums size on decolorization activity were investigated. The results indicated that only living cell showed the decolorization activity i.e. 78.6 ± 2.62%, while, no activity has been observed using autoclaved cells. The inoculums size of 8% v/v, showed maximum activity of 62.9 ± 3.00%. The isolate SAG1 was found to be more efficient in decolorizing the melanoidins from distillery effluent as compared to the reference culture Pseudomonas putida.

Changes in the quality and antioxidant properties of fresh-cut melon treated with the biopreservative culture Pseudomonas graminis CPA-7 during refrigerated storage

The strain CPA-7 of Pseudomonas graminis, isolated from apple by our group, was reported to reduce the population of foodborne pathogens in minimally processed fruit. The aim of this work was to investigate its effect on physical parameters, visual quality, enzymatic activity, vitamin C content and antioxidant activity of fresh-cut melon during refrigerated storage. CPA-7 treated or untreated fresh-cut melon was packaged under air or modified atmosphere (MAP) and stored at 5 and 10°C for 8 days. Results showed a loss of texture during storage, above all at 10°C. A good visual quality was observed for samples stored at 5°C, although samples treated with the antagonistic strain CPA-7 and stored in air presented the lowest score, whereas those stored at 10°C showed a poor visual appearance and a decrease of lightness. In general, CPA-7 treated samples presented higher POD values than untreated ones at 5°C, and lower values at 10°C. PPO increased during refrigerated storage regardless of the addition of the antagonist. Also, treatment with the antagonistic strain CPA-7 retained the antioxidant activity and vitamin C content of fresh-cut melon during refrigerated storage.

Characterization of para-Nitrophenol-Degrading Bacterial Communities in River Water by Using Functional Markers and Stable Isotope Probing.

Microbial degradation is a major determinant of the fate of pollutants in the environment. para-Nitrophenol (PNP) is an EPA-listed priority pollutant with a wide environmental distribution, but little is known about the microorganisms that degrade it in the environment. We studied the diversity of active PNP-degrading bacterial populations in river water using a novel functional marker approach coupled with [(13)C6]PNP stable isotope probing (SIP). Culturing together with culture-independent terminal restriction fragment length polymorphism analysis of 16S rRNA gene amplicons identified Pseudomonas syringae to be the major driver of PNP degradation in river water microcosms. This was confirmed by SIP-pyrosequencing of amplified 16S rRNA. Similarly, functional gene analysis showed that degradation followed the Gram-negative bacterial pathway and involved pnpA from Pseudomonas spp. However, analysis of maleylacetate reductase (encoded by mar), an enzyme common to late stages of both Gram-negative and Gram-positive bacterial PNP degradation pathways, identified a diverse assemblage of bacteria associated with PNP degradation, suggesting that mar has limited use as a specific marker of PNP biodegradation. Both the pnpA and mar genes were detected in a PNP-degrading isolate, P. syringae AKHD2, which was isolated from river water. Our results suggest that PNP-degrading cultures of Pseudomonas spp. are representative of environmental PNP-degrading populations.

Lotus effect-based coatings on marine steels to inhibit biofouling

A close observation of natural lotus leaves provided the inspiration to develop superhydrophobic surfaces on marine steels to inhibit biofouling. Low-alloyed steel such as DMR 249A is chosen for the present study and modified using mechanical polishing, shot blasting, grit blasting, glass bead blasting and pickling, and coated with a low-surface energy material such as silane. The resultant samples are found to be superhydrophobic with apparent advancing water contact angles around 150° and tilting angles less than 5°. The surface morphology and surface roughness are analysed using scanning electron microscopy and surface profilometer. Depending on the surface treatment employed, the morphology and surface roughness vary widely and they are correlated to the wetting behaviour of the samples. The surface-modified marine steels are exposed to the bacterial cultures of Pseudomonas sp. to elucidate their antifouling behaviour. The exposure studies revealed less bacterial adhesion in surface-modified samples as compared to the uncoated samples. The aim of this paper is to stress the unconventional use of superhydrophobic coatings based on silanes to inhibit biofouling on marine steels.

Mep72, a metzincin protease that is preferentially secreted by biofilms of Pseudomonas aeruginosa.

In this work, we compared the profile of proteins secreted by planktonic and biofilm cultures of Pseudomonas aeruginosa using two-dimensional difference gel electrophoresis (2D-DiGE). This revealed that a novel metzincin protease, Mep72, was secreted during biofilm growth. Subsequent Western blotting and reverse transcription-PCR (RT-PCR) analyses demonstrated that Mep72 was expressed only during biofilm growth. Mep72 has a tridomain structure comprised of a metzincin protease-like domain and two tandem carbohydrate-binding domains. Unlike the only other metzincin (alkaline protease; AprA) in P. aeruginosa, Mep72 is secreted through the type II pathway and undergoes processing during export. During this processing, the metzincin domain is liberated from the carbohydrate-binding domains. This processing may be self-catalyzed, since purified Mep72 autodegraded in vitro. This autodegradation was retarded in the presence of alginate (an extracellular matrix component of many P. aeruginosa biofilms). The expression of full-length mep72 in Escherichia coli was toxic. However, this toxicity could be alleviated by coexpression of mep72 with the adjacent gene, bamI. Mep72 and BamI were found to form a protein-protein complex in vitro. 2D-DiGE revealed that the electrophoretic mobility of several discrete protein spots was altered in the biofilm secretome of an mep72 mutant, including type III secretion proteins (PopD, PcrV, and ExoS) and a flagellum-associated protein (FliD). Mep72 was found to bind directly to ExoS and PcrV and to affect the processing of these proteins in the biofilm secretome. We conclude that Mep72 is a secreted biofilm-specific regulator that affects the processing of a very specific subset of virulence factors.

Survival in Sterile Soil and Atrazine Degradation of Pseudomonas sp. Strain ADP Immobilized on Zeolite

ABSTRACTIn laboratory settings, the ability of bacteria and fungi to degrade many environmental contaminants is well proven. However, the potential of microbial inoculants in soil remediation has not often been realized because catabolically competent strains rarely survive and proliferate in soil, and even if they do, they usually fail to express their desired catabolic potential. One method to address the survival problem is formulating the microorganisms with physical and chemical support systems. This study investigates the survival of Pseudomonas sp. strain ADP in sterile soil and its retention of atrazine-degrading functionality. Assessment was conducted with free and zeolite-immobilized bacteria incorporated into the soil. Pseudomonas sp. strain ADP remained viable for at least 10 weeks when stored at 15°C in sterile soil. Cell numbers increased for both free and zeolite-immobilized bacteria during this period, except for free cells when grown in Miller's Luria-Bertani medium, which exhibited constant cell numbers over the 10 weeks. Only the zeolite-immobilized cell retained full functionality to degrade atrazine after 10 weeks in sterile soil regardless of the medium used to culture Pseudomonas sp. strain ADP. Functionality was diminished in free-cell inoculations except when using an improved culture medium. Survival of zeolite-immobilized Pseudomonas sp. strain ADP separated from the soil matrix after 10 weeks’ incubation was significantly (p < .05) greater than in soil inoculated with free cells or in the soil fraction inoculated by release from zeolite-immobilized Pseudomonas sp. strain ADP.

Microbial bio-based plastics from olive-mill wastewater: Generation and properties of polyhydroxyalkanoates from mixed cultures in a two-stage pilot scale system

The operational efficiency of a two stage pilot scale system for polyhydroxyalkanoates (PHAs) production from three phase olive oil mill wastewater (OMW) was investigated in this study. A mixed anaerobic, acidogenic culture derived from a municipal wastewater treatment plant, was used in the first stage, aiming to the acidification of OMW. The effluent of the first bioreactor that was operated in continuous mode, was collected in a sedimentation tank in which partial removal of the suspended solids was taking place, and was then forwarded to an aerobic reactor, operated in sequential batch mode under nutrient limitation. In the second stage an enriched culture of Pseudomonas sp. was used as initial inoculum for the production of PHAs from the acidified waste. Clarification of the acidified waste, using aluminium sulphate which causes flocculation and precipitation of solids, was also performed, and its effect on the composition of the acidified waste as well as on the yields and properties of PHAs was investigated. It was shown that clarification had no significant qualitative or quantitative effect on the primary carbon sources, i.e. short chain fatty acids and residual sugars, but only on the values of total suspended solids and total chemical oxygen demand of the acidified waste. The type and thermal characteristics of the produced PHAs were also similar for both types of feed. However the clarification of the waste seemed to have a positive impact on final PHAs yield, measured as gPHAs/100g of VSS, which reached up to 25%. Analysis of the final products via nuclear magnetic resonance spectroscopy revealed the existence of 3-hydroxybutyrate (3HB) and 3-hydroxyoctanoate (HO) units, leading to the conclusion that the polymer could be either a blend of P3HB and P3HO homopolymers or/and the 3HB-co-3HO co-polymer, an unusual polymer occurring in nature with advanced properties.

The use of 16S rDNA sequencing in species diversity analysis for sputum of patients with ventilator-associated pneumonia

To study the species and amount of bacteria in sputum of patients with ventilator-associated pneumonia (VAP) by using 16S rDNA sequencing analysis, and to explore the new method for etiologic diagnosis of VAP. Bronchoalveolar lavage sputum samples were collected from 31 patients with VAP. Bacterial DNA of the samples were extracted and identified by polymerase chain reaction (PCR). At the same time, sputum specimens were processed for routine bacterial culture. The high flux sequencing experiment was conducted on PCR positive samples with 16S rDNA macro genome sequencing technology, and sequencing results were analyzed using bioinformatics, then the results between the sequencing and bacteria culture were compared. (1) 550 bp of specific DNA sequences were amplified in sputum specimens from 27 cases of the 31 patients with VAP, and they were used for sequencing analysis. 103 856 sequences were obtained from those sputum specimens using 16S rDNA sequencing, yielding approximately 39 Mb of raw data. Tag sequencing was able to inform genus level in all 27 samples. (2) Alpha-diversity analysis showed that sputum samples of patients with VAP had significantly higher variability and richness in bacterial species (Shannon index values 1.20, Simpson index values 0.48). Rarefaction curve analysis showed that there were more species that were not detected by sequencing from some VAP sputum samples. (3) Analysis of 27 sputum samples with VAP by using 16S rDNA sequences yielded four phyla: namely Acitinobacteria, Bacteroidetes, Firmicutes, Proteobacteria. With genus as a classification, it was found that the dominant species included Streptococcus 88.9% (24/27), Limnohabitans 77.8% (21/27), Acinetobacter 70.4% (19/27), Sphingomonas 63.0% (17/27), Prevotella 63.0% (17/27), Klebsiella 55.6% (15/27), Pseudomonas 55.6% (15/27), Aquabacterium 55.6% (15/27), and Corynebacterium 55.6% (15/27). (4) Pyrophosphate sequencing discovered that Prevotella, Limnohabitans, Aquabacterium, Sphingomonas might not be detected by routine bacteria culture. Among seven species which were identified by both methods, pyrophosphate sequencing yielded higher positive rate than that of ordinary bacteria culture [Streptococcus: 88.9% (24/27) vs. 18.5% (5/27), Klebsiella: 55.6% (15/27) vs. 18.5% (5/27), Acinetobacter: 70.4% (19/27) vs. 37.0% (10/27), Corynebacterium: 55.6% (15/27) vs. 7.4% (2/27), P<0.05 or P<0.01]. Sequencing positive rate was found to increase positive rate for culture of Pseudomonas [55.6% (15/27) vs. 25.9% (7/27), P=0.050]. No significant differences were observed between sequencing and ordinary bacteria culture for detection Staphylococcus [7.4% (2/27) vs. 11.1% (3/27)] and Neisseria bacteria genera [18.5% (5/27) vs. 3.7% (1/27), both P>0.05]. 16S rDNA sequencing analysis confirmed that pathogenic bacteria in sputum of VAP were complicated with multiple drug resistant strains. Compared with routine bacterial culture, pyrophosphate sequencing had higher positive rate in detecting pathogens. 16S rDNA gene sequencing technology may become a new method for etiological diagnosis of VAP.

Vertical and horizontal distributions of microbial abundances and enzymatic activities in propylene-glycol-affected soils

The natural microbial activity in the unsaturated soil is vital for protecting groundwater in areas where high loads of biodegradable contaminants are supplied to the surface, which usually is the case for airports using aircraft de-icing fluids (ADF) in the cold season. Horizontal and vertical distributions of microbial abundance were assessed along the western runway of Oslo Airport (Gardermoen, Norway) to monitor the effect of ADF dispersion with special reference to the component with the highest chemical oxygen demand (COD), propylene glycol (PG). Microbial abundance was evaluated by several biondicators: colony-forming units (CFU) of some physiological groups (aerobic and anaerobic heterotrophs and microscopic fungi), most probable numbers (MPN) of PG degraders, selected catabolic enzymatic activities (fluorescein diacetate (FDA) hydrolase, dehydrogenase, and β-glucosidase). High correlations were found between the enzymatic activities and microbial counts in vertical soil profiles. All microbial abundance indicators showed a steep drop in the first meter of soil depth. The vertical distribution of microbial abundance can be correlated by a decreasing exponential function of depth. The horizontal trend of microbial abundance (evaluated as total aerobic CFU, MPN of PG-degraders, and FDA hydrolase activity) assessed in the surface soil at an increasing distance from the runway is correlated negatively with the PG and COD loads, suggesting the relevance of other chemicals in the modulation of microbial growth. The possible role of potassium formate, component of runway de-icers, has been tested in the laboratory by using mixed cultures of Pseudomonas spp., obtained by enrichment with a selective PG medium from soil samples taken at the most contaminated area near the runway. The inhibitory effect of formate on the growth of PG degraders is proven by the reduction of biomass yield on PG in the presence of formate.

Glycine–Glomus–Phosphate Solubilizing Bacteria Interactions Lead to Fertilizer Phosphorus Economy in Soybean in a Himalayan Acid Alfisol

Effects of Glycine–Glomus–phosphate solubilizing bacteria (PSB) interactions were studied on productivity, nutrient dynamics, and root colonization in soybean in a phosphorus (P)–deficient Himalayan acidic Alfisol in a greenhouse experiment. Treatments consisted of three vesicular arbuscular mycorrhizae (VAM) cultures, VAML [VAM culture, Glomus mosseae, developed by CSK Himachal Pradesh Agricultural University, Palampur, India], VAMT [VAM culture, Glomus intraradices, developed by Centre for Mycorrhizal Research, The Energy and Resources Institute (TERI), New Delhi, India], and VAMI [VAM culture, Glomus mosseae, developed by Indian Agricultural Research Institute (IARI), New Delhi, India], and a local PSB culture (Pseudomonas striata) alone or in combination with or without 75% of recommended phosphorus pentoxide (P2O5) dose based on targeted yield concept following the soil-test crop response (STCR) precision model. Sole application of PSB or either of the above VAM cultures considerably enhanced VA-mycorrhizal root colonization and root weight besides crop productivity and nutrient uptake over control. A similar stimulatory effect with significant enhancement on mycorrhizal root colonization and root weight was observed with coinoculation of PSB and VAM cultures over the control. Dual inoculation of VAM and PSB cultures also resulted in significant improvement in grain and straw yield besides grain protein content, thereby revealing a synergistic interaction between VAM and PSB. Coinoculation with either of VAMT (Glomus intraradices) or VAMI (Glomus mosseae) + PSB + 75% P2O5 dose remained at par with sole application of 100% P2O5 dose with respect to crop productivity, nutrient content, nutrient uptake, and soil fertility status besides the greatest root colonization and root weight at flowering, indicating that Glycine–Glomus–PSB interactions in combination with 75% P2O5 dose based on STCR precision model lead to economization of fertilizer P by about one-fourth without impairing crop productivity and soil fertility in soybean in a Himalayan acidic Alfisol.

Bronchiolitis Obliterans Syndrome and Restrictive Allograft Syndrome: Do Specific Risk Factors Differ?

Purpose Chronic rejection is one of the major problems hampering long-term survival after lung transplantation (LTx). Recently, it became clear that patients with an irreversible decline in FEV1 can either develop an obstructive (Bronchiolitis Obliterans Syndrome, BOS) or a restrictive lung function defect (Restrictive allograft syndrome, RAS). Our aim was to investigate whether risk factors for BOS and RAS may be different. Methods and Materials A retrospective review of our patient cohort (LTx between 01/01/2001 and 01/11/2011) with a survival >90 days post LTx was performed (n=380). Patients with an irreversible decline in FEV1, not responding to azithromycin, and without another explanation were identified. BOS was diagnosed according to ISHLT criteria, RAS was diagnosed using TLC or FEV1/FVC ratio, combined with CAT-scan findings. Acute rejection (AR) and lymphocytic bronchiolitis (LB) were defined according to ISHLT criteria. Infections were defined as hospitalizations needing treatment with antibiotics. Colonization with Pseudomonas aeruginosa was defined as a positive culture of Pseudomonas in BAL fluid/sputum without need for treatment. Results 277 patients never experienced a persistent decline in FEV1 (stable, 69.8%). Within the remaining 103 patients, 24 developed RAS (23.3%) and 79 BOS (76.7%). Patients with RAS were significantly younger compared to BOS and stable patients (p Conclusions AR, LB, colonization with Pseudomonas, and infection are major risk factors for the later development of BOS and RAS. The exact mechanisms, remain to be established.

Draft Genome Sequences of Pseudomonas fluorescens BS2 and Pusillimonas noertemannii BS8, Soil Bacteria That Cooperate To Degrade the Poly-γ- d -Glutamic Acid Anthrax Capsule

A mixed culture of Pseudomonas fluorescens BS2 and Pusillimonas noertemannii BS8 degraded poly-γ-d-glutamic acid; when the 2 strains were cultured separately, no hydrolytic activity was apparent. Here we report the draft genome sequences of both soil isolates.

Identification of Antimony- and Arsenic-Oxidizing Bacteria Associated with Antimony Mine Tailing

Antimony (Sb) is a naturally occurring toxic element commonly associated with arsenic (As) in the environment and both elements have similar chemistry and toxicity. Increasing numbers of studies have focused on microbial As transformations, while microbial Sb interactions are still not well understood. To gain insight into microbial roles in the geochemical cycling of Sb and As, soils from Sb mine tailing were examined for the presence of Sb- and As-oxidizing bacteria. After aerobic enrichment culturing with AsIII (10 mM) or SbIII (100 μM), pure cultures of Pseudomonas- and Stenotrophomonas-related isolates with SbIII oxidation activities and a Sinorhizobium-related isolate capable of AsIII oxidation were obtained. The AsIII-oxidizing Sinorhizobium isolate possessed the aerobic arsenite oxidase gene (aioA), the expression of which was induced in the presence of AsIII or SbIII. However, no SbIII oxidation activity was detected from the Sinorhizobium-related isolate, suggesting the involvement of different mechanisms for Sb and As oxidation. These results demonstrate that indigenous microorganisms associated with Sb mine soils are capable of Sb and As oxidation, and potentially contribute to the speciation and mobility of Sb and As in situ.

A Culture-Independent Approach to Unravel Uncultured Bacteria and Functional Genes in a Complex Microbial Community

Most microorganisms in nature are uncultured with unknown functionality. Sequence-based metagenomics alone answers ‘who/what are there?’ but not ‘what are they doing and who is doing it and how?’. Function-based metagenomics reveals gene function but is usually limited by the specificity and sensitivity of screening strategies, especially the identification of clones whose functional gene expression has no distinguishable activity or phenotypes. A ‘biosensor-based genetic transducer’ (BGT) technique, which employs a whole-cell biosensor to quantitatively detect expression of inserted genes encoding designated functions, is able to screen for functionality of unknown genes from uncultured microorganisms. In this study, BGT was integrated with Stable isotope probing (SIP)-enabled Metagenomics to form a culture-independent SMB toolbox. The utility of this approach was demonstrated in the discovery of a novel functional gene cluster in naphthalene contaminated groundwater. Specifically, metagenomic sequencing of the 13C-DNA fraction obtained by SIP indicated that an uncultured Acidovorax sp. was the dominant key naphthalene degrader in-situ, although three culturable Pseudomonas sp. degraders were also present in the same groundwater. BGT verified the functionality of a new nag2 operon which co-existed with two other nag and two nah operons for naphthalene biodegradation in the same microbial community. Pyrosequencing analysis showed that the nag2 operon was the key functional operon in naphthalene degradation in-situ, and shared homology with both nag operons in Ralstonia sp. U2 and Polaromonas naphthalenivorans CJ2. The SMB toolbox will be useful in providing deep insights into uncultured microorganisms and unravelling their ecological roles in natural environments.

Abundance and diversity of culturable Pseudomonas constitute sensitive indicators for adverse long-term copper impacts in soil

For a field site heavily contaminated with copper more than 80 years ago, copper decreased the abundance of culturable Pseudomonas more than the relative abundance of Pseudomonas 16S rRNA genes. Moreover, the abundance and diversity of culturable Pseudomonas decreased more than those of culturable heterotrophic bacteria. This indicates that the Pseudomonas population is not resilient towards copper stress and that culturable Pseudomonas spp. comprise sensitive bio-indicators of adverse copper impacts in contaminated soils. Further this study shows that copper exposure decreases bacterial diversity at the species or sub-species level, while previous studies for the same soils showed no effect on the diversity of higher taxonomic ranks.

Biodegradação de paclobutrazol por Pseudomonas spp. em sistemas de solo saturados

Paclobutrazol is growth regulator of plants that has low mobility in soil and therefore has accumulated. The objective of this study was to investigate the paclobutrazol biodegradation in two soils from the Sao Francisco River Valley. The biodegradation experiments were conducted in batch using paclobutrazol and paclobutrazol added glycerol. The experiments were performed in sterile and nonsterile conditions using a mixed culture of Pseudomonas. The concentration of paclobutrazol was determined by high performance liquid chromatography. The biodegradation reached 43% in 14 days of experiments with only paclobutrazol and 70% in 28 days of experiments that contained glycerol and paclobutrazol.

A Novel Approach for Optimization of Crude Oil Bioremediation in Soil by the Taguchi Method

A design of experiment technique, the Taguchi method, has been used to optimize bioremediation of crude oil in contaminated soils. Mixed cultures of three strains of bacteria were used. Two sets of experiments were conducted with 8 factors in each set. In the first set of experiments the factors were aeration, bacteria, urea, glucose, humidity, compost, salt, and temperature. In the second set of experiments aeration and temperature were replaced with time and activated sludge, also molasses was used as additional carbon source. The contribution percentage of each factor was determined by analysis of variance. The oil removal was 64.4% and 68% for optimum conditions of the first and second set of experiments, respectively. Addition of urea and molasses showed low effect on oil removal, whereas aeration revealed 98% of oil removal from the contaminated soil. Also addition of activated sludge and compost did not substitute for mix culture of Pseudomonas.