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In this study, we report the ability of Stenotrophomonas maltophilia, Ochrobactrum anthropi, Pseudomonas mendocina, Microbacterium esteraromaticum and Pseudomonas aeruginosa to degrade multiple polycyclic aromatic hydrocarbons (PAHs) present in crude oil. The PAHs in the crude oil sample obtained from Digboi oil refinery, India were estimated to be naphthalene (10.0 mg L-1), fluorene (1.9 mg L-1), phenanthrene (3.5 mg L-1) and benzo(b)fluoranthene (6.5 mg L-1). Exposure of individual bacteria to crude oil showed high rate of biodegradation of specific PAHs by M. esteraromaticum, 81.4%-naphthalene; P. aeruginosa, 67.1%-phenanthrene and 61.0%-benzo(b)fluoranthene; S. maltophilia, 47.9%-fluorene in 45 days. However, consortium of these bacteria showed enhanced biodegradation of 89.1%-naphthalene, 63.8%-fluorene, 81% of phenanthrene and 72.8% benzo(b)fluoranthene in the crude oil. The degradation was further improved up to 10% by consortium on addition of 40 μg mL-1 rhamnolipid JBR-425 biosurfactant. These results suggest that the developed bacterial consortium has significant potential in PAH remediation.

Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P(3HB-co-4HB)) is a biodegradable plastic that is extensively utilized in many fields. In this work, P(3HB-co-4HB) powder was degraded by Pseudomonas mendocina for the preparation of low-molecular-mass (LMW) P(3HB-co-4HB). After degradation, the remaining P(3HB-co-4HB) powder was analyzed via gel permeation chromatography (GPC), differential scanning calorimetry (DSC), X-ray powder diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, and proton nuclear magnetic resonance (1H NMR) spectroscopy. The degradation of P(3HB-co-4HB) by P. mendocina occurred in two stages: the fast degradation stage (0–8 h) and the slow degradation stage (8–24 h). GPC analysis showed that the molecular weight of P(3HB-co-4HB) gradually decreased with degradation time. After 24 h of degradation, the weight-average molecular weight of P(3HB-co-4HB) was reduced to 4–5 kDa. DSC and XRD analyses both verified that the degree of crystallinity decreased with prolonged degradation time. The melting temperature of the degraded powder, however, remained unchanged. FTIR and 1H NMR analyses of the degraded powder showed that no new material was produced during degradation. Thus, the degradation of P(3HB-co-4HB) by P. mendocina could be used to produce LMW P(3HB-co-4HB) for use in various applications, such as the synthesis of amphiphilic block copolymers.

Aerobic denitrification is a process reducing the nitrate into gaseous nitrogen forms in the presence of oxygen gas, which makes the nitrification and denitrification performed simultaneously. However, little was known on the diversity of the culturable aerobic denitrifying bacteria in the surface water system. In this study, 116 strains of aerobic denitrifying bacteria were isolated from the sediment, water and biofilm samples in Liangshui River of Beijing. These bacteria were classified into 14 genera based on the 16 S rDNA, such as Pseudomonas, Rheinheimera, and Gemmobacter. The Pseudomonas sp., represented by the Pseudomonas stutzeri, Pseudomonas mendocina and Pseudomonas putida, composed the major culturable aerobic denitrifiers of the river, followed by Ochrobactrum sp. and Rheinheimera sp. The PCA plot showed the unclassified Pseudomonas sp. and Rheinheimera pacifica preferred to inhabit in biofilm phase while one unclassified Ochrobactrum sp. and Pseudomonas resinovorans had higher abundance in the sediment. In the overlying water, the Pseudomonas stutzeri and Ochrobactrum rhizosphaerae were found to have higher abundance, indicating these aerobic denitrifiers had different habitat-preferable characteristics among the 3 phases of river system. The findings may help select the niche to isolate the aerobic denitrifiers and facilitate the bioaugmentation-based purification of the nitrate polluted surface water.

Endosulfan contamination is one of the major concerns of soil ecosystem, which causes detrimental effects not only to humans but also to animals and plants. Therefore, the aim of this study was to isolate and identify a novel bacterial strain capable of degrading endosulfan in agriculture contaminated soils. A novel bacterial strain was isolated from the sugarcane field contaminated with endosulfan, and was named as ZAM1 strain. The ZAM1 bacterial strain was further identified as Pseudomonas mendocina based on the biochemical and molecular analysis. 16sRNA sequence analysis of ZAM1 strain shows maximum similarity with known endosulfan-degrading bacteria (Pseudomonas putida), respectively. Enrichment was carried out using the endosulfan as sole sulfur source. The ZAM1 strain was able to use α and β endosulfan as a sole sulfur source. Our results showed that ZAM1 strain degrades endosulfan >64.5% (50 mg/l) after 12 days of incubation. The residues were analyzed by GC–MS analysis and confirmed the formation of metabolites of dieldrin, 2 heptanone, methyl propionate, and endosulfan lactone compounds. Hence, these results indicate that the ZAM1 strain is a promising bacterial source for detoxification of endosulfan residues in the environment.

Root exudates are chemical compounds that are released from living plant roots and provide significant energy, carbon, nitrogen and phosphorus sources for microbes inhabiting the rhizosphere. The exudates shape the microflora associated with the plant, as well as influences the plant health and productivity. Therefore, a better understanding of the trophic link that is established between the plant and the associated bacteria is necessary. In this study, a comprehensive survey on the utilization of grapevine and rootstock related organic acids were conducted on a vineyard soil isolate which is Pseudomonas mendocina strain S5.2. Phenotype microarray analysis has demonstrated that this strain can utilize several organic acids including lactic acid, succinic acid, malic acid, citric acid and fumaric acid as sole growth substrates. Complete genome analysis using single molecule real-time technology revealed that the genome consists of a 5,120,146 bp circular chromosome and a 252,328 bp megaplasmid. A series of genetic determinants associated with the carbon utilization signature of the strain were subsequently identified in the chromosome. Of note, the coexistence of genes encoding several iron–sulfur cluster independent isoenzymes in the genome indicated the importance of these enzymes in the events of iron deficiency. Synteny and comparative analysis have also unraveled the unique features of d-lactate dehydrogenase of strain S5.2 in the study. Collective information of this work has provided insights on the metabolic role of this strain in vineyard soil rhizosphere. Electronic supplementary material The online version of this article (doi:10.1186/s13568-017-0437-7) contains supplementary material, which is available to authorized users.

We optimized the culture medium for 3-hydroxycarboxylic acid production by Pseudomonas mendocina DS-04-T-biodegraded polyhydroxybutyrate (PHB) using the Plackett-Burman design, steepest ascent method, and Box-Behnken design. The optimized concentrations of the constituents of the culture medium were as follows: PHB (7.57 g/L), NH4 Cl (5.0 g/L), KH2 PO4 (2.64 g/L), Na2 HPO4 ·12H2 O (12 g/L), MgSO4 ·7H2 O (0.5 g/L), and CaCl2 ·2H2 O (5 mg/L). The yield of 3-hydroxycarboxylic acid obtained using the optimized culture medium was 56.8 ± 1.64%, which was 2.5-fold higher than that obtained when the unoptimized culture medium was used.

Four methyl tert-butyl ether (MTBE)-degrading bacteria were isolated from a processing plant in Malaysia. Based on 16S rDNA sequences, the four isolates were identified as Exiguobacterium profundum P1M-2, Bacillus megaterium P1M-11, Alishewanella sp. P2A-12 and Pseudomonas mendocina P2M-8. Each of the isolates obtained optimum growth using a different source of nitrogen (0.1- 0.03 % yeast or peptone) and all four isolates were able to biodegrade 92.05-99.98 % of MTBE within seven days. Amongst the four isolates, the highest percentage of MTBE degradation was achieved using B. megaterium P1M-11. The highest growth on tert-butyl alcohol (TBA), tert-amyl alcohol (TAA) and 2-hydroxyisobutyric acid (2-HIBA) was also observed in B. megaterium P1M-11. This study suggests MTBE degradation by each of the bacteria can be enhanced by choosing the right nitrogen source. Furthermore, the ability of B. megaterium P1M-11 to grow on primary metabolites of MTBE and other structurally related ethers suggests the secretion of diverse degradative enzymes, making this isolate a good candidate to be applied in MTBE bioremediation strategies.

A novel toluene monooxygenase (TMO) six-gene cluster from Bradyrhizobium sp. BTAi1 having an overall 35, 36, and 38 % protein similarity with toluene o-xylene monooxygenase (ToMO) of Pseudomonas sp. OX1, toluene 4-monooxygenase (T4MO) of Pseudomonas mendocina KR1, and toluene-para-monooxygenase (TpMO) of Ralstonia pickettii PKO1, respectively, was cloned and expressed in Escherichia coli TG1, and its potential activity was investigated for aromatic hydroxylation and trichloroethylene (TCE) degradation. The natural substrate toluene was hydroxylated to p-cresol, indicating that the new toluene monooxygenase (T4MO·BTAi1) acts as a para hydroxylating enzyme, similar to T4MO and TpMO. Some shifts in regiospecific hydroxylations were observed compared to the other wild-type TMOs. For example, wild-type T4MO·BTAi1 formed catechol (88 %) and hydroquinone (12 %) from phenol, whereas all the other wild-type TMOs were reported to form only catechol. Furthermore, it was discovered that TG1 cells expressing wild-type T4MO·BTAi1 mineralized TCE at a rate of 0.67 ± 0.10 nmol Cl(-)/h/mg protein. Saturation and site directed mutagenesis were used to generate eight variants of T4MO·BTAi1 at alpha-subunit positions P101, P103, and H214: P101T/P103A, P101S, P101N/P103T, P101V, P103T, P101V/P103T, H214G, and H214G/D278N; by testing the substrates phenol, nitrobenzene, and naphthalene, positions P101 and P103 were found to influence the regiospecific oxidation of aromatics. For example, compared to wild type, variant P103T produced four fold more m-nitrophenol from nitrobenzene as well as produced mainly resorcinol (60 %) from phenol whereas wild-type T4MO·BTAi1 did not. Similarly, variants P101T/P103A and P101S synthesized more 2-naphthol and 2.3-fold and 1.6-fold less 1-naphthol from naphthalene, respectively.

Nitric oxide (NO) is one of the most important air pollutants in atmosphere mainly emitted from combustion source. A biotrickling filter was designed and operated to remove NO from an air stream using bacteria extracted from the sewage sludge of a municipal sewage treatment plant. To obtain the best operation conditions for the biotrickling filter, orthogonal experiments (L 9 (3 4 )) were designed. Inlet oxygen concentration was found to be the most significant factor of the biotrickling filter and has a significant negative effect on the system. The optimal conditions of the biotrickling filter occurred at a temperature of 40°C, a pH of 8.0 and a chemical oxygen demand of 165 mg/L in the recycled water with no oxygen in the system. The bacteria sample was detected by DNA sequencing technology and showed 93%–98% similarity to Pseudomonas mendocina. Moreover, a full gene sequencing results indicated the bacterium was a brand new strain and named as P. mendocina DLHK. This strain can transfer nitrate to organic nitrogen. The result suggested the assimilation nitrogen process in this system. Through the isotope experimental analysis, two intermediate products ( 15 NO and 15 N 2 O) were found. The results indicated the denitrification function and capability of the biotrickling filter in removing NO.

Highlights • Selective enrichment was used to isolate active biodegradative bacteria. • The role of chemotaxis in xenobiotic metabolism was elucidated. • Wastewater mesocosms were identified as a repository for biodegradative bacteria., Wastewater environment is a rich source of microorganisms with the capability for the degradation of malicious aromatic pollutants. Although wastewater could be regarded as both a resource and a problem, we intended to elucidate its beneficial aspect in this study sourcing for laccase-producing proteobacteria. Different wastewater samples, from selected wastewater treatment plants (WWTPs), were selectively enriched with some model compounds (vanillin, lignin and potassium hydrogen phthalate) to screen out bacterial isolates that possess excellent degradation potentials. Thereafter, positive isolates were screened for the production of laccase and degradation on phenolic (guaiacol, α-naphthol and syringaldazine) and non-phenolic (ABTS; 2,2 azino-bis -(3-ethylbenzothiazoline 6 sulphonic acid) and PFC; potassium ferrocyanoferrate) substrates characteristic of laccase oxidation. Remarkable laccase producers were identified based on their 16 S rRNA sequences and their secreted enzymes were subjected to substrate specificity test, employing laccase substrates; ABTS, PFC, guaiacol, α-naphthol, 2,6-dimethoxyphenol and pyrogallol. Results showed that wastewater and selective enrichment, in tandem, produced the gammaproteobacteria Pseudomonas aeruginosa DEJ16, Pseudomonas mendocina AEN16 and Stenotrophomonas maltophila BIJ16, which preferably oxidized the non-phenolic substrates. Units of extracellular laccase activity ranging between cca. 490 and cca. 600 U/mL were recorded for ABTS whereas outputs recorded from PFC catalysis ranged from cca. 320 to cca. 430 U/mL. Stenotrophomonas maltophila BIJ16 presented an unparalleled high laccase activity and had a responsive substrate specificity to aromatic and inorganic substrates, thereby suggesting its employment for in situ biodegradation studies. In conclusion, wastewater serves as an ideal milieu for the isolation of laccase producing bacteria.

Malachite green (MG) is a highly toxic, and recalcitrant tri-phenyl methane dye, which can be adsorbed on surface of chicken feathers, a non toxic poultry waste. This interaction can affect metabolism of chicken feathers by microbes in the environment. In this study, decolorization/degradation of MG and metabolism of MG adsorbed chicken feathers by Pseudomonas mendocina strain PM2 has been evaluated. Decolorization of different concentrations of MG by strain PM2 was studied. Degradation of MG was studied by UV–vis/FTIR analysis. Enzyme activities of selected enzymes were evaluated in order to ascertain their role in MG degradation. Microbial/phytotoxocity studies on MG and its degraded metabolites were carried out to confirm detoxification of MG after degradation. MG was adsorbed on surface of chicken feathers and its toxicity on strain PM2 and MG sensitive strain of Bacillus subtilis MTCC 441 was evaluated. Metabolism of native and MG adsorbed chicken feathers by strain PM2 and MTCC 441 was studied. Results showed that strain PM2 could decolorize up to 1800 mg/L MG. UV–vis/FTIR studies indicated degradation of MG by strain PM2. Assay of enzyme activity indicated involvement of MG reductase, lignin peroxidase and manganese peroxidase during degradation. Microbial/phytotoxicity study confirmed biodetoxification of MG after degradation. MG adsorbed chicken feathers were toxic to MTCC 441 and non toxic to strain PM2. Further, strain PM2 was able to metabolize MG adsorbed chicken feathers whereas MTCC 441 could not metabolize MG adsorbed chicken feathers. This study clearly indicates that MG adsorbed chicken feathers are difficult to be metabolized by MG sensitive strains.

A waste slip-stream is generated from the reclaiming process of monoethanolamine (MEA) based Post-Combustion Capture (PCC). It mainly consists of MEA itself, ammonium, heat-stable salts (HSS), carbamate polymers, and water. In this study, the waste quantity and nature are characterized for Fluor's Econamine FGSM coal-fired CO2 capture base case. Waste management options, including reuse, recycling, treatment, and disposal, are investigated due to the need for a more environmentally sound handling. Regulations, economic potential, and associated costs are also evaluated. The technical, economic, and regulation assessment suggests waste reuse for NOx scrubbing. Moreover, a high thermal condition is deemed as an effective technique for waste destruction, leading to considerations of waste recycling into a coal burner or incineration. As a means of treatment, three secondary-biological processes covering Complete-Mix Activated Sludge (CMAS), oxidation ditch, and trickling filter are designed to meet the wastewater standards in the United Arab Emirates (UAE). From the economic point of view, the value of waste as a NOx scrubbing agent is 6,561,600-7,348,992 USD/year. The secondary-biological treatment cost is 0.017-0.02 USD/ton of CO2, while the cost of an on-site incinerator is 0.031 USD/ton of CO2 captured. In conclusion, secondary biological treatment is found to be the most economical option.

This research investigated strategies used by the common aerobic soil bacterium Pseudomonas mendocina to acquire Fe associated with Fe(III)-bearing montmorillonite (MMT) clay. Given the known importance of Fe(III)-chelating siderophores, Fe-limited batch experiments were conducted using a wild-type (WT) strain that produces siderophores and a ΔpmhA mutant with a siderophore(−) phenotype. Growth measurements were coupled with a transcriptional biosensor assay that monitors the siderophore biosynthesis gene pmhA, measurements of cells’ reducing ability, and quantification of exopolymeric substance (EPS) production. WT cells actively grow when MMT is the sole Fe source, but sorption to MMT may decrease the concentration of dissolved Fe–siderophore complex accessible to cells. Cells also obtain Fe by reducing MMT-associated Fe(III), but because P. mendocina lacks a secreted/diffusible reductant, direct physical contact is required. Dual strategies for Fe acquisition—a reducing mechanism that requires contact and that is likely facilitated by biofilm production and a siderophore related mechanism that does not require contact—provide flexibility to address the environmental Fe challenge.

Two polyhydroxyalkanoate depolymerases, PHAase I and PHAase II, were purified to homogeneity from the culture supernatant of an effective PHA-degrading bacterium, Pseudomonas mendocina DS04-T. The molecular masses of PHAase I and PHAase II were determined by SDS-PAGE as 59.4 and 33.8 kDa, respectively. Their optimum pH values were 8.5 and 8, respectively. Enzymatic activity was optimal at 50 °C. Both purified enzymes could degrade PHB, PHBV, and P(3HB-co-4HB). Addition of Na(+) and K(+) slightly increased the rate of PHAase II. EDTA significantly inhibited PHAase II but not PHAase I. Mercaptoethanol and H2O2 also inhibited the activities of both enzymes.

The medium-chain-length polyhydroxyalkanoate (PHAMCL) synthase genes phaC1 and phaC2 of Pseudomonas mendocina NK-01 were cloned and inserted into expression plasmid pBBR1MCS-2 to form pBBR1MCS-C1 and pBBR1MCS-C2 which were expressed respectively in the PHAMCL-negative strain P. mendocina C7 whose PHAMCL synthesis operon was defined knock out. P. mendocina C7 derivatives P. mendocina C7C1 and C7C2 carrying pBBR1MCS-C1 and pBBR1MCS-C2 respectively were constructed. Fermentation and gel permeation chromatography (GPC) revealed that P. mendocina C7C1 had higher PHAMCL production rate but its PHAMCL had lower molecular weight than that of P. mendocina C7C2. Gas chromatograph/mass spectrometry (GC/MS) analysis revealed that the two PHAMCL had similarity in monomer composition with 3HD as the favorite monomer i.e. PhaC1 and PhaC2 had the same substrate specificity. Differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and X-ray diffraction (XRD) also revealed that the two PHAMCL had the same physical properties. P. mendocina NK-01was the first reported strain whose PHAMCL synthases PhaC1 and PhaC2 had the same substrate specificity.

The importance of our inner microbial communities for proper immune responses against invading pathogens is now well accepted, but the mechanisms underlying this protection are largely unknown. In this study, we used Caenorhabditis elegans to investigate such mechanisms. Since very little is known about the microbes interacting with C. elegans in its natural environment, we began by taking the first steps to characterize the C. elegans microbiota. We established a natural-like environment in which initially germfree, wild-type larvae were grown on enriched soil. Bacterial members of the adult C. elegans microbiota were isolated by culture and identified using 16S rRNA gene sequencing. Using pure cultures of bacterial isolates as food, we identified two, Bacillus megaterium and Pseudomonas mendocina , that enhanced resistance to a subsequent infection with the Gram-negative pathogen Pseudomonas aeruginosa . Whereas protection by B. megaterium was linked to impaired egg laying, corresponding to a known trade-off between fecundity and resistance, the mechanism underlying protection conferred by P. mendocina depended on weak induction of immune genes regulated by the p38 MAPK pathway. Disruption of the p38 ortholog, pmk-1 , abolished protection. P. mendocina enhanced resistance to P. aeruginosa but not to the Gram-positive pathogen Enterococcus faecalis . Furthermore, protection from P. aeruginosa was similarly induced by a P. aeruginosa gacA mutant with attenuated virulence but not by a different C. elegans -associated Pseudomonas sp. isolate. Our results support a pivotal role for the conserved p38 pathway in microbiota-initiated immune protection and suggest that similarity between microbiota members and pathogens may play a role in such protection.

Water, leaf, stone and sand samples from Sitakunda hot spring were studied and the microorganisms isolated from these samples were thoroughly characterized. The pH of the collected samples was found to range from 8.5 to 8.9 and the recorded temperature varied from 26.1 to 29.8 oC during winter season. BOD of the collected water samples ranged from 10 to 92 mg/L, while COD was almost zero. The total alkalinity and chlorine content ranged from 12 to 20 mg/L and 13.9 to 18.3 gm/L respectively. Quantitative enumeration of bacterial population showed the range of 8.0×10 2 to 3.8×10 5 cfu/ml or gm sample. All the 9 selected isolates were identified as Bacillus coagulans (S 1 B 4 ), Bacillus laterosporus (SB 18 ), Bacillus megaterium (S 2 B 11 ), Bacillus popilliae (S 4 P 1 ), Bacillus firmus (S 5 B 1 ), Citrobacter intermedius (S 4 B 17 ), Listeria monocytogenes (S 1 B 1 ) , Listeria denitrificans (S 2 B) and Pseudomonas mendocina (S 4 P 3 ). Bacillus laterosporus , Bacillus megaterium and Listeria denitrificans showed better growth in presence of methane gas in nutrient broth. By direct microscopic observation 7 genera of cyanobacteria ( Arthrospria, Microcoleus, Oscillatoria, Schyzothrix, Scytonema, Nostoc and Gloeocapsa ) and 4 genera of diatoms ( Gomphonema, Melosira, Navicula and Pinnularia ) were detected in the water sample DOI: http://dx.doi.org/10.3329/cujbs.v4i1.13394 The Chittagong Univ. J. B. Sci.,Vol. 4(1&2):107-117, 2009

The biodegradation behavior of polyhydroxybutyrate film in the presence of Pseudomonas mendocina DS04-T was studied and the degraded films were analyzed. The degradation was carried out in liquid basal medium in the presence of the strain for 52 h. Differential scanning calorimetry and scanning electron microscopy results revealed that the strain preferentially degraded the amorphous over the crystalline phase of the film during the initial stage of the degradation. Afterwards, the crystallized parts started to degraded. The center of the polyhydroxybutyrates pherocrystal was easily degraded. The thermogravimetric analysis revealed that the thermal stability of the degraded films decreases with the extent of degradation.

The screening of plant growth promoting rhizobacteria is a crucial step for their utilization as beneficial input in improving the crop productivity. This study was carried out to screen and evaluate the auxin producing rhizospheric isolated Pseudomonas strains for their potential to improve growth of Triticum aestivum (wheat) plant under laboratory and natural conditions. Three strains PNS-4, PNS-6 and PNS-15 were evaluated for auxin production by Salkowski’s method and further confirmed by high performance liquid chromatography (HPLC). The PNS-4, PNS-6 and PNS-15 strains were identified by I6S rRNA gene sequencing that showed maximum resemblance with Pseudomonas mendocina (99%), Pseudomonas alcaliphila (99%) and Pseudomonas sp. (99%) respectively. Selected strains were found to produce auxin with and without the amendment of exogenously applied L-tryptophan, a major precursor for auxin biosynthesis and an important constituent of plant root exudates. Efficacy of these strains on wheat plant growth was checked under laboratory and field conditions. All Pseudomonas species were found to improve the % seed germination and growth parameters (shoot length, root length, fresh weight and dry weight) of the wheat seedlings significantly (P = 0.05) as compared to the un-inoculated seedlings under laboratory condition. The biochemical parameters (total soluble protein content and endogenous auxin content) of the bacterial inoculated wheat seedling were also increased significantly than that of uninoculated ones. Under natural condition, seed bacterization also showed the significant effect (P = 0.05) on yield parameters (shoot length, number of tillers, spike length and weight of seeds in grams) of the wheat plants when compared with non-inoculated plants. Our results reported the three most promising Pseudomonas candidates and revealed the fact that experiments under laboratory and natural conditions may be helpful in selecting the best candidates as bio fertilizers for future agricultural practices.

The production conditions of 3-hydroxybutyrate (3-HB) monomers in submerged culture using Pseudomonas mendocina DS04-T as a degrading strain were optimized. The optimal culture medium constituents (w/v) were determined as follows: 0.5 % PHB, 0.15 % NH4Cl, 1.2 % Na2HPO4·12H2O, 0.3 % KH2PO4, 0.05 % MgSO4·7H2O, and 0.0005 % CaCl2·2H2O. The optimum parameters for liquid fermentation were as follows: temperature, 30 °C; inoculum content, 1.0 %; cultivation time, 18 h; initial pH, 7.0; volume of medium, 100 mL; and rotary speed, 180 rpm. Yield of 3-HB monomer and PHB depolymerase activity at optimized conditions were (56.4 ± 0.8) % and (57.4 ± 2.6) U/mL, respectively. The 3-HB monomer concentration obtained under optimized conditions was 1.5 times that obtained under the basic culture medium and initial conditions.

Background Gram-negative microorganisms are uncommon pathogens responsible for infective endocarditis. Pseudomonas mendocina, a Gram-negative water-borne and soil-borne bacterium, was first reported to cause human infection in 1992. Since then, it has rarely been reported as a human pathogen in the literature. We describe the first case of native valve infective endocarditis due to P. mendocina in the USA. Case presentation A 57-year-old white man presented with bilateral large leg ulcers, fever, and marked leukocytosis. His past medical history included gout and chronic alcohol use. P. mendocina was isolated from his blood cultures. A comprehensive review of P. mendocina infection in the literature was performed. A total of eight cases of P. mendocina infection were reported in the literature. More than two-thirds of the cases of P. mendocina septicemia were associated with native valve infective endocarditis. Thus, an echocardiogram was performed and demonstrated mitral valve endocarditis with mild mitral insufficiency. His leg wounds were debrided and were probably the source of P. mendocina bacteremia. Unlike Pseudomonas aeruginosa, P. mendocina is susceptible to third-generation cephalosporins. Our patient received a 6-week course of antimicrobial therapy with a favorable clinical outcome. Conclusions Our reported case and literature review illuminates a rare bacterial cause of infective endocarditis secondary to P. mendocina pathogen. Native cardiac valves were affected in all reported cases of infective endocarditis, and a majority of affected heart valves were left-sided. The antibiotics active against P. mendocina are different from those that are active against P. aeruginosa, and they notably include third-generation cephalosporins. The outcome of all reported cases of P. mendocina was favorable and no mortality was described.

Glucosylglycerols (GGs) are known as compatible solutes accumulated by some bacteria including cyanobacteria as well as higher plants for their adaptations to salt or desiccation stresses. Since being identified in Japanese sake, their physiological effects and potential applications on human health cares have been explored in the following 15 years. Several different synthesis methods have been successively developed for the production of GGs. However, the efficiency of GG synthesis, especially biological synthesis, is still low. With the recent advances in genome sequencing and synthetic biology tools, systematical screening of enzyme candidates and metabolic engineering approaches is necessary for improving GG synthesis efficiency. In this review, we will summarize GG structure information, protective effects on human skin and digestive system as well as industrial enzymes, together with their synthesis by chemical, enzymatic, and biological in vivo approaches in detail, and provide some prospects on improving GG production.

International audience; The universal Per-ARNT-Sim (PAS) domain functions as a signal transduction module involved in sensing diverse stimuli such as small molecules, light, redox state and gases. The highly evolvable PAS scaffold can bind a broad range of ligands, including haem, flavins and metal ions. However, although these ligands can support catalytic activity, to our knowledge no enzymatic PAS domain has been found. Here we report characterization of the first PAS enzyme: a haem-dependent oxidative N-demethylase. Unrelated to other amine oxidases, this enzyme contains haem, flavin mononucleotide, 2Fe-2S and tetrahydrofolic acid cofactors, and specifically catalyses the NADPH-dependent oxidation of dimethylamine. The structure of the α subunit reveals that it is a haem-binding PAS domain, similar in structure to PAS gas sensors. The dimethylamine substrate forms part of a highly polarized oxygen-binding site, and directly assists oxygen activation by acting as both an electron and proton donor. Our data reveal that the ubiquitous PAS domain can make the transition from sensor to enzyme, suggesting that the PAS scaffold can support the development of artificial enzymes.

Bacterial endophytes are a good source of plant growth promoting bacteria to design biofertilizers, but they can include pathogenic species. Since matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) is a reliable method for bacterial identification and all cultivable bacterial species causing human diseases are included in the Biotyper 3.0 database, we used this methodology to analyze a collection of endophytic bacteria isolated from maize roots in Canary Islands. We identified several human pathogens such as Bacillus cereus, Enterobacter cloacae, Kosakonia cowanii (formerly Entrobacter cowanii), Klebsiella pneumoniae, Pantoea agglomerans, Pseudomonas aeruginosa, Pseudomonas fulva, Pseudomonas mendocina, Pseudomonas mosselii, and Staphylococcus epidermidis. These results showed that MALDI-TOF MS is a useful technique in the process of characterization of endophytic bacteria allowing to discard strains belonging to pathogenic species that cannot be included in biofertilizers.

Microbial degradation of hydrocarbons is found to be an attractive process for remediation of contaminated habitats. However the poor bioavailability of hydrocarbons results in low biodegradation rates. Cyclodextrins are known to increase the bioavailability of variety of hydrophobic compounds. In the present work we purified the Cyclodextrin Glucanotransferase (CGTase) enzyme which is responsible for converting starch into cyclodextrins and studied its role on biodegradation of diesel oil contaminated soil. Purification of CGTase from Enterobacter cloacae was done which resulted in 6 fold increase in enzyme activity. The enzyme showed maximum activity at pH 7, temperature with a molecular weight of 66 kDa. Addition of purified CGTase to the treatment setup with Pseudomonas mendocina showed enhanced biodegradation of diesel oil () which was similar to the treatment setup when added with Pseudomonas mendocina and Enterobacter cloacae (). The residual diesel oil found in treatment setup added with Pseudomonas mendocina at end of the study was found to be . Immobilization of Pseudomonas mendocina on alginate containing starch also led to enhanced biodegradation of hydrocarbons in diesel oil at 336 hours.

A denitrifying bacterium aHD7 with nitrate removal efficiency of 91.67% (w/w) in 3 days cultured at 30 °C under stationary conditions was isolated from the activated sludge. The strain was characterized by biochemical tests and 16S rDNA sequence analysis, to be belonging to Pseudomonas. Factors on denitrification ability of aHD7, i.e., pH value, carbon source, C/N ratio, nitrogen source and initial nitrate concentration were investigated. Results showed that this strain could either use nitrate or nitrite as the electron acceptor, and the total inorganic nitrogen removal efficiency for sodium nitrite and potassium nitrate were up to 96.70% (w/w) and 95.93% (w/w) respectively. Denitrification was favored when the pH value was 6-9 and C/N ratio varied from 6 to 8, in which the optimal carbon source was ethanol. The strain had anammox activity, of which the average ammonium utilization rate reached up to 4.56 mg/l/d.

This research investigated the potential role of siderophores in aerobic microbial Fe acquisition from natural organic matter (NOM; XAD-8 isolate and reverse osmosis concentrate pre- and post-Chelex® treatment) through the use of a siderophore-producing Pseudomonas mendocina wild type (WT) bacterium and an engineered mutant (Mt) that was incapable of siderophore production. NOM had complex effects on microbial growth under Fe-limited conditions as measured by optical density, most likely because of the presence of other toxic (trace) metals such as Al, NOM binding interference with additional trace metal nutrients, and/or biofilm development. However, a bioassay for cellular Fe status showed that both WT and Mt readily acquired Fe naturally associated with NOM. Thus, while siderophores may be useful for Fe acquisition from NOM by P. mendocina, they do not appear to be essential for this process.

Crystal structures of toluene 4-monooxygenase hydroxylase in complex with reaction products and effector protein reveal active site interactions leading to regiospecificity. Complexes with phenolic products yield an asymmetric μ-phenoxo-bridged diiron center and a shift of diiron ligand E231 into a hydrogen bonding position with conserved T201. In contrast, complexes with inhibitors p-NH(2)-benzoate and p-Br-benzoate showed a μ-1,1 coordination of carboxylate oxygen between the iron atoms and only a partial shift in the position of E231. Among active site residues, F176 trapped the aromatic ring of products against a surface of the active site cavity formed by G103, E104 and A107, while F196 positioned the aromatic ring against this surface via a π-stacking interaction. The proximity of G103 and F176 to the para substituent of the substrate aromatic ring and the structure of G103L T4moHD suggest how changes in regiospecificity arise from mutations at G103. Although effector protein binding produced significant shifts in the positions of residues along the outer portion of the active site (T201, N202, and Q228) and in some iron ligands (E231 and E197), surprisingly minor shifts (

Summary The two-component system TmoS/TmoT controls the expression of the toluene-4-monooxygenase pathway in Pseudomonas mendocina RK1 via modulation of PtmoX activity. The TmoS/TmoT system belongs to the family of TodS/TodT like proteins. The sensor kinase TmoS is a 108 kDa protein composed of seven different domains. Using isothermal titration calorimetry we show that purified TmoS binds a wide range of aromatic compounds with high affinities. Tightest ligand binding was observed for toluene (KD = 150 nM), which corresponds to the highest affinity measured between an effector and a sensor kinase. Other compounds with affinities in the nanomolar range include benzene, the 3 xylene isomers, styrene, nitrobenzene or p-chlorotoluene. We demonstrate that only part of the ligands that bind to TmoS increase protein autophosphorylation in vitro and consequently pathway expression in vivo. These compounds are referred to as agonists. Other TmoS ligands, termed antagonists, failed to increase TmoS autophosphorylation, which resulted in their incapacity to stimulate gene expression in vivo. We also show that TmoS saturated with different agonists differs in their autokinase activities. The effector screening of gene expression showed that promoter activity of PtmoX and PtodX (controlled by the TodS/TodT system) is mediated by the same set of 22 compounds. The common structural feature of these compounds is the presence of a single aromatic ring. Among these ligands, toluene was the most potent inducer of both promoter activities. Information on the TmoS/TmoT and TodS/TodT system combined with a sequence analysis of family members permits to identify distinct features that define this protein family.

1-aminocyclopropane-1-carboxylate deaminase (ACC deaminase) produced by rhizobacteria could be remove theethylene precursor and stimulate plant growth. Aim of the work was investigation on effect of rhizosphere bacteria Pseudomonasmendocina containing plasmid carrying gene encoding ACC deaminase on resistance of tomato plant to salinity. Amplification ofacds gene in selected Pseudomonas was performed; the g e n e w a s c l o n e d i n Escherichia coli and was cloned subsequently in P.mendocina. Enzyme activity was determined in cloned Escherichia coli and cloned P. mendocina for confirmation of geneexpression. Effect of bacterial ACC deaminase on resistance of tomato plants to NaCl was studied in Pot and Greenhouse. In potexperiment, tomato plant treated by cloned P. mendocina was compared with plants treated by P. mendocina (without plasmid) andcontrol group. Salinity were established by adding 172 and 207 mM of NaCl to irrigated water. Greenhouse experiments wereconducted in similar groups of bacteria in 207 mM of NaCl. Results obtained from pot experiment revealed that plants treated bycloned P. mendocina in 172 mM of NaCl was showed increasing content of growth than ones treated by P. mendocina and controlas 11%, 18.4% growth for the shoot, 16.6%, 3.7% for roots and 9.6%, 27.5% for wet weight after five weeks, respectively. In 207mM of NaCl, the results were as 14.9 %, 9.7% for shoot, 94.3%, 15.7% for roots and 96.4%, 50.6% for wet weight, respectively. Ingreenhouse experiment, results in same parameter in 207 mM of NaCl were revealed as 63.7%, 7 times for shoot, 2.8, 14 times forroots and 66.1%, 154 times for wet weight, respectively. We concluded that recombinant P. mendocina producing ACC deaminaseby reduction of ethylene content of tomato plant in high salt concentrations could result in improvement of plant resistance tosalinity.

The abundance and structure of denitrifying bacterial community in different activated sludge samples were assessed, where the abundance of denitrifying functional genes showed nirS in the range of 10(4)-10(5), nosZ with 10(4)-10(6) and 16S rRNA gene in the range 10(9)-10(10) copy number per ml of sludge. The culturable approach revealed Pseudomonas sp. and Alcaligenes sp. to be numerically high, whereas culture independent method showed betaproteobacteria to dominate the sludge samples. Comamonas sp. and Pseudomonas fluorescens isolates showed efficient denitrification, while Pseudomonas mendocina, Pseudomonas stutzeri and Brevundimonas diminuta accumulated nitrite during denitrification. Numerically dominant RFLP OTUs of the nosZ gene from the fertilizer factory sludge samples clustered with the known isolates of betaproteobacteria. The data also suggests the presence of different truncated denitrifiers with high numbers in sludge habitat.

In late 2003, a new disease appeared in protected bean crops in southeastern Spain, causing a decrease of over 50% in production. Several samples of affected plants were collected and analyzed and the agent of this disease was identified as the bacterium Erwinia aphidicola, which had never been described as a pathogen previously. We attempted to determine the possible bacterium transmission through seeds, using 120 commercial bean seeds from the same batch as that used in an affected farm, and 120 seeds from the fruiting plants of the same farm. Seed coats, cotyledons and leaves of plants originating from them, were taken and analyzed. Several of the developed symptoms on plants from commercial and fruiting plant seeds were internervial chlorosis, necrotic pits and rough roots and they coincided with those observed on affected crops. Bacteria present in commercial seed cotyledons were isolated and analyzed by biochemical and molecular tests. Results confirmed the presence of Erwinia aphidicola in four analyzed seeds; moreover, Bacillus simplex/Bacillus muralis, Pseudomonas mendocina, Pseudomonas putida and Paenibacillus polymyxa were also identified.

Choline is abundant in association with eukaryotes and plays roles in osmoprotection, thermoprotection, and membrane biosynthesis in many bacteria. Aerobic catabolism of choline is widespread among soil proteobacteria, particularly those associated with eukaryotes. Catabolism of choline as a carbon, nitrogen, and/or energy source may play important roles in association with eukaryotes, including pathogenesis, symbioses, and nutrient cycling. We sought to generate choline analogues to study bacterial choline catabolism in vitro and in situ . Here we report the characterization of a choline analogue, propargylcholine, which inhibits choline catabolism at the level of Dgc enzyme-catalyzed dimethylglycine demethylation in Pseudomonas aeruginosa . We used genetic analyses and 13 C nuclear magnetic resonance to demonstrate that propargylcholine is catabolized to its inhibitory form, propargylmethylglycine. Chemically synthesized propargylmethylglycine was also an inhibitor of growth on choline. Bioinformatic analysis suggests that there are genes encoding DgcA homologues in a variety of proteobacteria. We examined the broader utility of propargylcholine and propargylmethylglycine by assessing growth of other members of the proteobacteria that are known to grow on choline and possess putative DgcA homologues. Propargylcholine showed utility as a growth inhibitor in P. aeruginosa but did not inhibit growth in other proteobacteria tested. In contrast, propargylmethylglycine was able to inhibit choline-dependent growth in all tested proteobacteria, including Pseudomonas mendocina , Pseudomonas fluorescens , Pseudomonas putida , Burkholderia cepacia , Burkholderia ambifaria , and Sinorhizobium meliloti . We predict that chemical inhibitors of choline catabolism will be useful for studying this pathway in clinical and environmental isolates and could be a useful tool to study proteobacterial choline catabolism in situ .

Two bacterial strains, i.e. Pseudomonas mendocina and Ochrobactrum sp. were isolated from bilge oil contaminated water of Mormugao harbour, Goa, India and grown in a culture medium with hexadecane as the sole carbon source. Pseudomonas mendocina was used in further studies as it was the dominant strain. This strain effectively degraded tetradecane, hexadecane and octadecane leaving a residual concentration of about 73 %, 54 % and 40 % respectively in 120 h. Sequence analysis of the dominant bands from the denaturing gradient gel electrophoresis profiles revealed the differences between the genera of bilge oil contaminated sea water and its enrichment culture on hexadecane indicating a shift in community structure based on the type of substrate available. Pseudomonas mendocina amplified for the following catabolic genes namely C23O, nid and ndo. Based on the catabolic gene study the potential of the bacterial strain isolated, i.e. Pseudomonas mendocina seems to be interesting as it will be able to degrade polyaromatic hydrocarbons as well. Physicochemical properties of Pseudomonas mendocina indicates production of exopolysaccharides based on the value of its isoelectric point.

When Pseudomonas mendocina NK-01 was cultivated in a 200-L fermentor using glucose as carbon source, 0.316 g L(-1) medium-chain-length polyhydroxyalkanoate (PHA(MCL)) and 0.57 g L(-1) alginate oligosaccharides (AO) were obtained at the end of the process. GC/MS was used to characterize the PHA(MCL), which was found to be a polymer mainly consisting of 3HO (3-hydroxyoctanoate) and 3HD (3-hydroxydecanoate). T (m) and T (g) values for the PHA(MCL) were 51.03 °C and -41.21 °C, respectively, by DSC. Its decomposition temperature was about 300 °C. The elongation at break was 700% under 12 MPa stress. MS and GPC were also carried out to characterize the AO which had weight-average molecular weights of 1,546 and 1,029 Da, respectively, for the two main components at the end of the fermentation process. MS analysis revealed that the AO were consisted of β-D-mannuronic acid and/or α-L-guluronic acid, and the β-D-mannuronic acid and/or α-L-guluronic acid residues were partially acetylated at position C2 or C3.

Pseudomonas mendocina was found to produce a unique homopolymer of poly(3-hydroxyoctanoate), P(3HO), rather than a copolymer, when grown on sodium octanoate as the sole carbon source. Although this polymer has been produced by other organisms, interestingly this is the first time an absolute homopolymer has been produced by a wild type organism. In addition, a detailed study on the effects of different extraction methods on the yield, molecular weight, thermal properties, and lipopolysaccharide content of P(3HO) has been carried out. The organism was able to accumulate P(3HO) up to 31.38% of its dry cell weight within 48 h in mineral salt medium. Characterization of the monomer was carried out using FTIR, GC-MS, (13)C, (1)H, and HSQC NMR spectroscopy. The polymer had a crystallinity of 37.5%, Young's modulus value of 11.6 MPa and contact angle of 77.3°. Microstructural studies of solvent cast polymer films revealed a smooth surface topography with a root-mean-square roughness value of 0.238 μm.

Near-infrared transflectance spectra of chicken breast muscle inoculated with Listeria innocua FH 2333, Pseudomonas fluorescens 2088, Pseudomonas putida 49128, Pseudomonas mendocina or Escherichia coli K12 were investigated in order to examine the potential of NIRS to detect microbes on a food matrix. Inoculated samples were stored at 36 °C for 24 h. Two control groups were used in this study, one of fresh uninoculated samples analysed directly after removal of the muscle from the carcass and another of uninoculated chicken stored at 36 °C for 24 h. The findings of this study suggest the NIR spectroscopy can detect and differentiate between inoculated and uninoculated chicken breast muscle. In addition, it can discriminate fresh from non-fresh (in this case samples that are inoculated or not and stored at 36 °C) samples. On the other hand, NIR spectroscopy failed to differentiate between the five different strains used as inoculates for this study. These results suggest that the make up of the microflora of chicken fillets is not the deciding factor in detecting and differentiating chicken fillet samples. Instead, it seems that the total quantity of microflora present in conjunction with the physical changes associated with spoilage and, in particular, the differences in microbial concentration and level of muscle decomposition of inoculated and uninoculated samples, may be responsible for the separations observed in this analysis. This reason could also be the deciding factor for the separations achieved between the fresh samples and the other two groups.

The size-dependent bioavailability of hematite (α-Fe(2)O(3)) nanoparticles to obligate aerobic Pseudomonas mendocina bacteria was examined using the natural siderophore-producing wild type strain and a siderophore(-) mutant strain. Results showed that Fe from hematite less than a few tens of nm in size appears to be considerably more bioavailable than Fe associated with larger particles. This increased bioavailability is related to the total available particle surface area, and depends in part on greater accessibility of the Fe to the chelating siderophore(s). Greater bioavailability is also related to mechanism(s) that depend on cell/nanomineral proximity, but not on siderophores. Siderophore(-) bacteria readily acquire Fe from particles

Site-directed saturation mutagenesis is a convenient method to fine tune enzyme activity and selectivity at known “hot spots”. The objective of this work was to investigate the influence of mutations in the tunnel entrance of toluene 4-monooxygenase (T4MO) from Pseudomonas mendocina KR1 on rate, regioselectivity, and enantio-selectivity, in comparison with mutations located in the active site. Residue TmoA D285, located at the tunnel entrance, distant from the active site was randomized, as well as residue I100 which is positioned in the vicinity of the diiron center. Both libraries were screened for regioselective hydroxylation of phenylethanol (PEA), and enantioselective oxidation of methyl p-tolyl sulfide and styrene. It was discovered that mutations at position 285 enhanced the rate of catalysis but did not affect the specificity. Substitutions D285I and D285Q were beneficial for the oxidation of the large and bulky substrates PEA and methyl p-tolyl sulfide (8–11-fold improvement), however variant D285S improved the activity rate for styrene oxidation (1.7-fold). Thus, the extent of improvement in rate, and the best amino acid replacement were substrate dependant. In contrast, mutations at position I100 influenced the activity rate and the selectivity for all three substrates tested. Combining the effects by mutating both “gates”, to the tunnel entrance along with the active site, resulted in an additive or even synergistic outcome on enzyme activity and selectivity.

A strain of Pseudomonas mendocina producing extracellular lipase was isolated from soil. The bacterium accumulates lipase in culture fluid when grown aerobically at 30 °C for 24 h in a medium composed of olive oil (1%) as substrate. Pseudomonas mendocina lipase was optimally active at pH 9.0, temperature of 50 °C and was found to be stable between pH 7.0-11.0. The lipase was inhibited by detergents such as SDS and Tween-80. The enzyme was stable in various organic solvents tested with maximum stability in chloroform followed by toluene and exhibited 1-3 regiospecificity for hydrolytic reaction. This lipase was capable of hydrolyzing a variety of lipidic substrates and is mainly active towards synthetic triglycerides and fatty acid esters that possess a butyryl group. Metal ions like Mg(2+), Ca(2+) and Na(+) stimulated lipase activity, whereas, Cu(2+), Mn(2+) and Hg(2+) ions caused inhibition.

Purpose In this study, we investigated the effects of elevated atmospheric CO2, drought, and inoculation with a plant-growth-promoting rhizobacterium (PGPR) or an arbuscular mycorrhizal (AM) fungus on microbial community composition and functional diversity in the rhizosphere of Lactuca sativa L. cv. Tafalla.

Three species of phosphate solubilizing bacteria viz, Pseudomonas mendocina Khsr2, Pseudomonas stutzeri Khsr3 and Pseudomonas putida Khsr4 were isolated from roots of weeds Lactuca dissecta D. Don, Solanum surattense Burm. f and Sonchus arvensis L. respectively growing in Khewra salt range (EC: 2.3 dS m−1; pH 8.6). Preliminary identification of bacterial isolate was made on the basis of morphological and biochemical characters and confirmed by partial 16S-rRNA gene sequencing. The genetic diversity among the isolates was evaluated by Randomly Amplified Polymorphic DNA finger printing and similarity matrix was measured. All the Pseudomonas sp. were capable of solubilizing phosphate, produced phytohormones: Indole-3-acetic acid, Gibberellic acid, Trans-zeatin riboside and Abscisic acid in culture media and were found to be efficient in stimulating root/shoot length and dry weight and proline contents of Zea mays L (advance germplasm line: Islamabad Gold) seedlings grown under normal and NaCl (20 dS m−1) stress. The strain Pseudomonas stutzeri Khsr3 appears to be a potential candidate as bio-inoculant for saline fields.

In the article the results of investigation of oxidation processes of crude-oil by bacteria of genus Pseudomonas: Pseudomonas mendocina H-3, Pseudomonas sp. H-7, Pseudomonas stutzeri H-10, Pseudomonas aeruginosa H-14, Pseudomonas alcaligenes H-15 and Pseudomonas sp H-16. It was shown that these microorganisms are capable to oxidize oil’s hydrocarbons and oil products. It means that the bacteria may be used for bioremediation of oil-pollutant soils.

Pseudomonas mendocina KR1 oxidizes the gasoline oxygenate methyl tertiary butyl ether (MTBE) to tertiary butyl alcohol (TBA) during growth on C5 -C8 n-alkanes. We have further explored oxidation of ether oxygenates by this strain to help identify the enzyme that catalyses these reactions. High levels of MTBE-oxidizing activity occurred in resting cells grown on C5 -C8 n-alkanes. Lower activities occurred in cells grown on longer-chain n-alkanes (C9 -C11 ) and 1°-alcohols (C5 -C10 ). N-octane-grown cells also oxidized tertiary amyl methyl ether (TAME) to tertiary amyl alcohol (TAA), but did not oxidize ethyl tertiary butyl ether (ETBE), TBA or TAA. A 39 kDa polypeptide in whole cell extracts of n-octane-grown cells strongly cross-reacted with an anti-AlkB polyclonal antiserum in an SDS-PAGE/immunoblot. This polypeptide was absent or less abundant in cells grown on dextrose, dextrose plus dicyclopropylketone or 1-octanol. N-octane-grown cells of Pseudomonas aeruginosa strains KSLA-473 and ATCC 17423 oxidized MTBE and TAME but not ETBE. N-hexadecane-grown cells of these strains and strain PAO1 did not oxidize any of the oxygenates tested. Our results indicate ether oxygenate-degrading activity in alkane-utilizing pseudomonads is consistently observed with close homologues of the GPo1 non-haem-iron alkane hydroxylases but is otherwise not a consistent catalytic feature of these diverse enzymes.