Characterization of the extended substrate spectrum of the class A ß-lactamase CESS-1 from Stenotrophomonas sp. and structure-based investigation into its substrate preference.

OBJECTIVES: Stenotrophomonas spp. intrinsically resistant to many ß-lactam antibiotics are found throughout the environment. CESS-1 identified in Stenotrophomonas sp. KCTC 12332 is an uncharacterized class A ß-lactamase. The goal of this study was to reveal biochemical and structural characteristics of CESS-1. METHODS: The hydrolytic activities of CESS-1 towards penicillins (penicillin G and ampicillin), cephalosporins (cephalexin, cefaclor, and cefotaxime), and carbapenems (imipenem and meropenem) was spectrophotometrically monitored. Structural information on E166Q mutants of CESS-1 acylated by cefaclor, cephalexin, or ampicillin were determined by X-ray crystallography. RESULTS: CESS-1 displayed hydrolytic activities toward penicillins and cephalosporins, with negligible activity toward carbapenems. Although cefaclor, cephalexin, and ampicillin have similar structures with identical R1 side chains, the catalytic parameters of CESS-1 toward them were distinct. The kcat values for cefaclor, cephalexin, and ampicillin were 1249.6 s-1, 204.3 s-1, and 69.8 s-1, respectively, with the accompanying KM values of 287.6 µM, 236.7 µM, and 28.8 µM, respectively. CONCLUSIONS: CESS-1 was able to discriminate between cefaclor and cephalexin with a single structural difference at C3 position: -Cl (cefaclor) and -CH3 (cephalexin). Structural comparisons among three E166Q mutants of CESS-1 acylated by cefaclor, cephalexin, or ampicillin, revealed that cooperative positional changes in the R1 side chain of substrates and their interaction with the ß5-ß6 loop affect the distance between Asn170 and the deacylating water at the acyl-enzyme intermediate state. This is directly associated with the differential hydrolytic activities of CESS-1 toward the three structurally similar ß-lactam antibiotics.

Stenotrophomonas cyclobalanopsidis sp. nov., isolated from the leaf spot disease of Cyclobalanopsis patelliformis.

A Gram-negative, facultatively anaerobic, motile bacterial strain, TPQG1-4T, was isolated from the leaf of Cyclobalanopsis patelliformis with spot disease. The isolate was investigated using the polyphasic taxonomic approach. 16S rRNA gene sequencing and analyzing revealed that the novel strain shares the highest sequence similarity with Stenotrophomonas lactitubi M15T (99.6%), Stenotrophomonas indicatrix WS40T (99.4%), Stenotrophomonas maltophilia IAM 12423T (99.2%) and Stenotrophomonas pavanii LMG 25348T (99.0%). In phylogenetic trees based on 16S rRNA gene sequences, the novel strain branched independently from other species of Stenotrophomonas. Average nucleotide identity values between the novel isolate and S. lactitubi M15T, S. indicatrix WS40T, S. maltophilia IAM 12423T, S. pavanii LMG 25348T, and Pseudomonas geniculata ATCC 19374T were 87.2%, 87.3%, 86.3%, 88.0%, and 81.3%, respectively, suggesting the isolate was a novel species of the genus Stenotrophomonas. The DNA G + C content of TPQG1-4T is 67.1 mol%. The major fatty acids were iso-C15:0 (25.4%) and anteiso-C15:0 (17.0%). The polar lipids of TPQG1-4T included phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, amino phospholipid and phospholipid. Based on phenotypic and genotypic characteristics, the strain represents a novel species in the genus Stenotrophomonas, for which the name Stenotrophomonas cyclobalanopsidis sp. nov. is proposed. The type strain is TPQG1-4T (= CFCC 15341T = LMG 31208T).

Production, purification and evaluation of biodegradation potential of PHB depolymerase of Stenotrophomonas sp. RZS7.

There are numerous reports on poly-ß-hydroxybutyrate (PHB) depolymerases produced by various microorganisms isolated from various habitats, however, reports on PHB depolymerase production by an isolate from plastic rich sites scares. Although PHB has attracted commercial significance, the inefficient production and recovery methods, inefficient purification of PHB depolymerase and lack of ample knowledge on PHB degradation by PHB depolymerase have hampered its large scale commercialization. Therefore, to ensure the biodegradability of biopolymers, it becomes imperative to study the purification of the biodegrading enzyme system. We report the production, purification, and characterization of extracellular PHB depolymerase from Stenotrophomonas sp. RZS7 isolated from a dumping yard rich in plastic waste. The isolate produced extracellular PHB depolymerase in the mineral salt medium (MSM) at 30°C during 4 days of incubation under shaking. The enzyme was purified by three methods namely ammonium salt precipitation, column chromatography, and solvent purification. Among these purification methods, the enzyme was best purified by column chromatography on the Octyl-Sepharose CL-4B column giving optimum yield (0.7993 Umg-1mL-1). The molecular weight of purified PHB depolymerase was 40 kDa. Studies on the assessment of biodegradation of PHB in liquid culture medium and under natural soil conditions confirmed PHB biodegradation potential of Stenotrophomonas sp. RZS7. The results obtained in Fourier-Transform Infrared (FTIR) analysis, High-Performance Liquid Chromatography (HPLC) study and Gas Chromatography Mass-Spectrometry (GC-MS) analysis confirmed the biodegradation of PHB in liquid medium by Stenotrophomonas sp. RZS7. Changes in surface morphology of PHB film in soil burial as observed in Field Emission Scanning Electron Microscopy (FESEM) analysis confirmed the biodegradation of PHB under natural soil environment. The isolate was capable of degrading PHB and it resulted in 87.74% biodegradation. A higher rate of degradation under the natural soil condition is the result of the activity of soil microbes that complemented the biodegradation of PHB by Stenotrophomonas sp. RZS7.

Endophytic Bacteria Potentially Promote Plant Growth by Synthesizing Different Metabolites and their Phenotypic/Physiological Profiles in the Biolog GEN III MicroPlateTM Test.

Endophytic bacteria, as the most promising components of effective, biofertilizers biostimulating and biocontrol preparations, should be very intensively obtained from various plants and studied in terms of the conditions determining the potential ability to promote plant growth. For this reason, endophytic bacteria have been isolated from both stems and roots of up to six systematically distant species of vascular plants: one species belonging to the seedless vascular plants (Monilophyta), and five seed plants (Spermatophyta). The 23 isolated strains represented nine genera: Delftia, Stenotrophomonas, Rhizobium, Brevundimonas, Variovorax, Achromobacter, Novosphingobium, Comamonas and Collimonas, notably which were closely related-belonging to the phylum Proteobacteria. Stenotrophomonas sp. strains showed the greatest ability to synthesize indole-3-acetic acid (IAA)-like compounds, while Achromobacter sp. strains produced the highest levels of siderophores. The presence of the nifH gene and nitrogen binding activity was demonstrated for 95% of the strains tested. Stenotrophomonas maltophila (ES2 strain) showed the highest metabolic activity based on Biolog GEN III test. The ability to solubilize phosphate was determined only for three tested strains from genus: Delftia, Rhizobium and Novosphingobium. The presented work demonstrated that the metabolic and phenotypic properties of plant growth-promoting endophytes are correlated with the genus of bacteria and are not correlated with the host plant species or part of plant (stem, root).

Microbial antimonate reduction with a solid-state electrode as the sole electron donor: A novel approach for antimony bioremediation.

The anaerobic antimonate [Sb(V)] reduction with a solid-state electrode serving as the sole electron donor was demonstrated by employing a bioelectrochemical system. The highest Sb(V) reduction efficiency was observed at the biocathode potential of -0.7 V versus standard hydrogen electrode using a cathode potential range from -0.5 V to -1.1 V. The scanning electron microscopy and energy dispersive X-ray spectroscopy indicated that both amorphous and crystallized Sb2O3 were formed as products of Sb(V) reduction. The irreversible recovery of bioelectrochemical Sb(V), when the cathode potential deviated from the optimal potential, was explained through the alteration in microbial communities, which was further elucidated by the next-generation sequencing of 16S rRNA gene amplicons. Chryseobacterium koreense and Stenotrophomonas nitritireducens were the dominant species of microbial consortia at Sb(V)-reducing biocathodes. This study revealed a novel option for bioremediation of Sb at underground contaminated sites, where the delivery of organic electron donors is limited or ineffective.

Volatiles of rhizobacteria Serratia and Stenotrophomonas alter growth and metabolite composition of Arabidopsis thaliana.

The emission of volatiles is a common, but mostly neglected, ability of bacteria that is important for inter- and intraspecific interactions. Currently, limited information is available on how the bacterial volatile (mVOC) signal is integrated into a plant's life at the physiological, transcriptional and metabolic level. Previous results provided evidence for volatile-dependent regulation of WRKY18, a pathogen-responsive transcription factor of Arabidopsis thaliana in co-culture with two rhizobacteria, Serratia plymuthica HRO-C48 and Stenotrophomonas maltophilia R3089. Dual cultures of these bacteria and A. thaliana; application of the common mVOC 2-phenyl-ethanol; extraction of metabolites of A. thaliana after exposure to bacterial volatiles; and analysis of the metabolomes (GC-TOF/MS) were carried out. The prominent microbial aromatic compound 2-phenyl-ethanol, emitted by both bacteria, negatively affects growth of A. thaliana wild type, whereas WRKY18 T-DNA insertion mutants were significantly more tolerant than wild-type seedlings. This paper also demonstrates for the first time the impact of the rhizobacterial volatiles on the metabolome of A. thaliana. Upon mVOC exposure the plants rearrange their metabolism by accumulation of e.g. amino acids and TCA intermediates that potentially allow plants to cope with and survive this stress. Our findings illustrate the high degree of complexity of metabolic rearrangements underlying the interactions of bacterial volatile elicitors and resulting plant responses. Furthermore, the impact of the volatile 2-phenyl-ethanol as a signal in the WRKY18-dependent pathway highlights this compound as an important molecular player.

Ectoine: a compatible solute in radio-halophilic Stenotrophomonas sp. WMA-LM19 strain to prevent ultraviolet-induced protein damage.

AIM: Thiss study was conducted to investigate the possible role of a compatible solute from radio-halophilic bacterium against desiccation and ultra-violet radiation-induced oxidative stress. METHODS AND RESULTS: Nine different radio-resistant bacteria were isolated from desert soil, where strain WMA-LM19 was chosen for detailed studies on the basis of its high tolerance to ultraviolet radiation among all these isolates. Here, 16S rRNA gene sequencing indicated the bacterium was closely related to Stenotrophomonas sp. (KT008383). A bacterial milking strategy was applied for extraction of intracellular compatible solutes in 70% (v/v) ethanol, which were purified by High Performance Liquid Chromatography (HPLC). The compound was characterized as ectoine by 1 H and 13 C Nuclear Magnetic Resonance (NMR), and Mass Spectrometry (MS). Ectoine inhibited oxidative damage to proteins and lipids in comparison to the standard ascorbic acid. It also demonstrated more efficient prevention (54·80%) against lysis to erythrocytes membrane by surface active agents than lecithin. Furthermore, a high level of ectoine-mediated protection of bovine serum albumin against ionizing radiation (1 500-2 000Jm-2 ) was observed, as indicated by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis. CONCLUSION: The results indicated that ectoine from Stenotrophomonas sp. WMA-LM19 can be used as a potential mitigator and radio-protective agent to overcome radiation- and salinity-mediated oxidative damages in extreme environment. SIGNIFICANCE AND IMPACT OF THE STUDY: Due to its anti-oxidant properties, ectoine from a radio-halophilic bacterium might be used in sunscreen formulation for protection against UV-induced oxidative stress.

Subsurface Endospore-Forming Bacteria Possess Bio-Sealant Properties.

Concrete is a strong and fairly inexpensive building substance, but has several disadvantages like cracking that allows corrosion, thus reducing its lifespan. To mitigate these complications, long-lasting microbial self-healing cement is an alternative that is eco-friendly and also actively repairs cracks. The present paper describes the detailed experimental investigation on compressive strength of cement mortars, mixed with six alkaliphilic bacteria, isolated from subsurface mica mines of high alkalinity. The experiments showed that the addition of alkaliphilic isolates at different cell concentrations (104 and 106 cells/ml) enhanced the compressive strength of cement mortar, because the rapid growth of bacteria at high alkalinity precipitates calcite crystals that lead to filling of pores and densifying the concrete mix. Thus, Bacillus subtilis (SVUNM4) showed the highest compressive strength (28.61%) of cement mortar at 104 cells/ml compared to those of other five alkaliphilic isolates (Brevibacillus sp., SVUNM15-22.1%; P. dendritiformis, SVUNM11-19.9%; B. methylotrophicus, SVUNM9-16%; B. licheniformis, SVUNM14-12.7% and S. maltophilia, SVUNM13-9.6%) and controlled cement mortar as well. This method resulted in the filling of cracks in concrete with calcite (CaCO3), which was observed by scanning electron microscopy (SEM). Our results showed that the alkaliphilic bacterial isolates used in the study are effective in self-healing and repair of concrete cracks.

Identification of biphenyl 2, 3-dioxygenase and its catabolic role for phenazine degradation in Sphingobium yanoikuyae B1.

Phenazines are important nitrogen-containing secondary metabolites that display a range of biological functionalities. However, these compounds have shown lethal effects on humans and, the fate of phenazine in the ecosystem remains uncertain. In this study, we investigated that Sphingobium yanoikuyae B1 could utilize phenazine as a sole carbon source for growth. Intermediate produced during phenazine degradation was purified and identified as 1, 2-dihydrogen 1, 2-dihydroxy phenazine. Biphenyl 2, 3-dioxygenase was determined to be the initial dioxygenase for phenazine degradation through gene cloning and whole cell transformation techniques. Phenazine was converted to 1, 2-dihydrogen 1, 2-dihydroxy phenazine through hydrogenation and hydroxylation, which then transformed to 2-hydroxy phenazine through spontaneous dehydration. ThebphA1fA2f, were evidenced to be the only genes encoding the initial dioxygenase for phenazine degradation. BphB (dihydrodiol dehydrogenase) and BphC (2,3-dihydroxybiphenyl 1,2-dioxygenase) did not exhibit any 1, 2-dihydrogen 1, 2-dihydroxy phenazine and 1, 2-dihydroxy phenazine degradation capability, suggesting no contribution in phenazine degradation. Phylogenetic analysis of the dioxygenases demonstrated enormous biodegradation potential in strain B1. In conclusion, this study opens up new possibilities in better understanding the phenazine degradation in the environment.

Biodegradation and dissolution of polyaromatic hydrocarbons by Stenotrophomonas sp.

The aim of this work was to study the biodegradation capabilities of a locally isolated bacterium, Stenotrophomonas sp. strain IITR87 to degrade the polycyclic aromatic hydrocarbons and also check the preferential biodegradation of polycyclic aromatic hydrocarbons (PAHs). From preferential substrate degradation studies, it was found that Stenotrophomonas sp. strain IITR87 first utilized phenanthrene (three membered ring), followed by pyrene (four membered ring), then benzo[α]pyrene (five membered ring). Dissolution study of PAHs with surfactants, rhamnolipid and tritonX-100 showed that the dissolution of PAHs increased in the presence of surfactants.

Astaxanthin preparation by fermentation of esters from Haematococcus pluvialis algal extracts with Stenotrophomonas species.

Natural astaxanthin (Ax) is an additive that is widely used because of its beneficial biochemical functions. However, the methods used to produce free Ax have drawbacks. Chemical saponification methods produce several by-products, and lipase-catalyzed hydrolysis methods are not cost effective. In this study, a bacterial strain of Stenotrophomonas sp. was selected to enzymatically catalyze the saponification of Ax esters to produce free all-trans-Ax. Through single-factor experiments and a Box-Behnken design, the optimal fermentation conditions were determined as follows: a seed culture age of 37.79 h, an inoculum concentration of 5.92%, and an initial broth pH of 6.80. Under these conditions, a fermentation curve was drawn, and the optimal fermentation time was shown to be 60 h. At 60 h, the degradation rate of the Ax esters was 98.08%, and the yield of free all-trans-Ax was 50.130 µg/mL. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:649-656, 2016.

Enhancement of the catalytic efficiency and thermostability of Stenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF.

In this study, we enhanced the catalytic efficiency and thermostability of keratinase KerSMD by replacing its N/C-terminal domains with those from a homologous protease, KerSMF, to degrade feather waste. Replacement of the N-terminal domain generated a mutant protein with more than twofold increased catalytic activity towards casein. Replacement of the C-terminal domain obviously improved keratinolytic activity and increased the k(cat)/K(m) value on a synthetic peptide, succinyl-Ala-Ala-Pro-Phe-p-nitroanilide, by 54.5%. Replacement of both the N- and C-terminal domains generated a more stable mutant protein, with a Tm value of 64.60 ± 0.65°C and a half-life of 244.6 ± 2 min at 60°C, while deletion of the C-terminal domain from KerSMD or KerSMF resulted in mutant proteins exhibiting high activity under mesophilic conditions. These findings indicate that the pre-peptidase C-terminal domain and N-propeptide are not only important for substrate specificity, correct folding and thermostability but also support the ability of the enzyme to convert feather waste into feed additives.

Indirect Manganese Removal by Stenotrophomonas sp. and Lysinibacillus sp. Isolated from Brazilian Mine Water.

Manganese is a contaminant in the wastewaters produced by Brazilian mining operations, and the removal of the metal is notoriously difficult because of the high stability of the Mn(II) ion in aqueous solutions. To explore a biological approach for removing excessive amounts of aqueous Mn(II), we investigated the potential of Mn(II) oxidation by both consortium and bacterial isolates from a Brazilian manganese mine. A bacterial consortium was able to remove 99.7% of the Mn(II). A phylogenetic analysis of isolates demonstrated that the predominant microorganisms were members of Stenotrophomonas, Bacillus, and Lysinibacillus genera. Mn(II) removal rates between 58.5% and 70.9% were observed for Bacillus sp. and Stenotrophomonas sp. while the Lysinibacillus isolate 13P removes 82.7%. The catalytic oxidation of Mn(II) mediated by multicopper oxidase was not properly detected; however, in all of the experiments, a significant increase in the pH of the culture medium was detected. No aggregates inside the cells grown for a week were found by electronic microscopy. Nevertheless, an energy-dispersive X-ray spectroscopy of the isolates revealed the presence of manganese in Stenotrophomonas sp. and Lysinibacillus sp. grown in K medium. These results suggest that members of Stenotrophomonas and Lysinibacillus genera were able to remove Mn(II) by a nonenzymatic pathway.

Production of 10-hydroxy-12,15(Z,Z)-octadecadienoic acid from α-linolenic acid by permeabilized Stenotrophomonas nitritireducens cells.

OBJECTIVE: To improve the production of 10-hydroxy-12,15(Z,Z)-octadecadienoic acid (HODA) from α-linolenic acid in Stenotrophomonas nitritireducens. RESULTS: Cells of the bacterium were permeabilized with 1.25% (v/v) methanol. The optimal conditions for HODA production by permeabilized cells were pH 7, 35 °C, 5% (v/v) DMSO, 50 g cells l(-1), and 22.5 g α-linolenic acid l(-1). Under these conditions, permeabilized cells produced 16.4 g HODA l(-1) after 2 h, with a conversion yield of 73 % (w/w) and a volumetric productivity of 8.2 g l(-1) h(-1). These values were 153 and 230 % of the values for non-permeabilized cells CONCLUSIONS: This is the highest concentration and volumetric and specific productivities of HODA reported thus far.

Quantitative analysis of volatile metabolites released in vitro by bacteria of the genus Stenotrophomonas for identification of breath biomarkers of respiratory infection in cystic fibrosis.

The aim of the present study was to characterize the volatile metabolites produced by genotypically diverse strains of the Stenotrophomonas genus in order to evaluate their potential as biomarkers of lung infection by non-invasive breath analysis. Volatile organic compounds (VOCs) emitted from 15 clinical and five environmental strains belonging to different genogroups of Stenotrophomonas maltophilia (n = 18) and Stenotrophomonas rhizophila (n = 2) cultured in Mueller-Hinton Broth (MHB) liquid media were analysed by gas chromatography mass spectrometry (GC-MS) and selected ion flow tube mass spectrometry (SIFT-MS). Several VOCs were detected in high concentration, including ammonia, propanol, dimethyl disulphide propanol and dimethyl disulphide. The GC-MS measurements showed that all 15 clinical strains produced similar headspace VOCs compositions, and SIFT-MS quantification showed that the rates of production of the VOCs by the genotypically distinct strains were very similar. All in vitro cultures of both the Stenotrophomonas species were characterised by efficient production of two isomers of methyl butanol, which can be described by known biochemical pathways and which is absent in other pathogens, including Pseudomonas aeruginosa. These in-vitro data indicate that methyl butanol isomers may be exhaled breath biomarkers of S. maltophilia lung infection in patients with cystic fibrosis.

Prevention of aflatoxin contamination by a soil bacterium of Stenotrophomonas sp. that produces aflatoxin production inhibitors.

A soil bacterium, designated strain no. 27, was found to produce aflatoxin-production inhibitors. The strain was identified as a species of the genus Stenotrophomonas, and was found to be closely related to Stenotrophomonas rhizophila. Two diketopiperazines, cyclo(L-Ala-L-Pro) and cyclo(L-Val-L-Pro), were isolated from the bacterial culture filtrate as main active components. These compounds inhibited aflatoxin production of Aspergillus parasiticus and Aspergillus flavus in liquid medium at concentrations of several hundred µM without affecting fungal growth. Both inhibitors inhibited production of norsorolinic acid, a biosynthetic intermediate involved in an early step of the aflatoxin biosynthetic pathway, and reduced the mRNA level of aflR, which is a gene encoding a key regulatory protein necessary for the expression of aflatoxin-biosynthetic enzymes. These results indicated that the inhibitors targets are present in early regulatory steps leading to AflR expression. Co-culture of strain no. 27 with aflatoxigenic fungi in liquid medium effectively suppressed aflatoxin production of the fungus without affecting fungal growth. Furthermore, application of the bacterial cells to peanuts in laboratory experiments and at a farmer's warehouse in Thailand by dipping peanuts in the bacterial cell suspension strongly inhibited aflatoxin accumulation. The inhibitory effect was dependent on bacterial cell numbers. These results indicated that strain no. 27 may be a practically effective biocontrol agent for aflatoxin control.

Delineation of Stenotrophomonas spp. by multi-locus sequence analysis and MALDI-TOF mass spectrometry.

The genus Stenotrophomonas is genetically and phenotypically heterogeneous. Of the nine species now accepted, only S. maltophilia is of clinical importance. Based on DNA-sequences of seven house keeping genes, it encompasses genogroups of DNA-similarity below 97% that predominantly comprise strains of environmental origin. Therefore, in order to unravel the uneven distribution of environmental isolates within genogroups and reveal genetic relationships within the genus, there is need for an easy and reliable approach for the identification and delineation of Stenotrophomonas spp. In this first study, a multi-locus sequence analysis (MLSA) with seven housekeeping genes (atpD, gapA, guaA, mutM, nuoD, ppsA and recA) was applied for analysis of 21 S. maltophilia of environmental origin, Stenotrophomonas spp. and related genera. The genotypic findings were compared with the results of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) analyses. Our MLSA provided reliable inter- and intra-species discrimination of all tested isolates that correlated with the MALDI-TOF mass spectrometry data. One distantly related genogroup of environmental S. maltophilia strains needs to be reclassified as S. rhizophila. However, there are still remaining delineated S. maltophilia genogroups of predominantly environmental origin. Our data provide further evidence that 'Pseudomonas'beteli is a heterotypic synonym of S. maltophilia. Based on MLSA and MALDI-TOF data, Stenotrophomonas sp. (DSM 2408) belongs to S. koreensis.

Degradative potential of Stenotrophomonas strain HPC383 having genes homologous to dmp operon.

A strain, Stenotrophomonas HPC383 is isolated from effluent treatment plant treating wastewater from pesticide industry; degrades various aromatic compounds (cresols, phenol, catechol, 4methyl-catechol and hydroquinone) and crude oil, as determined through HPLC and GC analysis. Culture HPC383 could degrade (%) various compounds (1 mM) from a mixture: phenol - 99, p-cresol - 100, 4-methylcatechol - 96 and hydroquinone - 43 within 48 h of incubation, whereas it took 7 days to degrade 94% of 0.5% crude oil. Gene locus dmpN, to identify phenol degrading capacity was determined by PCR followed by southern analysis. The sequenced DNA fragment exhibited 99% sequence similarity to phenol hydroxylase gene from Arthrobacter sp. W1 (FJ610336). Amino acid sequence analysis of phenol hydroxylase reveals it to belong to high-Ks (affinity constant) group. Application of HPC383 in bioremediation of aquatic and terrestrial sites contaminated with petrochemical has been suggested.

Genetic engineering of Stenotrophomonas strain YC-1 to possess a broader substrate range for organophosphates.

In this work, Stenotrophomonas sp. strain YC-1, a native soil bacterium that produces methyl parathion hydrolase (MPH), was genetically engineered to possess a broader substrate range for organophosphates (OPs). A surface anchor system derived from the truncated ice nucleation protein (INPNC) from Pseudomonas syringae was used to target organophosphorus hydrolase (OPH) onto the cell surface of strain YC-1, reducing the potential substrate uptake limitation. The surface localization of INPNC-OPH was verified by cell fractionation, Western blot, proteinase accessibility, and immunofluorescence microscopy. No growth inhibition was observed for the engineered strain, and suspended cultures retained almost 100% activity over a period of 2 weeks. Concomitant expression of OPH in strain YC-1 resulted in a recombinant strain capable of simultaneously degrading diethyl and dimethyl OPs. A mixture of six OP pesticides (0.2 mM each) could be degraded completely within 5 h. The broader substrate specificity in combination with the rapid degradation rate makes this engineered strain a promising candidate for in situ remediation of OP-contaminated sites.

Stenotrophomonas panacihumi sp. nov., isolated from soil of a ginseng field.

The study isolated a Gram-negative, rod-shaped, non-motile bacterium from the soil of a ginseng field in Daejeon, South Korea and characterized it to determine its taxonomic position. Phylogenetic analysis, based on the 16S rRNA gene sequence, revealed that strain MK06(T) belongs to the family Xanthomonadacea, and showed the highest degree of sequence similarity to Stenotrophomonas rhizophila e-p10(T) (98.6%), Xanthomonas campestris LMG 568T (98.0%), Stenotrophomonas maltophilia ATCC 1d3637(T) (97.3%), and Stenotrophomonas humi R-32729(T) (96.9%). Chemotaxonomic data revealed that strain MK06(T) possesses ubiquinone Q-8 as the predominant respiratory lipoquinone, which is common in the genus Stenotrophomonas, and that the predominant fatty acids were 15:0 iso (41.1%), 15:0 anteiso (12.6%), and 17:1 iso omega9c (8.6%). The results of physiological and biochemical tests clearly demonstrated that strain MK06(T) represents a distinct species and supported its affiliation with the genus Stenotrophomonas. Based on these data, MK06(T) (KCTC, 22893(T); JCM, 16536(T); KEMB, 9004-002(T)) should be classified as the type strain for a novel species, for which we propose the name Stenotrophomonas panacihumi sp. nov.