Mono-(2-Ethylhexyl) Phthalate Promotes Pro-Labor Gene Expression in the Human Placenta

Women exposed to phthalates during pregnancy are at increased risk for delivering preterm, but the mechanism behind this relationship is unknown. Placental corticotropin-releasing hormone (CRH) and cyclooxygenase-2 (COX-2) are key mediators of parturition and are regulated by the non-canonical NF-kB (RelB/p52) signaling pathway. In this study, we demonstrate that one of the major phthalate metabolites, mono-(2-ethylhexyl)-phthalate (MEHP), increased CRH and COX-2 mRNA and protein abundance in a dose-dependent manner in primary cultures of cytotrophoblast. This was coupled with an increase in nuclear import of RelB/p52 and its association with the CRH and COX-2 promoters. Silencing of NF-kB inducing kinase, a central signaling component of the non-canonical NF-kB pathway, blocked MEHP-induced upregulation of CRH and COX-2. These results suggest a potential mechanism mediated by RelB/p52 by which phthalates could prematurely induce pro-labor gene activity and lead to preterm birth.     

Degradation of Phthalate and Di-(2-Ethylhexyl)phthalate by Indigenous and Inoculated Microorganisms in Sludge-Amended Soil

The metabolism of phthalic acid (PA) and di-(2-ethylhexyl)phthalate (DEHP) in sludge-amended agricultural soil was studied with radiotracer techniques. The initial rates of metabolism of PA and DEHP (4.1 nmol/g [dry weight]) were estimated to be 731.8 and 25.6 pmol/g (dry weight) per day, respectively. Indigenous microorganisms assimilated 28 and 17% of the carbon in [¹⁴C]PA and [¹⁴C]DEHP, respectively, into microbial biomass. The rates of DEHP metabolism were much greater in sludge assays without soil than in assays with sludge-amended soil. Mineralization of [¹⁴C]DEHP to ¹⁴CO₂ increased fourfold after inoculation of sludge and soil samples with DEHP-degrading strain SDE 2. The elevated mineralization potential was maintained for more than 27 days. Experiments performed with strain SDE 2 suggested that the bioavailability and mineralization of DEHP decreased substantially in the presence of soil and sludge components. The microorganisms metabolizing PA and DEHP in sludge and sludge-amended soil were characterized by substrate-specific radiolabelling, followed by analysis of ¹⁴C-labelled phospholipid ester-linked fatty acids (¹⁴C-PLFAs). This assay provided a radioactive fingerprint of the organisms actively metabolizing [¹⁴C]PA and [¹⁴C]DEHP. The ¹⁴C-PLFA fingerprints showed that organisms with different PLFA compositions metabolized PA and DEHP in sludge-amended soil. In contrast, microorganisms with comparable ¹⁴C-PLFA fingerprints were found to dominate DEHP metabolism in sludge and sludge-amended soil. Our results suggested that indigenous sludge microorganisms dominated DEHP degradation in sludge-amended soil. Mineralization of DEHP and PA followed complex kinetics that could not be described by simple first-order equations. The initial mineralization activity was described by an exponential function; this was followed by a second phase that was described best by a fractional power function. In the initial phase, the half times for PA and DEHP in sludge-amended soil were 2 and 58 days, respectively. In the late phase of incubation, the apparent half times for PA and DEHP increased to 15 and 147 days, respectively. In the second phase (after more than 28 days), the half time for DEHP was 2.9 times longer in sludge-amended soil assays than in sludge assays without soil. Experiments with radiolabelled DEHP degraders suggested that a significant fraction of the ¹⁴CO₂ produced in long-term degradation assays may have originated from turnover of labelled microbial biomass rather than mineralization of [¹⁴C]PA or [¹⁴C]DEHP. It was estimated that a significant amount of DEHP with poor biodegradability and extractability remains in sludge-amended soil for extended periods of time despite the presence of microorganisms capable of degrading the compound (e.g., more than 40% of the DEHP added is not mineralized after 1 year).     

Heat Curing of a Sym Plasmid in a Fast-Growing Rhizobium sp. That Is Able to Nodulate Legumes and the Nonlegume Parasponia sp

Genes involved in nodulation of both legumes and the nonlegume Parasponia sp., as well as nitrogenase genes, reside on a large plasmid in a fast-growing Rhizobium sp. from Lablab purpureus. This plasmid can be cured by incubation at elevated temperatures and can be mobilized by the P1 group plasmid RP1::Tn501.     

Gamma Interferon Is a Major Suppressive Factor Produced by Activated Human Peripheral Blood Lymphocytes That Is Able To Inhibit Foamy Virus-Induced Cytopathic Effects

The activation of human peripheral blood lymphocytes by mitogens or by triggering the T-cell receptor with anti-CD3 antibodies leads to the production of a potent soluble inhibitory activity against foamy virus-induced cytopathic effects in vitro. The inhibitory activity acts in a species-specific manner. As a consequence, the isolation of foamy viruses from blood lymphocytes of infected humans is accelerated in a heterologous coculture system. Antibodies against gamma interferon (IFN-γ) are able to suppress most of the inhibitory activity, suggesting that IFN-γ is the dominant component.     

Isolation from Agricultural Soil and Characterization of a Sphingomonas sp. Able To Mineralize the Phenylurea Herbicide Isoproturon

A soil bacterium (designated strain SRS2) able to metabolize the phenylurea herbicide isoproturon, 3-(4-isopropylphenyl)-1,1-dimethylurea (IPU), was isolated from a previously IPU-treated agricultural soil. Based on a partial analysis of the 16S rRNA gene and the cellular fatty acids, the strain was identified as a Sphingomonas sp. within the α-subdivision of the proteobacteria. Strain SRS2 was able to mineralize IPU when provided as a source of carbon, nitrogen, and energy. Supplementing the medium with a mixture of amino acids considerably enhanced IPU mineralization. Mineralization of IPU was accompanied by transient accumulation of the metabolites 3-(4-isopropylphenyl)-1-methylurea, 3-(4-isopropylphenyl)-urea, and 4-isopropyl-aniline identified by high-performance liquid chromatography analysis, thus indicating a metabolic pathway initiated by two successive N-demethylations, followed by cleavage of the urea side chain and finally by mineralization of the phenyl structure. Strain SRS2 also transformed the dimethylurea-substituted herbicides diuron and chlorotoluron, giving rise to as-yet-unidentified products. In addition, no degradation of the methoxy-methylurea-substituted herbicide linuron was observed. This report is the first characterization of a pure bacterial culture able to mineralize IPU.     

A mono-2-ethylhexyl phthalate hydrolase from a Gordonia sp. that is able to dissimilate di-2-ethylhexyl phthalate

Gordonia sp. strain P8219, a strain able to decompose di-2-ethylhexyl phthalate, was isolated from machine oil-contaminated soil. Mono-2-ethylhexyl phthalate hydrolase was purified from cell extracts of this strain. This enzyme was a 32,164-Da homodimeric protein, and it effectively hydrolyzed monophthalate esters, such as monoethyl, monobutyl, monohexyl, and mono-2-ethylhexyl phthalate. The K(m) and V(max) values for mono-2-ethylhexyl phthalate were 26.9 +/- 4.3 microM and 18.1 +/- 0.9 micromol/min . mg protein, respectively. The deduced amino acid sequence of the enzyme exhibited less than 30% homology with those of meta-cleavage hydrolases which are serine hydrolases but exhibited no significant homology with the sequences of serine esterases. The pentapeptide motif GXSXG, which is conserved in serine hydrolases, was present in the sequence. The enzymatic properties and features of the primary structure suggested that this enzyme is a novel enzyme belonging to an independent group of serine hydrolases.     

Gordonia sinesedis sp. nov., a novel soil isolate

The taxonomic position of an actinomycete isolated from soil was evaluated using a polyphasic approach. The organism, strain J72, was found to have chemical and morphological properties consistent with its assignment to the genus Gordonia. A nearly complete 16S rDNA sequence of the strain was determined by direct sequencing of the amplified gene. The tested strain formed a distinct phylogenetic line within the evolutionary radiation occupied by the genus Gordonia and was most closely related to G. polyisoprenivorans DSM 44302^T. The phenotypic profile of strain 372 readily distinguishes it from representatives of the validly described species of Gordonia. The combined genotypic and phenotypic data show that strain J72 merits recognition as a new species of Gordonia. The name proposed for the new species is Gordonia sinesedis; the type strain is J72^T (= DSM 44455^T = NCIMB 13802^T). This revised version was published online in June 2006 with corrections to the Cover Date.     

A Novel Cellulosomal Scaffoldin from Acetivibrio cellulolyticus That Contains a Family 9 Glycosyl Hydrolase

A novel cellulosomal scaffoldin gene, termed cipV, was identified and sequenced from the mesophilic cellulolytic anaerobe Acetivibrio cellulolyticus. Initial identification of the protein was based on a combination of properties, including its high molecular weight, cellulose-binding activity, glycoprotein nature, and immuno-cross-reactivity with the cellulosomal scaffoldin of Clostridium thermocellum. The cipV gene is 5,748 bp in length and encodes a 1,915-residue polypeptide with a calculated molecular weight of 199,496. CipV contains an N-terminal signal peptide, seven type I cohesin domains, an internal family III cellulose-binding domain (CBD), and an X2 module of unknown function in tandem with a type II dockerin domain at the C terminus. Surprisingly, CipV also possesses at its N terminus a catalytic module that belongs to the family 9 glycosyl hydrolases. Sequence analysis indicated the following. (i) The repeating cohesin domains are very similar to each other, ranging between 70 and 90% identity, and they also have about 30 to 40% homology with each of the other known type I scaffoldin cohesins. (ii) The internal CBD belongs to family III but differs from other known scaffoldin CBDs by the omission of a 9-residue stretch that constitutes a characteristic loop previously associated with the scaffoldins. (iii) The C-terminal type II dockerin domain is only the second such domain to have been discovered; its predicted "recognition codes" differ from those proposed for the other known dockerins. The putative calcium-binding loop includes an unusual insert, lacking in all the known type I and type II dockerins. (iv) The X2 module has about 60% sequence homology with that of C. thermocellum and appears at the same position in the scaffoldin. (v) Unlike the other known family 9 catalytic modules of bacterial origin, the CipV catalytic module is not accompanied by a flanking helper module, e.g., an adjacent family IIIc CBD or an immunoglobulin-like domain. Comparative sequence analysis of the CipV functional modules with those of the previously sequenced scaffoldins provides new insight into the structural arrangement and phylogeny of this intriguing family of microbial proteins. The modular organization of CipV is reminiscent of that of the CipA scaffoldin from C. thermocellum as opposed to the known scaffoldins from the mesophilic clostridia. The phylogenetic relationship of the different functional modules appears to indicate that the evolution of the scaffoldins reflects a collection of independent events and mechanisms whereby individual modules and other constituents are incorporated into the scaffoldin gene from different microbial sources.     

Gordonia namibiensis sp. nov., a novel nitrile metabolising actinomycete recovered from an African sand

A polyphasic approach was used to establish the taxonomic position of two actinomycetes isolated from a Namibian soil and shown to utilise nitrile compounds as growth substrates. The organisms, strains NAM-BN063AT and NAM-BN063B, had chemical and morphological properties consistent with their assignment to the genus Gordonia. Direct 165 rRNA sequencing studies confirmed the taxonomic position of the strains following the generation of phylogenetic trees using four different algorithms. The strains consistently formed a distinct phylogenetic line within the evolutionary radiation occupied by gordoniae and were most closely related to Gordonia rubropertincta DSM 43197T. DNA:DNA relatedness studies indicated that the two organisms belonged to a genomic species that was readily distinguished from G. rubropertincta. The unique phenotypic profile of the strains sharply separated them from representatives of all of the validly described species of Gordonia. The combination of genotypic and phenotypic data indicates that the two strains should be classified in the genus Gordonia as a new species. The name proposed for this taxon is Gordonia namibiensis, the type strain is NAM-BN063AT (= DSM 44568T = NCIMB 13780T).     

Characterization of the Polyurethanolytic Activity of Two Alicycliphilus sp. Strains Able To Degrade Polyurethane and N-Methylpyrrolidone

Two bacterial strains (BQ1 and BQ8) were isolated from decomposed soft foam. These were selected for their capacity to grow in a minimal medium (MM) supplemented with a commercial surface-coating polyurethane (PU) (Hydroform) as the carbon source (MM-PUh). Both bacterial strains were identified as Alicycliphilus sp. by comparative 16S rRNA gene sequence analysis. Growth in MM-PUh showed hyperbolic behavior, with BQ1 producing higher maximum growth (17.8 ± 0.6 mg·ml⁻¹) than BQ8 (14.0 ± 0.6 mg·ml⁻¹) after 100 h of culture. Nuclear magnetic resonance, Fourier transform infrared (IR) spectroscopy, and gas chromatography-mass spectrometry analyses of Hydroform showed that it was a polyester PU type which also contained N-methylpyrrolidone (NMP) as an additive. Alicycliphilus sp. utilizes NMP during the first stage of growth and was able to use it as the sole carbon and nitrogen source, with calculated K_s values of about 8 mg·ml⁻¹. Enzymatic activities related to PU degradation (esterase, protease, and urease activities) were tested by using differential media and activity assays in cell-free supernatants of bacterial cultures in MM-PUh. Induction of esterase activity in inoculated MM-PUh, but not that of protease or urease activities, was observed at 12 h of culture. Esterase activity reached its maximum at 18 h and was maintained at 50% of its maximal activity until the end of the analysis (120 h). The capacity of Alicycliphilus sp. to degrade PU was demonstrated by changes in the PU IR spectrum and by the numerous holes produced in solid PU observed by scanning electron microscopy after bacterial culture. Changes in the PU IR spectra indicate that an esterase activity is involved in PU degradation.     

假单胞菌WBC-3甲基对硫磷水解酶性质的初步研究

从最近分离到的有机磷农药降解菌Pseudomonassp. WBC3中获得了甲基对硫磷水解酶（Methyl parathion hydrolase, MPH,EC 3183）。该酶在48h的培养物中分布比例分别为：上清液2.1%, 胞内862%和胞间质11.7%,说明MPH为胞内酶。经过Cmsepharose Fast Flow阳离子交换层析,获得电泳纯的酶。SDSPAGE和凝胶过滤层析表明,该酶为单体蛋白,分子量约为34kD。动力学分析显示该酶为非特异性有机磷降解酶,但最适底物为甲基对硫磷。在pH9～12范围,酶表现较高活力水平,最高活力的反应温度为40℃。根据各类金属离子和鳌合剂对酶活的影响,推测MPH为金属酶。     

Isolation and Characterization of Methanomethylovorans hollandica gen. nov., sp. nov., Isolated from Freshwater Sediment, a Methylotrophic Methanogen Able To Grow on Dimethyl Sulfide and Methanethiol

A newly isolated methanogen, strain DMS1^T, is the first obligately anaerobic archaeon which was directly enriched and isolated from a freshwater sediment in defined minimal medium containing dimethyl sulfide (DMS) as the sole carbon and energy source. The use of a chemostat with a continuous DMS-containing gas stream as a method of enrichment, followed by cultivation in deep agar tubes, resulted in a pure culture. Since the only substrates utilized by strain DMS1^T are methanol, methylamines, methanethiol (MT), and DMS, this organism is considered an obligately methylotrophic methanogen like most other DMS-degrading methanogens. Strain DMS1^T differs from all other DMS-degrading methanogens, since it was isolated from a freshwater pond and requires NaCl concentrations (0 to 0.04 M) typical of the NaCl concentrations required by freshwater microorganisms for growth. DMS was degraded effectively only in a chemostat culture in the presence of low hydrogen sulfide and MT concentrations. Addition of MT or sulfide to the chemostat significantly decreased degradation of DMS. Transient accumulation of DMS in MT-amended cultures indicated that transfer of the first methyl group during DMS degradation is a reversible process. On the basis of its low level of homology with the most closely related methanogen, Methanococcoides burtonii (94.5%), its position on the phylogenetic tree, its morphology (which is different from that of members of the genera Methanolobus, Methanococcoides, and Methanohalophilus), and its salt tolerance and optimum (which are characteristic of freshwater bacteria), we propose that strain DMS1^T is a representative of a novel genus. This isolate was named Methanomethylovorans hollandica. Analysis of DMS-amended sediment slurries with a fluorescence microscope revealed the presence of methanogens which were morphologically identical to M. hollandica, as described in this study. Considering its physiological properties, M. hollandica DMS1^T is probably responsible for degradation of MT and DMS in freshwater sediments in situ. Due to the reversibility of the DMS conversion, methanogens like strain DMS1^T can also be involved in the formation of DMS through methylation of MT. This phenomenon, which previously has been shown to occur in sediment slurries of freshwater origin, might affect the steady-state concentrations and, consequently, the total flux of DMS and MT in these systems.     

Streptococcus thermophilus Is Able To Produce a β-Galactosidase Active during Its Transit in the Digestive Tract of Germ-Free Mice

This work presents data on the application of a bacterial luciferase used to monitor gene expression of Streptococcus thermophilus in the digestive tract. The main result is that the bacterium was able to produce an active β-galactosidase in the digestive tract, although it did not multiply during its transit. This production was enhanced when lactose (the inducer) was added to the diet.     

Identification and Characterization of a Cold-Active Phthalate Esters Hydrolase by Screening a Metagenomic Library Derived from Biofilms of a Wastewater Treatment Plant

A cold-active phthalate esters hydrolase gene (designated dphB) was identified through functional screening of a metagenomic library derived from biofilms of a wastewater treatment plant. The enzyme specifically catalyzed the hydrolysis of dipropyl phthalate, dibutyl phthalate, and dipentyl phthalate to the corresponding monoalkyl phthalate esters at low temperatures. The catalytic triad residues of DphB were proposed to be Ser159, Asp251, and His281.     

Purification and Some Properties of Cyclo(Gly-Gly) Hydrolase from a Strain of Bacillus sp. No. 106

Bacillus sp. No. 106, which was isolated from soil, secreted an enzyme that hydrolyzed cyclo(Gly-Gly). The enzyme was purified to the ultracentrifugally homogeneous state and an activity more than 450-fold that of culture broth. The enzyme was activated by Na⁺, Mg²⁺, Ca²⁺, and Sr²⁺, and strongly inhibited by Ni²⁺, Cu²⁺, p-chloromercuribenzoate, and monoiodoacetic acid. The Km value for cyclo(Gly-Gly) was estimated to be 11.1 mm. The enzyme hydrolyzed only cyclo(Gly-Gly) among various diketopiperazines tested. Aslo, the enzyme was inert toward Gly-Gly, milk casein, and hemoglobin.     

Isolation and taxonomic study of a new canthaxanthin-containing bacterium, Gordonia jacobaea MV-1 sp. nov.

This article describes the isolation and taxonomic study of a coryneform isolate of a new Gordonia species (G. jacobaea), strain MV-1, which accumulates several carotenoids, including the ketocarotenoid trans-canthaxanthin. Identification of this new isolate by morphobiochemical methods did not allow unambiguous taxon assignment, but sequencing of the 16S rRNA gene clearly pointed to the genus Gordonia, Gordonia sputi being the closest fit. Differences in certain transversions/transitions in otherwise very well-conserved sequences of the described Gordonia species supported the proposal of this new taxon. The fact that both the best growth and best pigmentation were obtained with glucose, an inexpensive carbon source and at an industrially suitable temperature, suggests that this new bacterial strain may have good potential for the industrial production of canthaxanthin.     

Gordonia jinhuaensis sp. nov., a novel actinobacterium,isolated from a VBNC (viable but non-culturable) state in pharmaceutical wastewater

A Gram-stain positive, aerobic, non-motile and rod-shaped actinobacterial strain, designated as ZYR 51^T, was isolated from pharmaceutical wastewater in Jinhua city, Zhejiang province, Eastern China. Isolation was aided by using a resuscitation-promoting factor, suggesting the strain was recovered from a viable but non-culturable state. Strain ZYR 51^T was characterized by a polyphasic taxonomic approach. Growth was found to occur at 10–45 °C, pH 6.0–10.0 and 0–9 % NaCl (w/v). Based on the 16S rRNA gene sequence, phylogenetic analysis clearly demonstrated that strain ZYR 51^T belongs to the genus Gordonia and showed low level similarities (below 97 %) with all other members of this genus. The strain was found to possess meso-diaminopimelic acid (meso-DAP), along with MK-9(H₂) as the predominant menaquonine. Mycolic acids were found to be present. C_16:0 (34.9 %), 10-methyl C_18:0 (30.3 %), iso-C_18:0(8.2 %), and summed feature 3 (C_16:1 ω6c and/or C_16:1ω7c as define by MIDI; 18.8 %) were identified as the major cellular fatty acids. The polar lipid profile of strain ZYR 51^T was found to consist of diphosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannosides and some unknown lipids. The genomic DNA G+C content of strain ZYR 51^T was determined to be 67.7 mol%. The combined genotypic and phenotypic data showed that the strain represents a novel species of the genus Gordonia, for which the name Gordonia jinhuaensis sp. nov. is proposed, with the type strain is ZYR 51^T (=CGMCC 1.12827^T = NBRL B-59111^T = NBRC 110001^T).     

A Phytophthora sojae Glycoside Hydrolase 12 Protein Is a Major Virulence Factor during Soybean Infection and Is Recognized as a PAMP

We identified a glycoside hydrolase family 12 (GH12) protein, XEG1, produced by the soybean pathogen Phytophthora sojae that exhibits xyloglucanase and β-glucanase activity. It acts as an important virulence factor during P. sojae infection but also acts as a pathogen-associated molecular pattern (PAMP) in soybean (Glycine max) and solanaceous species, where it can trigger defense responses including cell death. GH12 proteins occur widely across microbial taxa, and many of these GH12 proteins induce cell death in Nicotiana benthamiana. The PAMP activity of XEG1 is independent of its xyloglucanase activity. XEG1 can induce plant defense responses in a BAK1-dependent manner. The perception of XEG1 occurs independently of the perception of ethylene-inducing xylanase. XEG1 is strongly induced in P. sojae within 30 min of infection of soybean and then slowly declines. Both silencing and overexpression of XEG1 in P. sojae severely reduced virulence. Many P. sojae RXLR effectors could suppress defense responses induced by XEG1, including several that are expressed within 30 min of infection. Therefore, our data suggest that PsXEG1 contributes to P. sojae virulence, but soybean recognizes PsXEG1 to induce immune responses, which in turn can be suppressed by RXLR effectors. XEG1 thus represents an apoplastic effector that is recognized via the plant’s PAMP recognition machinery.