Functional characterization of the transcription regulator WhiB1 from Gordonia sp. IITR100.

WhiB is a transcription regulator which has been reported to be involved in the regulation of cell morphogenesis, cell division, antibiotic resistance, stress, etc., in several members of the family Actinomycetes. The present study describes functional characterization of a WhiB family protein, WhiB1 (protein ID: WP_065632651.1), from Gordonia sp. IITR100. We demonstrate that WhiB1 affects chromosome segregation and cell morphology in recombinant Escherichia coli, Gordonia sp. IITR100 as well as in Rhodococcus erythropolis. Multiple sequence alignment suggests that WhiB1 is a conserved protein among members of the family Actinomycetes. It has been reported that overexpression of WhiB1 leads to repression of the biodesulfurization operon in recombinant E. coli, Gordonia sp. IITR100 and R. erythropolis. A WhiB1-mut containing a point mutation Q116A in the DNA binding domain of WhiB1 led to partial alleviation of repression of the biodesulfurization operon. We show for the first time that the WhiB family protein WhiB1 is also involved in repression of the biodesulfurization operon by directly binding to the dsz promoter DNA.

Methylation, sugar puckering and Z-form status of DNA from a heavy metal-acclimated freshwater Gordonia sp.

Heavy metal acclimation of bacteria is of particular interest in many aspects. It could add to our understanding of adaptation strategies applied by bacteria, as well as help us in devising ways to use such adaptive bacteria for bioremediation. In this study, we have explored the changes in the DNA of an aquatic Gordonia sp. acclimated to silver, cadmium, and lead. We have measured the changes in the DNA extracted from the acclimated bacteria by using ATR-FTIR coupled with unsupervised and supervised pattern recognition algorithms. Although whole-cell FTIR studies do reveal nucleic acid changes, the special care should be taken when considering marker nucleic acid bands in such spectra, as various other cell or tissue constituents also yield IR bands in the same region. An FTIR study on isolated DNA can be used to avoid this problem. The IR spectral profiles of the DNA molecules revealed significant changes in the backbone and sugar conformations of upon acclimation. We then further analyzed the DNA's global cytosine-methylation patterns of the heavy metal-acclimated bacteria. We aimed to find out whether epigenetic mechanisms operate in bacteria for survival and growth in inhibitory heavy metal concentrations or not. We found hypermethylation in Cd acclimation but hypomethylation for both Pb and Ag in Gordonia sp. Our results imply that changes in the conformational and methylation states of DNA seem to let bacteria to thrive in otherwise inhibitory conditions and mark the involvement of epigenetic modulation in acclimation processes.

Harnessing the catabolic versatility of Gordonia species for detoxifying pollutants.

The genus Gordonia includes variedly pigmented aerobic, non-motile, non-sporulating Gram positive (sometimes variable) coccoid forms and rods. Different isolates display distinguishing physiological traits and biochemical properties that are significant in remediation applications. Strains inherently prevalent in soils, seawater, sediments and wastewaters can degrade hydrocarbons. Immobilized cells and microbial consortia containing Gordonia species have been used for in situ applications. Hydrocarbon uptake in this Actinomycete is mediated by attachment to large droplets or by pseudosolubilization of substrates. Hydrocarbons so internalized are degraded by relevant enzymes that are innately present in this microorganism. Wild-type and recombinant strains also mediate desulfurization of polyaromatic sulfur heterocyclic compounds. This organism is metabolically capable of bringing about detoxification of phthalate esters. Two species namely, Gordonia polyisoprenivorans and Gordonia westfalica mediate degradation of rubber and the metabolic pathways involved in the process have been well-understood. Some members are able to transform nitriles into commercially valuable products and others degrade the explosive hexahydro-1,3,5-trinitro-1,3,5-triazine. Cholesterol, pyridine derivatives, fuel oxygenates, thiodiglycol, bis-(3-pentafluorophenylpropyl)-sulfide and 6:2 fluorotelomersulfonate are also biotransformed or degraded by Gordonia species. Some members of this genus are significant in the treatment of wastewaters including those that are rich in steroids and lignin. There are also several patents highlighting the commercial relevance of this genus. On account of its diverse catabolic properties, this Actinomycete has become important in bioremediation of polluted environments.

Analysis of desferrioxamine-like siderophores and their capability to selectively bind metals and metalloids: development of a robust analytical RP-HPLC method.

The Actinobacterium Gordonia rubripertincta CWB2 (DSM 46758) produces hydroxamate-type siderophores (188 mg L-1) under iron limitation. Analytical reversed-phase HPLC allowed determining a single peak of ferric iron chelating compounds from culture broth which was confirmed by the Fe-CAS assay. Elution profile and its absorbance spectrum were similar to those of commercial (des)ferrioxamine B which was used as reference compound. This confirms previously made assumptions and shows for the first time that the genus Gordonia produces desferrioxamine-like siderophores. The reversed-phase HPLC protocol was optimized to separate metal-free and -loaded oxamines. This allowed to determine siderophore concentrations in solutions as well as metal affinity. The metal loading of oxamines was confirmed by ICP-MS. As a result, it was demonstrated that desferrioxamine prefers trivalent metal ions (Fe3+ > Ga3+ > V3+ > Al3+) over divalent ones. In addition, we aimed to show the applicability of the newly established reversed-phase HPLC protocol and to increase the re-usability of desferrioxamines as metal chelators by immobilization on mesocellular silica foam carriers. The siderophores obtained from strain CWB2 and commercial desferrioxamine B were successfully linked to the carrier with a high yield (up to 95%) which was verified by the HPLC method. Metal binding studies demonstrated that metals can be bound to non-immobilized and to the covalently linked desferrioxamines, but also to the carrier material itself. The latter was found to be unspecific and, therefore, the effect of the carrier material remains a field of future research. By means of a reversed CAS assay for various elements (Nd, Gd, La, Er, Al, Ga, V, Au, Fe, As) it was possible to demonstrate improved Ga3+- and Nd3+-binding to desferrioxamine loaded mesoporous silica carriers. The combination of the robust reversed-phase HPLC method and various CAS assays provides new avenues to screen for siderophore producing strains, and to control purification and immobilization of siderophores.

Identification and characterization of a cell wall porin from Gordonia jacobaea.

Gordonia jacobaea is a bacterium belonging to the mycolata group characterized by its ability to produce carotenoids. Mycolic acids in the cell wall contribute to reducing the permeability of their envelopes requiring the presence of channel-forming proteins to allow the exchange of hydrophilic molecules with the surrounding medium. Identification and purification of the channel-forming proteins was accomplished by SDS-PAGE, Mass spectrometry and Mass peptide fingerprinting and the channel-forming activity was studied by reconstitution in lipid bilayers. Here, we describe for the first time the presence of a cell-wall protein from G. jacobaea with channel-forming activity. Our results suggest that this protein bears a low similarity to other hypothetical proteins from the genus Gordonia of uncharacterized functions. The channel has an average single-channel conductance of 800 pS in 1 M KCl, is moderately anion-selective, and does not show any voltage dependence for voltages between +100 and -100 mV. The channel characteristics suggest that this protein could be of relevance in the import and export of negatively charged molecules across the cell wall. This could contribute to design treatments for mycobacterial infections, as well as being of interest in biotechnology applications.

Gordonic Acid, a Polyketide Glycoside Derived from Bacterial Coculture of Streptomyces and Gordonia Species.

Despite numerous efforts to discover novel bioactive products from microorganisms, previously reported compounds are repetitively reisolated. A new polyketide glycoside, gordonic acid (1), isolated from the mixed culture of two Gram-positive bacteria, Gordonia sp. KMC005 and Streptomyces tendae KMC006, is reported. The structure of 1 was characterized as an acyclic polyene polyketide substituted with a ß-d-digitoxopyranose through NMR, HR-ESI-QTOF-MS, IR, and UV spectral data. The stereochemistry for 1 was determined by Mosher's method followed by 2D NOESY analysis and by NMR chemical shift calculations supported by DP4 analysis. Gordonic acid (1) showed weak activity against Micrococcus luteus and Enterococcus hirae.

Gordonia species as emerging causes of continuous-ambulatory-peritoneal-dialysis-related peritonitis identified by 16S rRNA and secA1 gene sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS).

We report here four cases of continuous ambulatory peritoneal dialysis-related peritonitis caused by three different species of Gordonia. The portal of entry was likely through Tenckhoff catheters. 16S rRNA and secA1 gene sequencing are so far the most reliable methods for the accurate identification of Gordonia species.

Amino acid-based biohybrids for nano-shellization of individual desulfurizing bacteria.

Amino acid-based biohybrids have been developed to self-assemble on the surface of desulfurizing bacteria to form nanothin and nanoporous shells. The shells not only endow the encapsulated cells with reusability, but also offer platforms to incorporate titania and magnetic nanoparticles to improve the desulfurizing activity and the separation efficiency.

[Mycolic acids--potential biomarkers of opportunistic infections caused by bacteria of the suborder Corynebacterineae]. / Kwasy mikolowe--potencjalne markery diagnostyki oportunistycznych zakazen mikroorganizmami z podrzedu Corynebacterineae.

Mycolic acids are one of the basic elements of the cell wall structure of bacteria belonging to the suborder Corynebacterineae, constituting from 20% to 40% of dry weight. Additionally, they show high structural diversity within each family and species. Nowadays, profiles of mycolic acids are widely described for the genus Mycobacterium, the causative agent of tuberculosis. However, the suborder Corynebacterineae also includes many representatives of opportunistic human pathogens, e.g. Dietzia, Gordonia, Nocardia and Rhodococcus. Currently, an increased infection risk caused by this group of microorganisms especially in immunocompromised patients has been observed. Better knowledge of mycolic acid profiles for Corynebacterineae may allow identification of mycolic acids as diagnostic markers in the detection of opportunistic bacterial infections. Modern techniques of chemical analysis, including mass spectrometry, may enable the development of new chemotaxonomic methods for the detection and differentiation of bacteria within the suborder Corynebacterineae.

Carotenoids in a Corynebacterineae, Gordonia terrae AIST-1: carotenoid glucosyl mycoloyl esters.

We isolated a strain of Corynebacterineae from surface seawater from the Inland Sea of Japan. This strain, AIST-1, was determined to be a strain of Gordonia terrae based on its 16S rRNA gene sequence. The colony was red-colored, and the pigments were identified to be carotenoid derivatives. The structures of two major carotenoids were (2'S)-deoxymyxol 1'-glucoside, a dihydroxyl derivative of gamma-carotene with 12 conjugated double bonds, and (2'S)-4-ketodeoxymyxol 1'-glucoside. Their glucosyl acyl esters and mycoloyl esters were also identified. While these carotenoid moieties have been found in only a few other bacteria, the carotenoid mycoloyl esters are novel carotenoid derivatives. The type strain of G. terrae NBRC 10016T also contained the same carotenoids, but the composition of the two carotenoid glucosides was low and the total carotenoid content was less than one tenth of that of strain AIST-1.

Metabolic cooperation of Gordonia sp. strain MTCC 4818 and Arthrobacter sp. strain WY in the utilization of butyl benzyl phthalate: effect of a novel co-culture in the degradation of a mixture of phthalates.

Degradation of butyl benzyl phthalate (BBP) by a co-culture of Gordonia sp. strain MTCC 4818 and Arthrobacter sp. strain WY was investigated. In the degradation of BBP by the co-culture, the limitations of the individual species in metabolizing BBP were overcome, leading to the development of a consortium capable of complete utilization of this ester. In the degradation of BBP by the co-culture, the presence of multiple esterases was demonstrated in both species by activity staining of non-denaturing gels, indicating their roles in the degradation process. The esterases were found to be inducible, with unique or broad substrate specificities towards BBP and its monoesters. Moreover, a number of catabolic enzymes other than esterases identified in the metabolism of BBP-degraded intermediates facilitated the co-culture-mediated degradation process. The versatility of the co-culture was further established by the rapid and complete degradation of a mixture of phthalate esters of environmental concern.

Exopolysaccharides produced by Gordonia alkanivorans enhance bacterial degradation activity for diesel.

Exopolysaccharides (EPS) produced by Gordonia alkanivorans CC-JG39 was used to stimulate cell floating, cell growth, and diesel biodegradation of indigenous or commercial-available, diesel-degrading bacteria. Addition of EPS-containing supernatant into the culture medium resulted in floatation of the non-floating bacteria and allowed a 40-45% and 38-42% increase in diesel degradation and cell growth, respectively. The EPS-stimulating effect on cell growth and diesel degradation positively correlated with the EPS dosage. Thus, the EPS may act as a biostimulant for bioremediation of oil-contaminated water or soil.

Surface-active compounds and their role in the access to hydrocarbons in Gordonia strains.

Three new bacterial strains (M22, BS25 and BS29) belonging to the Gordonia genus were isolated from a site chronically contaminated by diesel. Those Gordonia strains were able to grow using a wide range of straight and branched aliphatic hydrocarbons as carbon and energy sources and to produce at least two classes of surface-active compounds. Emulsifying agents were released in the culture medium when bacteria grew both on hydrocarbons and water-soluble substrates. Cell-bound biosurfactants, which reduce the surface tension, were produced on hydrocarbons; however, their production was significantly lower on water soluble substrates. The relationship of growth phase, surface-active compound production and cell-surface properties was analyzed in kinetic experiments on hydrocarbons. Gordonia sp. BS29 synthesized, and released extracellularly, bioemulsans during the exponential phase with n-hexadecane as carbon and energy source. The production of biosurfactants started in the exponential phase and their concentration increased during the following linear growth. Furthermore, the adhesion of bacterial cells to hydrocarbons decreased during growth. Our results led us to hypothesize a change in the mode by which Gordonia cells access the substrate during growth on hydrocarbons.

Isolation of Gordonia terrae from a patient with catheter-related bacteraemia.

A cornyeform bacterium was isolated from a blood culture from a 24-year-old man with familial hypertrophic non-obstructive cardiomyopathy, chronic abuse of anabolic steroids and prior admission to hospital because of clinical signs of sepsis. 16S rRNA gene analysis unambiguously identified Gordonia terrae.

Gordonia shandongensis sp. nov., isolated from soil in China.

The taxonomic position of strain SD29(T), isolated from soil, was clarified using a polyphasic taxonomic approach. The organism produced an elementary branching mycelium which fragmented into rod/coccus-shaped elements and it possessed meso-diaminopimelic acid, arabinose, galactose as diagnostic diamino acid and sugars, MK-9(H(2)) as predominant menaquinone, phospholipids of type PII and mycolic acid. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain SD29(T) was most closely related to Gordonia hydrophobica DSM 44015(T) and Gordonia sihwensis DSM 44576(T), forming a distinct but loosely related branch in the phylogenetic tree. A number of physiological properties readily separated the isolate from its nearest neighbours. It is evident from genotypic and phenotypic data that strain SD29(T) represents a novel species of the genus Gordonia, for which the name Gordonia shandongensis sp. nov. is proposed. The type strain is SD29(T) (=CGMCC 4.3492(T)=JCM 13907(T)).

Characterization of the dszABC genes of Gordonia amicalis F.5.25.8 and identification of conserved protein and DNA sequences.

Gordonia amicalis F.5.25.8 has the unique ability to desulfurize dibenzothiophene and to metabolize carbazole [Santos et al., Appl Microbiol Biotechnol 71:355-362, 2006]. Efforts to amplify the dsz genes from G. amicalis F.5.25.8 based on polymerase chain reaction (PCR) primers designed using the dsz gene sequences of Rhodococcus erythropolis IGTS8 were mostly unsuccessful. A comparison of the protein sequences of dissimilar desulfurization enzymes (DszABC, BdsABC, and TdsABC) revealed multiple conserved regions. PCR primers targeting some of the most highly conserved regions of the desulfurization genes allowed us to amplify dsz genes from G. amicalis F.5.25.8. DNA sequence data that include nearly the entirety of the desulfurization operon as well as the promoter region were obtained. The most closely related dsz genes are those of G. alkinovorans strain 1B at 85% identity. The PCR primers reported here should be useful in microbial ecology studies and the amplification of desulfurization genes from previously uncharacterized microbial cultures.

Gordonia soli sp. nov., a novel actinomycete isolated from soil.

A soil isolate, strain CC-AB07T, was characterized using a polyphasic approach. This organism had chemotaxonomic and morphological properties consistent with its classification in the genus Gordonia. 16S rRNA gene sequence analysis showed that the novel strain formed a monophyletic branch at the periphery of the evolutionary radiation occupied by the genus Gordonia, its closest neighbours being the type strains of Gordonia alkanivorans, Gordonia amicalis, Gordonia bronchialis, Gordonia desulfuricans, Gordonia polyisoprenivorans and Gordonia rhizosphera. The novel isolate was distinguished from all of these type strains using a range of phenotypic properties and by gyrB gene sequence analysis. It was evident from the genotypic and phenotypic data that strain CC-AB07T should be classified as representing a novel species in the genus Gordonia, for which the name Gordonia soli sp. nov. is proposed. The type strain is CC-AB07T (=BCRC 16810T=DSM 44995T).

Gordonia defluvii sp. nov., an actinomycete isolated from activated sludge foam.

Three strains of non-motile, Gram-positive, filamentous actinomycetes, isolates J4(T), J5 and J59, initially recognized microscopically in activated sludge foam by their distinctive branching patterns, were isolated by micromanipulation. The taxonomic positions of the isolates were determined using a polyphasic approach. Almost-complete 16S rRNA gene sequences of the isolates were aligned with corresponding sequences of representatives of the suborder Corynebacterineae and phylogenetic trees were inferred using three tree-making algorithms. The organisms formed a distinct phyletic line in the Gordonia 16S rRNA gene tree. The three isolates showed 16S rRNA gene sequence similarities within the range 96.9-97.2 % with their nearest phylogenetic neighbours, namely Gordonia bronchialis DSM 43247(T) and Gordonia terrae DSM 43249(T). Strain J4(T) was shown to have a chemotaxonomic profile typical of the genus Gordonia and was readily distinguished from representatives of the genus on the basis of Curie-point pyrolysis mass spectrometric data. The isolates shared nearly identical phenotypic profiles that distinguished them from representatives of the most closely related Gordonia species. It is evident from the genotypic and phenotypic data that the three isolates belong to a novel Gordonia species. The name proposed for this taxon is Gordonia defluvii sp. nov.; the type strain is J4(T) (=DSM 44981(T)=NCIMB 14149(T)).

Identification of a lipoarabinomannan-like lipoglycan in the actinomycete Gordonia bronchialis.

The cell envelopes of actinomycetes contain lipidated macroamphiphiles, of which the most extensively characterised are the lipoarabinomannans of mycobacteria and related bacteria. We have investigated the mycolic acid-containing actinomycete Gordonia bronchialis and identified the presence of a lipoarabinomannan-like lipoglycan. The extraction and purification procedures recovered a second amphiphilic fraction with properties suggesting a phosphatidylinositol mannoside, consistent with studies of other Gordonia species.

Gordonia otitidis sp. nov., isolated from a patient with external otitis.

The taxonomic positions of two clinically isolated actinomycetes were established using a polyphasic approach. The two strains, IFM 10032(T), isolated from ear discharge of a 28-year-old Japanese female patient with external otitis, and IFM 10148, isolated from pleural fluid of a 60-year-old Japanese male patient with bronchitis, possessed meso-diaminopimelic acid as the diagnostic amino acid, MK-9(H(2)) as the predominant menaquinone and mycolic acids ranging from 58 to 64 carbons. The 16S rRNA gene sequences of the two strains were most closely related to those of Gordonia aichiensis, Gordonia sputi and 'Gordonia jacobaea'. Differences in several phenotypic characteristics together with genotypic distinctiveness distinguish strains IFM 10032(T) and IFM 10148 from these three species. DNA-DNA hybridization results and the combination of genotypic and phenotypic data showed that the two strains belong to a single species, and merit recognition of a novel species within the genus Gordonia. The name proposed for this taxon is Gordonia otitidis sp. nov.; the type strain is IFM 10032(T) (=DSM 44809(T)=JCM 12355(T)=NBRC 100426(T)).