Adaptative biochemical pathways and regulatory networks in Klebsiella oxytoca BAS-10 producing a biotechnologically relevant exopolysaccharide during Fe(III)-citrate fermentation.

Abstract Background A bacterial strain previously isolated from pyrite mine drainage and named BAS-10 was tentatively identified as Klebsiella oxytoca. Unlikely other enterobacteria, BAS-10 is able to grow on Fe(III)-citrate as sole carbon and energy source, yielding acetic acid and CO2 coupled with Fe(III) reduction to Fe(II) and showing unusual physiological characteristics. In fact, under this growth condition, BAS-10 produces an exopolysaccharide (EPS) having a high rhamnose content and metal-binding properties, whose biotechnological applications were proven as very relevant. Results Further phylogenetic analysis, based on 16S rDNA sequence, definitively confirmed that BAS-10 belongs t…

Gene cloning and characterization of an alkane hydroxylase system in the long chain n-alkane- degrader Gordonia sp. SoCg

Cloning and sequencing of the dnaK region of Streptomyces coelicolor A3(2)

Abstract The dnaK homologue of Streptomyces coelicolor A3(2) strain M145 has been cloned and sequenced. Nucleotide sequence analysis of a 2.5-kb region revealed an open reading frame (ORF) encoding a predicted DnaK protein of 618 amino acids (Mr = 66 274). The dnaK coding sequence displays extreme codon bias and shows a strong preference for CGY and GGY, for Arg and Gly codons, respectively. The predicted DnaK sequence has a high Lys:Arg ratio which is not typical of streptomycete proteins. The region immediately downstream from dnaK contains an ORF for a GrpE-like protein; the predicted start codon of grpE overlaps the last two codons of dnaK, indicating that the two genes are translationa…

Attività e biodiversità microbica in sedimenti marini

Development and characterization of antimicrobial PCL fibres for surgical suture applications

Broad Spectrum Thiopeptide Recognition Specificity of theStreptomyces lividans TipAL Protein and Its Role in Regulating Gene Expression

Microbial metabolites isolated in screening programs for their ability to activate transcription of the tipA promoter (ptipA) in Streptomyces lividans define a class of cyclic thiopeptide antibiotics having dehydroalanine side chains ("tails"). Here we show that such compounds of heterogeneous primary structure (representatives tested: thiostrepton, nosiheptide, berninamycin, promothiocin) are all recognized by TipAS and TipAL, two in-frame translation products of the tipA gene. The N-terminal helix-turn-helix DNA binding motif of TipAL is homologous to the MerR family of transcriptional activators, while the C terminus forms a novel ligand-binding domain. ptipA inducers formed irreversible…

Endophytic Bacteria Associated with Origanum heracleoticum L. (Lamiaceae) Seeds

Seed-associated microbiota are believed to play a crucial role in seed germination, seedling establishment, and plant growth and fitness stimulation, due to the vertical transmission of a core microbiota from seeds to the next generations. It might be hypothesized that medicinal and aromatic plants could use the seeds as vectors to vertically transfer beneficial endophytes, providing plants with metabolic pathways that could influence phytochemicals production. Here, we investigated the localization, the structure and the composition of the bacterial endophytic population that resides in Origanum heracleoticum L. seeds. Endocellular bacteria, surrounded by a wall, were localized close to th…

SCO5208 encodes the histidinol phosphate phosphatase involved in histidine biosynthesis in Streptomyces coelicolor

Dbv4 is DNA-binding protein acting as a positive regulator of the glycopeptide antibiotic A40926 biosynthesis

Bacterial Community Structure of an IFAS-MBRs Wastewater Treatment Plant

TIn this work, the bacterial community putatively involved in BNR events of a UCT-MBMBR pilot plant was elucidated by both culture-dependent and metagenomics DNA analyses. The presence of bacterial isolates belonging to Bacillus (in the anoxic compartment) and to Acinetobacter, Stenotrophomonas, Rhodococcus, Escherichia and Aeromonas (in the aerobic compartment) is in agreement with the nitrification/denitrification processes observed in the plant. Moreover, the study of bacterial community structure by NGS revealed a microbial diversity suggesting a biochemical complexity which can be further explored and exploited to improve UCT-MBMBR plant performance.

LAPTOP: Lantibiotic production, Technology, optimization and improved process

The dnaK operon of Streptomyces coelicolor encodes a novel heat-shock protein which binds to the promoter region of the operon

Transcriptional studies have demonstrated that the dnaK gene of Streptomyces coelicolor A3(2) is contained within a 4.3 kb operon. The operon is transcribed from a single (transiently) heat-inducible promoter, dnaKp, that resembles the typical vegetative (sigma 70-recognized) eubacterial consensus promoter sequence. dnaK transcription was found to be heat-inducible at all stages of development in surface-grown cultures. In addition, at the normal growth temperature of 30 degrees C, dnaK transcript levels were shown to vary at different stages of development, being more abundant in young germinating cultures and in mycelium undergoing sporogenesis. The nucleotide sequence of the dnaK operon …

Omic-based strategies reveal novel links between primary metabolism and antibiotic production

PARTIAL SEQUENCING OF THE BETA-GLUCOSIDASE-ENCODING GENE FROM YEAST STRAINS ISOLATED FROM MUSTS AND WINES

The aim of the present work was the identification of the gene encoding for β-glucosidase and its partial sequencing in the strainsPichia anomala AL112,Hanseniaspora uvarum Y8 andSaccharomyces cerevisiae AL41. To this aim degenerated primers, designed on the basis of aminoacid similarities of four known yeast β-glucosidases, have been used in PCR amplifications. An expected fragment of about 200 bp was amplified from all the DNAs, cloned and sequenced. Sequence homology demonstrated for the first time the presence of a β-glucosidase encoding gene inHanseniaspora uvarum andSaccharomyces cerevisiae.

Preparazione e caratterizzazione di fibre a base di PCL con proprietà antimicrobiche per applicazioni chirurgiche

Antibiotic Resistance: Mechanisms and Impact

Plant growth promoting potential of bacterial endophytes from three terrestrial mediterranean orchid species

Orchids are highly dependent on symbiotic microorganisms during their entire life cycle. Whereas an important role in orchid seed germination and early plant development is well established for mycorrhizal fungi, the influence of endophytic bacteria on orchid growth has been less investigated. Here, we report the isolation of endophytic bacteria from different organs of three terrestrial Mediterranean orchid species (Spiranthes spiralis, Serapias vomeracea and Neottia ovata), the investigation of their potential Plant Growth-Promoting (PGP) traits and their interaction with the orchid mycorrhizal (OM) fungus Tulasnella calospora in vitro. Little overlap was found among endophytic bacteria i…

Orf4 is a DNA-binding protein acting as a positive regulator of A40926 in Nonomuraea sp. ATCC39727

Comparison of different primer sets for use in Automated Ribosomal Intergenic Spacer Analysis of complex bacterial communities.

ABSTRACT ITSF and ITSReub, constituting a new primer set designed for the amplification of the 16S-23S rRNA intergenic transcribed spacers, have been compared with primer sets consisting of 1406F and 23Sr (M. M. Fisher and E. W. Triplett, Appl. Environ. Microbiol. 65:4630-4636, 1999) and S-D-Bact-1522-b-S-20 and L-D-Bact-132-a-A-18 (L. Ranjard et al., Appl. Environ. Microbiol. 67:4479-4487, 2001), previously proposed for automated ribosomal intergenic spacer analysis (ARISA) of complex bacterial communities. An agricultural soil and a polluted soil, maize silage, goat milk, a small marble sample from the façade of the Certosa of Pavia (Pavia, Italy), and brine from a deep hypersaline anoxi…

EVA antimicrobial films prepared by melt processing

In vivo evaluation of the interaction between the Escherichia coli IGP synthase subunits using the Bacterial Two-Hybrid system

ABSTRACT Histidine biosynthesis is one of the most characterized metabolic routes for its antiquity and its central role in cellular metabolism; indeed, it represents a cross-road between nitrogen metabolism and de novo synthesis of purines. This interconnection is due to the activity of imidazole glycerol phosphate synthase, a heterodimeric enzyme constituted by the products of two his genes, hisH and hisF, encoding a glutamine amidotransferase and a cyclase, respectively. Despite their interaction was suggested by several in vitro experiments, their in vivo complex formation has not been demonstrated. On the contrary, the analysis of the entire Escherichia coli interactome performed using…

Differential proteomic analysis highlights metabolic strategies associated with balhimycin production in Amycolatopsis balhimycina chemostat cultivations

Abstract Background Proteomics was recently used to reveal enzymes whose expression is associated with the production of the glycopeptide antibiotic balhimycin in Amycolatopsis balhimycina batch cultivations. Combining chemostat fermentation technology, where cells proliferate with constant parameters in a highly reproducible steady-state, and differential proteomics, the relationships between physiological status and metabolic pathways during antibiotic producing and non-producing conditions could be highlighted. Results Two minimal defined media, one with low Pi (0.6 mM; LP) and proficient glucose (12 g/l) concentrations and the other one with high Pi (1.8 mM) and limiting (6 g/l; LG) glu…

AROMATIC AMINOACIDS AND ANTIBIOTIC PRODUCTION IN STREPTOMYCES COELICOLOR

Elucidating the molecular physiology of lantibiotic NAI-107 production in Microbispora ATCC-PTA-5024.

Background The filamentous actinomycete Microbispora ATCC-PTA-5024 produces the lantibiotic NAI-107, which is an antibiotic peptide effective against multidrug-resistant Gram-positive bacteria. In actinomycetes, antibiotic production is often associated with a physiological differentiation program controlled by a complex regulatory and metabolic network that may be elucidated by the integration of genomic, proteomic and bioinformatic tools. Accordingly, an extensive evaluation of the proteomic changes associated with NAI-107 production was performed on Microbispora ATCC-PTA-5024 by combining two-dimensional difference in gel electrophoresis, mass spectrometry and gene ontology approaches. R…

Inorganic phosphate is a trigger factor for Microbispora sp. ATCC-PTA-5024 growth and NAI-107 production

A putative membrane protein and an ABC-transporter of Planobispora rosea stimulate antibacterial activity in Streptomyces lividans

Expression in Streptomyces lividans of Nonomuraea genes cloned in an artificial chromosome

A bacterial artificial chromosomal library of Nonomuraea sp. ATCC39727 was constructed using Escherichia coli-Streptomyces artificial chromosome (ESAC) and screened for the presence of dbv genes known to be involved in the biosynthesis of the glycopeptide A40926. dbv genes were cloned as two large, partially overlapping, fragments and transferred into the host Streptomyces lividans, thus generating strains S. lividansColon, two colonsNmESAC50 and S. lividansColon, two colonsNmESAC57. The heterologous expression of Nonomuraea genes in S. lividans was successfully demonstrated by using combined RT-PCR and proteomic approaches. MALDI-TOF analysis revealed that a Nonomuraea ABC transporter is e…

Identification and characterization of new prolylendopeptidases (PEPs) from Actinomycetes

Applicazioni biotecnologiche di batteri del suolo

A large catabolic plasmid carries the genes for the dehalogenation of 4-chlorobenzoate in arthrobacter ramosus FG1

Exploring long chain n-alkane metabolism in Gordonia sp. strain SoCg

Many microorganisms are able to degrade aliphatic hydrocarbons and a relationship between n-alkane utilization and storage compound synthesis has been described in bacteria. The Gram positive GC-rich n-alkane degrader Gordonia sp. strain SoCg, isolated from a long-term accidentally contaminated beach in Sicily, is able to grow on long n-alkanes up to. It carries a single copy of the alkane hydroxylase gene alkB on its chromosome and its alk cluster revealed a genomic organization similar to other alk clusters of alkane-degrading Gram positive bacteria. The alk gene expression, analysed by Real-time RT-PCR, is induced by n-hexadecane and n-triacontane and coupled to alkane consumption. Inter…

Cloning and characterization of the histidine biosynthetic gene cluster of Streptomyces coelicolor A3(2).

Abstract Biochemical and genetic data indicate that in Streptomyces coelicolor A3(2) the majority of the genes involved in the biosynthesis of histidine are clustered in a small region of the chromosome [Carere et al., Mol. Gen. Genet. 123 (1973) 219–224; Russi et al., Mol. Gen. Genet. 123 (1973) 225–232]. To investigate the structural organization and the regulation of these genes, we have constructed genomic libraries from S. coelicolor A3(2) in pUC vectors. Recombinant clones were isolated by complementation of an Escherichia coli hisBd auxotroph. A recombinant plasmid containing a 3.4-kb fragment of genomic DNA was further characterized. When cloned in the plasmid vector, pIJ699, this f…

Regulation of antibiotic biosynthesis in Actinomycetes.

Further characterization of the histidine gene cluster of Streptomyces coelicolor A3(2): nucleotide sequence and transcriptional analysis of hisD.

We have further characterized the genomic region of Streptomyces coelicolor A3(2) that contains genes involved in the biosynthesis of histidine. A 2,357-base pair fragment contained in plasmid pSCH3328 that complemented hisD mutations has been sequenced. Computer analysis revealed an open reading frame that encodes a protein with significant homology to the Escherichia coli, Salmonella typhimurium and Mycobacterium smegmatis hisD product, Saccharomyces cerevisiae HIS4C, and Neurospora crassa his3 gene products. Two other contiguous open reading frames oriented divergently with respect to hisD did not show significant similarity with any of the his genes or to other sequences included in the…

NovelAmycolatopsis balhimycinabiochemical abilities unveiled by proteomics

Amycolatopsis balhimycina DSM5908 is an actinomycete producer of balhimycin, an analogue of vancomycin, the antibiotic of ‘last resort’ against multidrug-resistant Gram-positive pathogens. Most knowledge on glycopeptide biosynthetic pathways comes from studies on A. balhimycina as this strain, among glycopeptide producers, is genetically more amenable. The recent availability of its genome sequence allowed to perform differential proteomic analyses elucidating key metabolic pathways leading to antibiotic production in different growth conditions. To implement proteomic data on A. balhimycina derived from 2-DE approaches and to identify novel components, a combined approach based on protein …

Diversity of rhizobia nodulating wild shrubs of Sicily and some neighbouring islands

Abstract Legume shrubs have great potential for rehabilitation of semi-arid degraded soils in Mediterranean ecosystems as they establish mutualistic symbiosis with Nfxing rhizobia. Eighty-eight symbiotic rhizobia were isolated from seven wild legume shrubs native of Sicily (Southern Italy) and grouped in operational taxonomic units (OTU) by analysis of the ribosomal internal transcribed spacer (ITS) polymorphism. Partial sequencing of 16S rRNA gene of representative isolates of each OTU revealed that most Genisteae symbionts are related to Bradyrhizobium canariense, B. japonicum and B. elkanii. Teline monspessulana was the only Genistea nodulated by Mesorhizobium strains, and Anagyris foeti…

Amino acid biosynthesis in Streptomyces coelicolor

Diimidazolium organic salts and supramolecular gels exerting antimicrobial activity

Developmental control of the heat-shock stress regulon in Streptomyces coelicolor

In the differentiating eubacterium Streptomyces coelicolor, nutritional imbalances activate a developmental programme which involves the heat-shock stress regulon. In liquid batch cultures, the growth curve could be separated into four components: rapid growth 1 (RG1), transition (T), rapid growth 2 (RG2) and stationary (S). Patterns of gene expression in cultures subjected to heat shock in various phases were recorded on two-dimensional gels and analysed using advanced statistical methods. The responses of all heat-shock proteins (HSPs) were highly dependent upon the growth phase, thus demonstrating that the four phases of growth were physiologically distinct. For many HSPs, the levels of …

Streptomyces coelicolor Vesicles: Many Molecules To Be Delivered

ABSTRACT Streptomyces coelicolor is a model organism for the study of Streptomyces, a genus of Gram-positive bacteria that undergoes a complex life cycle and produces a broad repertoire of bioactive metabolites and extracellular enzymes. This study investigated the production and characterization of membrane vesicles (MVs) in liquid cultures of S. coelicolor M145 from a structural and biochemical point of view; this was achieved by combining microscopic, physical and -omics analyses. Two main populations of MVs, with different sizes and cargos, were isolated and purified. S. coelicolor MV cargo was determined to be complex, containing different kinds of proteins and metabolites. In particul…

Assessment and characterization of the bacterial community structure in advanced activated sludge systems

Abstract The present study is aimed to assess and characterize the structure of bacterial community in advanced activated sludge systems. In particular, activated sludge samples were collected from an Integrated Fixed-film Activated Sludge – Membrane Bioreactor pilot plant under a University of Cape Town configuration with in-series anaerobic (Noair)/anoxic (Anox)/aerobic (Oxy) reactors – and further analyzed. The achieved results – based on Next Generation Sequencing (NGS) of 16S rDNA amplicons – revealed that the bacterial biofilm (bf) communities on plastic carriers of Oxy and Anox reactors had a greater diversity compared to suspended (sp) bacterial flocs of Oxy, Anox and Noair. Indeed,…

Isolation of Gram positive n-alkane degraders from a hydrocarbon contaminated sicilian shoreline

Dbv4 is a DNA-binding protein regulating the expression of OxyA during A40926 biosynthesis

Influence of Drawing on the Antimicrobial and Physical Properties of Chlorhexidine-Compounded Poly(caprolactone) Monofilaments

In this work, we prepared poly(caprolactone) (PCL) monofilaments incorporating different amounts of the antiseptic chlorhexidine diacetate (CHX) and evaluated the effect of “online” drawing during the melt spinning or of an “offline” cold drawing on the antimicrobial and physical properties of PCL/CHX monofilaments. The results showed that the drawing conditions strongly affected the properties of the threads. In particular, the filaments exhibited antimicrobial activity for all the draw ratios (DR) adopted, but on increasing DR the antibacterial efficiency reduced. These results were correlated with the CHX surface concentration and with the release kinetics.

Dbv3 e Dbv4: due regolatori della biosintesi dell’antibiotico glicopeptidico A40926 di Nonomuraea ATCC39727

DBV4 IS A DNA BINDING PROTEIN ACTING AS A POSITIVE REGULATOR OF THE GLYCOPEPTIDE ANTIBIOTIC A40926 BIOSYNTHESIS

Tryptophan promotes morphological and physiological differentiation in Streptomyces coelicolor.

The molecular mechanisms regulating tryptophan biosynthesis in actinomycetes are poorly understood; similarly, the possible roles of tryptophan in the differentiation program of microorganism life-cycle are still underexplored. To unveil the possible regulatory effect of this amino acid on gene expression, an integrated study based on quantitative teverse transcription-PCR (qRT-PCR) and proteomic approaches was performed on the actinomycete model Streptomyces coelicolor. Comparative analyses on the microorganism growth in a minimal medium with or without tryptophan supplementation showed that biosynthetic trp gene expression in S. coelicolor is not subjected to a negative regulation by the …

Isolation of Gram-positive n-alkane degraders from a hydrocarbon-contaminated Mediterranean shoreline.

Aims: To investigate the petroleum hydrocarbon (HC)-degrading potential of indigenous micro-organisms in a sandy Mediterranean coast, accidentally contaminated with petroleum-derived HCs. Methods and Results: Using culturable methods, a population of Gram-positive n-alkane degraders was detected in the contaminated soil. Five isolates, identified as one Nocardia, two Rhodococcus and two Gordonia strains, were able to degrade medium- and long-chain n-alkanes up to C36 as assessed by growth assays and gas chromatography-mass spectrometry analysis. Diverging alkane hydroxylase-encoding genes (alkB) were detected by PCR, using degenerated primers, in all the strains; multiple sequences were obt…

Una collezione di rizobi da leguminose mediterranee per il recupero ambientale

Artificial chromosome libraries of Streptomyces coelicolor A3(2) and Planobispora rosea

Using an Escherichia coli-Streptomyces shuttle vector derived from a bacterial artificial chromosome (BAC), we developed methodologies for the construction of BAC libraries of filamentous actinomycetes. Libraries of Streptomyces coelicolor, the model actinomycete, and Planobispora rosea, a genetically intractable strain, were constructed. Both libraries have an average insert size of 60 kb, with maximal insert larger than 150 kb. The S. coelicolor library was evaluated by selected hybridisations to DraI fragments and by end sequencing of a few clones. Hybridisation of the P. rosea library to selected probes indicates a good representation of the P. rosea genome and that the library can be u…

Aromatic aminoacids and Calcium Dependent Antibiotic:Relation of primary metabolism to antibiotic production in Actinomycetes

Influence of Drawing on the Antimicrobial and Physical Properties of Chlorhexidine-Compounded Poly(caprolactone) Monofilaments

In this work, we prepared poly(caprolactone) (PCL) monofilaments incorporating different amounts of the antiseptic chlorhexidine diacetate (CHX) and evaluated the effect of "online" drawing during the melt spinning or of an "offline" cold drawing on the antimicrobial and physical properties of PCL/CHX monofilaments. The results showed that the drawing conditions strongly affected the properties of the threads. In particular, the filaments exhibited antimicrobial activity for all the draw ratios (DR) adopted, but on increasing DR the antibacterial efficiency reduced. These results were correlated with the CHX surface concentration and with the release kinetics.

From tryptophan metabolism to peptide antibiotic production in Streptomyces coelicolor

The tryptophan is a precursor of the lipopeptide calcium dependent antibiotic (CDA), produced from Streptomyces coelicolor and closely related to important antibiotics such as daptomycin. We have focused our attention on the correlation between CDA production and tryptophan metabolism in order to identify new strategies aimed at increasing the production of peptide antibiotics.

Polysaccharide-based silver nanoparticles synthesized by Klebsiella oxytoca DSM 29614 cause DNA fragmentation in E-coli cells

Silver nanoparticles (AgNPs), embedded into a specific exopolysaccharide (EPS), were produced by Klebsiella oxytoca DSM 29614 by adding AgNO3 to the cultures during exponential growth phase. In particular, under aerobic or anaerobic conditions, two types of silver nanoparticles, named AgNPs-EPS(aer) and the AgNPs-EPS(anaer), were produced respectively. The effects on bacterial cells was demonstrated by using Escherichia coli K12 and Kocuria rhizophila ATCC 9341 (ex Micrococcus luteus) as Gram-negative and Gram-positive tester strains, respectively. The best antimicrobial activity was observed for AgNPs-EPS(aer), in terms of minimum inhibitory concentrations and minimum bactericidal concentr…

Molecular analysis of lichen-associated bacterial communities.

The bacterial communities associated with 11 different lichen samples (belonging to eight different species) from different habitats were investigated. The culturable aerobic-heterotrophic fraction of the bacterial communities was isolated from nine lichen samples on protein-rich and sugar-rich/N-free media. Thirty-four bacterial isolates were purified and pooled into groups (phylotypes) by analysis of the ribosomal internal transcribed spacer polymorphism. Twenty five phylotypes were identified, each comprising between one and three isolates. One isolate of each phylotype was partially sequenced and the resulting 16S rRNA gene sequences were compared in a phylogenetic analysis. Three gener…

Differential proteomic analysis reveals novel links between primary metabolism and antibiotic production in Amycolatopsis balhimycina.

A differential proteomic analysis, based on 2-DE and MS procedures, was performed on Amycolatopsis balhimycina DSM5908, the actinomycete producing the vancomycin-like antibiotic balhimycin. A comparison of proteomic profiles before and during balhimycin production characterized differentially and constitutively expressed protein isoforms, which were associated to 203 ORFs in the A. balhimycina genome. These data, providing insights on the major metabolic pathways/molecular processes operating in this organism, were used to compile 2-DE reference maps covering 3-10, 4-7 and 4.5-5.5 pH gradients available over the World Wide Web as interactive web pages (http://www.unipa.it/ampuglia/Abal-prot…

Film di EVA con attività antibatterica

Differential proteomic analysis of an engineered Streptomyces coelicolor strain reveals metabolic pathways supporting growth on n-hexadecane

The alkB gene, encoding an alkane monooxygenase in the actinomycete Gordonia sp. SoCg, was expressed in the non-alkane-degrading actinomycete Streptomyces coelicolor M145. The resulting engineered strain, M145-AH, can grow on n-hexadecane as sole carbon source. To unravel proteins associated with growth on n-alkanes, proteome of M145-AH after 6, 24, and 48 h of incubation in the Bushnell-Haas (BH) mineral medium containing n-hexadecane as sole carbon source (H condition) and in BH without any carbon source (0 condition) were compared using 2D-differential gel electrophoresis. Proteome analysis revealed significant changes only at 48 h, showing 48 differentially abundant proteins identified …

Microbial bioresources for rehabilitation of natural and anthropogenic deserts

Arid lands and deserts cover one fifth of the earth’s land surface and they are currently increasing due to the global warming. Besides the natural desert settings, human activities lead to the desertification of certain areas: municipal landfills and caves are the most common examples of such anthropogenic deserts. Massive pollution and lack of natural soil profiles create harsh conditions and hinder the spontaneous rehabilitation of such areas. Phytoremediation is often used for renaturalization of such degraded soils. Two case studies are presented aiming at demonstrating that the plant-microbe interactions are essential for improving the soil quality and allowing the formation of crops.…

Amycolatopsis proteomic analysis to reveal primary metabolism genes involved in balhimycin production

Differential proteomics highlights metabolic changes associated with n-hexadecane utilization in a Streptomyces coelicolor strain expressing Gordonia sp. SoCg n-alkane monooxigenase.

Introduction: Alkanes are biodegraded to generate the corresponding primary alcohol trough alkane hydroxylases (AHs) consisting on an integral membrane alkane monooxygenase (AlkB) and two soluble proteins, rubredoxin and rubredoxin reductase. Recently, an alkB gene was reported to be involved in degradation of long chain n-alkanes in the actinobacterium Gordonia sp. SoCg. This gene was expressed in Streptomyces coelicolor M145 which is unable to degrade n-alkanes. Results: The engineered strain, M145-AH, can biotransform n-hexadecane into the corresponding 1-hexadecanol and it is able to grow on n-hexadecane as sole carbon source. Changes in global protein expression associated with n-hexad…

A phosphate-controlled regulator for the biosynthesis of the dalbavancin precursor A40926

Characterization of the regulatory genes for the production of the glycopeptide antibiotic A40926 by Nonomuraea ATCC 39727

Optimized RNA extraction and northern hybridization in streptomycetes.

Abstract Northern blot hybridization is a useful tool for analyzing transcript patterns. To get a picture of what really occurs in vivo, it is necessary to use a protocol allowing full protection of the RNA integrity and recovery and unbiased transfer of the entire transcripts population. Many protocols suffer from severe limitations including only partial protection of the RNA integrity and/or loss of small sized molecules. Moreover, some of them do not allow an efficient and even transfer in the entire sizes range. These difficulties become more prominent in streptomycetes, where an initial quick lysis step is difficult to obtain. We present here an optimized northern hybridization protoc…

Aminoacid metabolism and its impact on antibiotic production

TrpM, a Small Protein Modulating Tryptophan Biosynthesis and Morpho-Physiological Differentiation in Streptomyces coelicolor A3(2).

In the model actinomycete Streptomyces coelicolor A3(2), small open reading frames encoding proteins with unknown functions were identified in several amino acid biosynthetic gene operons, such as SCO2038 (trpX) in the tryptophan trpCXBA locus. In this study, the role of the corresponding protein in tryptophan biosynthesis was investigated by combining phenotypic and molecular analyses. The 2038KO mutant strain was characterized by delayed growth, smaller aerial hyphae and reduced production of spores and actinorhodin antibiotic, with respect to the WT strain. The capability of this mutant to grow on minimal medium was rescued by tryptophan and tryptophan precursor (serine and/or indole) su…

Gordonia sp. SoCg alkB gene confers the ability to degrade and use n-alkanes as carbon source in Gram positive bacteria

Gordonia sp. SoCg, a Gram positive strain able to grow on long chain n-alkanes1, possess a single copy of alkB2 gene, whose product is required for n-alkane hydroxylation3. An analysis of alkB flanking regions revealed five ORFs which were designed as orf1, rubA3, rubA4, rubB and alkU, according to the sequence 14 homology with that of known alk clusters3. In G. sp. SoCg the transcription of these genes was induced by long-chain and solid n-alkanes as revealed by quantitative RT-PCR, and the essential role of alkB in nalkane degradation was demonstrated by the construction of an alkB disruption mutant strain3. The SoCg alkB gene was successfully expressed in Streptomyces coelicolor M145 (M1…

PROTEOMIC ANALYSIS OF AMYCOLATOPSIS BALHIMYCINA TO REVEAL NOVEL LINKS BETWEEN PRIMARY AND SECONDARY METABOLISM.

A putative membrane protein and an ABC-transporter of Planobispora rosea induce antibacterial activity of Streptomyces lividans

Interactions among rhizobial strains and arbuscular mycorrhizal fungi on Spartium junceum

Regulation of the biosynthesis of the glycopeptide antibiotic A40926 in Nonomuraea ATCC39727

The actinomycete Nonomuraea produces the glycopeptide A40926, precursor of dalbavancin. Dalbavancin is a novel lipoglycopeptide agent against emerging resistant Gram-positive cocci with superior pharmacodynamics properties compared to vancomycin. Chemically, a glycopeptide antibiotic consists of a heptapeptide core constituted by proteinogenic and nonproteinogenic amino acids such as 3,5-dihydroxyphenylglycine (DPG) and 4-hydroxyphenylglycine (HPG). The heptapeptide is assembled by a nonribosomal peptide synthetase and modified by oxidative cross-linking of the electron-rich aromatic side chains, halogenation, sulfation, methylation, acylation, and glycosylation. The A40926 biosynthetic gen…

Two heterologously expressed Planobispora rosea proteins cooperatively induce Streptomyces lividans thiostrepton uptake and storage from the extracellular medium.

Abstract Background A bacterial artificial chromosomal library of Planobispora rosea, a genetically intractable actinomycete strain, was constructed using Escherichia coli-Streptomyces artificial chromosome (ESAC) and screened for the presence of genes known to be involved in the biosynthesis of antibiotics. Results One clone with a 40 kb insert showed antimicrobial activity against Gram positive bacteria. Insert sequence analysis and subcloning experiments revealed that the bioactivity was due to a 3.5 kb DNA fragment containing two open reading frames. These orfs encode two proteins with high similarity to a putative membrane protein of Streptomyces coelicolor and to the nogalamycin resis…

Effetto dello stiro sulle proprietà fisiche e antimicrobiche di fibre a base di PCL contenenti clorexidina

Amycolatopsis balhimycina gene expression under inorganic phosphate limitation

Genomic traits of Klebsiella oxytoca DSM 29614, an uncommon metal-nanoparticle producer strain isolated from acid mine drainages

Background Klebsiella oxytoca DSM 29614 - isolated from acid mine drainages - grows anaerobically using Fe(III)-citrate as sole carbon and energy source, unlike other enterobacteria and K. oxytoca clinical isolates. The DSM 29614 strain is multi metal resistant and produces metal nanoparticles that are embedded in its very peculiar capsular exopolysaccharide. These metal nanoparticles were effective as antimicrobial and anticancer compounds, chemical catalysts and nano-fertilizers. Results The DSM 29614 strain genome was sequenced and analysed by a combination of in silico procedures. Comparative genomics, performed between 85 K. oxytoca representatives and K. oxytoca DSM 29614, revealed th…

Proteomic investigations to unravel molecular physiology of the lantibiotic producer Microbispora sp. ATCC‐PTA‐5024

Tryptophan catabolism via kynurenine production in Streptomyces coelicolor: identification of three genes coding for the enzymes of tryptophan to anthranilate pathway

Most enzymes involved in tryptophan catabolism via kynurenine formation are highly conserved in Prokaryotes and Eukaryotes. In humans, alterations of this pathway have been related to different pathologies mainly involving the central nervous system. In Bacteria, tryptophan and some of its derivates are important antibiotic precursors. Tryptophan degradation via kynurenine formation involves two different pathways: the eukaryotic kynurenine pathway, also recently found in some bacteria, and the tryptophan-to-anthranilate pathway, which is widespread in microorganisms. The latter produces anthranilate using three enzymes also involved in the kynurenine pathway: tryptophan 2,3-dioxygenase (TD…

Proteomic analysis of the antibiotic producer Amycolatopsis balhimycina

Cloning of an alkane hydroxylase system in a long chain n-alkane- degrader Gordonia sp.

Five Gram-positive GC rich n-alkane degraders were isolated from a long-term accidentally contaminated beach in Sicily and identified as one Nocardia, two Rhodococcus and two Gordonia strains (Quatrini et al., 2008 J. Appl. Microbiol. 104:251-9). All the isolates were able to grow on long and very long chain n-alkanes up to C36. Diverging alkane-hydroxylase encoding genes (alkB) were detected by PCR using degenerated primers in all the strains. Multiple sequences were obtained from the Nocardia strain while only one alkB gene was detected in Rhodococcus and Gordonia. The aim of this work is to genetically characterize the alk cluster in one of the two Gordonia strains called SoCg. Pulsed Fi…

Isolation and characterization of Gordonia SoCg n-alkane degradation cluster

Gordonia strain SoCg is a Gram-positive GC rich n-alkane degrader, isolated from a long-term contaminated beach in Sicily for her ability to degrade long (dodecane C12) and very long chain n-alkanes up to hexatriacontane (C36) (P.Quatrini et al., J. Appl. Microbiol., 2007). PCR analysis, using degenerated primers, reveled that it carries one alkane-hydroxylase gene (alkB); PFGE and Southern analysis showed that this gene is localized on the chromosome. In order to isolate the alk cluster of Gordonia strain SoCg, an enriched gene bank was constructed in E.coli DH10B by isolating restriction fragments of the desired size from a preparative gel. One clone, containing a DNA insert of about 7 kb…

DICATIONIC IMIDAZOLIUM SALTS: TUNABLE ANTIMICROBIAL AND ANTITUMORAL CHEMIOTHERAPEUTIC LEADS

The chemical synthesis of novel chemotherapeutical leads is evolving thanks to possibility to design molecules with desired physical-chemical and, thus, biological properties. The imidazolium salts, recently proven effective to inhibit bacterial and/or cancer cell growth, posses an amphiphilic nature that is conferred by the imidazolium cation having a polar head generally coupled with aliphatic side chains. Thus, biological properties of imidazolium salts can be tuned through modifications involving the cation structure and/or the anion nature. By covalently linking two imidazolium rings, di-imidazolium salts were obtainedobtain differing in: i) kind of anions; ii) geometric isomerization …

LA BIODIVERSITÀ DEI RIZOBI SIMBIONTI DELLE LEGUMINOSE LEGNOSE MEDITERRANEE COME RISORSA FINALIZZATA AL RECUPERO DEGLI ECOSISTEMI

In vitro effects of AGNPs exopolysaccharide from Klebsiella Oxytoca DSM29614 on breast cancer cells

Analysis of the bacterial community associated with different species of lichens: preliminary results

I microrganismi di interesse biotecnologico

Phosphate-controlled regulator for the biosynthesis of the dalbavancin precursor A40926

ABSTRACT The actinomycete Nonomuraea sp. strain ATCC 39727 produces the glycopeptide A40926, the precursor of the novel antibiotic dalbavancin. Previous studies have shown that phosphate limitation results in enhanced A40926 production. The A40926 biosynthetic gene ( dbv ) cluster, which consists of 37 genes, encodes two putative regulators, Dbv3 and Dbv4, as well as the response regulator (Dbv6) and the sensor-kinase (Dbv22) of a putative two-component system. Reverse transcription-PCR (RT-PCR) and real-time RT-PCR analysis revealed that the dbv14 - dbv8 and the dbv30 - dbv35 operons, as well as dbv4 , were negatively influenced by phosphate. Dbv4 shows a putative helix-turn-helix DNA-bind…

Knock out of the sapA gene in Streptomyces lividans. A gene that encodes the Spores associated protein SapA

Bradyrhizobium sp. nodulating the Mediterranean shrub Spanish broom (Spartium junceum L.)

Aims: The molecular diversity of 25 strains of rhizobia, isolated in Sicily from root nodules of the Mediterranean shrubby legume Spanish broom (Spartium junceum L.), is presented in relation to the known rhizobial reference strains. Methods and Results: Our approach to the study of the S. junceum rhizobial diversity combined the information given by the 16S and the intergenic spacer (IGS) 16S–23S rDNA polymorphic region by obtaining them in a single polymerase chain reaction (PCR) step. The PCR fragment size of the S. junceum isolates was 2400–2500 bp and that of the reference strains varied from 2400 in Bradyrhizobium strains to 2800 in Sinorhizobium strains. Inter- and intrageneric lengt…

Two master switch regulators trigger A40926 biosynthesis in Nonomuraea sp. strain ATCC 39727

ABSTRACT The actinomycete Nonomuraea sp. strain ATCC 39727 produces the glycopeptide A40926, the precursor of dalbavancin. Biosynthesis of A40926 is encoded by the dbv gene cluster, which contains 37 protein-coding sequences that participate in antibiotic biosynthesis, regulation, immunity, and export. In addition to the positive regulatory protein Dbv4, the A40926-biosynthetic gene cluster encodes two additional putative regulators, Dbv3 and Dbv6. Independent mutations in these genes, combined with bioassays and liquid chromatography-mass spectrometry (LC-MS) analyses, demonstrated that Dbv3 and Dbv4 are both required for antibiotic production, while inactivation of dbv6 had no effect. In …

The Streptomyces coelicolor dnaK operon contains a second promoter driving the expression of the negative regulator hspR at physiological temperature

HspR (heat shock protein regulator) acts as a negative regulator of different genes in many bacteria. In Streptomyces coelicolor hspR gene is part and the transcriptional repressor of the dnaK operon which encodes the DnaK, GrpE, DnaJ chaperone machines and HspR itself. Our experiments led us to the discovery of a second promoter, internal to dnaK operon, located upstream hspR gene. Transcription from this promoter was detected at 30 degrees C indicating that hspR could play a key physiological role.

Antibiotics and Resistance: A Fatal Attraction

When penicillin and other classes of antibiotics were discovered and used to treat infectious diseases human morbidity and mortality due to infective microorganisms were successfully contrasted. However, almost as soon as antibacterial drugs were introduced in clinics, bacterial resistance spread. From a biological and microbiological viewpoint, antibacterial drug resistance is a fascinating aspect of molecular evolution, and resistome concept has been introduced to shed light on driving forces that control the spreading and maintenance of resistance genes within microbial populations. Despite the wide range of chemical complexity of antibiotics, there is a restricted number of modes of act…

Impact of aminoacid metabolism on antibiotic production

Preliminary results of EOLIFE99, a project concerning the conservation of four endangered plant species of Aeolian Archipelago (South Tyrrhenian Sea, Italy)

Effect of PCL/PEG-Based Membranes on Actinorhodin Production in Streptomyces coelicolor Cultivations

The actinomycetes, Gram-positive filamentous bacteria, are the most prolific source of natural occurring antibiotics. At an industrial level, antibiotics from actinomycete strains are produced by means of submerged fermentations, where one of the major factors negatively affecting bioproductivity is the pellet-shaped biomass growth. The immobilization of microorganisms on properly chosen supports prevents cell-cell aggregation resulting in improving the biosynthetic capability. Thus, novel porous biopolymer-based devices are developed by combining melt mixing and particulate leaching. In particular, polycaprolactone (PCL), polyethylene glycol (PEG), and sodium chloride (NaCl) with different…

Proteomic analysis of Amycolatopsis balhimycina to reveal primary metabolism proteins involved in balhimycin production

Unravelling the DNA sequences carried by Streptomyces coelicolor membrane vesicles

AbstractMembrane vesicles (MVs) are spherical particles with nanoscale dimensions and characterized by the presence of diverse cargos, such as nucleic acids, proteins, lipids, and cellular metabolites. Many examples of (micro)organisms producing MVs are reported in literature. Among them, bacterial MVs are of particular interest because they are now considered as the fourth mechanism of horizontal gene transfer. Streptomyces bacteria are well-known for their ecological roles and ability to synthesize bioactive compounds, with Streptomyces coelicolor being the model organism. It was previously demonstrated that it can produce distinct populations of MVs characterized by different protein and…

4-chlorobenzoate degrading Arthrobacter strains

Endophytic Bacteria and Essential Oil from Origanum vulgare ssp. vulgare Share Some VOCs with an Antibacterial Activity

Medicinal aromatic plants’ essential oils (EOs) are mixtures of volatile compounds showing antimicrobial activity, which could be exploited to face the emerging problem of multi-drug resistance. Their chemical composition can depend on the interactions between the plant and its endophytic microbiota, which is known to synthesize volatile organic compounds (VOCs). However, it is still not clear whether those volatile metabolites can contribute to the composition of the aroma profile of plants’ EOs. The aims of this study were to characterize medicinal plant O. vulgare ssp. vulgare bacterial endophyte VOCs, evaluating their ability to antagonize the growth of opportunistic human pathogens bel…

Antimicrobial activity of EVA blown films containing nisin

A proteomic investigation to explore biochemical capabilities of a lantibiotic producer Microbispora strain

Impact of aminoacid metabolism on antibiotic production in Streptomyces

Dbv4 is a DNA-binding protein acting as a positive regulator of the glycopeptide antibiotic A40926 biosynthesis

Stategie metaboliche indotte dal triptofano in Streptomyces coelicolor

A Genomic, Transcriptomic and Proteomic Look at the GE2270 Producer Planobispora rosea, an Uncommon Actinomycete.

We report the genome sequence of Planobispora rosea ATCC 53733, a mycelium-forming soil-dweller belonging to one of the lesser studied genera of Actinobacteria and producing the thiopeptide GE2270. The P. rosea genome presents considerable convergence in gene organization and function with other members in the family Streptosporangiaceae, with a significant number (44%) of shared orthologs. Patterns of gene expression in P. rosea cultures during exponential and stationary phase have been analyzed using whole transcriptome shotgun sequencing and by proteome analysis. Among the differentially abundant proteins, those involved in protein metabolism are particularly represented, including the G…

AN ABC-TRANSPORTER AND A MEMBRANE PROTEIN OF PLANOBISPORA ROSEA INDUCE ANTIBACTERIAL ACTIVITY IN STREPTOMYCES LIVIDANS

Pirin: A novel redox-sensitive modulator of primary and secondary metabolism in Streptomyces

Pirins are evolutionarily conserved iron-containing proteins that are found in all kingdoms of life, and have been implicated in diverse molecular processes, mostly associated with cellular stress. In the present study, we started from the evidence that the insertional inactivation of pirin-like gene SAM23877_RS18305 (pirA) by Phi C31 Att/Int system-based vectors in spiramycin-producing strain Streptomyces ambofaciens ATCC 23877 resulted in marked effects on central carbon and energy metabolism gene expression, high sensitivity to oxidative injury and repression of polyketide antibiotic production. By using integrated transcriptomic, proteomic and metabolite profiling, together with genetic…

Effect of osmotic and heat shock on the biosynthesis of glycopeptide antibiotics in actinomycetes

Molecular analysis of lichen-associated bacterial communities

The bacterial communities associated with 11 different lichen samples (belonging to eight different species) from different habitats were investigated. The culturable aerobic-heterotrophic fraction of the bacterial communities was isolated from nine lichen samples on protein-rich and sugar-rich/N-free media. Thirty-four bacterial isolates were purified and pooled into groups (phylotypes) by analysis of the ribosomal internal transcribed spacer polymorphism. Twenty five phylotypes were identified, each comprising between one and three isolates. One isolate of each phylotype was partially sequenced and the resulting 16S rRNA gene sequences were compared in a phylogenetic analysis. Three gener…

Rehabilitation of Mediterranean anthropogenic soils using symbiotic wild legume shrubs: Plant establishment and impact on the soil bacterial community structure.

Abstract Susceptibility to desertification in southern Europe is increasing and rehabilitation of desertification-threatened Mediterranean soils is a challenge due to the inhospitality of the environment. In particular, recovery of anthropogenic soils (mainly human-derived artefacts from housing construction and other inert materials or topsoil of terminal phase municipal landfills) cannot rely on spontaneous processes and low-cost/low-impact strategies are needed to prevent desertification. Mediterranean wild legume shrubs have great potential for soil recovery and conservation against desertification, thanks to drought resistance, and their symbiosis with N2-fixing rhizobia and arbuscular…

Effetti sulla comunità batterica di un suolo contaminato indotti dalla crescita di leguminose inoculate con rizobi e funghi Micorizzici: Analisi "Culture Independent" ed isolamenti su piastra

In vitro rescue of zygotic embryos of sour orange, Citrus aurantium L., and their detection based on RFLP analysis

Embryo development in vivo has been studied in four Citrus aurantium L. polyembryonic genotypes. Seeds were collected 65, 85, 105, 125 and 220 days after pollination (DAP). None of the immature seeds harvested 65 and 85 DAP contained visible embryos. A single embryo at a more advanced developmental stage was observed in the central position at the micropylar apex of the embryo sac in about 74% of seeds harvested at 105 DAP, while at 125 and 220 DAP the majority of seeds had two or more embryos at the same developmental stage crowded together. Restriction fragment length polymorphism (RFLP) analysis of low- and high-copy-number nuclear DNA was used to distinguish zygotic from nucellar seedli…

The beta-glucosidase-encoding gene from yeast strains isolated from sicilian musts and wines.

Combining in the melt physical and biological properties of poly(caprolactone) and chlorhexidine to obtain antimicrobial surgical monofilaments.

Bacterial infections on a sutured wound represent a critical problem, and the preparation of suture threads possessing antimicrobial properties is valuable. In this work, poly(caprolactone) (PCL) monofilaments were compounded at the concentration of 1, 2 and 4 % (w/w), respectively, to the antiseptic chlorhexidine diacetate (CHX). The incorporation was carried out in the melt by a single-step methodology, i.e. “online” approach. Mechanical tests revealed that the incorporation of CHX does not significantly change tensile properties of PCL fibres as the thermal profile adopted to prepare the compounded fibres does not compromise the antibacterial activity of CHX. In fact, CHX confers to comp…

Knock-out of the sapA gene in Streptomyces lividans

Inorganic phosphate is a trigger factor for Microbispora sp. ATCC-PTA-5024 growth and NAI-107 production

Background NAI-107, produced by the actinomycete Microbispora sp. ATCC-PTA-5024, is a promising lantibiotic active against Gram-positive bacteria and currently in late preclinical-phase. Lantibiotics (lanthionine-containing antibiotics) are ribosomally synthesized and post-translationally modified peptides (RiPPs), encoded by structural genes as precursor peptides. The biosynthesis of biologically active compounds is developmentally controlled and it depends upon a variety of environmental stimuli and conditions. Inorganic phosphate (Pi) usually negatively regulates biologically-active molecule production in Actinomycetes, while it has been reported to have a positive control on lantibiotic…

The Streptomyces coelicolor Small ORF trpM Stimulates Growth and Morphological Development and Exerts Opposite Effects on Actinorhodin and Calcium-Dependent Antibiotic Production

In actinomycetes, antibiotic production is often associated with a morpho-physiological differentiation program that is regulated by complex molecular and metabolic networks. Many aspects of these regulatory circuits have been already elucidated and many others still deserve further investigations. In this regard, the possible role of many small open reading frames (smORFs) in actinomycete morpho-physiological differentiation is still elusive. In Streptomyces coelicolor, inactivation of the smORF trpM (SCO2038) – whose product modulates L-tryptophan biosynthesis – impairs production of antibiotics and morphological differentiation. Indeed, it was demonstrated that TrpM is able to interact w…

Phosphate-controlled regulators for the biosynthesis of the dalbavancin precursor A40926

Involvement of an Alkane Hydroxylase System of Gordonia sp. Strain SoCg in Degradation of Solid n-Alkanes▿

ABSTRACT Enzymes involved in oxidation of long-chain n -alkanes are still not well known, especially those in Gram-positive bacteria. This work describes the alkane degradation system of the n -alkane degrader actinobacterium Gordonia sp. strain SoCg, which is able to grow on n -alkanes from dodecane (C 12 ) to hexatriacontane (C 36 ) as the sole C source. SoCg harbors in its chromosome a single alk locus carrying six open reading frames (ORFs), which shows 78 to 79% identity with the alkane hydroxylase (AH)-encoding systems of other alkane-degrading actinobacteria. Quantitative reverse transcription-PCR showed that the genes encoding AlkB (alkane 1-monooxygenase), RubA3 (rubredoxin), RubA4…

The endophytic microbiota of Citrus limon is transmitted from seed to shoot highlighting differences of bacterial and fungal community structures

AbstractCitrus limon (L.) Burm. F. is an important evergreen fruit crop whose rhizosphere and phyllosphere microbiota  have been characterized, while seed microbiota is still unknown. Bacterial and fungal endophytes were isolated from C. limon surface-sterilized seeds. The isolated fungi—belonging to Aspergillus, Quambalaria and Bjerkandera genera—and bacteria—belonging to Staphylococcus genus—were characterized for indoleacetic acid production and phosphate solubilization. Next Generation Sequencing based approaches were then used to characterize the endophytic bacterial and fungal microbiota structures of surface-sterilized C. limon seeds and of shoots obtained under aseptic conditions fr…

Regulation of the biosynthesis of the dalbavancin precursor A40926

L’uso di piante mediterranee per salvare i suoli marginali o a rischio desertificazione.

Effect of cold drawing on the antimicrobial and physical properties of PCL/CHX monofilaments

From genome to phenome and back: understanding the high metabolic veratility of Bulkholderia cepacia complex

Characterization of two Streptomyces coelicolor A3(2) small orfs from the major locus of histidine biosynthesis.

Transcriptional analysis of the biosynthetic gene cluster encoding the glycopeptide antibiotic A40926

Microbispora sp. proteomic analysis to study metabolic pathway changes during lantibiotic production

Regulation of the biosynthesis of dalbavancin

Having a look at Microbispora sp. ATCC-PTA-5024, a lantibiotic producer, "from the cradle to the grave" at the proteome level

Microbial technologies for the discovery of novel bioactive metabolites

Soil microbes represent an important source of biologically active compounds. These molecules present original and unexpected structure and are selective inhibitors of their molecular targets. At Biosearch Italia, discovery of new bioactive molecules is mostly carried out through the exploitation of a proprietary strain collection of over 50000 strains, mostly unusual genera of actinomycetes and uncommon filamentous fungi. A critical element in a drug discovery based on microbial extracts is the isolation of unexploited groups of microorganisms that are at the same time good producers of secondary metabolites. Molecular genetics can assist in these efforts. We will review the development an…

Struttura e funzione della cellula procariote

Isolation and characterization of sea urchin P. lividus microbiota from coelomic fluid

An integrated proteomic and metabolomic study to evaluate the effect of nucleus-cytoplasm interaction in a diploid citrus cybrid between sweet orange and lemon.

Key message: Our results provide a comprehensive overview how the alloplasmic condition might lead to a significant improvement in citrus plant breeding, developing varieties more adaptable to a wide range of conditions. Abstract: Citrus cybrids resulting from somatic hybridization hold great potential in plant improvement. They represent effective products resulting from the transfer of organelle-encoded traits into cultivated varieties. In these cases, the plant coordinated array of physiological, biochemical, and molecular functions remains the result of integration among different signals, which derive from the compartmentalized genomes of nucleus, plastids and mitochondria. To dissect …

Composition and geographic variation of the bacterial microbiota associated with the coelomic fluid of the sea urchin Paracentrotus lividus

AbstractIn the present work, culture-based and culture-independent investigations were performed to determine the microbiota structure of the coelomic fluid of Mediterranean sea urchin Paracentrotus lividus individuals collected from two distinct geographical sites neighboring a high-density population bay and a nature reserve, respectively. Next Generation Sequencing analysis of 16S rRNA gene (rDNA) showed that members of the Proteobacteria, Bacteroidetes and Fusobacteria phyla, which have been previously reported to be commonly retrieved from marine invertebrates, dominate the overall population of microorganisms colonizing this liquid tissue, with minority bacterial genera exhibiting rem…

Streptomyces coelicolor extracellular vesicles

This study may reveal the importance of extracellular vesicles in the physiology of S. coelicolor and may also have important biotechnological implications.

Genetic instability in calamondin (Citrus madurensis Lour.) plants derived from somatic embryogenesis induced by diphenylurea derivative

Somatic embryos were regenerated in vitro from calamondin style-stigma explants cultured in the presence of N (6)-benzylaminopurine (BAP) cytokinin and three synthetic phenylurea derivatives, N-(2-chloro-4-pyridyl)-N-phenylurea (4-CPPU), N-phenyl-N'-benzothiazol-6-ylurea (PBU) and N,N'-bis-(2,3-methilendioxyphenyl)urea (2,3-MDPU). The phenylurea derivative compounds tested at micromolar level (12 muM) were able to induce a percentage of responsive explants significantly higher from that obtained with BAP and hormone-free (HF) conditions. In order to verify the genetic stability of the regenerants, 27 plants coming from different embryogenic events were randomly selected from each different …

A breakdown in macromolecular synthesis preceding differentiation in Streptomyces coelicolor A3(2)

Summary: A transitory cessation of growth was recorded in Streptomyces coelicolor A3(2) at the end of vegetative mycelium formation on solid medium. In the same phase a striking reduction in protein and nucleic acid synthesis was detected. Growth and macromolecular synthesis resumed, nearly reaching the original values, when morphological differentiation occurred. It is concluded that a physiological stress occurs within the bacterial population just before the onset of the morphological differentiation.

Identification of a large circular plasmid in Arthrobacter ramosus FG1 encoding the dechlorination of 4-chlorobenzoate

The cypsela (achene) of Echinacea purpurea as a diffusion unit of a community of microorganisms

AbstractEchinacea purpurea is a plant cultivated worldwide for its pharmaceutical properties, mainly related to the stimulation of the immune system in the treatment of respiratory infections. The cypselas (fruits) of E. purpurea were examined in order to investigate the presence, localization and potential function(s) of endophytic microorganisms. Electron and confocal microscopy observations showed that three different components of microorganisms were associated to cypselas of E. purpurea: (i) one endocellular bacterial component in the cotyledons, enclosed within the host membrane; (ii) another more generic bacterial component adhering to the external side of the perianth; and (iii) a f…

Genomic Analysis of Endophytic Bacillus-Related Strains Isolated from the Medicinal Plant Origanum vulgare L. Revealed the Presence of Metabolic Pathways Involved in the Biosynthesis of Bioactive Compounds

Multidrug-resistant pathogens represent a serious threat to human health. The inefficacy of traditional antibiotic drugs could be surmounted through the exploitation of natural bioactive compounds of which medicinal plants are a great reservoir. The finding that bacteria living inside plant tissues, (i.e., the endophytic bacterial microbiome) can influence the synthesis of the aforementioned compounds leads to the necessity of unraveling the mechanisms involved in the determination of this symbiotic relationship. Here, we report the genome sequence of four endophytic bacterial strains isolated from the medicinal plant Origanum vulgare L. and able to antagonize the growth of opportunistic pa…

Phosphate control of balhimycin biosynthesis in Amycolatopsis balhimycina

Evoluzione della comunità batterica di un suolo di discarica sottoposto a fitoremediation con specie di leguminose in associazione con rizobi e funghi micorrizici

Identification of a large catabolic plasmid in Arthrobacter ramosus FG1

Gram positive long chain n-alkane degraders from a hydrocarbon contaminated Sicilian shoreline.

Proprietà di film antimicrobici a base di EVA preparati tramite filmatura in bolla

Primary and secondary metabolism gene expression in Amycolatopsis balhimycina

Complementation of the hisBpx mutation in Streptomyces coelicolor A3(2)”.

Incorporation of nisin in poly (ethylene-co-vinyl acetate) films by melt processing: a study on the antimicrobial properties

Both industry and academia have shown a growing interest in materials with antimicrobial properties suitable for food packaging applications. In this study, we prepared and characterized thin films of ethylene-co-vinyl acetate (EVA) copolymer with antimicrobial properties. The films were prepared with a film blowing process by incorporating a nisin preparation as an antimicrobial agent in the melt. Two grades of EVA containing 14 and 28% (wt/wt) vinyl acetate (EVA 14 and EVA 28, respectively) and two commercial formulations of nisin with different nominal activities were used. The effect of the nisin concentration also was evaluated. The films with the highest antimicrobial activity were th…

The Histidinol Phosphate Phosphatase Involved in Histidine Biosynthetic Pathway Is Encoded by SCO5208 (hisN) in Streptomyces coelicolor A3(2)

Through the screening of a Streptomyces coelicolor genomic library, carried out in a histidinol phosphate phosphatase (HolPase) deficient strain, SCO5208 was identified as the last unknown gene involved in histidine biosynthesis. SCO5208 is a phosphatase, and it can restore the growth in minimal medium in this HolPase deficient strain when cloned in a high or low copy number vector. Moreover, it shares sequence homology with other HolPases recently identified in Actinobacteria. During this work a second phosphatase, SCO2771, sharing no homologies with SCO5208 and all so far described phosphatases was identified. It can complement HolPase activity mutation only at high copy number. Sequence …

Novel Sources of Biodiversity and Biomolecules from Bacteria Isolated from a High Middle Ages Soil Sample in Palermo (Sicily, Italy)

: The urban plan of Palermo (Sicily, Italy) has evolved throughout Punic, Roman, Byzantine, Arab, and Norman ages until it stabilized within the borders that correspond to the current historic center. During the 2012 to 2013 excavation campaign, new remains of the Arab settlement, directly implanted above the structures of the Roman age, were found. The materials investigated in this study derived from the so-called Survey No 3, which consists of a rock cavity of subcylindrical shape covered with calcarenite blocks: it was probably used to dispose of garbage during the Arabic age and its content, derived from daily activities, included grape seeds, scales and bones of fish, small animal bon…

The seed endosphere of Anadenanthera colubrina is inhabited by a complex microbiota, including Methylobacterium spp. and Staphylococcus spp. with potential plant-growth promoting activities

Background and aims: Plant seeds are emerging micro–habitats, whose importance as reservoir and vector of beneficial microbes just begins to be recognized. Here we aimed to characterize the bacterial microbiota of the Anadenanthera colubrina seed endosphere, with special focus to beneficial traits and to the colonization pattern. Methods: Cultivation–dependent (isolation from surface–sterilized seeds) and cultivation–independent (pyrosequencing of 16S rRNA gene from metagenomic seed DNA) analyses, functional tests and microscopical investigations (fluorescence in situ hybridization coupled with confocal laser scanning microscopy (FISH-CLSM) were performed. Results: We isolated several Methy…

Bacterial community structure and removal performances in IFAS-MBRs: A pilot plant case study

Abstract The paper reports the results of an experimental campaign carried out on a University of Cape Town (UCT) integrated fixed-film activated sludge (IFAS) membrane bioreactor (MBR) pilot plant. The pilot plant was analysed in terms of chemical oxygen demand (COD) and nutrients removal, kinetic/stoichiometric parameters, membrane fouling and sludge dewaterability. Moreover, the cultivable bacterial community structure was also analysed. The pilot plant showed excellent COD removal efficiency throughout experiments, with average value higher than 98%, despite the slight variations of the influent wastewater. The achieved nitrification efficiency was close to 98% for most of the experimen…

Artificial chromosomes for antibiotic-producing actinomycetes.

Bacteria belonging to the order Actinomycetales produce most microbial metabolites thus far described, several of which have found applications in medicine and agriculture. However, most strains were discovered by their ability to produce a given molecule and are, therefore, poorly characterized physiologically and genetically. Thus, methodologies for genetic manipulation of actinomycetes are not available and efficient tools have been developed for just a few strains. This constitutes a serious limitation to applying molecular genetics approaches to strain development and structural manipulation of microbial metabolites. To overcome this hurdle, we have developed bacterial artificial chrom…

Microbispora sp. ATCC-PTA-5024 proteomic analysis to study metabolic pathway changes before and during lantibiotic production

Heterologous expression of Nonomuraea sp. ATCC39727 genes in Streptomyces lividans

Identification of SCP2165, a new SCP2-derived plasmid of Streptomyces coelicolor A3(2).

Aims:  Characterization of SCP2165, a plasmid identified in the Gram-positive bacterium Streptomyces coelicolor A3(2). Methods and Results:  Pulsed-field gel electrophoresis (PFGE) of mycelia of a S. coelicolor strain embedded in low melting agarose revealed the presence of a plasmid. Restriction enzyme mapping and sequence analysis of a 2·1 kb fragment revealed that this plasmid could be SCP2. SCP2 and its spontaneous derivative SCP2* are self-transmissible plasmids and have chromosome mobilizing ability (c.m.a.). SCP2* has a c. 1000-fold increased c.m.a. compared with SCP2. Interestingly the plasmid, named SCP2165, shows a c.m.a. from 5 × 10−2 to 1 × 10−1 which is 50–100-fold higher than …

Antimicrobial properties and release profile of monofilaments incorporating chlorhexidine for surgical applications