Due to the diffusion of severe pathogens, everyday life is exposed to the risks of contracting severe diseases. For this reason, efficient antimicrobial surfaces are of paramount importance. In this work we present the first evidences of a new technique to obtain an antibacterial ultra high molecular weight polyethylene based on a non- stoichiometric, visible light responsive, titanium oxide coating. The coating was obtained through a process in which titanium ions, resulting from laser ablation of a corresponding target, were accelerated and implanted on the samples. The samples were tested against a Staphylo- coccus aureus strain, in order to assay their antimicrobial efficacy. Results show that this treatment strongly dis- courages bacterial colonization of the treated surfaces.

We show that, in particular experimental conditions, the time course of the radiant fluxes, measured from a bioluminescent emission of a Vibrio harveyi related strain, collapse after suitable rescaling onto the Gumbel distribution of extreme value theory. We argue that the activation times of the strain luminous emission follow the universal behavior described by this statistical law, in spite of the fact that no extremal process is known to occur.

We present an effective dynamical model for the onset of bacterial bioluminescence, one of the most studied quorum sensing-mediated traits. Our model is built upon simple equations that describe the growth of the bacterial colony, the production and accumulation of autoinducer signal molecules, their sensing within bacterial cells, and the ensuing quorum activation mechanism that triggers bioluminescent emission. The model is directly tested to quantitatively reproduce the experimental distributions of photon emission times, previously measured for bacterial colonies of Vibrio jasicida, a luminescent bacterium belonging to the Harveyi clade, growing in a highly drying environment. A distinctive and novel feature of the proposed model is bioluminescence 'quenching' after a given time elapsed from activation. Using an advanced fitting procedure based on the simulated annealing algorithm, we are able to infer from the experimental observations the biochemical parameters used in the model. Such parameters are in good agreement with the literature data. As a further result, we find that, at least in our experimental conditions, light emission in bioluminescent bacteria appears to originate from a subtle balance between colony growth and quorum activation due to autoinducers diffusion, with the two phenomena occurring on the same time scale. This finding is consistent with a negative feedback mechanism previously reported for Vibrio harveyi.

The molecular mechanisms altered by the traditional mutation and screening approach during the improvement of antibiotic-producing microorganisms are still poorly understood although this information is essential to design rational strategies for industrial strain improvement. In this study, we applied comparative genomics to identify all genetic changes occurring during the development of an erythromycin overproducer obtained using the traditional mutate-and- screen method.Compared with the parental Saccharopolyspora erythraea NRRL 2338, the genome of the overproducing strain presents 117 deletion, 78 insertion and 12 transposition sites, with 71 insertion/deletion sites mapping within coding sequences (CDSs) and generating frame-shift mutations. Single nucleotide variations are present in 144 CDSs. Overall, the genomic variations affect 227 proteins of the overproducing strain and a considerable number of mutations alter genes of key enzymes in the central carbon and nitrogen metabolism and in the biosynthesis of secondary metabolites, resulting in the redirection of common precursors toward erythromycin biosynthesis. Interestingly, several mutations inactivate genes coding for proteins that play fundamental roles in basic transcription and translation machineries including the transcription anti-termination factor NusB and the transcription elongation factor Efp. These mutations, along with those affecting genes coding for pleiotropic or pathway-specific regulators, affect global expression profile as demonstrated by a comparative analysis of the parental and overproducer expression profiles. Genomic data, finally, suggest that the mutate-and-screen process might have been accelerated by mutations in DNA repair genes.This study helps to clarify the mechanisms underlying antibiotic overproduction providing valuable information about new possible molecular targets for rationale strain improvement.

The growing resistance of many strains of bacteria to antibiotics and antiseptics is becoming a serious problem in medicine. Nano-silver is one of the most prominent products in medicine because it exhibits unusual physicochemical properties and a strong biological activity. In this work an innovative silver deposition technology was applied to temporary polyurethane catheters for haemodialysis. The working conditions of catheters were reproduced through laboratory equipment that ensured the flow of deionized water and simulated body fluid inside the lumina at corporeal temperature. The growth and the adhesion of Staphylococcus aureus on the surface of the device were studied through fluorescence microscopy. ICP-AES was adopted to calculate the amount of silver released from the substrate. The stability of the coating during the whole working life of the device was demonstrated through thermo-gravimetric analysis.

In this study, the evidence of electron-dense magnetic inclusions with polyhedral shape in the cytoplasm of Harveyi clade Vibrio strain PS1, a bioluminescent bacterium living in symbiosis with marine organisms, led us to investigate the behavior of this bacterium under exposure to static magnetic fields ranging between 20 and 2000 Gauss. When compared to sham-exposed, the light emission of magnetic field-exposed bacteria growing on solid medium at 18°C ±0.1°C was increased up to two-fold as a function of dose and growth phase. Stimulation of bioluminescence by magnetic field was more pronounced during the post-exponential growth and stationary phase, and was lost when bacteria were grown in the presence of the iron chelator deferoxamine, which caused disassembly of the magnetic inclusions suggesting their involvement in magnetic response. As in luminescent Vibrio spp. bioluminescence is regulated by quorum sensing, possible effects of magnetic field exposure on quorum sensing were investigated. Measurement of mRNA levels by reverse transcriptase real time-PCR demonstrated that luxR regulatory gene and luxCDABE operon coding for luciferase and fatty acid reductase complex were significantly up-regulated in magnetic field-exposed bacteria. In contrast, genes coding for a type III secretion system, whose expression was negatively affected by LuxR, were down-regulated. Up-regulation of luxR paralleled with down-regulation of small RNAs that mediate destabilization of luxR mRNA in quorum sensing signaling pathways. The results of experiments with the well-studied Vibrio campbellii strain BB120 (originally classified as Vibrio harveyi) and derivative mutants unable to synthesize autoinducers suggest that the effects of magnetic fields on quorum sensing may be mediated by AI-2, the interspecies quorum sensing signal molecule.

Rifampin chemoprophylaxis against Neisseria meningitidis infections led to the onset of rifampin resistance in clinical isolates harboring point mutations in the rpoB gene, coding for the RNA polymerase β chain. These resistant strains are rare in medical practice, suggesting their decreased fitness in the human host. In this study, we isolated rifampin-resistant rpoB mutants from hypervirulent serogroup C strain 93/4286 and analyzed their different properties, including the ability to grow/survive in different culture media and in differentiated THP-1 human monocytes and to compete with the wild-type strain in vitro. Our results demonstrate that different rpoB mutations (H553Y, H553R, and S549F) may have different effects, ranging from low- to high-cost effects, on bacterial fitness in vitro. Moreover, we found that the S549F mutation confers temperature sensitivity, possibly explaining why it is observed very rarely in clinical isolates. Comparative high-throughput RNA sequencing analysis of bacteria grown in chemically defined medium demonstrated that the low-cost H553Y substitution resulted in global transcriptional changes that functionally mimic the stringent response. Interestingly, many virulence-associated genes, including those coding for meningococcal type IV pili, porin A, adhesins/invasins, IgA protease, two-partner secretion system HrpA/HrpB, enzymes involved in resistance to oxidative injury, lipooligosaccharide sialylation, and capsular polysaccharide biosynthesis, were downregulated in the H553Y mutant compared to their level of expression in the wild-type strain. These data might account for the reduced capacity of this mutant to grow/survive in differentiated THP-1 cells and explain the rarity of H553Y mutants among clinical isolates.

Polymorphonuclear neutrophil leucocytes (PMNs) are a critical part of innate immune defence against bac- terial pathogens, and only a limited subset of microbes can escape killing by these phagocytic cells. Here we show that Neisseria meningitidis, a leading cause of septicaemia and meningitis, can avoid killing by PMNs and this is dependent on the ability of the bacterium to acquire L-glutamate through its GltT uptake system. We demonstrate that the uptake of available L- glutamate promotes N. meningitidis evasion of PMN reactive oxygen species produced by the oxidative burst. In the meningococcus, L-glutamate is converted to glutathione, a key molecule for maintaining intrac- ellular redox potential, which protects the bacterium from reactive oxygen species such as hydrogen per- oxide. We show that this mechanism contributes to the ability of N. meningitidis to cause bacteraemia, a criti- cal step in the disease process during infections caused by this important human pathogen.

The spreading of pathogens represents a serious threat for human beings. Consequently, efficient antimicrobial surfaces are needed in order to reduce risks of contracting severe diseases. In this work we present the first evidences of a new technique to obtain a highly antibacterial Ultra High Molecular Weight Polyethylene (UHMWPE) based on a non-stoichiometric titanium oxide coating, visible-light responsive, obtained through ion implantation.

Both academic and industrial laboratories currently use constraint - based reconstruction methods to predict optimal genetic modifications aiming at improving the yield of chemical production. Streptomyces ambofaciens, a prolific producer of bioactive compounds has been studied with different modeling tools . The interest is linked to its ability to produce a wide range of secondary metabolites such as spiramycin, kinamycin, antimycin and stambomycins, and novel polyketides with antibacterial and antiproliferative activities. In this study, the metabolic pattern of Streptomyces ambofaciens has been globally explored: a set of candidate overexpression gene targets supposed to lead to spiramycin overproduction have been evidenced through metabolic modeling. Model predictions were experimentally validated by genetic manipulation of the ethylmalonyl-CoA metabolic node, providing evidence that spiramycin productivity may be increased by enhancing the carbon flow through this pathway. The goal was achieved by over - expressing the ccr paralog srm4 in an ad hoc engineered plasmid. The first metabolic reconstruction of S. ambofaciens and the successful experimental validation of model predictions have been described and the validity and the importance of in silico modeling tools for the overproduction of molecules with a biotechnological interest demonstrated. As a result, the proposed metabolic reconstruction represents a solid platform for the future exploitation of S. ambofaciens biotechnological potential.

Bacteria synthesise an extremely rich repertoire of odorous compounds. Unfortunately this feature is not much known and bacteria are more often associated with the bad smell of rotting material. The focus of this review is on the capability of bacteria to synthesize odorous volatiles, either when they are free-living in soil and aquatic environments or when they establish symbiotic association with plants and animals, with specific reference to humans. In particular, the biological and ecological significance of bacterial volatiles and their biotechnological perspectives are covered in this present work.

We discuss the mechanisms of bacterial Quorum Sensing, the biophysical phenomenon point- ing out a social behavior in bacteria, highlighting thus the very complex structure of these sys- tems. Actually, bacterial bioluminescence is an example of a quorum sensing mediated prop- erty. We show that the distribution of the activation times of the bioluminescent emission follows the universal behavior described by the Gumbel distribution of extreme value statis- tics. We provide further evidence on the system size scaling of bioluminescence, showing that the relation between cell density and total number of photons radiated by bacteria is highly non-linear.

We present the study and the analyses of a transmission line for radiofrequency (RF) irradiation of bacteria belonging to Vibrio harveyi-related strain PS1, a bioluminescent bacterium living in symbiosis with many marine organisms. The bioluminescence represents a new biologic indicator which is useful for studying the behaviour of living samples in the presence of RF waves due to the modern communication systems. A suitable transmission line, used as an irradiating cell and tested up to the maximum frequency used by the global system for mobile communications and universal mobile telecommunications system transmissions, was characterized. In this experiment, the RF voltage applied to the transmission line was 1V. Due to short dimensions of the line and the applied high frequencies, standing waves were produced in addition to progressing waves and the electric field strength varies particularly along the longitudinal direction. The magnetic field map was not strongly linked to the electric one due to the presence of standing waves and of the outgoing irradiation. RF fields were measured by two homemade suitable probes able to diagnostic fields of high frequency. The field measurements were performed without any specimens inside the line. Being our sample made of living matter, the real field was modified and its value was estimated by a simulation code. The bioluminescence experiments were performed only at 900MHz for two different measured electric fields, 53 and 140V/m. The light emission was measured right from the beginning and after 7 and 25 h. Under RF irradiation, we found that the bioluminescence activity decreased. Compared with the control sample, the diminution was 6.8% and 44% after 7 and 25 h of irradiation, respectively, both with the low or high field. No changes of the survival factor for all the samples were observed. Besides, to understand the emission processes, we operated the deconvolution of the spectra by two Gaussian curves. The Gaussian peaks were approximately centered at 460 nm and 490 nm. The 490 nm peak was higher than the control one. Under RF, the 490 nm peak decreased compared to the 460 nm one. The decreasing was stronger for the sample in the higher field. The ratio of the emission area of the 490 nm to 460 nm was 5 for the control sample. It decreased up to 1.6 for the samples under RF. The bioluminescence improves the DNA repair by photoreactivation, and there is evidence that photolyase is preferentially activated by blue/violet light. Our finding suggests that RF exposure may stimulate DNA repair by shifting the emission spectra from blue/green (490 nm) to blue/violet (460 nm). VC 2012 American Institute of Physics.

Following its introduction in 1967, rifampicin has become a mainstay of therapy in the treatment of tuberculosis, leprosy and many other widespread diseases. Its potent antibacterial activity is due to specific inhibition of bacterial RNA polymerase. However, resistance to rifampicin was reported shortly after its introduction in the medical practice. Studies in the model organism Escherichia coli helped to define the molecular mechanism of rifampicin-resistance demonstrating that resistance is mostly due to chromosomal mutations in rpoB gene encoding the RNA polymerase β chain. These studies also revealed the amazing potential of the molecular genetics to elucidate the structure-function relationships in bacterial RNA polymerase. The scope of this paper is to illustrate how rifampicin-resistance has been recently exploited to better understand the regulatory mechanisms that control bacterial cell physiology and virulence, and how this information has been used to maneuver, on a global scale, gene expression in bacteria of industrial interest. In particular, we reviewed recent literature regarding: (i) the effects of rpoB mutations conferring rifampicin-resistance on transcription dynamics, bacterial fitness, physiology, metabolism and virulence; (ii) the occurrence in nature of "mutant-type" or duplicated rifampicin-resistant RNA polymerases; and (iii) the RNA polymerase genetic engineering method for strain improvement and drug discovery.

We have previously shown that during the late stages of the infectious process serogroup B meningococci (MenB) are able to escape the phagosome of in vitro-infected human epithelial cells. Then, they multiply in the cytosolic environment and spread intracellularly and to surrounding cells by exploiting the microtubule cytoskeleton as suggested by results of infections in the presence of microtubule inhibitors and the evidence of "nanotubes" connecting neighboring cells. In this study, by using microtubule binding assays with purified microtubule "asters" and "bundles" and microtubule "bundles" synthesized in vitro we demonstrate that the MenB capsule directly mediates the interaction between bacteria and microtubules. The direct interaction between the microtubules and the MenB capsular polysaccharide was confirmed by co-immunoprecipitation experiments. Unexpectedly, serogroup C meningococci (MenC), which have a capsular polysaccharide that differs from that of MenB only by its anomeric linkage, α(2→9) instead of α(2→8), were not able to interact with the microtubules, and the lack of interaction was not due to capsular polysaccharide O-acetylation that takes place in most of MenC strains but not in MenB. Moreover, we demonstrate that the MenB capsular polysaccharide inhibits tubulin polymerization in vitro. Thus, at variance with MenC, MenB may interfere with microtubule dynamics during cell infection.

Abstract: Strain SPC-1T was isolated from the phyllosphere of Cynara cardunculus L. var. sylvestris (Lamk) Fiori (wild cardoon), a Mediterranean native plant considered the wild ancestor of the globe artichoke and cultivated cardoon. This Gram-negative, catalase-positive, oxidase-negative, non-spore-forming, rod-shaped and non-motile strain secreted copious amounts of an exopolysaccharide and formed slimy, viscous, orange-pigmented colonies and grew optimally at around pH 6.0-6.5 and 26-30°C in the presence of 0-0.5% NaCl. Phylogenetic analysis based on comparisons of 16S rRNA gene sequences demonstrated that SPC-1T clustered together with species of the genus Sphingomonas sensu strictu. The G+C content of the DNA (66.1 mol%), the presence of Q-10 as the predominant ubiquinone, sym-homospermidine as the predominant polyamine, and 2-hydroxymiristic acid (C14:0 2-OH) as the major hydroxylated fatty acid, the absence of 3-hydroxy fatty acids and the presence of sphingoglycolipid supported this taxonomical position. 16S rRNA gene sequence analysis showed that SPC-1 was most closely related to Sphingomonas hankookensis ODN7T, Sphingomonas insulae DS-28T and Sphingomonas panni C52T (98.19%, 97.91% and 97.11% similarity, respectively). However, DNA-DNA hybridization analysis did not reveal any relatedness at the species level. Further differences were apparent in biochemical traits, and fatty acid, quinone and polyamine profiles leading us to conclude that strain SPC-1T (JCM 17498; ITEM 13494) represents a new species of Sphingomonas, for which the name Sphingomonas cynarae sp. nov. is proposed. A component analysis of the exopolysaccharide (named SPC-1T EPS) suggested that it represents a novel type of sphingan containing glucose, rhamnose, mannose and galactose, while glucuronic acid, which is commonly found in sphingans, was not detected.

Spiramycin is a macrolide antibiotic and antiparasitic that is used to treat toxoplasmosis and various other infections of soft tissues. In the current study, we evaluated the effects of α-cyclodextrin, β-cyclodextrin, or methyl-β-cyclodextrin supplementation to a synthetic culture medium on biomass and spiramycin production by Streptomyces ambofaciens ATCC 23877. We found a high stimulatory effect on spiramycin production when the culture medium was supplemented with 0.5% (w/v) methyl-βcyclodextrin, whereas α-cyclodextrin or β-cyclodextrin weakly enhanced antibiotic yields. As the stimulation of antibiotic production could be because of spiramycin complexation with cyclodextrins with effects on antibiotic stability and/or efflux, we analyzed the possible formation of complexes by physical−chemical methods. The results of Job plot experiment highlighted the formation of a nonhost@guest complex methyl-β-cyclodextrin@ spiramycin I in the stoichiometric ratio of 3:1 while they excluded the formation of complex between spiramycin I and α- orβ-cyclodextrin. Fourier-transform infrared spectroscopy measurements were then carried out to characterize the methyl-β-cyclodextrin@spiramycin I complex and individuate the chemical groups involved in the binding mechanism. These findings may help to improve the spiramycin fermentation process, providing at the same time a new device for better delivery of the antibiotic at the site of infection by methyl-β-cyclodextrin complexation, as it has been well-documented for other bioactive molecules.

In contrast to the widely accepted consensus of the existence of a single RNA polymerase in bacteria, several actinomycetes have been recently shown to possess two forms of RNA polymerases due the to co-existence of two rpoB paralogs in their genome. However, the biological significance of the rpoB duplication is obscure. In this study we have determined the genome sequence of the lipoglycopeptide antibiotic A40926 producer Nonomuraea gerenzanensis ATCC 39727, an actinomycete with a large genome and two rpoB genes, i.e. rpoB(S) (the wild-type gene) and rpoB(R) (the mutant-type gene). We next analyzed the transcriptional and metabolite profiles in the wild-type gene and in two derivative strains over-expressing either rpoB(R) or a mutated form of this gene to explore the physiological role and biotechnological potential of the "mutant-type" RNA polymerase. We show that rpoB(R) controls antibiotic production and a wide range of metabolic adaptive behaviors in response to environmental pH. This may give interesting perspectives also with regard to biotechnological applications.

Bacteria undergoing environmental effects is extremely interesting for structural, mechanistic, and evolutionary implications. Luminescent bacteria that have evolved in a specific ambient have developed particular responses and their behavior can give us new suggestions on the task and production of luciferina proteins. To analyze the UV interaction under controlled laboratory conditions, we used photoluminescent bacterial strains belonging to a new species evolutionarily close to Vibrio harveyi sampled from a coastal cave with a high radon content that generates ionizing radiation. The survival of the bacterial strains was analyzed, in the light and in the dark, following a variety of genotoxic treatments including UV radiation exposure. The strains were irradiated by a germicide lamp. The results demonstrated that most of the strains exhibited a low rate of survival after the UV exposure. After irradiation by visible light following the UV exposure, all strains showed a high capability of photoreactivation when grown. This capability was quite unexpected because these bacteria were sampled from a dark ambient without UV radiation. This leads us to hypothesize that the photoreactivation in these bacteria might have been evolved to repair DNA lesions also induced by different radiation sources other than UV (e.g., x-ray) and that the luminescent bacteria might use their own light emission to carry out the photoreactivation. The high capability of photoreactivation of these bacteria was also justified by the results of deconvolution. The deconvolution was applied to the emission spectra and it was able to show evidence of different light peaks. The presence of the visible peak could control the photolysis enzyme.