Search

The mechanisms and regulation of RNA degradation in mycobacteria have been subject to increased interest following the identification of interplay between RNA metabolism and drug resistance. Mycobacteria encode multiple ribonucleases that are predicted to participate in mRNA degradation and/or processing of stable RNAs. RNase E is an endoribonuclease hypothesized to play a major role in mRNA degradation due to its essentiality in mycobacteria and its role in mRNA degradation in gram- negative bacteria. Here, we defined the impact of RNase E on mRNA degradation rates transcriptome- wide in the non-pathogenic modelMycolicibacterium smegmatis. RNase E played a rate-limiting role in the degradation of at least 89% of protein-coding genes, with leadered transcripts generally being more affected by RNase E repression than leaderless transcripts. There was an apparent global slowing of transcription in response to knockdown of RNase E, suggesting thatM. smegmatisregulates transcription in responses to changes in mRNA degradation. This compensation was incomplete, as the abundance of most transcripts increased upon RNase E knockdown. We assessed the sequence preferences for cleavage by RNase E transcriptome-wide in bothM. smegmatisandM. tuberculosis, and found a consistent bias for cleavage in C-rich regions. Purified RNase E had a clear preference for cleavage immediately upstream of cytidines, distinct from the sequence preferences of RNase E in gram-negatives. We furthermore report a high-resolution map of mRNA cleavage sites inM. tuberculosis, which occur primarily within the RNase E-preferred sequence context, confirming RNase E as a broad contributor toM. tuberculosistranscriptome structure.

In the last decades, several anodizing processes for Mg alloys have been proposed to achieve a good wear and corrosion resistance combination. In particular, Electro-Chemical Oxidation (ECO) showed an improved dense and compact anodized layer compared to other anodizing processes carried out above the dielectric breakdown voltage, such as Plasma Electrolytic Oxidation (PEO). However, the influence of the ECO treatment on the tribological behavior and cyclic mechanical performance of Mg alloys has not been investigated yet. This paper reports on the influence of ECO on dry sliding behavior (vs. 100Cr6 bearing steel (block-on-ring contact geometry)) and rotating bending fatigue performance of the rare earth (RE)-containing Mg alloy EV31A-T6, comparing it with both untreated EV31A-T6 and PEO-treated EV31A-T6, used as benchmarks. The ECO-treated alloy showed improved tribological behavior (critical load for coating failure one order of magnitude higher and coefficient of friction 40% lower than for PEO) and fatigue strength (no decrease for ECO-treated samples compared to the untreated alloy, while PEO-treated samples induced a 15% decrease) due to the increased compactness and lower defectivity of the anodized layer, induced by the minimization of destructive arc discharges during coating growth. In addition, the ECO treatment significantly improved wear resistance compared to the untreated alloy, avoiding, at the same time, the decrease in fatigue strength, which typically occurs after PEO. Therefore, the ECO process can be applied to improve wear resistance without decreasing the fatigue strength of high-performance components.

Considering the increasing interest for Al alloys in the packaging industry, where component lightweighting allows higher process efficiency, as well as the increasing use of additively manufactured parts, which may not show a satisfactory tribological behaviour in the as-built condition, anodising by Electrochemical Oxidation (ECO) and Plasma Electrolytic Oxidation (PEO) was applied to as-built A357 (AlSi7Mg0.6) produced by Laser Powder Bed Fusion (LPBF). ECO allowed to compensate surface and surface-connected defects in LPBF A357 and produced a thicker, less-defective anodic oxide than PEO, providing higher wear resistance whilst maintaining a lower Coefficient of Friction (COF) in dry sliding tests against 100Cr6 steel. Polishing before ECO only resulted in negligible friction and wear improvements, even if it improved the compactness of the anodic oxide.

Mycobacterium abscessuscauses severe, virtually incurable disease in young patients with cystic fibrosis. Little is known inM. abscessusabout the roles of small regulatory RNAs (sRNA) in gene expression regulation. Here, we show that the sRNA B11 controls gene expression and virulence-associated phenotypes in this pathogen. B11 deletion from the smooth strain ATCC_19977 produced a rough colony morphology, increased pro-inflammatory signaling and virulence inin-vivoinfection models, and increased resistance to clinically relevant antibiotics. Examination of clinical isolate cohorts revealed some isolates with B11 mutations or reduced expression. We used RNAseq and proteomics to investigate the effects of B11 on gene expression and test the impact of two mutations found in clinical isolates. Approximate 230 genes were differentially expressed in the B11 deletion mutant. Strains with the clinical B11 mutations showed similar expression trends to the deletion mutant but of a lesser magnitude, suggesting partial loss of function. Among genes upregulated in the B11 mutant, there was a strong enrichment for genes with B11-complementary sequences in their predicted ribosome binding sites (RBS), consistent with a model of translational repression via base-pairing of B11 to RBSs. Comparing the proteomes similarly revealed that upregulated proteins were strongly enriched for B11-complementary sequences in their RBS, consistent with B11 functioning as a negative regulator through direct binding of target mRNAs. Intriguingly, the genes upregulated in the absence of B11 included components of the ESX-4 secretion system, known to be critical forM. abscessusvirulence. One of these genes had a B11-complementary sequence at its RBS, and fusing the UTR of this gene to a reporter was sufficient to make the reporter suppressible by B11. Taken together, our data show that B11 may act as either a negative or positive regulator with pleiotropic effects on gene expression and clinically important phenotypes inM. abscessus. The presence of hypomorphic B11 mutations in clinical strains supports the idea that lower B11 activity may be advantageous forM. abscessusin some clinical contexts. To our knowledge, this is the first report of the role of an sRNA inM. abscessus.

International audience; The conservation of excavated wrecks or abandoned aircraft is of cultural and historical interest. A first step in understanding the challenges posed by this type of cultural heritage and in finding solutions for its conservation is to document the constituent materials and their subsequent alterations. In the project PROCRAFT, aircraft wrecks from six nations involved in WWII were collected and then studied. Analysis of the elemental composition together with archival research allowed identification of the wrought aluminium alloys used in the non-structural and structural parts of the aircraft. Three manufacturing processes were recognised: rolling, extrusion and forging. The corrosion protection treatments applied during manufacturing of the aircraft parts were also examined, which revealed cladding with a thin aluminium foil, anodisation or the application of primer/paint. Based on typical examples of the alterations that occurred in the metal components, we demonstrate the importance of this original protection against the aggressiveness of the environment.

Epilithic bacteria play a fundamental role in the conservation of cultural heritage (CH) materials. On stones, bacterial communities cause both degradation and bioprotection actions. Bronze biocorrosion in non-burial conditions is rarely studied. Only few studies have examined the relationship between bacteria communities and the chemical composi- tion of patinas (surface degradation layers). A better comprehension of bacterial communities growing on our CH is fundamental not only to understand the related decay mechanisms but also to foresee possible shifts in their compo- sition due to climate change. The present study aims at (1) characterizing bacterial communities on bronze andmarble statues; (2) evaluating the differences in bacterial communities' composition and abundance occurring between differ- ent patina types on different statues; and (3) providing indications about a representative bacterial community which can be used in laboratory tests to better understand their influence on artefact decay. Chemical and biological charac- terization ofdifferent patinas were carried out by sampling bronze and marble statues in Bologna and Ravenna (Italy), using EDS/Raman spectroscopy and MinION-based 16SrRNA sequencing. Significant statistical differences were found in bacterial composition between marble and bronze statues, and among marble patinas in different statues and in the same statue. Marble surfaces showed high microbial diversity and were characterized mainly by Cyanobacteria, Proteobacteria and Deinococcus-Thermus. Bronze patinas showed low taxa diversity and were dominated by copper-resistant Proteobacteria. The copper biocidal effect is evident in greenish marble areas affected by the leaching of copper salts, where the bacterial community is absent. Here, Ca and Cu oxa- lates are present because of the biological reaction of living organisms to Cu ions, leading to metabolic product secre- tions, such as oxalic acid. Therefore, a better knowledge on the interaction between bacteria communities and patinas has been achieved.

The acidity of soils significantly reduces the productivity of legumes mainly because of the detrimental effects of hydrogen ions on the legume plants, leading to the establishment of an inefficient symbiosis and poor biological nitrogen fixation. We recently reported the analysis of the fully sequenced genome of Rhizobium favelukesii LPU83, an alfalfa-nodulating rhizobium with a remarkable ability to grow, nodulate and compete in acidic conditions. To gain more insight into the genetic mechanisms leading to acid tolerance in R. favelukesii LPU83, we constructed a transposon mutant library and screened for mutants displaying a more acid-sensitive phenotype than the parental strain. We identified mutant Tn833 carrying a single-transposon insertion within LPU83_2531, an uncharacterized short ORF located immediately upstream from ubiF homolog. This gene encodes a protein with an enzymatic activity involved in the biosynthesis of ubiquinone. As the transposon was inserted near the 3′ end of LPU83_2531 and these genes are cotranscribed as a part of the same operon, we hypothesized that the phenotype in Tn833 is most likely due to a polar effect on ubiF transcription. We found that a mutant in ubiF was impaired to grow at low pH and other abiotic stresses including 5 mM ascorbate and 0.500 mM Zn(2+). Although the ubiF mutant retained the ability to nodulate alfalfa and Phaseolus vulgaris, it was unable to compete with the R. favelukesii LPU83 wild-type strain for nodulation in Medicago sativa and P. vulgaris, suggesting that ubiF is important for competitiveness. Here, we report for the first time an ubiF homolog being essential for nodulation competitiveness and tolerance to specific stresses in rhizobia. GRAPHICAL ABSTRACT: [Image: see text]

Research background. In recent decades, laccases (p-diphenol-dioxygen oxidoreductases; EC 1.10.3.2) have attracted the attention of researchers due to their wide range of biotechnological and industrial applications. Laccases can oxidize a variety of organic and inorganic compounds, making them suitable as biocatalysts in biotechnological processes. Even though the most traditionally used laccases in the industry are of fungal origin, bacterial laccases have shown an enormous potential given their ability to act on several substrates and in multiple conditions. The present study aims to characterize a plasmid-encoded laccase-like multicopper oxidase (LMCO) from Ochrobactrum sp. BF15, a bacterial strain previously isolated from polluted soil. Experimental approach. We used in silico profile hidden Markov models to identify novel laccase-like genes in Ochrobactrum sp. BF15. For laccase characterization, we performed heterologous expression in Escherichia coli, purification and activity measurement on typical laccase substrates. Results and conclusions. Profile hidden Markov models allowed us to identify a novel LMCO, named Lac80. In silico analysis of Lac80 revealed the presence of three conserved copper oxidase domains characteristic of three-domain laccases. We successfully expressed Lac80 heterologously in E. coli, allowing us to purify the protein for further activity evaluation. Of thirteen typical laccase substrates tested, Lac80 showed lower activity on 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), pyrocatechol, pyrogallol and vanillic acid, and higher activity on 2,6-dimethoxyphenol. Novelty and scientific contribution. Our results show Lac80 as a promising laccase for use in industrial applications. The present work shows the relevance of bacterial laccases and highlights the importance of environmental plasmids as valuable sources of new genes encoding enzymes with potential use in biotechnological processes., Pozadina istraživanja. Posljednjih su desetljeća lakaze (p-difenol:dioksigen oksidoreduktaze; EC 1.10.3.2) privukle pažnju znanstvenika zbog raznolikosti moguće primjene u biotehnologiji i industriji. Lakaze mogu oksidirati različite organske i anorganske spojeve u biokatalizatore prikladne za biotehnološke procese. Iako se u industriji najčešće koriste lakaze iz plijesni, lakaze iz bakterija pokazale su veliki potencijal primjene jer djeluju na više raznih supstrata u različitim uvjetima. Svrha je ovoga rada bila opisati plazmidno kodiranu višebakrenu oksidazu sličnu lakazama iz bakterije Ochrobactrum sp. BF15, prethodno izolirane iz zagađenog tla. Eksperimentalni pristup. Za identifikaciju novih gena za oksidazu sličnu lakazi u bakteriji Ochrobactrum sp. BF15 koristili smo in silico profil skrivenih Markovljevih modela. Da bismo okarakterizirali lakazu, proveli smo heterolognu ekspresiju gena u bakteriji Escherichia coli, pročistili enzim i mjerili njegovu aktivnost u supstratima karakterističnim za lakazu. Rezultati i zaključci. Dobiveni profili skrivenih Markovljevih modela omogućili su identifikaciju nove oksidaze slične lakazi, koju smo nazvali Lac80. In silico analiza nove Lac80 pokazala je prisutnost triju konzerviranih domena bakrene oksidaze karakterističnih za lakaze s tri domene. Uspješno smo proveli heterolognu ekspresiju Lac80 u E. coli, čime nam je omogućeno pročišćavanje proteina za daljnju analizu aktivnosti. Od trinaest ispitanih supstrata tipičnih za lakazu, Lac80 je imao slabiju aktivnost u 2,2’-azino-bis(3-etilbenzotiazolin-6-sulfonskoj kiselini) (ABTS), pirokateholu, pirogalolu i vanilinskoj kiselini, te veću aktivnost u 2,6-dimetoksifenolu. Novina i znanstveni doprinos. Dobiveni rezultati pokazuju da Lac80 može imati obećavajuću primjenu u industriji. U ovom je radu prikazan značaj lakaza iz bakterija te je naglašena važnost plazmida izoliranih iz okoliša kao vrijednih izvora novih gena koji kodiraju enzime za primjenu u biotehnološkim procesima.

Objective Restriction-Modification (R-M) systems are ubiquitous in bacteria and were considered for years as rudimentary immune systems that protect bacterial cells from foreign DNA. Currently, these R-M systems are recognized as important players in global gene expression and other cellular processes such us virulence and evolution of genomes. Here, we report the role of the unique DNA methyltransferase in Mycobacterium smegmatis, which shows a moderate degree of sequence similarity to MamA, a previously characterized methyltransferase that affects gene expression in Mycobacterium tuberculosis and is important for survival under hypoxic conditions. Results We found that depletion of mamA levels impairs growth and produces elongated cell bodies. Microscopy revealed irregular septation and unevenly distributed DNA, with large areas devoid of DNA and small DNA-free cells. Deletion of MSMEG_3214, a predicted endonuclease-encoding gene co-transcribed with mamA, restored the WT growth phenotype in a mamA-depleted background. Our results suggest that the mamA-depletion phenotype can be explained by DNA cleavage by the apparent cognate restriction endonuclease MSMEG_3214. In addition, in silico analysis predicts that both MamA methyltransferase and MSMEG_3214 endonuclease recognize the same palindromic DNA sequence. We propose that MamA and MSMEG_3214 constitute a previously undescribed R-M system in M. smegmatis.

The application of protective coatings for the conservation of outdoor metallic artworks is currently considered the best approach actively employed. This paper focuses on the evaluation of the protectiveness of an optimised fluorosilane (FA-MS) coating with low occupational hazard impact and high inhibition efficiency. FA-MS coating was applied on a typical historical as-cast bronze (Cu-Sn-Zn-Pb), artificially patinated by synthetic acid rain dropping, to simulate natural unsheltered patinas. Subsequently, the patinated bronze was coated by brushing and then artificially aged by synthetic rain dropping. In order to investigate the coating-patina system, a multi-analytical approach was applied including scanning electron microscopy coupled with elemental analysis (FEG-SEM with EDS), microstructural analysis of cross-sections produced by Focused Ion Beam milling (FIB) and X-ray Photoelectron Spectroscopy (XPS). The coating homogenously covered the whole patinated surface producing a transparent layer few μms thick. The weathering solutions were also analysed for the assessment of metallic cations release. FA-MS showed excellent protective performances for the patinated bronze exposed to simulated outdoor conditions. Very high values of inhibiting efficiency in terms of metal release in the weathering solutions (up to 99%) were measured. In order to investigate all the necessary requirements for application in conservation field, the durability of FA-MS was also tested on a real bronze artistic casting, in terms of limiting the alloy composition changes (measured by in situ XRF) over time. A good durability of the coating was observed during the ongoing field exposure test. Lastly, reversibility of FA-MS was investigated by applying different mechanical cleaning methods, such as brushing and blasting. The reversibility was assessed by the evaluation of the morphology (colour change, surface roughness and optical microscopy) and the composition (SEM-EDS and Py-GC-MS chromatography) of the cleaned surfaces. Cleaning by brushing proved to be an efficient method for removing this coating without significantly altering the bronze patina.

This study characterizes the mechanical performance of the AlSi10Mg alloy produced by powder bed fusion-laser beam (PBF-LB) subjected to two combined cycles consisting of multilayer coating deposition (electroless nickel (Ni-P) + diamond-like carbon (DLC)) and heat treatment. In particular, the DLC deposition phase replaces the artificial aging step in the T5 and T6 heat treatments, obtaining the following post-production cycles: (i) Ni-P + DLC deposition and (ii) rapid solution (SHTR) (10 min at 510 °C) before Ni-P + DLC deposition. Microstructural characterization shows no appreciable modifications in the morphology and dimensions of the hard Si-rich phase of the eutectic network and secondary spheroidal Si phase. However, overaging phenomena induced by DLC coating deposition and differences in elastic-plastic properties between the multilayer coating and the PBF-LB AlSi10Mg substrate lead to a reduction in tensile strength by up to 31% and a significant decrease in ductility by up to 58%. In contrast, higher resistance to crack opening thanks to improved surface hardness and residual compressive stresses of the coating and reduced defect sensitivity of the substrate increase the fatigue resistance by 54% in T5-coated alloy and 24% in T6R-coated alloy. Moreover, the coating remains well adherent to the substrate during fatigue testing, not becoming a source of fatigue cracks.

The San Carlo Colossus, known as San Carlone, is a monument constituted by an internal stone pillar support to which a wrought iron structure is attached. Embossed copper sheets are fixed to the iron structure to give the final shape to the monument. After more than 300 years of outdoor exposure, this statue represents an opportunity for an in-depth investigation of long-term galvanic coupling between wrought iron and copper. Most iron elements of the San Carlone appeared in good conservation conditions with scarce evidence of galvanic corrosion. In some cases, the same iron bars presented some portions in good conservation conditions and other nearby portions with active corrosion. The aim of the present study was to investigate the possible factors correlated with such mild galvanic corrosion of wrought iron elements despite the widespread direct contact with copper for more than 300 years. Optical and electronic microscopy and compositional analyses were carried out on representative samples. Furthermore, polarisation resistance measurements were performed both on-site and in a laboratory. The results revealed that the iron bulk composition showed a ferritic microstructure with coarse grains. On the other hand, the surface corrosion products were mainly composed of goethite and lepidocrocite. Electrochemical analyses showed good corrosion resistance of both the bulk and surface of the wrought iron, and galvanic corrosion is not occurring probably due to the iron’s relatively noble corrosion potential. The few areas where iron corrosion was observed are apparently related to environmental factors, such as the presence of thick deposits and to the presence of hygroscopic deposits that create localized microclimatic conditions on the surface of the monument.

Despite the existence of well-characterized, canonical mutations that confer high-level drug resistance to Mycobacterium tuberculosis (Mtb), there is evidence that drug resistance mechanisms are more complex than simple acquisition of such mutations. Recent studies have shown that Mtb can acquire non-canonical resistance-associated mutations that confer survival advantages in the presence of certain drugs, likely acting as stepping-stones for acquisition of high-level resistance. Rv2752c/rnj, encoding RNase J, is disproportionately mutated in drug-resistant clinical Mtb isolates. Here we show that deletion of rnj confers increased tolerance to lethal concentrations of several drugs. RNAseq revealed that RNase J affects expression of a subset of genes enriched for PE/PPE genes and stable RNAs and is key for proper 23S rRNA maturation. Gene expression differences implicated two sRNAs and ppe50-ppe51 as important contributors to the drug tolerance phenotype. In addition, we found that in the absence of RNase J, many short RNA fragments accumulate because they are degraded at slower rates. We show that the accumulated transcript fragments are targets of RNase J and are characterized by strong secondary structure and high G+C content, indicating that RNase J has a rate-limiting role in degradation of highly structured RNAs. Taken together, our results demonstrate that RNase J indirectly affects drug tolerance, as well as reveal the endogenous roles of RNase J in mycobacterial RNA metabolism.

The paper deals with monitoring and analyzing the indoor environmental parameters through remote data collection to evaluate the pollution and moisture infiltration effects on aircraft heritage conservation. First, based on the meteorological and pollution data, the moisture penetration and airborne pollution infiltration into indoor spaces of a heritage site (hangar) with stored historic aircrafts are determined. The hangar under investigation is located in the Aviation museum Kbely, Prague, Czech Republic. The determination is performed by wet/dry cycles (fluctuations) evaluation and applying ISO 11844 methodology to outdoor pollution infiltration into the interior. Next, a time of wetness (ToW) is determined indoors according to ISO 9223, rather as an environmental than a surface parameter as dewing and exceeding high humidity level (approxl RH 80% at T>0 °C) are considered. The actual moisture adsorption onto polluted surfaces of aircraft artifacts is then dependent on the hygroscopic corrosion products developed. Such an adsorption prolongs actual surface ToW. In addition to ToW, even the deposition rate of indoor pollutants, particularly sulphur dioxide and chlorides, are considered and the atmosphere corrosivity is estimated by applying the ISO standardized statistical models for aluminium. The resulting iso-corrosivity figures out the aggressiveness of the hangar environment from the point of view of aircraft material susceptibility to corrosion and degradation.

Few systematic studies on the correlation between alloy microstructure and mechanical failure of the AlSi10Mg alloy produced by laser-based powder bed fusion (L-PBF) are available in the literature. This work investigates the fracture mechanisms of the L-PBF AlSi10Mg alloy in as-built (AB) condition and after three different heat treatments (T5 (4 h at 160 °C), standard T6 (T6B) (1 h at 540 °C followed by 4 h at 160 °C), and rapid T6 (T6R) (10 min at 510 °C followed by 6 h at 160 °C)). In-situ tensile tests were conducted with scanning electron microscopy combined with electron backscattering diffraction. In all samples the crack nucleation was at defects. In AB and T5, the interconnected Si network fostered damage at low strain due to the formation of voids and the fragmentation of the Si phase. T6 heat treatment (T6B and T6R) formed a discrete globular Si morphology with less stress concentration, which delayed the void nucleation and growth in the Al matrix. The analysis empirically confirmed the higher ductility of the T6 microstructure than that of the AB and T5, highlighting the positive effects on the mechanical performance of the more homogeneous distribution of finer Si particles in T6R.

Antimicrobial resistance represents a major global health concern and environmental bacteria are considered a source of resistance genes. Carbapenems are often used as the last antibiotic option to treat multidrug-resistant bacteria. Metallo-β-lactamases (MBLs) are able to render resistance to almost all β-lactam antibiotics, including carbapenems. Unfortunately, there are no inhibitors against MBLs for clinical use. Subclass B2 MBLs are the only enzymes working as strict carbapenemases, under-represented, encoded in chromosome genes and only functional as mono-zinc enzymes. Despite current efforts in MBLs inhibitor development, B2 carbapenemase activity is especially difficult to suppress, even in vitro. In this study we characterized BioF, a novel subclass B2 MBL identified in a new environmental Pseudomonas sp. strain isolated from an on-farm biopurification system (BPS). Although blaBioF is most likely a chromosomal gene, it is found in a genomic island and may represent a step previous to the horizontal transmission of B2 genes. The new B2 MBL is active as a mono-zinc enzyme and is a potent carbapenemase with incipient activity against some cephalosporins. BioF activity is not affected by excess zinc and is only inhibited at high metal chelator concentrations. The discovery and characterization of B2 MBL BioF as a potent carbapenemase in a BPS bacterial isolate emphasizes the importance of exploring antibiotic resistances existing in the environmental microbiota under the influence of human activities before they could emerge clinically.

Next-generation sequencing technologies facilitate the analysis of multiple important properties of transcriptomes in addition to gene expression levels. Here, we describe a method for mapping RNA 5' ends in Mycobacterium tuberculosis and Mycobacterium smegmatis, which allows the determination of transcription start sites (TSSs), comparative analysis of promoter usage under different conditions, and mapping of endoribonucleolytic cleavage sites. We describe in detail the procedures for constructing RNA sequencing libraries appropriate for RNA 5' end mapping using an Illumina sequencing platform, as well as bioinformatic procedures for data analysis.

Silicon bronze is widely used for monuments: corrosion of Cu-3Si-1Mn was studied with respect to Cu-5Sn-5Zn-5Pb during ageing tests (runoff/full immersion) in acid rain. Surface characterisation (FEG-SEM/EDS elaborated through PCA, TEM, Raman, FTIR, profilometry), metal release and EIS measurements were performed. Under runoff conditions, where mechanical action plays a role, the alloys showed comparable corrosion resistance. In full immersion the corrosion rate of Cu-3Si-1Mn was slightly higher than that of Cu-5Sn-5Zn-5Pb, due to the different nature of the corrosion products. Results showed that, compared to Sn oxide/hydroxide, Si tends to form less protective amorphous products, detaching from the surface.

Selective Laser Melting (SLM) is increasingly employed for producing complex, customised aluminium alloys parts for engineering applications in automotive, aerospace and aircraft. However, an important limitation of these alloys is their poor tribological properties in sliding conditions, which can be improved by surface treatments such as anodizing. Processing conditions during SLM strongly affect both surface roughness and microstructure (leading to hierarchical microstructure and very fine distribution of eutectic Si network in Al-Si alloys), hence influencing the growth of the anodic oxide. In this study, Electro-Chemical Oxidation (ECO) at high voltage was applied to the A357 (AlSi7Mg) alloy produced by SLM, in order to improve its tribological behaviour. Sand cast A357, heat-treated to T6 condition was investigated as a reference for comparison. Dry sliding tests were carried out in block-on-ring configuration against a quenched and tempered SAE52100 (100Cr6) bearing steel rotating cylinder, using the anodized alloys as stationary blocks. The results of this study allowed the comparison of microstructural features and dry sliding behaviour of the oxide layers grown on both the conventional sand cast and SLM A357 alloy.

The nitrogen-fixing alpha-proteobacterium Sinorhizobium meliloti genome codifies at least 50 response regulator (RR) proteins mediating different and, in many cases, unknown processes. RR-mutant library screening allowed us to identify genes potentially implicated in survival to acid conditions. actJ mutation resulted in a strain with reduced growth rate under mildly acidic conditions as well as a lower capacity to tolerate a sudden shift to lethal acidic conditions compared with the parental strain. Mutation of the downstream gene actK, which encodes for a histidine kinase, showed a similar phenotype in acidic environments suggesting a functional two-component system. Interestingly, even though nodulation kinetics, quantity, and macroscopic morphology of Medicago sativa nodules were not affected in actJ and actK mutants, ActK was required to express the wild-type nitrogen fixation phenotype and ActJK was necessary for full bacteroid development and nodule occupancy. The actJK regulatory system presented here provides insights into an evolutionary process in rhizobium adaptation to acidic environments and suggests that actJK-controlled functions are crucial for optimal symbiosis development. Copyright © 2021 Elsevier B.V. All rights reserved.

In this work, Plasma Electrolytic Oxidation (PEO) coatings were produced on AZ91D Mg alloy, using as electrolyte an aqueous solution containing both silicates and phosphates. SiC particles, or borosilicate glass particles or a combination of them (SiC + borosilicate glass) were suspended into the electrolyte. The PEO-treated samples were characterized through Scanning Electron Microscope (SEM) (both on the surface and in cross section), X-Ray Diffraction (XRD) and X-Ray Photoelectron Spectroscopy (XPS) in order to study the morphology, thickness, adhesion, structure and composition of the coating as well as particle distribution. The corrosion behavior was analyzed by potentiodynamic polarization and electrochemical impedance spectroscopy (EIS). The wear resistance was evaluated by dry sliding tests vs. AISI 52100 bearing steel (block-on-ring contact geometry). The results showed that both SiC and borosilicate glass particles were incorporated into the PEO layers. Glass particles contributed to enhance corrosion resistance, whereas SiC particles decreased it. On the other hand, SiC particles improved the wear resistance (due to enhanced load support), leading to an increase of the friction coefficient (due to an increase of the abrasive component of friction). The best combination of properties was obtained with the addition of glass particles for 3 min treatment time. In this way, both the corrosion and wear resistance were increased, minimizing the detrimental effects of SiC particles on the corrosion properties and the friction coefficient.

Application of protective coatings is the most widely used conservation treatment for outdoor bronzes. Eco-friendly and non-hazardous coatings are currently needed for conservation of outdoor bronze monuments. To fulfil this need, the M-ERA.NET European research project B-IMPACT (Bronze-IMproved non-hazardous PAtina CoaTings) aimed at assessing the protectiveness of innovative coatings for historical and modern bronze monuments exposed outdoors. In this project, two bronze substrates (historical Cu-Sn-Zn-Pb and modern Cu-Si-Mn alloys) were artificially patinated, by acid rain solution using dropping test and by “liver of sulphur” procedure (K2S aqueous solution) to obtain black patina, respectively. Subsequently, the application of several newly developed protective coatings was carried out and their performance was investigated by preliminary electrochemical tests. In the following steps of the work, the assessment of the best-performing coatings was carried out and their performance was compared to Incralac, one of the most widely used protective coatings in conservation practice. A multi-analytical approach was adopted, considering artificial ageing (carried out in representative conditions, including exposure to rain runoff, stagnant rain and UV radiation) and metal release, as well as visual aspect (so as to include aesthetical impact among the coating selection parameters) and morphological and structural evolution of the coated surfaces due to simulated outdoor exposure. Lastly, also the health impact of selected coatings was assessed by occupational hazard tests. The removability and re-applicability of the best-performing coatings were also assessed. The best alternatives to the conventional Incralac exhibited were: (i) fluoroacrylate blended with methacryloxy-propyl-trimethoxy-silane (FA-MS) applied on patinated Cu-Sn-Zn-Pb bronze and (ii) 3-mercapto-propyl-trimethoxysilane (PropS-SH) applied on patinated Cu-Si-Mn bronze.

International audience; A 3-mercapto-propyl-trimethoxysilane coating (PropS-SH) applied on Cu-Si-Mn bronze, patinated by “liver of sulphur”, was investigated as a non-toxic alternative to Incralac®, usually applied on outdoor artistic bronzes. Electrochemical testing was performed in synthetic acid rain. Exposure to temperature/UV cycles and accelerated corrosion test simulating unsheltered exposure to rainwater was also carried out. The exposed samples were characterised by FEG-SEM coupled with EDS on FIB cross-sections and XPS on free surfaces. The black patina without protective coating was scarcely protective against bronze corrosion and easily transformed into cuprous oxide. PropS-SH coating fully preserved the black patina microstructure and phase constituents (cuprous oxide and cuprous sulphide). The PropS-SH coating also resulted more protective than Incralac® when aged under run-off conditions. Selective dissolution of copper from the silicon bronze alloy was observed on both uncoated and Incralac®-coated bronze, leading to the formation of an internal Si-rich corrosion layer.

This work shows microstructural evidences of morphological transformation from as-hardened martensite to C-expanded martensite as a result of the low-temperature plasma carburizing treatment carried out in AISI 420 martensitic stainless steel. Treatments were carried out at different treatment temperatures (350, 400, 450, and 500 °C, for a 12 h fixed time) and times (12, 24, 36, and 48 h, at 400 °C fixed temperature). Confocal laser scanning microscopy and SEM results show martensite coarsening in the treated surfaces regarding the respective substrate bulks, which is related to the surface C-alloying, thus being dependent on the treatment time and temperature. Hardness measurement and XRD results strongly suggest that the martensite coarsening is related to the transformation from lath- to plate-type martensite due to the enhancement of the martensite C content, in accordance with the Zener ordering theory. For samples carburized at relatively low temperatures (namely 350, and 400 °C), as C diffuses in paraequilibrium, strain-induced transformation mechanism would govern the changes in the initial martensite morphology as a result of its C content increase. On the other hand, as the chromium carbide precipitation occurs for samples treated at relatively high temperatures (higher than 450 °C for 12 h), and long times (longer than 36 h at 400 °C), C atoms would leave the supersaturated (expanded) martensite phase solid solution, leading the plate martensite to transform back to lath martensite.

Recently, research gives emphasis to eco-friendly and sustainable approaches for the preservation of cultural heritage that could offer advantages in terms of compatibility, durability and safety. Hence, a biological treatment, based on a specific fungal strain of Beauveria bassiana, is exploited for the stabilization of soluble and/or active bronze corrosion products, converting them into copper oxalates. The chemical stability of the latter represents a real improvement for the long-term preservation of bronze, especially in case of exposure to acid rain. However, the corrosion behaviour of bronze differs from that of pure copper due to the presence of additional alloying elements. In natural environments, the selective dissolution of copper leads to a relative tin-enrichment within the corrosion layers, mostly in unsheltered areas exposed to rainwater runoff. To understand the influence of tin-enrichment on the formation of oxalates, pure tin and artificially tin-enriched bronze coupons were treated with this novel biological system and, in the case of bronze coupons, exposed to accelerated ageing. Tin enrichment and accelerated ageing were performed through runoff tests. Before and after treatment and ageing, the sample surface was characterized through Fourier transform infrared (FTIR) and Raman spectroscopies, scanning electron microscopy coupled to energy dispersive spectroscopy (SEM-EDS). Metals released in the ageing solutions were analysed through atomic absorption spectrometry (AAS). The analytical results allowed to better understand the response of unsheltered areas from outdoor bronze monuments to the biological treatment proposed.

This work focuses on the comparison of microstructure and tribological behaviour (dry sliding vs. 100Cr6 steel) of anodic layers produced on wrought AA6082-T5 by hard anodizing (HA), plasma electrolytic oxidation (PEO, sometimes also termed MAO, Micro-Arc Oxidation) and novel electro-chemical oxidation (ECO), derived from PEO minimising its disruptive plasma discharge. The results showed that PEO and HA do not decrease the coefficient of friction of AA6082, whilst ECO does (particularly after sealing with a phosphate-based solution), thanks to its smooth surface and decreased stability of Fe–O based transfer layers. All the anodizing treatments improve wear resistance of AA6082: ECO showed the highest wear resistance, due to the beneficial combination of compact microstructure, high and uniform thickness, high microhardness and adhesion.

This study focuses on components for a pressing device involved in a sliding contact. In particular, the tribosystem consists of a pin (AISI A2 tool steel) and a guide plate (UNS C95500 aluminium bronze) coupled under a counterformal line contact. Firstly, the main wear mechanisms in the pin/guide plate contact were identified by failure analysis on real components after field operation, in order to define the conditions for lab-scale dry sliding tests (block-on-ring). Adhesive wear of bronze (leading to material transfer on steel) and two-body abrasion due to hard carbides from tool steel were the main wear mechanisms in field operation. Secondly, lab-scale tests were carried out for studying the influence of (i) applied load, (ii) sliding speed and (iii) bronze microstructure (as-cast condition or precipitation hardened) on tribological behavior. Tribological laboratory tests showed that the coefficient of friction of the bronze/steel couple was more influenced by applied load and sliding speed than by bronze heat treatment conditions. The wear behavior of bronze is mainly influenced by its work hardening capacity as well as by its ability to form a stable and protective copper oxide surface layer.

Magnesium alloys are used in the motorsport and aerospace fields because of their high specific strength. However, due to their low corrosion resistance, protective surface treatments, such as conversion coating or electroless plating, are necessary when they are used in humid or corrosive environments. The present study aimed at evaluating the effect of plasma electrolytic oxidation (PEO), followed by the deposition of a polymeric layer by powder coating, on the rotating bending fatigue behaviour of the wrought magnesium alloy ZK60A-T5. The specimens were extracted from forged wheels of racing motorbikes and were PEO treated and powder coated. Microstructural characterization was carried out by optical (OM) and scanning electron microscopy (SEM) to analyse both the bulk material and the multilayer, consisting of the anodic oxide interlayer with the powder coating top layer (about 40 &micro, m total thickness). Rotating bending fatigue tests were carried out to obtain the S&ndash, N curve of PEO-treated specimens. The results of the rotating bending tests evidenced fatigue strength equal to 104 MPa at 106 cycles and 90 MPa at 107 cycles. The results of the investigation pointed out that PEO led to a reduction in fatigue strength between 14% and 17% in comparison to the untreated alloy. Fracture surface analyses of the fatigue specimens, carried out by SEM and by 3D digital microscopy, highlighted multiple crack initiation sites at the interface between the PEO layer and substrate, induced by the concurrent effects of coating defects, local tensile stresses in the substrate, and increased roughness at the substrate&ndash, coating interface.

International audience; When attempting to sufficiently protect outdoor bronze monuments from corrosion, searches for an effective solution are usually based on coating applications which have a high anticorrosive efficiency. In order to correctly assess the level of protection provided by such coatings, adherence (practical adhesion) measurements need to be performed for the proper evaluation of the deterioration of coating systems with aging. Although a coupled study of adherence with aging would be of great interest, very few such studies are available. In this work, a methodological approach is proposed for the evaluation of coatings applied to metallic cultural heritage monuments of, based on the use of a three-point bending test. Adherence characterization of different protective coatings has been performed both on bare and on traditionally black-patinated bronze coupons (Cu–Sn alloy with 5.9 wt% Sn), which were used as basic model substrates. The investigated coatings were Incralac®, silane, sol–gel oxysilane, and a silane-modified polymethacrylate (an adhesion promoter for fluoropolymer). The results of measurements which were obtained before and after accelerated aging in concentrated acid rain made it possible to more easily differentiate between the various adherence levels of different coating systems. Coupled with adherence measurements, the results of systematic optical and SEM observation of the different failure morphologies are also presented. In the case of the coated bare alloy, adhesive failures were mainly observed. The silane (PropS-SH) coating showed the best adherence. In the case of the patinated bronze test specimens, only cohesive failures occurred. Adherence is directly related to the cohesion of the black patina rather than that of the applied coating. It was observed that aging reduces the level of the adherence.

The ability of Mycobacterium tuberculosis to infect, proliferate, and survive during long periods in the human lungs largely depends on the rigorous control of gene expression. Transcriptome-wide analyses are key to understanding gene regulation on a global scale. Here, we combine 5′-end-directed libraries with RNAseq expression libraries to gain insight into the transcriptome organization and post-transcriptional mRNA cleavage landscape in mycobacteria during log phase growth and under hypoxia, a physiologically relevant stress condition. Using the model organism Mycobacterium smegmatis, we identified 6,090 transcription start sites (TSSs) with high confidence during log phase growth, of which 67% were categorized as primary TSSs for annotated genes, and the remaining were classified as internal, antisense, or orphan, according to their genomic context. Interestingly, over 25% of the RNA transcripts lack a leader sequence, and of the coding sequences that do have leaders, 53% lack a strong consensus Shine-Dalgarno site. This indicates that like M. tuberculosis, M. smegmatis can initiate translation through multiple mechanisms. Our approach also allowed us to identify over 3,000 RNA cleavage sites, which occur at a novel sequence motif. To our knowledge, this represents the first report of a transcriptome-wide RNA cleavage site map in mycobacteria. The cleavage sites show a positional bias toward mRNA regulatory regions, highlighting the importance of post-transcriptional regulation in gene expression. We show that in low oxygen, a condition associated with the host environment during infection, mycobacteria change their transcriptomic profiles and endonucleolytic RNA cleavage is markedly reduced, suggesting a mechanistic explanation for previous reports of increased mRNA half-lives in response to stress. In addition, a number of TSSs were triggered in hypoxia, 56 of which contain the binding motif for the sigma factor SigF in their promoter regions. This suggests that SigF makes direct contributions to transcriptomic remodeling in hypoxia-challenged mycobacteria. Taken together, our data provide a foundation for further study of both transcriptional and posttranscriptional regulation in mycobacteria.

International audience; n this research, different Schiff bases were synthesized and tested as bronze corrosion inhibitors in concentrated synthetic acid rain at pH 3.3. Some of them showed good inhibiting effects, as detected by EIS and polarization curve recording. The most efficient inhibitor (2-(salicylideneimino) thiophenol, SITP) was also tested with ad-dition of sodium dihydrogen phosphate (P). P improved the surface film resistance against breakdown phe-nomena. XPS and SEM investigations showed that SITP and SITP + P significantly hinder decuprification. Moreover, SITP forms a protective layer on bronze, composed of both Cu(I)-SITP complexes and adsorbed in-hibitor molecules. Cu(II) phosphates were also detected with P addition.

Plasma Electrolytic Oxidation (PEO) was applied to extruded ZM21 Mg alloys to improve their corrosion resistance in a chloride-containing environment. PEO was carried out in DC mode and voltage control in a fluoride-free electrolyte. Potentiodynamic polarization tests in 3.5 wt.% NaCl aqueous solution and neutral salt spray (NSS) tests were carried out. Microstructural and profilometric characterization, as well as NSS tests were performed in different conditions: (i) bare ZM21, (ii) PEO-treated ZM21, (iii) powder-coated ZM21 (without PEO interlayer), and (iv) PEO-treated ZM21 with powder coating top layer (carboxyl-functionalized polyester resin). The PEO + powder coating double layer was identified as the best-performing corrosion protection.

The town of Sumhuram was founded by the South Arabian Kingdom of Hadramawt ca. the end of the 3rd century BCE. Strategically located on the coast of what is now Dhofar in the Southwestern Sultanate of Oman, for several hundred years it controlled the trade route connecting Arabia with India on one side and with the Mediterranean, via the Red Sea, on the other. There is abundant archaeological evidence that copper/bronze items were produced at the site. To shed light on the metallurgical technologies and alloys used, the composition and microstructure of 29 metal items dating between the late 3rd century BCE and the late 4th century CE were investigated by means of SEM-EDS and Optical Microscopy. These investigations are discussed alongside other data previously collected from the site. The results indicate the general use of alloys with a high Pb content, and the apparent absence of a correlation between the objects’ shape/function and their composition. No diachronic evolution is evident in the metallurgical procedures. Comparable results are rare in the literature; our hope is that this work will stimulate further investigation in the metallurgy of the area, which could either supply coherent evidence or firmly contradict it.

Le ricerche di metallurgia antica, o archeometallurgia, sono dedicate allo studio della storia passata dei materiali metallici (in termini di materie prime, tecnologie di produzione e finalità di utilizzo), della storia presente (definita dallo stato attuale di degrado) e della storia futura (determinata dalle strategie di conservazione e controllo del degrado.