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Duralumin-type alloys have been employed for structural and non-structural parts of aircraft since the early 1920s up to now. In the European project “PROtection and Conservation of Heritage AirCRAFT” (PROCRAFT), up to 34 aircraft wrecks from World War II from different nations (United States of America, United Kingdom, France, Germany, Italy) were identified. On most of them, the collection of numerous fragments was possible, thus constituting a large sampling group. A metallographic analysis was carried out, and elemental composition, microstructure and hardness were measured. This article focuses firstly on the elemental composition of this group of historical Al alloys. From these data, some fine characteristics specific to the manufacturing countries will be investigated through principal component analysis. Then, on a restricted group constituted of German alloys, we will discuss how some manufacturing parameters, such as micro-alloying and thermo-mechanical treatments, influence the mechanical properties of the assessed aircraft components. Other influencing parameters, linked to the archaeological nature of the parts, are considered and addressed based on a specific study of an alloy collected on a crashed JU88 aircraft originating from the German company Junkers.

This work reports the study of metal fragments from Roman pipes excavated from the archaeological site of Pompeii and currently preserved in the deposits of the National Archaeological Museum of Naples (MANN). The Roman pipe, called the tibia, is a reed wind musical instrument similar to the Greek aulos. It can be made of wood, bone, and/or metal. Materials consisting of metal Cu-based alloys were excavated from archaeological burial environments. This research aims to identify the composition of the alloys, characterize the corrosion patinas, and identify any ancient surface treatments on the fragments. Non-invasive and micro-invasive techniques were used to achieve this aim, i.e., optical microscopy, Raman spectroscopy, attenuated total reflectance Fourier-transform infrared spectrophotometry, scanning electron microscopy, and energy dispersive spectrometry. This research contributes to a deeper understanding of the materials and manufacturing techniques used for these instruments, as well as the degradation processes occurring over the centuries.

Abstract The development of innovative solutions for the conservation and the protection of historic aircraft is a recent and important issue raised in the cultural heritage field and represents the goal of the PROCRAFT (Protection and Conservation of Heritage Aircraft) project. Assessing and documenting the constituent materials as well as their degradation state is necessary to determine and understand factors inducing aluminium alloy corrosion, in order to develop tailored conservation treatments and identify effective protective coatings. The first step of the project was the identification of the constituent materials of aircraft wrecks. Al-Cu-Mg alloys—Duralumin and Super Duralumin, with a higher content of Mg—were the most employed alloys for structural and non-structural parts. These materials undergo a wide range of alterations, amongst which pitting, exfoliation and galvanic corrosion. In the present work, results of the characterisation of wrecks from a North American Republic P-47D Thunderbolt, crashed in Italy in 1945, are reported and compared to wrecks from a French Breguet 765 Sahara n°504 64-PH, built in 1958. The constituent alloys resulted to be similar to the modern 2024 and 2017A alloy. In order to assess the representativeness of these modern alloys in simulating the corrosion behaviour of the historic ones, exfoliation susceptibility (ASTM G34) as well as non-destructive and destructive electrochemical tests in 0.1 M NaCl were carried out on both historic and modern alloys. These results contributed to the selection of representative substrates for the development of protective coatings as well as to the expansion of the dataset on composition and microstructure of historic Al alloys for aircraft.

The development of Healthcare-Associated Infections (HAIs) represents an increasing threat to patient health. In this context, Pseudomonas aeruginosa is responsible for various HAIs, determining about 20% of the infections in hospitalized patients, which makes it one of the most effective pathogens due to its strong ability to form biofilms. Using Cu-based materials as foils on high-touch surfaces can help to prevent and mitigate P. aeruginosa contamination in biohazardous settings. However, the antibiofilm properties of Cu-based surfaces against P. aeruginosa may vary due to frequent touches combined with indoor environmental exposure. The main aim of this study is to investigate the impact of accelerated ageing, mimicking a high-touch frequency by cyclic exposure to artificial sweat solution as well as to temperature and relative humidity variations, on the efficacy of Cu-based thin foils against P. aeruginosa biofilms. Three Cu-based materials (rolled and annealed Phosphorous High-Conductivity (PHC) Cu, Cu15Zn brass, and Cu18Ni20Zn nickel silver) were evaluated. The ageing process enhanced the antibiofilm properties, due to an increment in Cu ion release: aged PHC Cu and Cu15Zn exhibited the highest Cu ion release and hence the highest biofilm inhibition (decrease in colony forming unit (CFU)) in comparison to their pristine counterpart, while aged Cu18Ni20Zn displayed the lowest biofilm formation reduction, despite showing the highest aesthetic and morphological stability. The Cu-based surface, which highlited the highest biofilm formation inhibition due to accelerated ageing, was Cu15Zn.

Mycobacterium abscessus causes severe disease in patients with cystic fibrosis. Little is known in M. abscessus about the roles of small regulatory RNAs (sRNA) in gene regulation. 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 strain, increased pro-inflammatory signaling and virulence in multiple infection models, and increased resistance to antibiotics. Examination of clinical isolate cohorts identified 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 mutations found in clinical isolates. Over 200 genes were differentially expressed in the deletion mutant. Strains with the clinical B11 mutations showed expression trends similar to the deletion mutant, 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 B11 functioning as a negative regulator that represses translation via base-pairing to RBSs. Comparing the proteomes similarly revealed that upregulated proteins were strongly enriched for B11-complementary sequences. Intriguingly, genes upregulated in the absence of B11 included components of the ESX-4 secretion system, critical for M. abscessus virulence. Many of these genes had B11-complementary sequences at their RBSs, which we show is sufficient to mediate repression by B11 through direct binding. Altogether, our data show that B11 acts as a direct negative regulator and mediates (likely indirect) positive regulation with pleiotropic effects on gene expression and clinically important phenotypes in M. abscessus. The presence of hypomorphic B11 mutations in clinical strains is consistent with the idea that lower B11 activity may be advantageous for M. abscessus in some clinical contexts. This is the first report on an sRNA role in M. abscessus.

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.

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.

Abstract 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 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.

IncP-1, IncP-7 and IncP-9 plasmids often carry genes encoding enzymes involved in the degradation of man-made and natural contaminants, thus contributing to bacterial survival in polluted environments. However, the lack of suitable molecular tools often limits the detection of these plasmids in the environment. In this study, PCR followed by Southern blot hybridization detected the presence of plasmid-specific sequences in total community (TC-) DNA or fosmid DNA from samples originating from different environments and geographic regions. A novel primer system targeting IncP-9 plasmids was developed and applied along with established primers for IncP-1 and IncP-7. Screening TC-DNA from biopurification systems (BPS) which are used on farms for the purification of pesticide-contaminated water revealed high abundances of IncP-1 plasmids belonging to different subgroups as well as IncP-7 and IncP-9. The novel IncP-9 primer-system targeting the rep gene of nine IncP-9 subgroups allowed the detection of a high diversity of IncP-9 plasmid specific sequences in environments with different sources of pollution. Thus polluted sites are "hot spots" of plasmids potentially carrying catabolic genes.