Selective Pressure by Rifampicin Modulates Mutation Rates and Evolutionary Trajectories of Mycobacterial Genomes

Resistance to the frontline antibiotic rifampicin constitutes a challenge to the treatment and control of tuberculosis. Here, we analyzed the mutational landscape of Mycobacterium smegmatis during long-term evolution with increasing concentrations of rifampicin, using a mutation accumulation assay combined with whole-genome sequencing. Antibiotic treatment enhanced the acquisition of mutations, doubling the genome-wide mutation rate of the wild-type cells. While antibiotic exposure led to extinction of almost all wild-type lines, the hypermutable phenotype of the ΔnucS mutant strain (noncanonical mismatch repair deficient) provided an efficient response to the antibiotic, leading to high rates of survival. This adaptative advantage resulted in the emergence of higher levels of rifampicin resistance, an accelerated acquisition of drug resistance mutations in rpoB (β RNA polymerase), and a wider diversity of evolutionary pathways that led to drug resistance. Finally, this approach revealed a subset of adaptive genes under positive selection with rifampicin that could be associated with the development of antibiotic resistance. IMPORTANCE Rifampicin is the most important first-line antibiotic against mycobacterial infections, including tuberculosis, one of the top causes of death worldwide. Acquisition of rifampicin resistance constitutes a major global public health problem that makes the control of the disease challenging. Here, we performed an experimental evolution assay under antibiotic selection to analyze the response and adaptation of mycobacteria, leading to the acquisition of rifampicin resistance. This approach explored the total number of mutations that arose in the mycobacterial genomes under long-term rifampicin exposure, using whole-genome sequencing. Our results revealed the effect of rifampicin at a genomic level, identifying different mechanisms and multiple pathways leading to rifampicin resistance in mycobacteria. Moreover, this study detected that an increase in the rate of mutations led to enhanced levels of drug resistance and survival. In summary, all of these results could be useful to understand and prevent the emergence of drug-resistant isolates in mycobacterial infections.This research was funded by MCIN/AEI/10.13039/501100011033, grant PID2020-112865RB-I00, and Instituto de Salud Carlos III, grant FIS PI17/00159 (ISCIII/FEDER, UE). E.C.-S. is the recipient of a PFIS predoctoral research fellowship (FI18/00036) cofinanced by the Instituto de Salud Carlos III and the European Social Fund. A.C.-G. acknowledges financial support from the Spanish State Research Agency, AEI/10.13039/501100011033, through the “Severo Ochoa” Program for Centers of Excellence in R&D (SEV-2013-0347, SEV-2017-0712). Editorial assistance was provided by Stuart L. Rulten. Statistical consultancy was provided by Applied Statistical Department-SGAI-CSIC.S

Alq mutation increases fruit set rate and allows the maintenance of fruit yield under moderate saline conditions

[EN] Arlequin (Alq) is a gain-of-function mutant whose most relevant feature is that sepals are able to become fruit-like organs due to the ectopic expression of the ALQ-TAGL1 gene. The role of this gene in tomato fruit ripening was previously demonstrated. To discover new functional roles for ALQ-TAGL1, and most particularly its involvement in the fruit set process, a detailed characterization of Alq yield-related traits was performed. Under standard conditions, the Alq mutant showed a much higher fruit set rate than the wild type. A significant percentage of Alq fruits were seedless. The results showed that pollination-independent fruit set in Alq is due to early transition from flower to fruit. Analysis of endogenous hormones in Alq suggests that increased content of cytokinins and decreased level of abscisic acid may account for precocious fruit set. Comparative expression analysis showed relevant changes of several genes involved in cell division, gibberellin metabolism, and the auxin signalling pathway. Since pollination-independent fruit set may be a very useful strategy for maintaining fruit production under adverse conditions, fruit set and yield in Alq plants under moderate salinity were assessed. Interestingly, Alq mutant plants showed a high yield under saline conditions, similar to that of Alq and the wild type under unstressed conditions.This work was supported by the research grants AGL2015-64991-C3-3-R and AGL2015-64991-C3-1-R from the Spanish Ministry of Economy and Competitiveness (MINECO/FEDER). The PhD grant to CRA (BES-2013-063778) was funded by the Spanish Ministry of Economy and Competitiveness.The authors thank Dr Isabel Lopez-Diaz and Dr Esther Carrera for their help in hormone quantification carried out at the Plant Hormone Quantification Service, IBMCP,Valencia, Spain. The authors thank David Harry Rhead for reviewing the manuscript in the English language.Ribelles Alfonso, C.; García Sogo, B.; Yuste-Lisbona, FJ.; Atarés Huerta, A.; Castañeda, L.; Capel, C.; Lozano, R.... (2019). Alq mutation increases fruit set rate and allows the maintenance of fruit yield under moderate saline conditions. Journal of Experimental Botany. 70(20):5731-5744. https://doi.org/10.1093/jxb/erz342S57315744702

Genotoxicity Effects of Ceramic Coatings Applied on Metallic Substrates using Single Cell Gel Electrophoresis Assay In Vitro

Abstract. Sol-Gel coatings are a good choice for protection and bioactivation of metals used as dentistry and standard surgical implant materials. These films should both prevent degradation of the substrates by wear or corrosion, and bioactivate the material for inducing the formation of a hydroxyapatite (HA) rich layer onto the material surface, thereby permitting a natural bonding to living tissues. The aim of this work was to estimate the clastogenicity in vitro by Single Cell Gel Electrophoresis Assay (SCGE) or "comet" assay of coatings of TiN applied by magnetron sputtering and of hybrid layers obtained by Sol-Gel containing glass, glass-ceramic and HA particles on stainless steel AISI 304. Six test specimens were prepared: AISI 304 Stainless Steel coated with an hybrid silica single film (SF), applied by sol-gel process, AISI 304 SS coated with double film with bioactive glass (DFG), glass-ceramics (DFGC) and HA (DFHA) particles, AISI 304 SS coated with TiN multi films (MFTiN) applied by PVD and bare AISI 304 SS (304SS). Significantly lower DNA migration (p>0.005) was observed in the cells of the cultures corresponding to the samples coated with SF, DFG, DFGC, DFHA and MFTiN respect to the bare 304 SS. The comparison between negative control and the same coated samples did not reveal any statistically significant difference (p>0.005) in clastogenicity in vitro evaluated by SCGE

Eficacia de la pelmatoscopia a través de capacitación especializada e implementación de oficinas pelmatoscópicas en el hospital “Hermilio Valdizán Medrano” y el centro de salud “Carlos Showing Ferrari”. Huánuco, mayo 2003 – abril 2004

The study demonstrated that the techniques used for the taking of plantar impression are not the suitable ones, the same ones that do not guarantee reliability from the legal medical point of view; on the other hand, the process of identification of new born is not fulfilled efficiently, so this process is left unfinished, fulfilling only with the first stage consisting of the capture of the plantar impression on the part of the personnel of health and his corresponding file in clinical history; the expert's report is not realized on the part of the police personnel in the moment of the discharge of the patient; steps that must be realized according to the stipulated thing in the Manual of Proceeding of Personal Identification. Opposite to this problems, it was tried to demonstrated the effectiveness of the pelmatoscopía by means of the use of the suitable skills and the processing logistic resources through a specializing training and the office implementation pelmatoscópicas in the institutions in study. The type and level of investigation are descriptive-explanatory and transversal. Between the main results it was obtained that 100% of the samples picked up (15 pelmatoscópicas sheets) of the personnel who received the training, and at the same time they have the requirements established by the effective norms for a good taking of papiloscópicas impressions; therefore, there is a team of obstetricians professionals that are in the capacity to assume the taking of plantar impression in the identification of the neonatus.El en estudio se demostró que las técnicas utilizadas para la toma de impresión plantar no son las adecuadas, las mismas que no garantizan confiabilidad desde el punto de vista médico legal; por otro lado, el proceso de identificación del recién nacido no se cumple eficientemente, pues este proceso se deja inconcluso, cumpliendo solamente con la primera fase, consistente en la toma de la impresión plantar por parte del personal de salud y su correspondiente archivo en la historia clínica; sin realizarse el peritaje por parte del personal policial en el momento del alta del paciente; pasos que deben ser realizados de acuerdo a lo estipulado en el Manual de Procedimientos de Identificación Personal. Frente a esta problemática, se pretendió demostrar la eficacia de la pelmatoscopía mediante el uso de las técnicas adecuadas y el procesamiento de los recursos logísticos a través de una capacitación especializada y la implementación de oficinas pelmatoscópicas en las instituciones en estudio. El tipo y nivel de investigación es el descriptivo explicativo y transversal. Entre los principales resultados se obtuvo que el 100 % de las muestras recogidas (15 hojas pelmatoscópicas) del personal que recibió la capacitación cumplen con los requisitos establecidos por las normas vigentes para una buena toma de impresiones papiloscópicas; por tanto, existe un equipo de profesionales de obstetricia que está en la capacidad de asumir la toma de impresión plantar en la identificación del neonato

A study of the interplay between ionized gas and star clusters in the central region of NGC 5253 with 2D spectroscopy

ABRIDGED: A detailed 2D study of the central region of NGC5253 has been performed to characterize the stellar and ionized gas structure as well as the extinction distribution, physical properties and kinematics of the ionized gas in the central ~210pc x 130pc. We utilized optical integral field spectroscopy (IFS) data obtained with FLAMES. A detailed extinction map for the ionized gas in NGC5253 shows that the largest extinction is associated with the prominent Giant HII region. There is an offset of ~0.5" between the peak of the optical continuum and the extinction peak in agreement with findings in the infrared. We found that stars suffer less extinction than gas by a factor of 0.33. The [SII]l6717/[SII]l6731 map shows an electron density (N_e) gradient declining from the peak of emission in Ha (790cm^-3) outwards, while the argon line ratio traces areas with $N_e~4200 - 6200cm^(-3). The area polluted with extra nitrogen, as deduced from the excess [NII]/Ha, extends up to distances of 3.3" (~60pc) from the maximum pollution, which is offset by ~1.5" from the peak of continuum emission. Wolf-Rayet features are distributed in an irregular pattern over a larger area (~100pc x 100pc) and associated with young stellar clusters. We measured He^+ abundances over most of the field of view and values of He^++/H^+<~0.0005 in localized areas which do not coincide, in general, with the areas presenting W-R emission or extra nitrogen. The line profiles are complex. Up to three emission components were needed to reproduce them. One of them, associated with the giant HII region, presents supersonic widths and [NII] and [SII] emission lines shifted up to 40km/s with respect to Ha. Similarly, one of the narrow components presents offsets in the [NII] line of <~20km/s. This is the first time that maps with such velocity offsets for a starburst galaxy have been presented.Comment: accepted in Astronomy & Astrophysics. 21 pages, 22 figure

Charge density wave in layered La1-xCexSb2

The layered rare-earth diantimonides RSb2 are anisotropic metals with generally low electronic densities whose properties can be modified by substituting the rare earth. LaSb2 is a nonmagnetic metal with a low residual resistivity presenting a low-temperature magnetoresistance that does not saturate with the magnetic field. It has been proposed that the latter can be associated to a charge density wave (CDW), but no CDW has yet been found. Here we find a kink in the resistivity above room temperature in LaSb2 (at 355 K) and show that the kink becomes much more pronounced with substitution of La by Ce along the La1−xCexSb2 series. We find signatures of a CDW in x-ray scattering, specific heat, and scanning tunneling microscopy (STM) experiments in particular for x ≈ 0.5. We observe a distortion of rare-earth–Sb bonds lying in-plane of the tetragonal crystal using x-ray scattering, an anomaly in the specific heat at the same temperature as the kink in resistivity and charge modulations in STM. We conclude that LaSb2 has a CDW which is stabilized in the La1−xCexSb2 series due to substitutional disorderThis work was supported by the Spanish MINECO (FIS2014-54498-R, MAT2011-27470-C02-02, and CSD-2009-00013), by the European Union (Graphene Flagship Contract No. CNECT-ICT-604391 and COST MP1201 action), and by the Comunidad de Madrid through programs Nanofrontmag-CM (S2013/MIT-2850) and MAD2D-CM (S2013/MIT-3007).We acknowledge MINECO and CSIC for financial support and for provision of synchrotron radiation facilities and would like to thank the SpLine BM25 staff for assistance in using the beamlin

Functionalization of photosensitized silica nanoparticles for advanced photodynamic therapy of cancer

BODIPY dyes have recently attracted attention as potential photosensitizers. In this work, commercial and novel photosensitizers (PSs) based on BODIPY chromophores (haloBODIPYs and orthogonal dimers strategically designed with intense bands in the blue, green or red region of the visible spectra and high singlet oxygen production) were covalently linked to mesoporous silica nanoparticles (MSNs) further functionalized with PEG and folic acid (FA). MSNs approximately 50 nm in size with different functional groups were synthesized to allow multiple alternatives of PS-PEG-FA decoration of their external surface. Different combinations varying the type of PS (commercial Rose Bengal, Thionine and Chlorine e6 or custom-made BODIPY-based), the linkage design, and the length of PEG are detailed. All the nanosystems were physicochemically characterized (morphology, diameter, size distribution and PS loaded amount) and photophysically studied (absorption capacity, fluorescence efficiency, and singlet oxygen production) in suspension. For the most promising PS-PEG-FA silica nanoplatforms, the biocompatibility in dark conditions and the phototoxicity under suitable irradiation wavelengths (blue, green, or red) at regulated light doses (10–15 J/cm2) were compared with PSs free in solution in HeLa cells in vitro