Genetic regulation of the ompX porin of Salmonella Typhimurium in response to hydrogen peroxide stress.

BACKGROUND: Salmonella Typhimurium is a Gram-negative pathogen that causes a systemic disease in mice resembling typhoid fever. During its infective cycle, S. Typhimurium is phagocytized by macrophages and proliferates inside a Salmonella-containing vacuole where Salmonella is exposed and survives oxidative stress induced by H2O2 through modulation of gene expression. After exposure of Salmonella to H2O2, the expression of the porin-encoding gene ompX increases, as previously shown by microarray analysis. Expression of ompX mRNA is regulated at a post-transcriptional level by MicA and CyaR sRNAs in aerobiosis. In addition, sequence analysis predicts a site for OxyS sRNA in ompX mRNA. RESULTS: In this work we sought to evaluate the transcriptional and post-transcriptional regulation of ompX under H2O2 stress. We demonstrate that ompX expression is induced at the transcriptional level in S. Typhimurium under such conditions. Unexpectedly, an increase in ompX gene transcript and promoter activity after challenges with H2O2 does not translate into increased protein levels in the wild-type strain, suggesting that ompX mRNA is also regulated at a post-transcriptional level, at least under oxidative stress. In silico gene sequence analysis predicted that sRNAs CyaR, MicA, and OxyS could regulate ompX mRNA levels. Using rifampicin to inhibit mRNA expression, we show that the sRNAs (MicA, CyaR and OxyS) and the sRNA:mRNA chaperone Hfq positively modulate ompX mRNA levels under H2O2-induced stress in Salmonella during the exponential growth phase in Lennox broth. CONCLUSIONS: Our results demonstrate that ompX mRNA is regulated in response to H2O2 by the sRNAs CyaR, MicA and OxyS is Salmonella Typhimurium.

Elucidating the multiple structures of pipecolic acid by rotational spectroscopy.

The complex conformational space of the non-proteinogenic cyclic amino acid pipecolic acid has been explored in the gas phase for the first time. Solid pipecolic acid samples were vaporized by laser ablation and expanded in a supersonic jet where the rotational spectral signatures owing to nine different conformers were observed by Fourier transform microwave spectroscopy. All species were identified by comparison of the experimental rotational and nuclear quadrupole coupling constants with those predicted theoretically. Observation of type-III conformers, leading to a difference when compared against the conformational behavior of the analog amino acid proline, has been interpreted by an increment in steric hindrance when increasing the number of carbons present in the ring.

The last link of the x-aminobutyric acid series: the five conformers of ß-aminobutyric acid.

ß-Aminobutyric acid is a non-proteinogenic amino acid that is known to protect plants against various pathogens. Its structure is midway between α-aminobutyric acid and γ-aminobutyric acid. The structural differences in the position of the amino group in the conformational stabilization of ß-aminobutyric acid have been studied by laser ablation Fourier transform microwave spectroscopy. Five conformers have been detected, and their rotational and quadrupole coupling constants of the 14N nucleus determined. Three of the conformers, including the most stable structure, are stabilized by a non-bifurcated N-HO[double bond, length as m-dash]C intramolecular hydrogen bond. The other two conformers possess a NH-O intramolecular interaction. In this work we highlight that ß-aminobutyric acid shows the conformational peculiarities of α-aminobutyric acid and γ-aminobutyric acid completing the rotational spectroscopic study of the x-aminobutyric acid series.

Recent Advances in Remineralization Therapies for Caries Lesions.

Remineralization of caries lesions is naturally achieved by salivary ions, and it can be enhanced by external factors or elements such as fluoride. Numerous studies have demonstrated the remineralizing efficacy of fluoride therapies as well as the limitations with some groups of the population. Consequently, developing new remineralization therapies to close this gap in efficacy has been a priority for the last 2 decades. In this review, we summarize and briefly discuss some of the latest advances in remineralization therapies. Most new therapies try to enhance the effect of fluoride by adding other potentially active ingredients to the formulation, such as calcium, phosphate, stannous, xylitol, and arginine. Other remineralization strategies have focused on creating remineralizing scaffolds within the lesions (e.g., self-assembling peptides). While several of the new remineralization strategies have progressed significantly in recent years, for most of them, the evidence is still insufficient to assess their true clinical potential.

The role of amino acid side chains in stabilizing dipeptides: the laser ablation Fourier transform microwave spectrum of Ac-Val-NH2.

The steric effects imposed by the isopropyl group of valine in the conformational stabilization of the capped dipeptide N-acetyl-l-valinamide (Ac-Val-NH2) have been studied by laser ablation molecular beam Fourier transform microwave (LA-MB-FTMW) spectroscopy. The rotational and quadrupole coupling constants of the two 14N nuclei determined in this work show that this dipeptide exists as a mixture of C7 and C5 conformers in the supersonic expansion. The conformers are stabilized by a C[double bond, length as m-dash]OH-N intramolecular hydrogen bond closing a seven- or a five-membered ring, respectively. The observation of both conformers is in good agreement with previous results on the related dipeptides containing different residues, confirming that the polarity/non-polarity of the side chains of the amino acid is responsible for the conformational locking/unlocking. The voluminous isopropyl group is not able to prevent the less stable C5 conformer from forming but it destabilizes the C[double bond, length as m-dash]OH-N interaction.

The effect of inoculum source and fluid shear force on the development of in vitro oral multispecies biofilms.

AIMS: Saliva has been previously used as an inoculum for in vitro oral biofilm studies. However, the microbial community profile of saliva is markedly different from hard- and soft-tissue-associated oral biofilms. Here, we investigated the changes in the biofilm architecture and microbial diversity of in vitro oral biofilms developed from saliva, tongue or plaque-derived inocula under different salivary shear forces. METHODS AND RESULTS: Four inoculum types (saliva, bacteria harvested from the tongue, toothbrush and curette-harvested plaque) were collected and pooled. Biofilms (n ≥ 15) were grown for 20 h in cell-free human saliva flowing at three different shear forces. Stained biofilms were imaged using a confocal laser scanning microscope. Biomass, thickness and roughness were determined by image analysis and bacterial community composition analysed using Ion Torrent. All developed biofilms showed a significant reduction in observed diversity compared with their respective original inoculum. Shear force altered biofilm architecture of saliva and curette-collected plaque and community composition of saliva, tongue and curette-harvested plaque. CONCLUSIONS: Different intraoral inocula served as precursors of in vitro oral polymicrobial biofilms which can be influenced by shear. SIGNIFICANCE AND IMPACT OF THE STUDY: Inoculum selection and shear force are key factors to consider when developing multispecies biofilms within in vitro models.

Enhancing Intraoral Fluoride Retention in Older Adults: A Randomized Crossover Study.

INTRODUCTION: Previous studies have shown that a calcium prerinse can increase intraoral fluoride retention from a fluoride rinse. To explore the potential of this approach to control root caries, we assessed intraoral fluoride bioavailability after a calcium prerinse in older adults with normal to low salivary flow rates. METHODS: In a 2-period crossover trial (NCT04239872), 20 participants (65-80 y old), with low or normal salivary flow rate, rinsed for 1 min with a 0.05% NaF mouth rinse (226 ppm F, F only) or with this rinse immediately after a 1-min rinse with 150 mM calcium lactate (Ca→F). Dental biofilm and saliva samples were collected before and up to 2 h after the rinse(s). Fluoride concentrations in saliva (whole and clarified) and dental biofilm (fluid and solid phases) were blindly determined. Data were statistically analyzed by a mixed-effects model for the effect of treatment, time, and their interaction (α = 5%). RESULTS: The Ca→F group resulted in significantly higher fluoride concentrations in all variables analyzed, for almost all of the collection time points. The effect was greater in the biofilm solids and whole saliva (compatible with the formation of calcium fluoride deposits) and still significant (P < 0.001) after 2 h in the biofilm fluid and clarified saliva, suggesting that fluoride stored in insoluble particles was released, increasing free fluoride. CONCLUSION: The use of a calcium prerinse before a fluoride rinse was able to prolong intraoral fluoride bioavailability in older adults. KNOWLEDGE TRANSFER STATEMENT: A calcium prerinse increased intraoral fluoride bioavailability in older individuals. This approach could be used to improve root caries control without the need to increase the fluoride concentration in dental products.

Alternatives for the administration of oral antineoplastics in patients with swallowing difficulties.

Oncology patients often experience swallowing difficulties, which can compromise adherence to treatment and consequently reduce its effectiveness. Improper handling of these hazardous drugs can lead to the risk of inhalation of particles or other exposures endangering the health of the persons involved such as nurses and pharmacists. The aim of this review is to analyse and update the recommendations for the manipulation of oral antineoplastic drugs in patients with swallowing difficulties. A literature review of articles, websites, guidelines and other documents published up to about the conditions of handling and administration of oral antineoplastic agents in oncology and oncohaematology was carried out. A table of 110 active principles was compiled. The information was grouped according to the name of the drug, instructions for oral and nasogastric tube administration and suggested recommendations. Among the drugs reviewed, 66.4% were suitable for dissolution. Although there is a lot of information in the literature, the nonstop development of new oncological drugs requires continuous updating. Therefore, we have collected the most recent data to provide a consultation tool for healthcare professionals and patients with swallowing difficulties.

This review can be used by all types of healthcare professionals, especially nurses, who handle oncological medicinal drugs. In addition, the safest handling methods for the worker have been recommended.

Conformers of ß-aminoisobutyric acid probed by jet-cooled microwave and matrix isolation infrared spectroscopic techniques.

ß-aminoisobutyric acid (BAIBA) has been studied in isolation conditions: in the gas phase and trapped into a cryogenic N2 matrix. A solid sample of the compound was vaporized by laser ablation and investigated through their rotational spectra in a supersonic expansion using two different spectroscopic techniques: broadband chirped pulse Fourier transform microwave spectroscopy and conventional molecular beam Fourier transform microwave spectroscopy. Four conformers with structures of two types could be successfully identified by comparison of the experimental rotational and (14)N nuclear quadruple coupling constants with those predicted theoretically: type A, bearing an OH⋯N intramolecular hydrogen bond and its carboxylic group in the trans geometry (H-O-C=O dihedral ∼180°), and type B, having an NH⋯O bond and the cis arrangement of the carboxylic group. These two types of conformers could also be trapped from the gas phase into a cryogenic N2 matrix and probed by Fourier transform infrared (IR) spectroscopy. In situ irradiation of BAIBA isolated in N2 matrix of type B conformers using near-IR radiation tuned at the frequency of the O-H stretching 1st overtone (∼6930 cm(-1)) of these forms allowed to selectively convert them into type A conformers and into a new type of conformers of higher energy (type D) bearing an NH⋯O=C bond and a O-H "free" trans carboxylic group.

Are we ready for definitive clinical guidelines on xylitol/polyol use?

In the past decades, numerous studies have looked at the anticaries effects of polyols, particularly xylitol, and a great many studies have focused on xylitol's antimicrobial properties. Researched vehicles have mostly included chewing gums, followed by lozenges/candies, toothpastes, and others (e.g., syrup). Good evidence supports the claims that xylitol is non-cariogenic and has a dose-/frequency-dependent antimicrobial effect on dental plaque/mutans streptococci, and that polyol use is very safe. However, interpretation of caries data has been controversial, due in part to variability in study designs, formulations/dosages tested, and outcomes reported (e.g., many caries studies have a "no gum" control, limiting the interpretation of the polyol's benefit; few studies have compared different polyols side-by-side, or in adults). Even when the level/strength of high-quality anticaries evidence is still limited, most recent systematic reviews have consistently concluded that the habitual use of sucrose-free xylitol or polyol-combination chewing gum/ lozenges is an effective adjunct in coronal caries prevention. Consequently, many health organizations worldwide are supporting this recommendation for at-risk populations. However, most experts agree that well-designed, placebo-controlled randomized clinical trials (RCTs) (focusing on efficacy, feasibility, adherence, dosage, vehicle, synergism with other preventive strategies, and cost) are still needed in target populations worldwide to reach definitive caries-preventive/therapeutic recommendations.

Internal rotation analysis of the microwave and millimeter wave spectra of fluoral (CF3CHO).

The rotational spectrum (4-40 GHz and 50-330 GHz) has been measured and analyzed for trifluoroacetaldehyde, also known as fluoral (CF3CHO), which is one of the degradation products of the fluorinated contaminants emitted into the atmosphere. The complexity of the spectroscopic analysis of this molecule arises from the strong coupling between the internal rotation motion of CF3 group and the overall rotation of the molecule. The value obtained for its coupling constant (ρ = 0.91723481(49)) is comparable to the corresponding value of methanol (CH3OH, ρ = 0.81), which is known for its complex spectrum. A total of 12,322 transitions of the ground, the first and second excited torsional states (ΔE1υt = 62.0183(13)cm-1; ΔE2υt = 120.3315(13)cm-1) with J ≤ 50 were included in the analysis that was performed employing the rho-axis-method (RAM), and the RAM36 code. A fit within experimental error (root mean square deviation equals to 35 kHz) has been achieved for this dataset using 47 parameters of the RAM torsion-rotation Hamiltonian. In the course of the analysis, it became evident that for such high ρ value, as it is determined for fluoral, a larger than usual torsional basis set at the first diagonalization step of the two-step diagonalization procedure is required for achieving a fit within experimental error.

Rapid probe of the nicotine spectra by high-resolution rotational spectroscopy.

Nicotine has been investigated in the gas phase and two conformational forms were characterized through their rotational spectra. Two spectroscopic techniques have been used to obtain the spectra: a new design of broadband Fourier transform microwave (FTMW) spectroscopy with an in-phase/quadrature-phase-modulation passage-acquired-coherence technique (IMPACT) and narrowband FTMW spectroscopy with coaxially oriented beam-resonator arrangement (COBRA). The rotational, centrifugal distortion and hyperfine quadrupole coupling constants of two conformers of nicotine have been determined and found to be in N-methyl trans configurations with the pyridine and pyrrolidine rings perpendicular to one another. The quadrupole hyperfine structure originated by two (14)N nuclei has been completely resolved for both conformers and used for their unambiguous identification.

Effect of gap geometry on secondary caries wall lesion development.

OBJECTIVE: To investigate how the size of the space between restoration and dentinal wall of the tooth affects the development of secondary caries lesions, especially wall lesions. METHODS: Tooth-resin composite specimens were mounted on custom-made gap model stages and divided into 4 groups (n = 10): group 1 with a 30-µm gap throughout both enamel and dentin, group 2 with a 30-µm enamel gap and 530-µm dentinal gap, group 3 with 525-µm gaps in both enamel and dentin, and group 4 with 525-µm and 1,025-µm gaps in enamel and dentin, respectively. Specimens were gas sterilized and incubated in a cycling microbial caries model for 8 days and analyzed with confocal microscopy for lesion size at the enamel outer lesion (EOL), enamel wall lesion (EWL), dentin wall lesion A (DWL-A) next to the dentin-enamel junction (DEJ) and dentin wall lesion B (DWL-B) at 750 µm from the DEJ. RESULTS: No difference in EOL or EWL size was found between the groups. DWL-A and DWL-B were larger in group 3 than groups 1 and 2. A larger DWL-B was found in group 3 than group 4. CONCLUSIONS: The presence of additional space at the dentinal wall area did not affect secondary caries development as long as the enamel gap was small. However, with enamel gaps of approximately 500 µm, the presence of the additional gap space at the dentinal wall led to the development of smaller dentinal wall lesions at the deeper parts of the simulated cavity. In uniform gaps, the size of the interface was positively correlated with the size of the dentinal wall lesions.

Comprehensive rotational study and astronomical search for cyclopropanecarboxaldehyde.

CONTEXT: At least a dozen molecules with a formyl group (HCO) have been observed to date in the interstellar medium (ISM), suggesting that other such species exist and remain to be discovered. However, there is still a lack of high-resolution spectroscopic data for simple molecular species of this type that could provide a basis for their detection. AIMS: Cyclopropanecarboxaldehyde, c-C3H5CHO, is a small molecule containing a formyl group and is therefore an interesting candidate for astrophysical detection. The rotational spectrum of cyclopropanecarboxaldehyde has been observed before, but its experimental rotational parameters are not precise enough to allow its detection in the millimetre-wave domain. METHODS: We measured the rotational spectrum of cyclopropanecarboxaldehyde in the frequency ranges 31.5-50 GHz and 72-116.5 GHz using the GACELA (GAS CEll for Laboratory Astrophysics) broadband high-resolution rotational spectrometer constructed at the Yebes Observatory. The spectroscopic study was supported by high-level theoretical calculations which were used in the identification of the vibrational excited states of cyclopropanecarboxaldehyde. RESULTS: Our analysis of the rotational spectrum of cyclopropanecarboxaldehyde allowed us to obtain accurate rotational parameters for the ground state of both cis and trans isomers, which were used to derive sufficiently reliable predictions up to 300 GHz. In addition to the ground states, we identified 12 and 6 vibrationally excited states for the trans and cis isomers, respectively, including fundamental modes, multiple excitation quanta, and combination states. We find that the gas phase concentration of the trans isomer is almost 1.2 times larger than that of the cis one. These new experimental rotational parameters were employed to search for cyclopropanecarboxaldehyde in the warm molecular clouds Orion KL and Sgr B2(N) using the spectral surveys captured by ALMA (Orion) and IRAM 30 m (Sgr) at 1 mm and 3 mm, respectively.

First detection of doubly deuterated methyl acetylene (CHD2CCH and CH2DCCD)☠.

We report the first detection in space of the two doubly deuterated isotopologues of methyl acetylene. The species CHD2CCH and CH2DCCD were identified in the dense core L483 through nine and eight, respectively, rotational lines in the 72-116 GHz range using the IRAM 30m telescope. The astronomical frequencies observed here were combined with laboratory frequencies from the literature measured in the 29-47 GHz range to derive more accurate spectroscopic parameters for the two isotopologues. We derive beam-averaged column densities of (2.7 ± 0.5) × 1012 cm-2 for CHD2CCH and (2.2 ± 0.4) × 1012 cm-2 for CH2DCCD, which translate to abundance ratios CH3CCH/CHD2CCH = 34 ± 10 and CH3CCH/CH2DCCD = 42 ± 13. The doubly deuterated isotopologues of methyl acetylene are only a few times less abundant than the singly deuterated ones, concretely around 2.4 times less abundant than CH3CCD. The abundances of the different deuterated isotopologues with respect to CH3CCH are reasonably accounted for by a gas-phase chemical model in which deuteration occurs from the precursor ions C3H6D+ and C3H5D+, when the ortho-to-para ratio of molecular hydrogen is sufficiently low. This points to gas-phase chemical reactions, rather than grain-surface processes, as responsible for the formation and deuterium fractionation of CH3CCH in L483. The abundance ratios CH2DCCH/CH3CCD = 3.0 ± 0.9 and CHD2CCH/CH2DCCD = 1.25 ± 0.37 observed in L483 are consistent with the statistically expected values of three and one, respectively, with the slight overabundance of CHD2CCH compared to CH2DCCD being well explained by the chemical model.

Detection of deuterated methylcyanoacetylene, CH2DC3N, in TMC-1.

We report the first detection in space of the single deuterated isotopologue of methylcyanoacetylene, CH2DC3N. A total of fifteen rotational transitions, with J = 8-12 and Ka = 0 and 1, were identified for this species in TMC-1 in the 31.0-50.4 GHz range using the Yebes 40m radio telescope. The observed frequencies were used to derive for the first time the spectroscopic parameters of this deuterated isotopologue. We derive a column density of (8.0 ± 0.4) × 1010 cm-2. The abundance ratio between CH3C3N and CH2DC3N is ∼22. We also theoretically computed the principal spectroscopic constants of 13C isotopologues of CH3C3N and CH3C4H and those of the deuterated isotopologues of CH3C4H for which we could expect a similar degree of deuteration enhancement. However, we have not detected either CH2DC4H nor CH3C4D nor any 13C isotopologue. The different observed deuterium ratios in TMC-1 are reasonably accounted for by a gas phase chemical model where the low temperature conditions favor deuteron transfer through reactions with H2D+.

O-bearing complex organic molecules at the cyanopolyyne peak of TMC-1: detection of C2H3CHO, C2H3OH, HCOOCH3, and CH3OCH3.

We report the detection of the oxygen-bearing complex organic molecules propenal (C2H3CHO), vinyl alcohol (C2H3OH), methyl formate (HCOOCH3), and dimethyl ether (CH3OCH3) toward the cyanopolyyne peak of the starless core TMC-1. These molecules are detected through several emission lines in a deep Q-band line survey of TMC-1 carried out with the Yebes 40m telescope. These observations reveal that the cyanopolyyne peak of TMC-1, which is the prototype of cold dark cloud rich in carbon chains, contains also O-bearing complex organic molecules like HCOOCH3 and CH3OCH3, which have been previously seen in a handful of cold interstellar clouds. In addition, this is the first secure detection of C2H3OH in space and the first time that C2H3CHO and C2H3OH are detected in a cold environment, adding new pieces in the puzzle of complex organic molecules in cold sources. We derive column densities of (2.2 ± 0.3) × 1011 cm™2, (2.5 ± 0.5) × 1012 cm-2, (1.1 ± 0.2) × 1012 cm-2, and (2.5 ± 0.7) × 1012 cm-2 for C2H3CHO, C2H3OH, HCOOCH3, and CH3OCH3, respectively. Interestingly, C2H3OH has an abundance similar to that of its well known isomer acetaldehyde (CH3CHO), with C2H3OH/CH3CHO ~ 1 at the cyanopolyyne peak. We discuss potential formation routes to these molecules and recognize that further experimental, theoretical, and astronomical studies are needed to elucidate the true mechanism of formation of these O-bearing complex organic molecules in cold interstellar sources.

Discovery of the propargyl radical (CH2CCH) in TMC-1: one of the most abundant radicals ever found and a key species for cyclization to benzene in cold dark clouds.

We present the first identification in interstellar space of the propargyl radical (CH2CCH). This species was observed in the cold dark cloud TMC-1 using the Yebes 40m telescope. The six strongest hyperfine components of the 20,2-10,1 rotational transition, lying at 37.46 GHz, were detected with signal-to-noise ratios in the range 4.6-12.3 σ. We derive a column density of 8.7 × 1013 cm-2 for CH2CCH, which translates to a fractional abundance relative to H2 of 8.7 × 10-9. This radical has a similar abundance to methyl acetylene, with an abundance ratio CH2CCH/CH3CCH close to one. The propargyl radical is thus one of the most abundant radicals detected in TMC-1, and it is probably the most abundant organic radical with a certain chemical complexity ever found in a cold dark cloud. We constructed a gas-phase chemical model and find calculated abundances that agree with, or fall two orders of magnitude below, the observed value depending on the poorly constrained low-temperature reactivity of CH2CCH with neutral atoms. According to the chemical model, the propargyl radical is essentially formed by the C + C2H4 reaction and by the dissociative recombination of C3Hn + ions with n = 4-6. The propargyl radical is believed to control the synthesis of the first aromatic ring in combustion processes, and it probably plays a key role in the synthesis of large organic molecules and cyclization processes to benzene in cold dark clouds.

Space and laboratory discovery of HC3 S.

We report the detection in TMC-1 of the protonated form of C3S. The discovery of the cation HC3S+ was carried through the observation of four harmonically related lines in the Q band using the Yebes 40m radiotelescope, and is supported by accurate ab initio calculations and laboratory measurements of its rotational spectrum. We derive a column density N(HC3S+) = (2.0 ± 0.5) × 1011 cm-2, which translates to an abundance ratio C3S/HC3S+ of 65 ± 20. This ratio is comparable to the CS/HCS+ ratio (35 ± 8) and is a factor of about ten larger than the C3O/HC3O+ ratio previously found in the same source. However, the abundance ratio HC3O+/HC3S+ is 1.0 ± 0.5, while C3O/C3S is just ~ 0.11. We also searched for protonated C2S in TMC-1, based on ab initio calculations of its spectroscopic parameters, and derive a 3σ upper limit of N(HC2S+)≤ 9×1011 cm-2 and a C2S/HC2S+ ≥ 60. The observational results are compared with a state-of-the-art gas-phase chemical model and conclude that HC3S+ is mostly formed through several pathways: proton transfer to C3S, reaction of S+ with c-C3H2, and reaction between neutral atomic sulfur and the ion C3H+ 3.