Label-free detection and profiling of individual solution-phase molecules.

Most chemistry and biology occurs in solution, in which conformational dynamics and complexation underlie behaviour and function. Single-molecule techniques1 are uniquely suited to resolving molecular diversity and new label-free approaches are reshaping the power of single-molecule measurements. A label-free single-molecule method2-16 capable of revealing details of molecular conformation in solution17,18 would allow a new microscopic perspective of unprecedented detail. Here we use the enhanced light-molecule interactions in high-finesse fibre-based Fabry-Pérot microcavities19-21 to detect individual biomolecules as small as 1.2 kDa, a ten-amino-acid peptide, with signal-to-noise ratios (SNRs) >100, even as the molecules are unlabelled and freely diffusing in solution. Our method delivers 2D intensity and temporal profiles, enabling the distinction of subpopulations in mixed samples. Notably, we observe a linear relationship between passage time and molecular radius, unlocking the potential to gather crucial information about diffusion and solution-phase conformation. Furthermore, mixtures of biomolecule isomers of the same molecular weight and composition but different conformation can also be resolved. Detection is based on the creation of a new molecular velocity filter window and a dynamic thermal priming mechanism that make use of the interplay between optical and thermal dynamics22,23 and Pound-Drever-Hall (PDH) cavity locking24 to reveal molecular motion even while suppressing environmental noise. New in vitro ways of revealing molecular conformation, diversity and dynamics can find broad potential for applications in the life and chemical sciences.

A culture-free biphasic approach for sensitive and rapid detection of pathogens in dried whole-blood matrix.

Blood stream infections (BSIs) cause high mortality, and their rapid detection remains a significant diagnostic challenge. Timely and informed administration of antibiotics can significantly improve patient outcomes. However, blood culture, which takes up to 5 d for a negative result, followed by PCR remains the gold standard in diagnosing BSI. Here, we introduce a new approach to blood-based diagnostics where large blood volumes can be rapidly dried, resulting in inactivation of the inhibitory components in blood. Further thermal treatments then generate a physical microscale and nanoscale fluidic network inside the dried matrix to allow access to target nucleic acid. The amplification enzymes and primers initiate the reaction within the dried blood matrix through these networks, precluding any need for conventional nucleic acid purification. High heme background is confined to the solid phase, while amplicons are enriched in the clear supernatant (liquid phase), giving fluorescence change comparable to purified DNA reactions. We demonstrate single-molecule sensitivity using a loop-mediated isothermal amplification reaction in our platform and detect a broad spectrum of pathogens, including gram-positive methicillin-resistant and methicillin-susceptible Staphylococcus aureus bacteria, gram-negative Escherichia coli bacteria, and Candida albicans (fungus) from whole blood with a limit of detection (LOD) of 1.2 colony-forming units (CFU)/mL from 0.8 to 1 mL of starting blood volume. We validated our assay using 63 clinical samples (100% sensitivity and specificity) and significantly reduced sample-to-result time from over 20 h to <2.5 h. The reduction in instrumentation complexity and costs compared to blood culture and alternate molecular diagnostic platforms can have broad applications in healthcare systems in developed world and resource-limited settings.

Pachymeningeal disease: a systematic review and metanalysis.

BACKGROUND: Pachymeningeal disease (PMD) is a newly recognized pattern of brain metastasis (BrM) failure that specifically occurs following surgery with adjuvant stereotactic radiosurgery (SRS) and has unique prognostic implications relative to leptomeningeal disease (LMD). Here, we report its prevalence, prognostic implications, and associated risk factors. METHODS: A literature search was performed in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses on PUBMED and Cochrane from January 2000 to June 2023. RESULTS: We identified 12 studies that included a total of 3992 BrM patients, 659 (16.5%) of whom developed meningeal disease (MD) following surgery plus adjuvant SRS, including either PMD or LMD. The mean prevalence of MD across studies was 20.9% (7.9-38.0%), with PMD accounting for 54.6% of this prevalence and LMD comprising the remaining 45.4%. Mean of the median overall survivals following diagnosis of PMD and LMD was 10.6 months and 3.7 months p = 0.007, respectively, a significant difference. Only 2 risk factors for PMD were reported in ≥ 2 studies and also identified as statistically significant per our meta-analysis: infratentorial location and controlled systemic disease status. CONCLUSION: While PMD has a superior prognosis to LMD, it is nevertheless a critical oncologic event associated with significant mortality and remains poorly recognized. PMD is predominantly observed in patients with controlled systemic disease status and infratentorial location. Future treatment strategies should focus on reducing surgical seeding and sterilizing surgical cavities.

Tunable fiber Fabry-Perot cavities with high passive stability.

We present three high finesse tunable monolithic fiber Fabry-Perot cavities (FFPCs) with high passive mechanical stability. The fiber mirrors are fixed inside slotted glass ferrules, which guarantee an inherent alignment of the resonators. An attached piezoelectric element enables fast tuning of the FFPC resonance frequency over the entire free-spectral range for two of the designs. Stable locking of the cavity resonance is achieved for sub-Hertz feedback bandwidths, demonstrating the high passive stability. At the other limit, locking bandwidths up to tens of kilohertz, close to the first mechanical resonance, can be obtained. The root-mean-square frequency fluctuations are suppressed down to ∼2% of the cavity linewidth. Over a wide frequency range, the frequency noise is dominated by the thermal noise limit of the system's mechanical resonances. The demonstrated small footprint devices can be used advantageously in a broad range of applications like cavity-based sensing techniques, optical filters or quantum light-matter interfaces.

Culture-free biphasic approach for sensitive detection of Escherichia coli O157:H7 from beef samples.

Foodborne illnesses are a major threat to public health also leading to significant mortality and financial and reputational damage to industry. It is very important to detect pathogen presence in food products early, rapidly, and accurately to avoid potential outbreaks and economic loss. However, "gold standard" culture methods, including enrichment of pathogens, can take up to several days. Moreover, the food matrix often interferes with nucleic acid amplification methods of detection, requiring DNA extraction from the sample for successful molecular detection of pathogens. Here, we introduce a "biphasic" amplification method that can achieve high sensitivity detection with background noise from ground beef food samples without culture or other extraction methods in 2.5 h. Homogenized ground beef is dried resulting in an increase in porosity of the dried food matrix to allowing amplification enzymes and primers to access the target DNA and initiate the reaction within the dried food matrix. Using Loop Mediated Isothermal Amplification, we demonstrate the detection of 1-3 cfu of Escherichia coli bacteria in 30 mg of dried food matrix. Our approach significantly lowers the time to result to less than a few hours and have a pronounced impact on reduction of instrumentation complexity and costs.

3D printed millimeter-wave beam-steering reflector using dielectric fluids.

A three-dimensional printed beam-steering reflector surface with dielectric fluids as the tuning agent is presented. The reflector is made using ECO-ABS with six rows of 19 parallel channels of square cross-sections. The permittivity of the ECO-ABS was measured at 2.55 with a loss tangent of 0.053. A conductor is placed at the back of the dielectric. The squared channels are filled with either distilled water or air. The effective permittivity within the reflector changes according to the material used to fill the channels. As an incident wave propagates through the printed dielectric, the configuration of air-water channels shapes the exiting phase front of the wave by locally controlling its phase delay. The resulting phase profile created by the air-water configuration leads to a steered beam. Numerical full-wave simulations show steerable angles ranging from -42∘ to 23° for a set of air-water configurations at 30 GHz. A prototype was fabricated and tested for the same configurations. Experiments confirm a wide range of angles starting at -40∘ up to 20°.

Conversion of "Customizable Units" into N-Alkyl Amino Acids and Generation of N-Alkyl Peptides.

An efficient conversion of hydroxyproline "customizable" units into new amino acids with a variety of N-alkyl substituents is described. The process is versatile and can afford valuable N-methyl amino acids and N, O-acetals. In addition, it allows the introduction of N-homoallylic substituents and N-chains with terminal ester, ketone, or cyano groups. These chains could be used for peptide extension or conjugation to other molecules (e.g., by olefin metathesis, peptide ligation, etc.). The transformation is carried out in just two (for R = CH2OAc) or three steps (scission of the pyrrolidine ring, manipulation of the α-chain, and the N-substituent) under mild, metal-free conditions, affording products with high optical purity.

Metal-Free, Site-Selective Peptide Modification by Conversion of "Customizable" Units into ß-Substituted Dehydroamino Acids.

Our site-selective modification of serine or threonine units in peptides allows the generation of ß-substituted dehydroamino acids, which increase peptide resistance to hydrolysis and may improve their biological properties. Both the terminal and internal positions can be modified, and different customizable units can be activated separately. Remarkably, high Z selectivity is achieved, even at internal positions. The conversion involves a one-pot oxidative radical scission/phosphorylation process by using the low-toxicity (diacetoxyiodo)benzene/iodine system as the scission reagent. The resulting α-amino phosphonates undergo a Horner-Wadsworth-Emmons reaction to produce the dehydroamino acid derivatives (in a Z/E ratio of usually >98:2) under mild and metal-free conditions.

Site-selective modification of peptides: From "customizable units" to novel α-aryl and α-alkyl glycine derivatives, and components of branched peptides.

The creation of peptide libraries by site-selective modification of a few peptide substrates would increase the efficiency of discovery processes, but still is a real synthetic challenge. The site-selective modification of small peptides at serine or threonine residues, by using a short scission-addition procedure, allows the preparation of peptides with unnatural α-aryl glycines. In a similar way, the scission of hydroxyproline residues is the key step in the production of optically pure α-alkyl glycines which are precursors or components of branched peptides. With these versatile processes, a single peptide can be transformed into a variety of peptide derivatives. The process takes place under mild conditions, and good global yields are obtained. © 2015 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 104: 650-662, 2015.

[Resistance exercise improves serum lipids in adult women]. / Reducción del riesgo cardiovascular en mujeres adultas mediante ejercicio físico de sobrecarga.

BACKGROUND: Sedentariness is one of the main cardiovascular risk factors. Aerobic exercise can reduce the risk; however, resistance training seems to be more effective in reducing cardiovascular risk. AIM: To determine the metabolic effects of a 12 weeks resistance training program of high intensity and low volume in two groups of sedentary adult women. MATERIAL AND METHODS: Forty women aged between 30 and 60 years were randomly assigned to an experimental group and a control group. Four kinds of strength exercises, using their own body weight were chosen. Plasma lipid profile and body composition were measured at baseline and at the end of the intervention. RESULTS: A reduction in total cholesterol and triacylglycerol and an increase in HDL cholesterol was observed in the experimental group after the training program. CONCLUSIONS: The high intensity and low volume training improved plasma lipid profile in this group of women.

Phylogeographic analysis of introns and mitochondrial DNA in the clam Ruditapes decussatus uncovers the effects of Pleistocene glaciations and endogenous barriers to gene flow.

Studies on the phylogeography of species inhabiting the Mediterranean and the nearby coasts of the NE Atlantic Ocean (MEDAT) have found subdivision and/or phylogeographic structure in one or more of the Atlantic, western Mediterranean and eastern Mediterranean basins. This structure has been explained as the result of past population fragmentation caused by Pleistocene sea level changes and current patterns of marine circulation. However, the increasing use of nuclear markers has revealed that these two factors alone are not enough to explain the phylogeographic patterns, and an additional role has been suggested for endogenous barriers to gene flow or natural selection. In this article we examined the role of these factors in Ruditapes decussatus, a commercial clam species native to MEDAT. A genetic analysis of 11 populations was carried out by examining 6 introns with a PCR-RFLP technique. We found subdivision in three regions: Atlantic (ATL), western Mediterranean plus Tunisia (WMED), and Aegean and Adriatic seas (AEGAD). Two introns (Ech and Tbp) showed alleles that were restricted to AEGAD. Sequencing a subsample of individuals for these introns indicated that AEGAD-specific alleles were separate clades, thus revealing a phylogeographic brake at the WMED-AEGAD boundary. Sequencing of the mitochondrial COI locus confirmed this phylogeographic break. Dating of the AEGAD mitochondrial haplotypes and nuclear alleles with a Bayesian MCMC method revealed that they shared common ancestors in the Pleistocene. These results can be explained in the framework of Pleistocene sea level drops and patterns of gene flow in MEDAT. An additional observation was a lack of differentiation at COI between the ATL and WMED, in sharp contrast with 4 introns that showed clear genetic subdivision. Neutrality tests did not support the hypothesis of a selective sweep acting on mtDNA to explain the contrasting levels of differentiation between mitochondrial and nuclear markers across the ATL-WMED transition, and we argue that the difference between markers is best explained by the existence of an endogenous genetic barrier, rather than by a physical barrier to larval migration alone.

Predictive models for health outcomes due to SARS-CoV-2, including the effect of vaccination: a systematic review.

BACKGROUND: The interaction between modelers and policymakers is becoming more common due to the increase in computing speed seen in recent decades. The recent pandemic caused by the SARS-CoV-2 virus was no exception. Thus, this study aims to identify and assess epidemiological mathematical models of SARS-CoV-2 applied to real-world data, including immunization for coronavirus 2019 (COVID-19). METHODOLOGY: PubMed, JSTOR, medRxiv, LILACS, EconLit, and other databases were searched for studies employing epidemiological mathematical models of SARS-CoV-2 applied to real-world data. We summarized the information qualitatively, and each article included was assessed for bias risk using the Joanna Briggs Institute (JBI) and PROBAST checklist tool. The PROSPERO registration number is CRD42022344542. FINDINGS: In total, 5646 articles were retrieved, of which 411 were included. Most of the information was published in 2021. The countries with the highest number of studies were the United States, Canada, China, and the United Kingdom; no studies were found in low-income countries. The SEIR model (susceptible, exposed, infectious, and recovered) was the most frequently used approach, followed by agent-based modeling. Moreover, the most commonly used software were R, Matlab, and Python, with the most recurring health outcomes being death and recovery. According to the JBI assessment, 61.4% of articles were considered to have a low risk of bias. INTERPRETATION: The utilization of mathematical models increased following the onset of the SARS-CoV-2 pandemic. Stakeholders have begun to incorporate these analytical tools more extensively into public policy, enabling the construction of various scenarios for public health. This contribution adds value to informed decision-making. Therefore, understanding their advancements, strengths, and limitations is essential.

mRNA-Seq and microarray development for the Grooved Carpet shell clam, Ruditapes decussatus: a functional approach to unravel host-parasite interaction.

BACKGROUND: The Grooved Carpet shell clam Ruditapes decussatus is the autochthonous European clam and the most appreciated from a gastronomic and economic point of view. The production is in decline due to several factors such as Perkinsiosis and habitat invasion and competition by the introduced exotic species, the manila clam Ruditapes philippinarum. After we sequenced R. decussatus transcriptome we have designed an oligo microarray capable of contributing to provide some clues on molecular response of the clam to Perkinsiosis. RESULTS: A database consisting of 41,119 unique transcripts was constructed, of which 12,479 (30.3%) were annotated by similarity. An oligo-DNA microarray platform was then designed and applied to profile gene expression in R. decussatus heavily infected by Perkinsus olseni. Functional annotation of differentially expressed genes between those two conditionswas performed by gene set enrichment analysis. As expected, microarrays unveil genes related with stress/infectious agents such as hydrolases, proteases and others. The extensive role of innate immune system was also analyzed and effect of parasitosis upon expression of important molecules such as lectins reviewed. CONCLUSIONS: This study represents a first attempt to characterize Ruditapes decussatus transcriptome, an important marine resource for the European aquaculture. The trancriptome sequencing and consequent annotation will increase the available tools and resources for this specie, introducing the possibility of high throughput experiments such as microarrays analysis. In this specific case microarray approach was used to unveil some important aspects of host-parasite interaction between the Carpet shell clam and Perkinsus, two non-model species, highlighting some genes associated with this interaction. Ample information was obtained to identify biological processes significantly enriched among differentially expressed genes in Perkinsus infected versus non-infected gills. An overview on the genes related with the immune system on R. decussatus transcriptome is also reported.

Genetic and phenoptypic differentiation of zebra mussel populations colonizing Spanish river basins.

Zebra mussel populations in Ebro and Mijares Rivers (northern Spain) were analyzed to study the mechanisms by which this aquatic species deals with pollution. Variability analyses of mitochondrial cytochrome oxidase I gene and of one nuclear microsatellite were performed for ten populations from the Ebro River and one from the Mijares River. Comparison of these results with those from five additional European populations indicated that the Spanish populations constitute a homogeneous gene pool. Transcriptome analyses of gill samples from a subset of the Spanish populations showed changes on expression levels that correlated with variations in general fitness and loads of heavy metals. The less polluted upstream Ebro populations showed overexpression of mitochondrial and cell proliferation-related genes compared to the more polluted, downstream Ebro populations. Our data indicate that heavy metals were the main factors explaining these transcriptomic patterns, and that zebra mussel is resilient to pollutants (like mercury and organochlorine compounds) proved to be extremely toxic to vertebrates. We propose that zebra mussel populations sharing a common gene pool may acclimate to different levels and forms of pollution through modulations in their transcriptomic profile, although direct selection on genes showing differential expression patterns cannot be ruled out.

Electromagnetic deactivation spectroscopy of human coronavirus 229E.

An investigation of the deactivation of pathogens using electromagnetic waves in the microwave region of the spectrum is achieved using custom-built waveguide structures. The waveguides feature sub-wavelength gratings to allow the integration of an air cooling system without disturbing the internal propagating fields. The waveguides are tapered to accommodate an experimental sample internally with sufficient surrounding airflow. The proposed methodology allows for precise control over power densities due to the well-defined fundamental mode excited in each waveguide, in addition to temperature control of the sample due to microwave exposure over time. Human coronavirus (HCoV-229E) is investigated over the 0-40 GHz range, where a peak 3-log viral reduction is observed in the 15.0-19.5 GHz sub-band. We conclude HCoV-229E has an intrinsic resonance in this range, where nonthermal structure damage is optimal through the structure-resonant energy transfer effect.

Label-free observation of individual solution phase molecules.

The vast majority of chemistry and biology occurs in solution, and new label-free analytical techniques that can help resolve solution-phase complexity at the single-molecule level can provide new microscopic perspectives of unprecedented detail. Here, we use the increased light-molecule interactions in high-finesse fiber Fabry-Pérot microcavities to detect individual biomolecules as small as 1.2 kDa with signal-to-noise ratios >100, even as the molecules are freely diffusing in solution. Our method delivers 2D intensity and temporal profiles, enabling the distinction of sub-populations in mixed samples. Strikingly, we observe a linear relationship between passage time and molecular radius, unlocking the potential to gather crucial information about diffusion and solution-phase conformation. Furthermore, mixtures of biomolecule isomers of the same molecular weight can also be resolved. Detection is based on a novel molecular velocity filtering and dynamic thermal priming mechanism leveraging both photo-thermal bistability and Pound-Drever-Hall cavity locking. This technology holds broad potential for applications in life and chemical sciences and represents a major advancement in label-free in vitro single-molecule techniques.

Translation, adaptation, and validation of the MeNTS score in colombian population: The MeNTS Col score.

Objective: To evaluate and validate the medically necessary and time sensitive score by testing the variables, in order to create a surgical preoperative score for procedure prioritization in COVID-19 pandemic in Colombia. Methods: A multicenter retrospective cross-sectional study of instrument validation with a cultural adaptation and translation into the Spanish language was carried out in Bogota, Colombia. Patients over 18 years of age who had undergone elective procedures of general surgery and subspecialties were included. The translation of the medically necessary and time sensitive score into Spanish was performed independently by two bilingual surgeons fluent in both English and Spanish. A final version of the Spanish questionnaire (MeNTS Col) for testing was then produced by an expert committee. After translation and cultural adaptation, it was submitted to evaluate the psychometric properties of the medically necessary and time sensitive score. Cronbach's α was used to represent and evaluate the internal consistency and assess reliability. Results: A total of 172 patients were included, with a median age of 54 years; of which 96 (55.8%) patients were females. The vast majority of patients were treated for general surgery (n = 60) and colon and rectal surgery (n = 31). The evaluation of the internal consistency of the scale items in Spanish version was measured, and values of 0.5 for 0.8 were obtained. In the reliability and validation process, Cronbach's α values in all items remained higher than 0.7. The new MeNTS Col model was analyzed, and a result of 0.91 was obtained. Conclusions: The Spanish version of the medically necessary and time sensitive, the MeNTS Col score, and its respective Spanish translation perform similarly to the original version. Therefore, they can be useful and reproducible in Latin American countries.

Conformation and chiral effects in α,ß,α-tripeptides.

Short α,ß,α-tripeptides comprising a central chiral trisubstituted ß(2,2,3)*-amino acid residue form unusual γ-turns and δ-turns in CDCl(3) and DMSO-d(6) solutions but do not form ß-turns. Thermal coefficients of backbone amide protons, 2D-NMR spectra, and molecular modeling revealed that these motifs were strongly dependent on the configuration (chiral effect) of the central ß-amino acid residue within the triad. Accordingly, SSS tripeptides adopted an intraresidual γ-turn like (C6) arrangement in the central ß-amino acid, whereas SRS diastereomers preferred an extended δ-turn (C9) conformation. A different SRS-stabilizing bias was observed in the crystal structures of the same compounds, which shared the extended δ-turn (C9) found in solution, but incorporated an additional extended ß-turn (C11) to form an overlapped double turn motif.

Preparation of modified peptides: direct conversion of α-amino acids into ß-amino aldehydes.

A direct method for the transformation of α-amino acids into ß-amino aldehydes was developed, and applied to the modification of the C-terminal residue of peptides. The method takes place in good yields and under mild conditions. The application of this methodology to the preparation of small peptides with γ-amino alcohol units, which are precursors of analogues of peptaibol antibiotics, is also described.

Active optics for dynamical correction of fluctuations of atmospheric refraction on a differential optical absorption spectroscopy device.

We have designed and developed a feedback mechanism for continuous monitoring in a long-pass differential optical absorption spectroscopy (LP-DOAS) setup. This allows one to correct photo-thermal deflection due to the local fluctuations refraction index of the air. For this purpose, using an unbalanced beam splitter, a small fraction of the collected DOAS signal is imaged onto a low-cost CCD camera using a biconvex lens, while the other portion of the signal is coupled into a fiber optic for trace gas detection. By monitoring the registered signal at the CCD camera, a feedback mechanism acting on the transversal position of the lens is able to compensate an arbitrary transversal displacement of the collected signal at the focal plane of the receiver telescope, allowing an optimal coupling into the optical fiber.