Extracellular matrix protein N-glycosylation mediates immune self-tolerance in Drosophila melanogaster.

In order to respond to infection, hosts must distinguish pathogens from their own tissues. This allows for the precise targeting of immune responses against pathogens and also ensures self-tolerance, the ability of the host to protect self tissues from immune damage. One way to maintain self-tolerance is to evolve a self signal and suppress any immune response directed at tissues that carry this signal. Here, we characterize the Drosophila tuSz1 mutant strain, which mounts an aberrant immune response against its own fat body. We demonstrate that this autoimmunity is the result of two mutations: 1) a mutation in the GCS1 gene that disrupts N-glycosylation of extracellular matrix proteins covering the fat body, and 2) a mutation in the Drosophila Janus Kinase ortholog that causes precocious activation of hemocytes. Our data indicate that N-glycans attached to extracellular matrix proteins serve as a self signal and that activated hemocytes attack tissues lacking this signal. The simplicity of this invertebrate self-recognition system and the ubiquity of its constituent parts suggests it may have functional homologs across animals.

Investigación evaluativa de un protocolo para urgencias de salud mental en el primer nivel de atención.

Introducción: Esta investigación parte de algunos obstáculos en la implementación de la Ley Nacional de Salud Mental, especialmente en el primer nivel de atención y en lo relativo al manejo de urgencias, con o sin riesgo cierto e inminente. La escasez de protocolos de intervención en salud mental y la percepción de que podrían limitar la lectura singular de cada situación crítica son desafíos identificados en la práctica. No obstante, los protocolos pueden ser herramientas organizativas valoradas en circunstancias complejas. En la bibliografía se destaca la necesidad de acompañar su implementación con capacitación para evitar enfoques exclusivamente burocráticos y deshumanizados. Materiales y métodos: Para abordar esta brecha, se realizó esta investigación,en la que se diseñó un protocolo de actuación en urgencias de salud mental y una capacitación acorde para el personal de salud del primer nivel de atención de un  territorio del área metropolitana de Buenos Aires. El objetivo de la investigación fue evaluar su efectividad. Se utilizaron métodos cuali-cuantitativos con el fin de caracterizar las intervenciones previas a la implementación del protocolo, comparándolas con las posteriores a la capacitación y aplicación del protocolo. Resultados: Se halló que su implementación favoreció respuestas más adecuadas en contextos críticos y mejoró el conocimiento acerca de las estrategias de resolución que pueden efectuarse en el marco de la Ley Nacional de Salud Mental, reforzando asimismo factores protectores de la salud mental de los trabajadores de la salud, tales como el acceso a una estrategia que colabore en la organización de la tarea y el trabajo en equipo.

SifR is an Rrf2-family quinone sensor associated with catechol iron uptake in Streptococcus pneumoniae D39.

Streptococcus pneumoniae (pneumococcus) is a Gram-positive commensal and human respiratory pathogen. How this bacterium satisfies its nutritional iron (Fe) requirement in the context of endogenously produced hydrogen peroxide is not well understood. Here, we characterize a novel virulence-associated Rrf2-family transcriptional repressor that we term SifR (streptococcal IscR-like family transcriptional repressor) encoded by spd_1448 and conserved in Streptococci. Global transcriptomic analysis of a ΔsifR strain defines the SifR regulon as genes encoding a candidate catechol dioxygenase CatE, an uncharacterized oxidoreductase YwnB, a candidate flavin-dependent ferric reductase YhdA, a candidate heme-based ferric reductase domain-containing protein and the Piu (pneumococcus iron uptake) Fe transporter (piuBCDA). Previous work established that membrane-anchored PiuA binds FeIII-bis-catechol or monocatechol complexes with high affinity, including the human catecholamine stress hormone, norepinephrine. We demonstrate that SifR senses quinone via a single conserved cysteine that represses its regulon when in the reduced form. Upon reaction with catechol-derived quinones, we show that SifR dissociates from the DNA leading to regulon derepression, allowing the pneumococcus to access a catechol-derived source of Fe while minimizing reactive electrophile stress induced by quinones. Consistent with this model, we show that CatE is an FeII-dependent 2,3-catechol dioxygenase with broad substrate specificity, YwnB is an NAD(P)H-dependent quinone reductase capable of reducing the oxidized and cyclized norepinephrine, adrenochrome, and YhdA is capable of reducing a number of FeIII complexes, including PiuA-binding transport substrates. These findings are consistent with a model where FeIII-catechol complexes serve as significant nutritional Fe sources in the host.

Multiresidue method for the determination of critically and highly important classes of antibiotics and their metabolites in agricultural soils and sewage sludge.

In this paper, a method is proposed for the determination of antibiotics classified by the World Health Organization as critically important (four macrolides and three quinolones) and highly important (one tetracycline, one diaminopyridine, and three sulfonamides) and eight of their metabolites. The method is based on ultrasound-assisted extraction, dispersive solid-phase extraction clean-up, and analytical determination by liquid chromatography-tandem mass spectrometry. Variables affecting each stage of the analytical method were thoroughly optimised. The method was validated for its application to sewage sludge from different treatment stages (non-treated sludge: primary and secondary sludge; and treated sludge: digested sludge and compost) and to agricultural soil. Limits of quantification were in the range of 0.03-7.50 ng g-1 dry weight (dw) for most of the compounds. Accuracy values were in the range of 70-102%. Precision was below 17%. The application of the method to real samples revealed that macrolides and fluoroquinolones were the antibiotic classes at the highest concentrations in all types of samples. The lowest concentrations of antibiotics were measured in compost (highest concentration: 27 ng g-1 dw, corresponding to norfloxacin) and soil samples (highest concentration: 93 ng g-1 dw, corresponding to a metabolite of clarithromycin). The proposed method is the first developed to date for the determination of multiclass antibiotics and their main metabolites in sludge from different treatment stages. The method can provide a useful tool for obtaining information about antibiotics in sewage sludge prior to its application to agricultural soils and in agricultural soils.

Adsorption of perfluoroalkyl substances on polyamide microplastics: Effect of sorbent and influence of environmental factors.

Microplastics (MPs) and perfluoroalkyl substances (PFASs) are two types of pollutants coexisting in the environment. Their co-exposure is a source of increasing concern. MPs present in the natural environment suppose an ideal surface for the sorption of hazardous contaminants. This study investigates the adsorption behaviour of six PFASs on polyamide (PA) MPs. Adsorption experiments under various internal (PA and PFASs dosage, PA particle size) and environmental (pH, ionic strength, dissolved organic matter) factors were carried out. Isotherm results (from 0.1 to 25 mg/L of PFASs) showed that the maximum adsorption capacity of the selected PFASs on the PA was as follows: perfluorooctanesulfonic acid (PFOS, 0.873 mg/g) > perfluorooctanoic acid (0.235 mg/g) > perfluoroheptanoic acid (0.231 mg/g) > perfluorohexanoic acid (0.201 mg/g) > perfluoropentanoic acid (0.192 mg/g) > perfluorobutanoic acid (0.188 mg/g) (pH 5.88, 0% salinity and 0% of dissolved organic matter). The PFOS has more tendency to be sorbed onto PA than perfluorocarboxilic acids. The MP characterization by scanning electron microscopy, X ray diffraction and Fourier transform infrared spectroscopy showed changes in the PA surface after adsorption assays. Pore filling, hydrophobic interactions and hydrogen bonds governed sorption process. The sorption capacity of PFASs was crucially affected by the PA size (from 19.6% to 99.9% for 3 mm and 50 µm particle size, respectively). The process was not significantly influenced by salinity while the dissolved organic matter exerted a negative effect (decrease from 100% to 26% for PFOS in presence of 25 mg/L of humic acid). Finally, adsorption rates of PFASs were quantified in real water matrices (influent and effluent wastewater, surface and tap water samples). The results revealed interactions between PA and PFASs and evidenced the role of PA as a vector to transport PFASs in the aquatic environment.

Uptake and depuration of three common antibiotics in benthic organisms: Sea cucumber (Holothuria tubulosa), snakelocks anemone (Anemonia sulcata) and beadlet anemone (Actinia equina).

Antibiotics are widely used drugs in human and veterinary medicine, which has attracted great attention in relation to the development of bacterial resistance, currently a problem of great concern for governments and states, as it is related to the resurgence of infectious diseases already eradicated. Understanding the bioaccumulation of antibiotics in aquatic organisms is an important key to understanding their risk assessment. The present study was designed to study the bioaccumulation of target antibiotics in relevant organisms inhabiting benthic marine environments. The uptake and elimination of ciprofloxacin (CIP), sulfamethoxazole (SMX) and trimethoprim (TMP) were investigated in sea cucumbers (Holothuria tubulosa), snakelock anemone (Anemonia sulcata) and beadlet anemone (Actinia equina) under controlled laboratory conditions. The results show that antibiotics have a particular trend over time during all periods of absorption and depuration. The tissue distribution of antibiotics in sea cucumber is strongly influenced by the structure of the compounds, while CIP is concentrated in the body wall; TMP is concentrated in the digestive tract. Two different approaches were used to estimate bioconcentration factors (BCFs) in different animal models, based on toxicokinetic data and measured steady-state concentrations. The BCF ranges were 456-2731 L/kg, 6-511 L/kg and 9-100 L/kg for TMP, CIP and SMX, respectively. The estimated BCF values obtained classify TMP as cumulative in A. equina and H. tubulosa, underlining the potential bioconcentration in these marine organisms. A correlation was observed between the BCFs of the target antibiotics and the octanol-water distribution coefficient (Dow) (r2 > 0.7). The animal-specific BCF followed the order of beadlet anemone > sea cucumber > snakelock anemone.

Thermodynamic and Kinetic Investigation of the Adsorption and Desorption of Trimethoprim and Its Main Metabolites in Mediterranean Crop Soils.

The adsorption-desorption processes of organic pollutants into the soil are one of the main factors influencing their potential environmental risks and distribution in the environment. In the present work, the adsorption-desorption behavior of an antibiotic, trimethoprim (TMP), and two of its main metabolites, 3-desmethyltrimethoprim (DM-TMP) and 4-hydroxytrimethoprim (OH-TMP), were assessed in three Mediterranean agricultural soils with different physicochemical characteristics. Results showed that the adsorption kinetic is performed in two steps: external sorption and intraparticle diffusion. The adsorptions of the studied compounds in soils were similar and fitted to the three models but were better fitted to a linear model. In the case of DM-TMP and OH-TMP, their adsorptions were positively correlated with the soil organic matter. In addition, desorption was higher in less organic matter soil (from 1.3 to 30.9%). Furthermore, the desorptions measured for the TMP metabolites were lower than those measured in the case of TMP (from 2.0 and 4.0% for OH-TMP and DM-TMP, respectively, to 9.0% for TMP).

Bioorthogonal, Fluorogenic Targeting of Voltage-Sensitive Fluorophores for Visualizing Membrane Potential Dynamics in Cellular Organelles.

Electrical potential differences across lipid bilayers play foundational roles in cellular physiology. Plasma membrane voltage is the most widely studied; however, the bilayers of organelles like mitochondria, lysosomes, nuclei, and the endoplasmic reticulum (ER) also provide opportunities for ionic compartmentalization and the generation of transmembrane potentials. Unlike plasma membranes, organellar bilayers, cloistered within the cell, remain recalcitrant to traditional approaches like patch-clamp electrophysiology. To address the challenge of monitoring changes in organelle membrane potential, we describe the design, synthesis, and application of the LUnAR RhoVR (Ligation Unquenched for Activation and Redistribution Rhodamine-based Voltage Reporter) for optically monitoring membrane potential changes in the ER of living cells. We pair a tetrazine-quenched RhoVR for voltage sensing with a transcyclooctene (TCO)-conjugated ceramide (Cer-TCO) for targeting to the ER. Bright fluorescence is observed only at the coincidence of the LUnAR RhoVR and TCO in the ER, minimizing non-specific, off-target fluorescence. We show that the product of the LUnAR RhoVR and Cer-TCO is voltage-sensitive and that the LUnAR RhoVR can be targeted to an intact ER in living cells. Using the LUnAR RhoVR, we use two-color, ER-localized, fast voltage imaging coupled with cytosolic Ca2+ imaging to validate the electroneutrality of Ca2+ release from internal stores. Finally, we use the LUnAR RhoVR to directly visualize functional coupling between the plasma-ER membranes in patch clamped cell lines, providing the first direct evidence of the sign of the ER potential response to plasma membrane potential changes. We envision that the LUnAR RhoVR, along with other existing organelle-targeting TCO probes, could be applied widely for exploring organelle physiology.

Postoperative Packing of Perianal Abscess Cavities (PPAC2): randomized clinical trial.

BACKGROUND: Perianal abscess is common. Traditionally, postoperative perianal abscess cavities are managed with internal wound packing, a practice not supported by evidence. The aim of this randomized clinical trial (RCT) was to assess if non-packing is less painful and if it is associated with adverse outcomes. METHODS: The Postoperative Packing of Perianal Abscess Cavities (PPAC2) trial was a multicentre, RCT (two-group parallel design) of adult participants admitted to an NHS hospital for incision and drainage of a primary perianal abscess. Participants were randomized 1:1 (via an online system) to receive continued postoperative wound packing or non-packing. Blinded data were collected via symptom diaries, telephone, and clinics over 6 months. The objective was to determine whether non-packing of perianal abscess cavities is less painful than packing, without an increase in perianal fistula or abscess recurrence. The primary outcome was pain (mean maximum pain score on a 100-point visual analogue scale). RESULTS: Between February 2018 and March 2020, 433 participants (mean age 42 years) were randomized across 50 sites. Two hundred and thirteen participants allocated to packing reported higher pain scores than 220 allocated to non-packing (38.2 versus 28.2, mean difference 9.9; P < 0.0001). The occurrence of fistula-in-ano was low in both groups: 32/213 (15 per cent) in the packing group and 24/220 (11 per cent) in the non-packing group (OR 0.69, 95 per cent c.i. 0.39 to 1.22; P = 0.20). The proportion of patients with abscess recurrence was also low: 13/223 (6 per cent) in the non-packing group and 7/213 (3 per cent) in the packing group (OR 1.85, 95 per cent c.i. 0.72 to 4.73; P = 0.20). CONCLUSION: Avoiding abscess cavity packing is less painful without a negative morbidity risk. REGISTRATION NUMBER: ISRCTN93273484 (https://www.isrctn.com/ISRCTN93273484). REGISTRATION NUMBER: NCT03315169 (http://clinicaltrials.gov).

Perianal abscess is a common, painful condition due to infection and swelling around the anus caused by blockage of the anal glands. The treatment of perianal abscess has stayed the same for over 50 years. An operation is performed under general anaesthetic to cut the skin and drain the infection. This is followed by continued internal dressing (packing) of the remaining cavity (hole) until the skin has healed over. Packing changes are needed multiple times a week for several weeks. Packing is the accepted treatment as it is believed to reduce the chance of the abscess coming back, and also reduces the chance of perianal fistula forming. There are no medical studies to support this idea. Perianal fistula (an abnormal passage between the skin around the anus, and the inside of the anal canal or rectum) is a long-term condition, which causes pain, and pus (and sometimes faeces) discharge, and often needs another operation (or multiple operations) to fix it. This trial was performed to demonstrate if no packing of a perianal abscess would result in a reduction of pain, with no increase in unwanted abscess recurrences and fistulas, in comparison to the standard treatment of packing. The trial recruited 433 people, who were randomly chosen to enter one of two groups; one to have their wound packed and the other to have no packing of the wound. After being discharged from hospital following surgery, the patients attended or were visited by a community nurse for the dressing to be changed or wound packed. Each patient provided information on pain from their wound, including worst pain each day and pain before, during, and after the changing of their dressing or packing. This and other information was gathered for the first 10 days after surgery and periodically until 6 months after surgery. The no-packing group experienced much less pain than the packing group. There was no difference in abscess recurrence and fistula formation between the non-packing and packing groups. The findings demonstrate that no packing of perianal abscess wounds after drainage operation is the best treatment.

Development and validation of a highly effective analytical method for the evaluation of the exposure of migratory birds to antibiotics and their metabolites by faeces analysis.

The widespread occurrence of antibiotics in the environment may exert a negative impact on wild organisms. In addition, they can become environmental reservoirs, through the ingestion of food or contaminated water, and vectors for antibiotic-resistant bacteria. This fact is even more important in migratory birds that can promote their dissemination across continents. In this work, a multiresidue analytical method suitable for the determination of five families of antibiotics and their main metabolites in waterbird faeces has been developed and validated. The target compounds include environmentally significant sulfonamides, macrolides, fluoroquinolones, tetracyclines and antifolates. Sample treatment involves ultrasound-assisted extraction with methanol and dispersive solid-phase extraction clean-up with C18. Analytical determination was carried out by liquid chromatography-tandem mass spectrometry. The most significant parameters affecting sample extraction and extract clean-up were optimised by means of experimental designs. Good linearity (R2 > 0.994), accuracy (from 41 to 127%), precision (relative standard deviation lower than 24%) and limits of quantification (lower than 2 ng g-1 (dry weight, dw)) were obtained for most of the compounds. The method was applied to the determination of the selected compounds in 27 faeces samples from three common migratory waterbird species. Nine antibiotics and three of their metabolites were detected in the analysed samples. Fluoroquinolones and macrolides were the antibiotics most frequently detected. The highest concentrations corresponded to norfloxacin (up to 199 ng g-1 dw).

Ultrasound-assisted extraction as an easy-to-perform analytical methodology for monitoring ibuprofen and its main metabolites in mussels.

Non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to be the main pharmaceutical class accumulated in seafood. Among them, ibuprofen (IBU) is of special concern as it is used worldwide to treat common pain, does not require a medical prescription, it is often taken in a high daily dose, and has been reported to cause potential adverse effects on aquatic organisms. IBU is highly transformed into hydroxy- and carboxy-metabolites and/or degradation products generated not only after its administration but also during wastewater treatment or in the environment. These compounds can be present in the environment at higher concentrations than IBU and present higher toxicity. In this work, a low-cost and affordable routine analytical method was developed and validated for the first-time determination of IBU and its main metabolites in mussels. The method is based on ultrasound-assisted extraction (UAE), clean-up by dispersive solid-phase extraction (d-SPE) and analytical determination by liquid chromatography-tandem mass spectrometry. Box-Behnken experimental design was used for method optimisation to better evaluate the influence and interactions of UAE and d-SPE variables. Extraction recoveries were in the range from 81 to 115%. Precision, expressed as relative standard deviation, was lower than 7%. Method detection limits were in the range from 0.1 to 1.9 ng g-1 dry weight. The method was successfully applied to wild mussels.

Antibiotic adsorption by natural and modified clay minerals as designer adsorbents for wastewater treatment: A comprehensive review.

Increased antibiotic use worldwide has become a major concern because of their health and environmental impacts. Since most antibiotic residues can hardly be removed from wastewater using conventional treatments, alternative methods receive great attention. Adsorption is one of the most efficient and cost-effective treatment methods for antibiotics. Among the adsorbents, clay minerals have garnered increasing attention due to their unique properties including availability, high specific surface area, low cost, cation exchange capacity, and good removal efficiency. This paper reviews the recent progress made in the use of natural and modified clay minerals for the removal of antibiotics from water. First, the sources, occurrence, removal and health effects of the antibiotics commonly encountered in water bodies are described. Antibiotic concentration levels and average removal efficiencies measured in conventional activated sludge treatment systems worldwide are also provided to better address the problem. Second, the review explores the characteristics of clay minerals as adsorbent of antibiotics and the factors affecting the adsorption. The review identifies and discusses the future trends and strategies used to increase the adsorption capacity of clay minerals by modification and combination techniques (intercalation of novel functional groups such as organocations, biopolymers and metal pillared-clay minerals, combination with biochar or thermal activation). The quantitative comparisons of clay minerals' ability for antibiotic removal are given. Some natural clay minerals have good removal potential for antibiotics, with maximum adsorption capacities over 100 mg/g. For most other adsorbents, surface modifications and combination techniques resulted in improved adsorption properties (including higher surface area, enhanced adsorption capacity, increased stability and mechanical strength). Finally, the application of these adsorbents at pilot scale, using real wastewater samples, their reuse, economic analysis and life cycle assessment are other issues that have been considered.

Comparison of Different Techniques for the Determination of Platinized Cytostatic Drugs in Urine Samples.

Platinum-based cytostatic drugs are one of the most widely used cancer treatments. They are excreted via the urinary tract and can reach the environment through wastewater, posing a risk to human health due to their side effects. Four identification and quantification techniques, including liquid chromatography (LC) separation coupled to (i) a diode array ultraviolet (UV(DAD)) (ii), mass spectrometer in single ion monitoring mode (LC-MS) and (iii) multiple reaction monitoring mode (LC-MS/MS) and (iv) derivatization with diethyldithiocarbamate prior to LC-MS/MS analysis, have been optimized and compared for the multiresidue determination of main platinized cytostatic drugs (cisplatin, carboplatin, and oxaliplatin) in urine samples. Parameters that affect the efficiency of the chromatographic separation and analytical determination of different methods (column, mobile phase, wavelength, precursor ions, fragmentor, and product ions) were optimized. Analytical features, such as matrix effect, sensitivity, precision, selectivity, and linearity, were calculated. In terms of selectivity, the derivatization technique was discarded since it was only applicable to the platinated sum. A high dilution of the sample with LC-UV(DAD) was needed to reduce the matrix effect. Overall, the LC-MS/MS method presented the best analytical features (% RSD ≤ 12.8%, R2 ≥ 0.991, or method-detection limits between 0.01-1 µg mL-1). The selected method was applied to the quantification of platinized cytostatic drugs in hospital urine samples from oncologic patients.

An Overview of Analytical Methods to Determine Pharmaceutical Active Compounds in Aquatic Organisms.

There is increasing scientific evidence that some pharmaceuticals are present in the marine ecosystems at concentrations that may cause adverse effects on the organisms that inhabit them. At present, there is still very little scientific literature on the (bio)accumulation of these compounds in different species, let alone on the relationship between the presence of these compounds and the adverse effects they produce. However, attempts have been made to optimize and validate analytical methods for the determination of residues of pharmaceuticals in marine biota by studying the stages of sample treatment, sample clean-up and subsequent analysis. The proposed bibliographic review includes a summary of the most commonly techniques, and its analytical features, proposed to determine pharmaceutical compounds in aquatic organisms at different levels of the trophic chain in the last 10 years.

Controlling the Structure, Properties and Surface Reactivity of Clickable Azide-Functionalized Au25 (SR)18 Nanocluster Platforms Through Regioisomeric Ligand Modifications.

To fine-tune structure-property correlations of thiolate-protected gold nanoclusters through post-assembly surface modifications, we report the synthesis of the o, m, and p regioisomeric forms of the anionic azide-functionalized [Au25 (SCH2 CH2 -C6 H4 -N3 )18 ]1- platform. They can undergo cluster-surface strain-promoted alkyne-azide cycloaddition (CS-SPAAC) chemistry with complementary strained-alkynes. Although their optical properties are similar, the electrochemical properties appear to correlate with the position of the azido group. The ability to conduct CS-SPAAC chemistry without altering the parent nanocluster structure is different as the isomeric form of the surface ligand is changed, with the [Au25 (SCH2 CH2 -p-C6 H4 -N3 )18 ]1- isomer having the highest reaction rates, while the [Au25 (SCH2 CH2 -o-C6 H4 -N3 )18 ]1- isomer is not stable following CS-SPAAC. Single-crystal X-ray diffraction provide the molecular structure of the neutral forms of the three regioisomeric clusters, [Au25 (SCH2 CH2 -o/m/p-C6 H4 -N3 ]0 , which illustrates correlated structural features of the central core as the position of the azido moiety is changed.

[Use of Information and Communication Technologies and health personnel from Argentine during COVID-19 pandemic]. / Uso de Tecnologías de la Información y la Comunicación y personal de salud en Argentina durante la pandemia por COVID-19

INTRODUCTION: Since the beginning of the COVID-19 pandemic, the importance of mental health actions for health personnel was emphasized and the use of Information and Communication Technologies (ICTs) was recommended. Given this unprecedented panorama, the objective of this research was to evaluate the effectiveness of an intervention with ICTs for health personnel from first-level care networks in Berazategui, Buenos Aires, Argentina. METHOD: It was an exploratory qualitative research and evaluative design. The selection of subjects included intervened health personnel who agreed to participate. Satisfaction surveys were implemented, records of the intervening team and their meetings were analyzed. RESULTS: The intervention was effective: improvements in communication and interaction, self-care and a change to a proactive and creative attitude at work, decreased anxiety and stress were detected. As a protective factor, the fact of having a solid socio-affective network was highlighted. The effectiveness was characterized by influencing a repositioning of the health personnel with respect to the context, the task and the work ties. CONCLUSION: The use of ICTs was advantageous, enabled access to mental health care and generated the feeling of continuous support, strengthening the socio-affective network.

Cellular Mn/Zn Ratio Influences Phosphoglucomutase Activity and Capsule Production in Streptococcus pneumoniae D39.

Capsular polysaccharide (CPS) is a major virulence determinant for many human-pathogenic bacteria. Although the essential functional roles for CPS in bacterial virulence have been established, knowledge of how CPS production is regulated remains limited. Streptococcus pneumoniae (pneumococcus) CPS expression levels and overall thickness change in response to available oxygen and carbohydrate. These nutrients in addition to transition metal ions can vary significantly between host environmental niches and infection stage. Since the pneumococcus must modulate CPS expression among various host niches during disease progression, we examined the impact of the nutritional transition metal availability of manganese (Mn) and zinc (Zn) on CPS production. We demonstrate that increased Mn/Zn ratios increase CPS production via Mn-dependent activation of the phosphoglucomutase Pgm, an enzyme that functions at the branch point between glycolysis and the CPS biosynthetic pathway in a transcription-independent manner. Furthermore, we find that the downstream CPS protein CpsB, an Mn-dependent phosphatase, does not promote aberrant dephosphorylation of its target capsule-tyrosine kinase CpsD during Mn stress. Together, these data reveal a direct role for cellular Mn/Zn ratios in the regulation of CPS biosynthesis via the direct activation of Pgm. We propose a multilayer mechanism used by the pneumococcus in regulating CPS levels across various host niches. IMPORTANCE Evolving evidence strongly indicates that maintenance of metal homeostasis is essential for establishing colonization and continued growth of bacterial pathogens in the vertebrate host. In this study, we demonstrate the impact of cellular manganese/zinc (Mn/Zn) ratios on bacterial capsular polysaccharide (CPS) production, an important virulence determinant of many human-pathogenic bacteria, including Streptococcus pneumoniae. We show that higher Mn/Zn ratios increase CPS production via the Mn-dependent activation of the phosphoglucomutase Pgm, an enzyme that functions at the branch point between glycolysis and the CPS biosynthetic pathway. The findings provide a direct role for Mn/Zn homeostasis in the regulation of CPS expression levels and further support the ability of metal cations to act as important cellular signaling mediators in bacteria.

Imaging Reversible Mitochondrial Membrane Potential Dynamics with a Masked Rhodamine Voltage Reporter.

Mitochondria are the site of aerobic respiration, producing ATP via oxidative phosphorylation as protons flow down their electrochemical gradient through ATP synthase. This negative membrane potential across the inner mitochondrial membrane (ΔΨm) represents a fundamental biophysical parameter central to cellular life. Traditional, electrode-based methods for recording membrane potential are impossible to implement on mitochondria within intact cells. Fluorescent ΔΨm indicators based on cationic, lipophilic dyes are a common alternative, but these indicators are complicated by concentration-dependent artifacts and the requirement to maintain dye in the extracellular solution to visualize reversible ΔΨm dynamics. Here, we report the first example of a fluorescent ΔΨm reporter that does not rely on ΔΨm-dependent accumulation. We redirected the localization of a photoinduced electron transfer (PeT)-based indicator, Rhodamine Voltage Reporter (RhoVR), to mitochondria by masking the carboxylate of RhoVR 1 as an acetoxymethyl (AM) ester. Once within mitochondria, esterases remove the AM ester, trapping RhoVR inside of the mitochondrial matrix, where it can incorporate within the inner membrane and reversibly report on changes in ΔΨm. We show that this Small molecule, Permeable, Internally Redistributing for Inner membrane Targeting Rhodamine Voltage Reporter, or SPIRIT RhoVR, localizes to mitochondria across a number of different cell lines and responds reversibly to changes in ΔΨm induced by exceptionally low concentrations of the uncoupler FCCP without the need for exogenous pools of dye (unlike traditional, accumulation-based rhodamine esters). SPIRIT RhoVR is compatible with multi-color imaging, enabling simultaneous, real-time observation of cytosolic Ca2+, plasma membrane potential, and reversible ΔΨm dynamics.

Validation of ICD-10-CM codes for injuries complicating pregnancy, childbirth and the puerperium: a medical record review.

INTRODUCTION: The purpose of this study was to estimate the positive predictive value (PPV) of International Classification of Diseases, 10th Revision, Clinical Modification (ICD-10-CM) codes for injury, poisoning, physical or sexual assault complicating pregnancy, childbirth and the puerperium (PCP) to capture injury encounters within both hospital and emergency department claims data. METHODS: A medical record review was conducted on a sample (n=157) of inpatient and emergency department claims from one Kentucky healthcare system from 2015 to 2017, with any diagnosis in the ICD-10-CM range O9A.2-O9A.4. Study clinicians reviewed medical records for the sampled cases and used an abstraction form to collect information on documented presence of injury and PCP complications. The study estimated the PPVs and the 95% CIs of O9A.2-O9A.4 codes for (1) capturing injuries and (2) capturing injuries complicating PCP. RESULTS: The estimated PPV for the codes O9A.2-O9A.4 to identify injury in the full sample was 79.6% (95% CI 73.3% to 85.9%) and the PPV for capturing injuries complicating PCP was 72.0% (95% CI 65.0% to 79.0%). The estimated PPV for an inpatient principal diagnosis O9A.2-O9A.4 to capture injuries was 90.7% (95% CI 82.0% to 99.4%) and the PPV for capturing injuries complicating PCP was 88.4% (95% CI 78.4% to 98.4%). The estimated PPV for any mention of O9A.2-O9A.4 in emergency department data to capture injuries was 95.2% (95% CI 90.6% to 99.9%) and the PPV for capturing injuries complicating PCP was 81.0% (95% CI 72.4% to 89.5%). DISCUSSION: The O9A.2-O9A.4 codes captured high percentage true injury cases among pregnant and puerperal women.

The risk and predictors of mortality in octogenarians undergoing emergency laparotomy: a multicentre retrospective cohort study.

OBJECTIVES: This study aims to evaluate the risk of postoperative mortality in octogenarians undergoing emergency laparotomy. METHODS: In compliance with STROCSS guideline for observational studies, we conducted a multicentre retrospective cohort study. All consecutive patients aged over 80 with acute abdominal pathology requiring emergency laparotomy between April 2014 and August 2019 were considered eligible for inclusion. The primary outcome measure was 30-day postoperative mortality, and the secondary outcome measures were in-hospital mortality and 1-year mortality. Statistical analyses included simple descriptive statistics, binary logistic regression analyses, and Kaplan-Meier survival statistics. RESULTS: A total of 523 octogenarians were eligible for inclusion. Emergency laparotomy in octogenarians was associated with 21.8% (95% CI 18.3-25.6%) 30-day postoperative mortality, 22.6% (95% CI 19.0-26.4%) in-hospital mortality, and 40.2% (95% CI 35.9-44.5%) 1-year mortality. Binary logistic regression analysis identified ASA status (OR, 2.49; 95% CI 1.82-3.38, P < 0.0001) and peritoneal contamination (OR, 2.00; 95% CI 1.30-3.08, P = 0.002) as predictors of 30-day postoperative mortality. The ASA status (OR, 1.92; 95% CI 1.50-2.46, P < 0.0001), peritoneal contamination (OR, 1.57; 95% CI 1.07-2.48, P = 0.020), and presence of malignancy (OR, 2.06; 95% CI 1.36-3.10, P = 0.001) were predictors of 1-year mortality. Log-rank test showed significant difference in postoperative survival rates among patients with different ASA status (P < 0.0001) and between patients with and without peritoneal contamination (P = 0.0011). CONCLUSIONS: Emergency laparotomies in patients older than 80 years with ASA status more than 3 in the presence of peritoneal contamination carry a high risk of immediate postoperative and 1-year mortality. This should be taken into account in communications with patients and their relatives, consent process, and multidisciplinary decision-making process for operative or non-operative management of such patients.