Risk factors for transfer from Respiratory Intermediate Care Unit to Intensive Care Unit in COVID-19

BackgroundPatients hospitalized for COVID-19-related pneumonia often need several degrees of ventilatory support, which are performed between Respiratory Intermediate Care Units (RICUs) and Intensive Care Units (ICUs), and which depend on the severity of acute respiratory distress syndrome. There is no firm consensus on transfer predictors from the RICU to the ICU.MethodsIn this retrospective observational single center study, we evaluated 96 COVID-19 patients referred to the RICU for acute respiratory failure (ARF) according to their transferal to the ICU or their stay at the RICU. We compared demographic data, baseline laboratory profile, and final clinical outcomes to identify early risk factors for transfer.ResultsThe best predictors for transfer to the ICU were elevated C-reactive protein and lymphopenia. The mortality rate was lower in the RICU than in the ICU, where transferred patients who died were mostly younger men and with less comorbidities than those in the RICU.ConclusionsFew inflammatory markers can predict the need for transfer from the RICU to the ICU. Due to the ongoing COVID-19 pandemic, we urge better clinical stratification by early and meaningful profiles in patients admitted to the RICU who are at risk of transferal to the ICU.     

急性反应蛋白在老龄大鼠大肠杆菌肺炎中的变化

目的　观察老龄大肠杆菌肺炎模型大鼠急性反应蛋白 (APP)的变化。方法　复制大肠杆菌肺炎模型 ,分为青年对照组和青年模型组 ,老龄对照组和老龄模型组。观察肺脏含水量、肺泡灌洗液中白细胞、中性粒细胞计数和纤维结合蛋白 (Fn)以及血清 C反应蛋白 (CRP)、铜蓝蛋白(CP)含量变化。结果　老龄大鼠肺炎的肺组织损伤较青年大鼠严重。青年模型组和老龄模型组肺脏含水量、肺泡灌洗液中性粒细胞和血清 CRP、CP含量均高于青年对照组和老龄对照组 (P<0 .0 1 ) ,肺泡灌洗液和血清中的 Fn,老龄模型组和青年模型组显著低于老龄对照组和青年对照组 (P<0 .0 1 )。结论　肺脏中性粒细胞和血清中 APP的变化参与肺炎的发生发展。     

Neurologic evaluation of dysphagia

Unexplained dysphagia is often caused by unrecognized neurologic disease. A previous article (Buchholz 1987) discussed the many neurologic diseases that may cause dysphagia. This article reviews a neurologist's approach to dysphagia of possible neurologic origin. As with most medical problems, a careful history provides more information about dysphagia than any other data-gathering technique. Therefore, history-taking is discussed first, followed by review of pertinent aspects of the general and neurologic examinations and ancillary testing.     

Visualizing monolayers with a water-soluble fluorophore to quantify adsorption,desorption, and the double layer

Contrast in confocal microscopy of phase-separated monolayers at the air–water interface can be generated by the selective adsorption of water-soluble fluorescent dyes to disordered monolayer phases. Optical sectioning minimizes the fluorescence signal from the subphase, whereas convolution of the measured point spread function with a simple box model of the interface provides quantitative assessment of the excess dye concentration associated with the monolayer. Coexisting liquid-expanded, liquid-condensed, and gas phases could be visualized due to differential dye adsorption in the liquid-expanded and gas phases. Dye preferentially adsorbed to the liquid-disordered phase during immiscible liquid–liquid phase coexistence, and the contrast persisted through the critical point as shown by characteristic circle-to-stripe shape transitions. The measured dye concentration in the disordered phase depended on the phase composition and surface pressure, and the dye was expelled from the film at the end of coexistence. The excess concentration of a cationic dye within the double layer adjacent to an anionic phospholipid monolayer was quantified as a function of subphase ionic strength, and the changes in measured excess agreed with those predicted by the mean-field Gouy–Chapman equations. This provided a rapid and noninvasive optical method of measuring the fractional dissociation of lipid headgroups and the monolayer surface potential.Lipid monolayers have long been used as a simplified model for cell membranes (1–4), especially for the study of 2D phase separation that may underlie the raft hypothesis of cell membrane organization (5–8). Monolayers are also of interest in the physics of ordering and dynamic processes in two dimensions such as the structure and flow of hexatic phases (9–11). Additionally, all air-breathing mammals have a lipid–protein monolayer lining the lung alveoli to minimize surface tension effects on breathing; deficiency or disruption of this monolayer leads to potentially fatal respiratory distress syndrome in infants and adults (12, 13). The general acceptance of complex monolayer and bilayer phase behavior, especially phase coexistence (14–16) and critical phenomena (7, 8, 17, 18), has relied on the visualization of fluorescently tagged lipids segregated between phases in the monolayer. This segregation, due to differences in local molecular organization, provides the necessary contrast to visualize even subtle differences in packing density, molecular tilt, and short- and long-range ordering. Since the technique was introduced (14, 19–21), visualization of the distribution of labeled lipids has settled numerous debates over the molecular organization of monolayers (2, 4, 22, 23) and bilayers (8, 24).However, fluorescently tagged lipids, like many of the other lipids in monolayers and bilayers, are effectively insoluble in the surrounding aqueous subphase and are trapped in the monolayer or bilayer. As a result, expelling the fluorescent lipid from one phase means concentrating the it in another phase, which can result in fluorescence quenching, alterations in the structures of both the less-ordered (dye-rich) and more-ordered (dye-poor) domains, or perturbations in the mechanical properties of the monolayer (25). Increased fluorescence detector sensitivities and improved quantum efficiencies of fluorophores (26) have gradually decreased the overall mole fraction of fluorescent lipid required for visualization from 2 mol% in early experiments (14, 19–21) to ∼0.1–0.5 mol% in recent work (27). Unknown concentration and quenching effects limit analysis to qualitative assessment of the fluorescence intensity distributions. The only alternative to fluorescently labeled lipids has been Brewster angle microscopy (BAM), in which the contrast in the image results from small variations in refractive index and/or interfacial thickness between different monolayer areas due to different phases, molecular density, or molecular tilt (3, 28–30).Here we demonstrate that the selective adsorption of a water-soluble fluorophore from the subphase to the monolayer provides similar contrast to that of insoluble lipid dyes in confocal microscope images of liquid-expanded (LE), liquid-condensed (LC), and gas phase coexistence. The soluble dye provides this contrast by differential adsorption, preferring the least-ordered phases in the monolayer. The water-soluble fluorophore also provides contrast between coexisting immiscible liquid phases; the liquid-ordered (l_o) and liquid-disordered (l_d) phases can be differentiated even in the vicinity of critical points (18, 24). The optical sectioning of the confocal microscope (31) allows the interfacial fluorescence of the monolayer-adsorbed dye to be visualized and rejects the fluorescence from the subphase, which is not possible with conventional wide-field microscopy. Isotherms, domain shapes, and size distributions are similar to those with fluorescently tagged lipids and to BAM images with no dye (29, 30). Unlike insoluble lipid dyes, the water-soluble dye concentration is set by equalizing the chemical potentials of the dye in the subphase with that in the monolayer to establish a dynamic equilibrium between the monolayer and the subphase. The chemical potential of the dye in the monolayer likely depends on the temperature, surface pressure, and the local ordering of the monolayer. As a result, the dye is not concentrated in the LE (or l_d) phase as the fraction of LC (or l_o) phase increases. Instead, the dye concentration in the monolayer adjusts to constantly equalize the chemical potentials. We find different absolute adsorptions depending on temperature, monolayer composition, and surface pressure, which may be helpful in determining the composition of the disordered phases.Moreover, the contrast in the images can be quantified to determine the local fluorophore concentrations. Deconvolution of the confocal microscopy images of monolayers in contact with a subphase of soluble dye allowed for the quantification of the adsorption and desorption kinetics of the fluorophore. The rate of dye adsorption can be described with classic Langmuir adsorption kinetics and saturates in a few minutes. In addition to determining the quantity of dye coadsorbed with the monolayer, optical sectioning can also determine the distribution of labeled species distributed between the subphase and the monolayer. This allowed us to determine the surface potential and fractional ionization of an anionic dipalmitoylphosphatidylglycerol (DPPG) monolayer. We compared the measured excess cationic dye within the electric double layer with the predictions of the Gouy–Chapman theory combined with a mass-action model for the dissociation equilibrium of DPPG (32, 33). There was excellent agreement between theory and experiment for an optimal headgroup pK_a of 2.4 over a range of subphase ionic strengths. Conventional monolayer surface potential measurements using the Kelvin vibrating capacitive probe give only the combined surface potentials of the monolayer–water and monolayer–air interfaces (33); it can be difficult to separate the two potentials quantitatively.We expect that the approach presented here can be extended to quantify the distribution of a variety of labeled proteins, polymers, and other materials of interest in the vicinity of monolayers at the air–water interface, using confocal microscopy (34–37). The method may also be amenable to determining local fluorophore concentrations near liquid–liquid and liquid–solid interfaces to further address issues of ion distributions and modifications to Gouy–Chapman theory (38).     

The genome of Clostridium kluyveri, a strict anaerobe with unique metabolic features

Clostridium kluyveri is unique among the clostridia; it grows anaerobically on ethanol and acetate as sole energy sources. Fermentation products are butyrate, caproate, and H₂. We report here the genome sequence of C. kluyveri, which revealed new insights into the metabolic capabilities of this well studied organism. A membrane-bound energy-converting NADH:ferredoxin oxidoreductase (RnfCDGEAB) and a cytoplasmic butyryl-CoA dehydrogenase complex (Bcd/EtfAB) coupling the reduction of crotonyl-CoA to butyryl-CoA with the reduction of ferredoxin represent a new energy-conserving module in anaerobes. The genes for NAD-dependent ethanol dehydrogenase and NAD(P)-dependent acetaldehyde dehydrogenase are located next to genes for microcompartment proteins, suggesting that the two enzymes, which are isolated together in a macromolecular complex, form a carboxysome-like structure. Unique for a strict anaerobe, C. kluyveri harbors three sets of genes predicted to encode for polyketide/nonribosomal peptide synthetase hybrides and one set for a nonribosomal peptide synthetase. The latter is predicted to catalyze the synthesis of a new siderophore, which is formed under iron-deficient growth conditions.     

Validation studies for germ-free Smad3-/- mice as a bio-assay to test the causative role of fecal microbiomes in IBD

ABSTRACT

While the association between microbiomes and inflammatory bowel disease (IBD) is well known, establishing causal relationships between the two is difficult in humans. Germ-free (GF) mice genetically susceptible to IBD can address this question. Smad3^-/- mice with defective transforming growth factor ß signaling are a model of IBD and colon cancer. They develop IBD upon colonization with Helicobacter under specific pathogen-free conditions, suggesting a role of the microbiome in IBD in this model. Thus, we rederived Smad3^-/- mice GF to determine the potential of using these mice for testing the causative role of microbiomes in IBD. We found that fecal microbiomes from mice with IBD cause more severe gut inflammation in GF Smad3^-/- and wild type mice compared to microbiomes from healthy mice and that Helicobacter induces gut inflammation within the context of other microbiomes but not by itself. Unexpectedly, GF Smad3^+/+ and Smad3^+/- mice given IBD microbiomes develop IBD despite their lack of disease in SPF conditions upon Helicobacter infection. This was not unique to the background strain of our Smad3 model (129); both wild type C57BL/6 and 129 strains developed IBD upon fecal transfer. However, wild type Swiss Webster stock was not susceptible, indicating that the genetic background of recipient mice influences the severity of IBD following fecal transfer. Our data suggest that the microbiome is an independent risk factor contributing to IBD development, and careful characterization of new GF models is needed to understand potential sources of confounding factors influencing microbiome studies in these mice.     

Phase I and pharmacological study of cytarabine and tanespimycin in relapsed and refractory acute leukemia

Background

In preclinical studies the heat shock protein 90 (Hsp90) inhibitor tanespimycin induced down-regulation of checkpoint kinase 1 (Chk1) and other client proteins as well as increased sensitivity of acute leukemia cells to cytarabine. We report here the results of a phase I and pharmacological study of the cytarabine + tanespimycin combination in adults with recurrent or refractory acute leukemia.

Design and Methods

Patients received cytarabine 400 mg/m²/day continuously for 5 days and tanespimycin infusions at escalating doses on days 3 and 6. Marrow mononuclear cells harvested before therapy, immediately prior to tanespimycin, and 24 hours later were examined by immunoblotting for Hsp70 and multiple Hsp90 clients.

Results

Twenty-six patients were treated at five dose levels. The maximum tolerated dose was cytarabine 400 mg/m²/day for 5 days along with tanespimycin 300 mg/m² on days 3 and 6. Treatment-related adverse events included disseminated intravascular coagulation (grades 3 and 5), acute respiratory distress syndrome (grade 4), and myocardial infarction associated with prolonged exposure to tanespimycin and its active metabolite 17-aminogeldanamycin. Among 21 evaluable patients, there were two complete and four partial remissions. Elevations of Hsp70, a marker used to assess Hsp90 inhibition in other studies, were observed in more than 80% of samples harvested 24 hours after tanespimycin, but down-regulation of Chk1 and other Hsp90 client proteins was modest.

Conclusions

Because exposure to potentially effective concentrations occurs only for a brief time in vivo, at clinically tolerable doses tanespimycin has little effect on resistance-mediating client proteins in relapsed leukemia and exhibits limited activity in combination with cytarabine.     

Variable genetic architectures produce virtually identical molecules in bacterial symbionts of fungus-growing ants

Small molecules produced by Actinobacteria have played a prominent role in both drug discovery and organic chemistry. As part of a larger study of the actinobacterial symbionts of fungus-growing ants, we discovered a small family of three previously unreported piperazic acid-containing cyclic depsipeptides, gerumycins A–C. The gerumycins are slightly smaller versions of dentigerumycin, a cyclic depsipeptide that selectively inhibits a common fungal pathogen, Escovopsis. We had previously identified this molecule from a Pseudonocardia associated with Apterostigma dentigerum, and now we report the molecule from an associate of the more highly derived ant Trachymyrmex cornetzi. The three previously unidentified compounds, gerumycins A–C, have essentially identical structures and were produced by two different symbiotic Pseudonocardia spp. from ants in the genus Apterostigma found in both Panama and Costa Rica. To understand the similarities and differences in the biosynthetic pathways that produced these closely related molecules, the genomes of the three producing Pseudonocardia were sequenced and the biosynthetic gene clusters identified. This analysis revealed that dramatically different biosynthetic architectures, including genomic islands, a plasmid, and the use of spatially separated genetic loci, can lead to molecules with virtually identical core structures. A plausible evolutionary model that unifies these disparate architectures is presented.Fungus-growing ants of the tribe Attini originated ∼50 Mya in the Amazon and have since evolved into more than 200 species that collectively are the major herbivores of the New World Tropics (1, 2). A typical agricultural system contains at least four interacting organisms: the ants, their fungal crop (phylum Basidiomycota), a symbiotic bacterium (Actinobacteria), and a specialized fungal pathogen (phylum Ascomycota). The ants maintain their fungal gardens by providing plant material to the fungal crop, whereas the bacterial symbionts, which often localize with specialized anatomical structures on the ant, provide small-molecule chemical defenses that suppress microfungal pathogens in the genus Escovopsis and other potential competitors (3, 4). In an earlier study on Apterostigma dentigerum, we characterized dentigerumycin (1) as the major antifungal agent produced by the symbiotic bacterium that selectively inhibited the Escovopsis pathogen (4). Our subsequent studies on the molecular diversity to be found in this well-defined ecological niche featured both a metabolomics approach to structurally characterize the molecules produced and a genomic approach to characterize the biosynthetic genes that produced them.To initiate a systematic study on small molecules and their biosynthetic gene clusters (BGCs) found in the symbiotic bacteria, we returned to isolates from both Apterostigma spp. and the more highly derived Trachymyrmex cornetzi with a focus on dentigerumycin-like molecules. This study reports three previously unidentified cyclic depsipeptides named gerumycins A–C, which are structurally related to the originally discovered selective antifungal molecule dentigerumycin, the first three finished ant-associated Pseudonocardia genomes, and the three BGCs encoding the cyclic depsipeptides. The BGCs are encoded within variable genetic architectures in the three producing bacteria and are most plausibly accommodated by a model in which each has been recently acquired by the respective symbiont. The results, which describe both plasmid and chromosomal genomic islands, illustrate a useful strategy for symbionts of the fungus-growing ants to acquire and evolve new molecules.