Evolutionary origin, population diversity, and diagnostics for a cryptic hybrid pathogen.

Cryptic fungal pathogens pose significant identification and disease management challenges due to their morphological resemblance to known pathogenic species while harboring genetic and (often) infectionrelevant trait differences. The cryptic fungal pathogen Aspergillus latus, an allodiploid hybrid originating from Aspergillus spinulosporus and an unknown close relative of Aspergillus quadrilineatus within section Nidulantes, remains poorly understood. The absence of accurate diagnostics for A. latus has led to misidentifications, hindering epidemiological studies and the design of effective treatment plans. We conducted an in-depth investigation of the genomes and phenotypes of 44 globally distributed isolates (41 clinical isolates and three type strains) from Aspergillus section Nidulantes. We found that 21 clinical isolates were A. latus; notably, standard methods of pathogen identification misidentified all A. latus isolates. The remaining isolates were identified as A. spinulosporus (8), A. quadrilineatus (1), or A. nidulans (11). Phylogenomic analyses shed light on the origin of A. latus, indicating one or two hybridization events gave rise to the species during the Miocene, approximately 15.4 to 8.8 million years ago. Characterizing the A. latus pangenome uncovered substantial genetic diversity within gene families and biosynthetic gene clusters. Transcriptomic analysis revealed that both parental genomes are actively expressed in nearly equal proportions and respond to environmental stimuli. Further investigation into infection-relevant chemical and physiological traits, including drug resistance profiles, growth under oxidative stress conditions, and secondary metabolite biosynthesis, highlight distinct phenotypic profiles of the hybrid A. latus compared to its parental and closely related species. Leveraging our comprehensive genomic and phenotypic analyses, we propose five genomic and phenotypic markers as diagnostics for A. latus species identification. These findings provide valuable insights into the evolutionary origin, genomic outcome, and phenotypic implications of hybridization in a cryptic fungal pathogen, thus enhancing our understanding of the underlying processes contributing to fungal pathogenesis. Furthermore, our study underscores the effectiveness of extensive genomic and phenotypic analyses as a promising approach for developing diagnostics applicable to future investigations of cryptic and emerging pathogens.

Fungal-fungal co-culture: a primer for generating chemical diversity.

Covering: 2002 to 2020In their natural environment, fungi must compete for resources. It has been hypothesized that this competition likely induces the biosynthesis of secondary metabolites for defence. In a quest to discover new chemical diversity from fungal cultures, a growing trend has been to recapitulate this competitive environment in the laboratory, essentially growing fungi in co-culture. This review covers fungal-fungal co-culture studies beginning with the first literature report in 2002. Since then, there has been a growing number of new secondary metabolites reported as a result of fungal co-culture studies. Specifically, this review discusses and provides insights into (1) rationale for pairing fungal strains, (2) ways to grow fungi for co-culture, (3) different approaches to screening fungal co-cultures for chemical diversity, (4) determining the secondary metabolite-producing strain, and (5) final thoughts regarding the fungal-fungal co-culture approach. Our goal is to provide a set of practical strategies for fungal co-culture studies to generate unique chemical diversity that the natural products research community can utilize.

Opportunities and Limitations for Assigning Relative Configurations of Antibacterial Bislactones using GIAO NMR Shift Calculations.

Four new bislactones, dihydroacremonol (1), clonostachyone (2), acremodiol B (3), and acremodiol C (4), along with one known compound, hymeglusin (5), were isolated from cultures of two fungal strains (MSX59876 and MSX59260). Both strains were identified based on phylogenetic analysis of molecular data as Clonostachys spp.; yet, they biosynthesized a suite of related, but different, secondary metabolites. Given the challenges associated with elucidating the structures and configurations of bislactones, GIAO NMR calculations were tested as a complement to traditional NMR and HRESIMS experiments. Fortuitously, the enantiomer of the new natural product (4) was known as a synthetic compound, and the predicted configuration from GIAO NMR calculations (i.e., for the relative configuration) and optical rotation calculations (i.e., for the absolute configuration) matched those of the synthesis product. These results engendered confidence in using similar procedures, particularly the mixture of GIAO NMR shift calculations coupled with an orthogonal technique, to predict the configuration of 1-3; however, there were important limitations, which are discussed for each of these. The metabolites displayed antimicrobial activities, with compounds 1 and 4 being the most potent against Staphylococcus aureus with MICs of 1 and 4 µg/mL, respectively.

Projected Future Changes in Tropical Cyclones Using the CMIP6 HighResMIP Multimodel Ensemble.

Future changes in tropical cyclone properties are an important component of climate change impacts and risk for many tropical and midlatitude countries. In this study we assess the performance of a multimodel ensemble of climate models, at resolutions ranging from 250 to 25 km. We use a common experimental design including both atmosphere-only and coupled simulations run over the period 1950-2050, with two tracking algorithms applied uniformly across the models. There are overall improvements in tropical cyclone frequency, spatial distribution, and intensity in models at 25 km resolution, with several of them able to represent very intense storms. Projected tropical cyclone activity by 2050 generally declines in the South Indian Ocean, while changes in other ocean basins are more uncertain and sensitive to both tracking algorithm and imposed forcings. Coupled models with smaller biases suggest a slight increase in average TC 10 m wind speeds by 2050.

Variation Among Biosynthetic Gene Clusters, Secondary Metabolite Profiles, and Cards of Virulence Across Aspergillus Species.

Aspergillus fumigatus is a major human pathogen. In contrast, Aspergillus fischeri and the recently described Aspergillus oerlinghausenensis, the two species most closely related to A. fumigatus, are not known to be pathogenic. Some of the genetic determinants of virulence (or "cards of virulence") that A. fumigatus possesses are secondary metabolites that impair the host immune system, protect from host immune cell attacks, or acquire key nutrients. To examine whether secondary metabolism-associated cards of virulence vary between these species, we conducted extensive genomic and secondary metabolite profiling analyses of multiple A. fumigatus, one A. oerlinghausenensis, and multiple A. fischeri strains. We identified two cards of virulence (gliotoxin and fumitremorgin) shared by all three species and three cards of virulence (trypacidin, pseurotin, and fumagillin) that are variable. For example, we found that all species and strains examined biosynthesized gliotoxin, which is known to contribute to virulence, consistent with the conservation of the gliotoxin biosynthetic gene cluster (BGC) across genomes. For other secondary metabolites, such as fumitremorgin, a modulator of host biology, we found that all species produced the metabolite but that there was strain heterogeneity in its production within species. Finally, species differed in their biosynthesis of fumagillin and pseurotin, both contributors to host tissue damage during invasive aspergillosis. A. fumigatus biosynthesized fumagillin and pseurotin, while A. oerlinghausenensis biosynthesized fumagillin and A. fischeri biosynthesized neither. These biochemical differences were reflected in sequence divergence of the intertwined fumagillin/pseurotin BGCs across genomes. These results delineate the similarities and differences in secondary metabolism-associated cards of virulence between a major fungal pathogen and its nonpathogenic closest relatives, shedding light onto the genetic and phenotypic changes associated with the evolution of fungal pathogenicity.

Innovative pain management solutions in animals may provide improved wound pain reduction during debridement in humans: An opinion informed by veterinary literature.

Painful animal husbandry procedures are routinely performed in a range of livestock species without analgesia. Recently, innovative strategies have been developed to address wound pain in these animals. In particular, a farmer-applied "spray and stay" approach that is administered directly to open wounds was developed (Tri-Solfen® Medical Ethics Pty Ltd., Melbourne, Victoria, Australia). This strategy anaesthetises the wounds immediately upon their formation, with long-lasting effect. This development, described as a "pain management revolution," has become firmly established in the Australian livestock industries and has global potential. The positive outcomes of this approach provide insights and highlight potential benefits that may be accrued from its use in human wound care, providing rapid-onset wound analgesia and/or anaesthetising wounds prior to cleansing and debridement procedures. If these benefits are realised from a clinician and patient perspective for wound debridement as an initial indication, it could provide new horizons in pain management for a spectrum of wound-related procedures. Evidence from use in animal husbandry does support the concept that multimodal anaesthesia holds great potential in the field of wound management across many procedures.

Appropriate use of dressings containing nanocrystalline silver to support antimicrobial stewardship in wounds.

Antimicrobial resistance is an ever-increasing global concern, with the era of untreatable infection becoming a reality. Wound care is no exception, with increasing issues of antibiotic-resistant infections across different wound types and care settings. Antibiotic resistance and stewardship have been the priority for most strategic interventions so far; however, in wound care, alternative or supplementary strategies using antiseptics should be considered. Antiseptics such as silver can provide effective cidal activity across a broad range of wound pathogens, assuming they are used at the correct level for an appropriate duration. Evidence summarised in this manuscript suggests that effective antiseptics, such as nanocrystalline silver, have an increasing body of evidence in support of their use to minimise transmission of antibiotic-resistant organisms as part of institutional infection control procedures and, in addition, through appropriate early use and stewardship on local wound infections, in conjunction with local procedures, to minimise the need for systemic antibiotic therapy. Engagement, alignment, and collaboration between wound care professionals and wider related teams and governments on antimicrobial stewardship, and the potential role of antiseptics within this, will help to generate further evidence for such interventions in the fight against antimicrobial-resistant infections in wound care.

A 2000 patient retrospective assessment of a new strategy for burn wound management in view of infection prevention and treatment.

Infections in burn patients are still the principal cause of complications in burn injuries. The aim of this study is to assess a new strategy for burn wound management in view of infection prevention and treatment in the experience of the Burn Treatment Center in Siemianowice Slaskie. The applied methodology involved the analysis of patient records describing the hospital's epidemiological situation between 2014 and 2016. The analysis also included the use and cost of antibiotics, silver-containing dressings, and other antiseptics relative to the number of sepsis cases, including those caused by Pseudomonas aeruginosa, as well as the mortality ratio. The total costs of prevention and treatment of infections were reduced, while the use of silver-containing dressings and antiseptics increased. The number of patients with sepsis decreased, including cases caused by P. aeruginosa, and the mortality ratio was reduced. Introducing a strategy for burn wound-oriented infection prevention and treatment in burn patients provides a number of benefits. It is also cost-effective. Using locally applied active dressings and antiseptics can be a welcome choice for often-unnecessary antibiotic therapy of a suspected or existing burn wound infection.

The Role of Topical Antiseptic Agents Within Antimicrobial Stewardship Strategies for Prevention and Treatment of Surgical Site and Chronic Open Wound Infection.

Scope and Significance: The topical use of antiseptics for wound care has a role in an antimicrobial stewardship strategy. However, the details of this role need clarification. Further clinical research into the use of topical antiseptics in wound care would lower the risk of furthering antibiotic resistance and contribute to more effective antibiotic use. As part of this research, experimental and surveillance data are needed on the resistance and tolerance patterns associated with topical antiseptic use in wound infections. Objective: The development of antibiotic resistance presents global challenges in terms of patient harm and increased healthcare costs. The treatment of "at risk" and infected wounds contributes to this conundrum. Synergies between antibiotics and antiseptics and their appropriate combined use need exploration. Approach: A review of available evidence on the appropriateness of antiseptics as a fundamental component of antimicrobial stewardship strategies has been undertaken. Innovation: Opening up new ways of thinking and identifying gaps of knowledge will lead to optimizing justification of antimicrobial choices and combinations. This may lead to changes in practice in terms of solutions for the prevention and treatment of wound infection. Conclusion: Antiseptics are an integral part of antimicrobial stewardship strategies for the prevention and treatment of surgical site and chronic open wound infections.

A systematic review and meta-analysis of clinical outcomes associated with nanocrystalline silver use compared to alternative silver delivery systems in the management of superficial and deep partial thickness burns.

OBJECTIVE: The purpose of this systematic review and meta-analysis was to assess the clinical effectiveness of nanocrystalline silver compared to alternative silver delivery systems (silver sulphadiazine [SSD] and silver nitrate) in adults and children with superficial and deep partial thickness burns. METHODS: PubMed, EMBASE, Cochrane and other databases were searched to identify relevant randomised controlled trials and observational studies. RESULTS: Eight studies that assessed both nanocrystalline silver and SSD and one study that compared nanocrystalline silver vs. silver nitrate were identified. Nanocrystalline silver compared to SSD/silver nitrate was associated with a statistically significant reduction in infections (odds ratio [OR] 0.21, 95% CI 0.07-0.62, p=0.005), length of stay in hospital (mean difference -4.74 (95% CI -5.79 to -3.69, p=0.00001) and surgical procedures (OR 0.40, 95% CI 0.28-0.56, p=0.00001). Three studies that reported on pain had lower pain scores with nanocrystalline silver use than with SSD/silver nitrate; a high level of heterogeneity precluded pooling estimates. CONCLUSION: This comprehensive systematic review and meta-analysis of the available evidence suggest that the use of nanocrystalline silver dressings results in shorter length of stay in hospital, less pain, fewer surgical procedures and reduced infection rates compared to silver sulphadiazine/silver nitrate.

Quantifying, Visualizing, and Monitoring Lead Optimization.

Although lead optimization (LO) is by definition a process, process-centric analysis and visualization of this important phase of pharmaceutical R&D has been lacking. Here we describe a simple statistical framework to quantify and visualize the progression of LO projects so that the vital signs of LO convergence can be monitored. We refer to the resulting visualizations generated by our methodology as the "LO telemetry" of a project. These visualizations can be automated to provide objective, holistic, and instantaneous analysis and communication of LO progression. This enhances the ability of project teams to more effectively drive LO process, while enabling management to better coordinate and prioritize LO projects. We present the telemetry of five LO projects comprising different biological targets and different project outcomes, including clinical compound selection, termination due to preclinical safety/tox, and termination due to lack of tractability. We demonstrate that LO progression is accurately captured by the telemetry. We also present metrics to quantify LO efficiency and tractability.

Novel spiroketal pyrrolidine GSK2336805 potently inhibits key hepatitis C virus genotype 1b mutants: from lead to clinical compound.

Rapid clinical progress of hepatitis C virus (HCV) replication inhibitors, including these selecting for resistance in the NS5A region (NS5A inhibitors), promises to revolutionize HCV treatment. Herein, we describe our explorations of diverse spiropyrrolidine motifs in novel NS5A inhibitors and a proposed interaction model. We discovered that the 1,4-dioxa-7-azaspiro[4.4]nonane motif in inhibitor 41H (GSK2236805) supported high potency against genotypes 1a and 1b as well as in genotype 1b L31V and Y93H mutants. Consistent with this, 41H potently suppressed HCV RNA in the 20-day RNA reduction assay. Pharmacokinetic and safety data supported further progression of 41H to the clinic.

Discovery of a potent boronic acid derived inhibitor of the HCV RNA-dependent RNA polymerase.

A boronic acid moiety was found to be a critical pharmacophore for enhanced in vitro potency against wild-type hepatitis C replicons and known clinical polymorphic and resistant HCV mutant replicons. The synthesis, optimization, and structure-activity relationships associated with inhibition of HCV replication in a subgenomic replication system for a series of non-nucleoside boron-containing HCV RNA-dependent RNA polymerase (NS5B) inhibitors are described. A summary of the discovery of 3 (GSK5852), a molecule which entered clinical trials in subjects infected with HCV in 2011, is included.

Hepatitis C replication inhibitors that target the viral NS4B protein.

We describe the preclinical development and in vivo efficacy of a novel chemical series that inhibits hepatitis C virus replication via direct interaction with the viral nonstructural protein 4B (NS4B). Significant potency improvements were realized through isosteric modifications to our initial lead 1a. The temptation to improve antiviral activity while compromising physicochemical properties was tempered by the judicial use of ligand efficiency indices during lead optimization. In this manner, compound 1a was transformed into (+)-28a which possessed an improved antiviral profile with no increase in molecular weight and only a modest elevation in lipophilicity. Additionally, we employed a chimeric "humanized" mouse model of HCV infection to demonstrate for the first time that a small molecule with high in vitro affinity for NS4B can inhibit viral replication in vivo. This successful proof-of-concept study suggests that drugs targeting NS4B may represent a viable treatment option for curing HCV infection.

Atlantic Meridional Overturning Circulation slowdown cooled the subtropical ocean.

[1] Observations show that the upper 2 km of the subtropical North Atlantic Ocean cooled throughout 2010 and remained cold until at least December 2011. We show that these cold anomalies are partly driven by anomalous air-sea exchange during the cold winters of 2009/2010 and 2010/2011 and, more surprisingly, by extreme interannual variability in the ocean's northward heat transport at 26.5°N. This cooling driven by the ocean's meridional heat transport affects deeper layers isolated from the atmosphere on annual timescales and water that is entrained into the winter mixed layer thus lowering winter sea surface temperatures. Here we connect, for the first time, variability in the northward heat transport carried by the Atlantic Meridional Overturning Circulation to widespread sustained cooling of the subtropical North Atlantic, challenging the prevailing view that the ocean plays a passive role in the coupled ocean-atmosphere system on monthly-to-seasonal timescales.

Coupling of CO2 and ice sheet stability over major climate transitions of the last 20 million years.

The carbon dioxide (CO2) content of the atmosphere has varied cyclically between approximately 180 and approximately 280 parts per million by volume over the past 800,000 years, closely coupled with temperature and sea level. For earlier periods in Earth's history, the partial pressure of CO2 (pCO2) is much less certain, and the relation between pCO2 and climate remains poorly constrained. We use boron/calcium ratios in foraminifera to estimate pCO2 during major climate transitions of the past 20 million years. During the Middle Miocene, when temperatures were approximately 3 degrees to 6 degrees C warmer and sea level was 25 to 40 meters higher than at present, pCO2 appears to have been similar to modern levels. Decreases in pCO(2) were apparently synchronous with major episodes of glacial expansion during the Middle Miocene (approximately 14 to 10 million years ago) and Late Pliocene (approximately 3.3 to 2.4 million years ago).

Minor groove DNA binders as antimicrobial agents. 1. Pyrrole tetraamides are potent antibacterials against vancomycin resistant Enterococci [corrected] and methicillin resistant Staphylococcus aureus.

A new series of short pyrrole tetraamides are described whose submicromolar DNA binding affinity is an essential component for their strong antibacterial activity. This class of compounds is related to the linked bis-netropsins and bis-distamycins, but here, only one amino-pyrrole-carboxamide unit and an amidine tail is connected to either side of a central dicarboxylic acid linker. The highest degree of DNA binding, measured by compound-induced changes in UV melting temperatures of an AT-rich DNA oligomer, was observed for flat, aromatic linkers with no inherent bent, i.e., terephthalic acid or 1,4-pyridine-dicarboxylic acid. However, the antibacterial activity is critically linked to the size of the N-alkyl substiutent of the pyrrole unit. None of the tetraamides with the commonly used methyl-pyrrole showed antibacterial activity. Isoamyl- or cyclopropylmethylene-substituted dipyrrole derivatives have the minimum inhibitory concentrations in the submicromolar range. In vitro toxicity against human T-cells was studied for all compounds. The degree to which compounds inhibited cell growth was neither directly correlated to DNA binding affinity nor directly correlated to antibacterial activity but seemed to depend strongly on the nature of the N-alkyl pyrrole substituents.