A Standard-Cell-Based Neuro-Inspired Integrate-and-Fire Analog-to-time converter for Biological and Low-Frequency Signals - Comparison with Analog Version.

Continuous-time asynchronous data converters namely, analog-to-digital converters and analog-to-time converters, can be beneficial for certain types of applications, such as, processing of biological signals with sparse information. A particular case of these converters is the integrate-and-fire converter (IFC) that is inspired by the neural system. If it is possible to develop a standard-cell-based (SCB) IFC circuit to perform well in advanced technology nodes, it will benefit from the simplicity of SCB circuit designs and can be implemented in widely available field-programmable gate arrays (FPGAs). This way, this paper proposes two IFC circuits designed and prototyped in a 130 nm CMOS standard process. The first is a novel SCB open-loop dynamic IFC. The latter, is a closed-loop analog IFC with conventional blocks. This paper presents a through comparison between the two IFC circuits. They have a power dissipation of 59 µW and 53 µW, and an energy per pulse of 18 pJ and 1060 pJ, SCB and analog IFC, respectively. The SCB IFC has one of the lowest energy per pulse consumption reported for IFC circuits. The analog IFC, being fully differential, is to our knowledge the first of its kind. Moreover, they do not require an external clock. They can convert signals with a peak-to-peak amplitude from 1.6 mV to 28 mV and 0.6 mV to 2.4 mV, and a frequency range of 2 Hz to 42 kHz and 10 Hz to 4 kHz, SCB and analog IFC, respectively. Presenting low normalized RMS conversion plus reconstruction errors, below 5.2 %. The maximum pulse density (average firing-rate) is 3300 kHz, for the SCB and 50 kHz, for the analog IFC.

In the flow of molecular miniaturized fungal diagnosis.

The diagnosis of fungal infections presents several challenges and limitations, stemming from the similarities in symptomatology, diversity of underlying pathogenic species, complexity of fungal biology, and scarcity of rapid, affordable, and point-of-care approaches. In this review, we assess technological advances enabling the conversion of cutting-edge laboratory molecular diagnostic methods to cost-effective microfluidic devices. The most promising strategies toward the design of DNA sequence-based fungal diagnostic systems, capable of capturing and deciphering the highly informative DNA of the pathogen and adapted for resource-limited settings, are discussed, bridging fungal biology, molecular genetics, microfluidics, and biosensors.

Mutation in the 26S proteasome regulatory subunit rpn2 gene in Plasmodium falciparum confers resistance to artemisinin.

Introduction: Malaria parasites increasingly develop resistance to all drugs available in the market, hampering the goal of reducing malaria burden. Methods: Herein, we evaluated the impact of a single-nucleotide variant, E738K, present in the 26S proteasome regulatory subunit rpn2 gene, identified in Plasmodium chabaudi resistant parasites. Plasmids carrying a functional rpn2 interspecies chimeric gene with 5' recombination region from P. falciparum and 3' from P. chabaudi were constructed and transfected into Dd2 P. falciparum parasites. Results and discussion: The 738K variant parasite line presented increased parasite survival when subjected to dihydroartemisinin (DHA), as well as increased chymotrypsin-like activity and decreased accumulation of polyubiquitinated proteins. We thus conclude that the ubiquitin-proteasome pathway, including the 738K variant, play an important role in parasite response to DHA, being the first report of a mutation in a potential DHA drug target enhancing parasite survival and contributing to a significant advance in the understanding the biology of artemisinin resistance.

The Year in Cardiothoracic and Vascular Anesthesia: Selected Highlights From 2023.

This special article is the 16th in an annual series for the Journal of Cardiothoracic and Vascular Anesthesia. The authors thank the editor-in-chief, Dr. Kaplan, and the editorial board for the opportunity to continue this series, namely the research highlights of the past year in the specialty of cardiothoracic and vascular anesthesiology. The major themes selected for 2023 are outlined in this introduction, and each highlight is reviewed in detail in the main article. The literature highlights in the specialty for 2023 begin with an update on perioperative rehabilitation in cardiothoracic surgery, with a focus on novel methods to best assess patients in the preoperative and postoperative periods, and the impact of rehabilitation on outcomes. The second major theme is focused on cardiac surgery, with the authors discussing new insights into inhaled pulmonary vasodilators, coronary revascularization surgery, and discussion of causes of coronary graft failure after surgery. The third theme is focused on cardiothoracic transplantation, with discussions focusing on bridge-to-transplantation strategies. The fourth theme is focused on mechanical circulatory support, with discussions focusing on both temporary and durable support. The fifth and final theme is an update on medical cardiology, with a focus on outcomes of invasive approaches to heart disease. The themes selected for this article are only a few of the diverse advances in the specialty during 2023. These highlights will inform the reader of key updates on various topics, leading to improved perioperative outcomes for patients with cardiothoracic and vascular disease.

The flavodiiron protein from Syntrophomonas wolfei has five domains and acts both as an NADH:O2 or an NADH:H2 O2 oxidoreductase.

Flavodiiron proteins (FDPs) are a family of enzymes with a significant role in O2 /H2 O2 and/or NO detoxification through the reduction of these species to H2 O or N2 O, respectively. All FDPs contain a minimal catalytic unit of two identical subunits, each one having a metallo-ß-lactamase-like domain harboring the catalytic diiron site, and a flavodoxin-like domain. However, more complex and diverse arrangements in terms of domains are found in this family, of which the class H enzymes are among the most complex. One of such FDPs is encoded in the genome of the anaerobic bacterium Syntrophomonas wolfei subsp. wolfei str. Goettingen G311. Besides the core domains, this protein is predicted to have three additional ones after the flavodoxin core domain: two short-chain rubredoxins and a NAD(P)H:rubredoxin oxidoreductase-like domain. This enzyme, FDP_H, was produced and characterized and the presence of the predicted cofactors was investigated by a set of biochemical and spectroscopic methodologies. Syntrophomonas wolfei FDP_H exhibited a remarkable O2 reduction activity with a kcat = 52.0 ± 1.2 s-1 and a negligible NO reduction activity (~ 100 times lower than with O2 ), with NADH as an electron donor, that is, it is an oxygen-selective FDP. In addition, this enzyme showed the highest turnover value for H2 O2 reduction (kcat = 19.1 ± 2.2 s-1 ) ever observed among FDPs. Kinetic studies of site-directed mutants of iron-binding cysteines at the two rubredoxin domains demonstrated the essential role of these centers since their absence leads to a significant decrease or even abolishment of O2 and H2 O2 reduction activities.

Update in lung transplantation: anesthetic considerations.

The field of lung transplantation (LTx) has expanded rapidly since its inception in the early 1960s with the work of James Hardy and colleagues at the University of Mississippi from the work of local single specialty physicians into an international multidisciplinary specialty. Advancements throughout the next several decades have led to the completion of over 70,000 lung transplants worldwide. The unique challenges presented by patients with end-stage lung disease have both evolved and remained consistent since then, yet these challenges are being answered with major improvements and advancements in perioperative care in the 21st century. The current practice of LTx medicine is fundamentally multidisciplinary, and members of the LTx team includes surgeons, physicians, and allied health staff. The integration of anesthesiologists into the LTx team as well as the multidisciplinary nature of LTx necessitates anesthetic considerations to be closely incorporated into emerging surgical, medical, and systems techniques for patient care. This review discusses a host of emerging strategies across the spectrum of LTx, including efforts to expand the donor pool, utilization of perioperative extracorporeal life support, perioperative echocardiography, and anesthetic techniques to mitigate primary graft dysfunction that have all contributed to improved long term outcomes in LTx patients.

Metabolic reconstruction of the human pathogen Candida auris: using a cross-species approach for drug target prediction.

Candida auris is an emerging human pathogen, associated with antifungal drug resistance and hospital candidiasis outbreaks. In this work, we present iRV973, the first reconstructed Genome-scale metabolic model (GSMM) for C. auris. The model was manually curated and experimentally validated, being able to accurately predict the specific growth rate of C. auris and the utilization of several sole carbon and nitrogen sources. The model was compared to GSMMs available for other pathogenic Candida species and exploited as a platform for cross-species comparison, aiming the analysis of their metabolic features and the identification of potential new antifungal targets common to the most prevalent pathogenic Candida species. From a metabolic point of view, we were able to identify unique enzymes in C. auris in comparison with other Candida species, which may represent unique metabolic features. Additionally, 50 enzymes were identified as potential drug targets, given their essentiality in conditions mimicking human serum, common to all four different Candida models analysed. These enzymes represent interesting drug targets for antifungal therapy, including some known targets of antifungal agents used in clinical practice, but also new potential drug targets without any human homolog or drug association in Candida species.

Advances in the treatment and prevention of HIV: what you need to know.

The global epidemic of HIV/AIDs has seen many advances in the development of effective treatments, including antiretroviral therapy that provides increasing sustained viral suppression, robust immune reconstitution and fewer side effects than before. Early HIV treatment regimens were notoriously complex, comprising up to 22 pills that needed to be taken at different times of the day. However, the advent of a single fixed dose combination drug formation simplified the treatment regimen so this could be taken once daily. Novel drugs are constantly being developed to provide better tolerated medications with robust, sustained viral suppression and immune reconstitution; these include long-acting injectables and implants, and preventative treatments for pre-exposure prophylaxis. This article provides an overview of emerging therapeutics for the treatment and prevention of HIV infection.

In Candida glabrata, ERMES Component GEM1 Controls Mitochondrial Morphology, mtROS, and Drug Efflux Pump Expression, Resulting in Azole Susceptibility.

Mitochondrial dysfunction or morphological abnormalities in human pathogenic fungi are known to contribute to azole resistance; however, the underlying molecular mechanisms are unknown. In this study, we investigated the link between mitochondrial morphology and azole resistance in Candida glabrata, which is the second most common cause of human candidiasis worldwide. The ER-mitochondrial encounter structure (ERMES) complex is thought to play an important role in the mitochondrial dynamics necessary for mitochondria to maintain their function. Of the five components of the ERMES complex, deletion of GEM1 increased azole resistance. Gem1 is a GTPase that regulates the ERMES complex activity. Point mutations in GEM1 GTPase domains were sufficient to confer azole resistance. The cells lacking GEM1 displayed abnormalities in mitochondrial morphology, increased mtROS levels, and increased expression of azole drug efflux pumps encoded by CDR1 and CDR2. Interestingly, treatment with N-acetylcysteine (NAC), an antioxidant, reduced ROS production and the expression of CDR1 in Δgem1 cells. Altogether, the absence of Gem1 activity caused an increase in mitochondrial ROS concentration, leading to Pdr1-dependent upregulation of the drug efflux pump Cdr1, resulting in azole resistance.

The Year in Cardiothoracic and Vascular Anesthesia: Selected Highlights from 2022.

This special article is the 15th in an annual series for the Journal of Cardiothoracic and Vascular Anesthesia. The authors thank the editor-in-chief Dr. Kaplan and the editorial board for the opportunity to continue this series, namely the research highlights of the past year in the specialties of cardiothoracic and vascular anesthesiology. The major themes selected for 2022 are outlined in this introduction, and each highlight is reviewed in detail in the main body of the article. The literature highlights, in the specialties for 2022, begin with an update on COVID-19 therapies, with a focus on the temporal updates in a wide range of therapies, progressing from medical to the use of extracorporeal membrane oxygenation and, ultimately, with lung transplantation in this high-risk group. The second major theme is focused on medical cardiology, with the authors discussing new insights into the life cycle of coronary disease, heart failure treatments, and outcomes related to novel statin therapy. The third theme is focused on mechanical circulatory support, with discussions focusing on both right-sided and left-sided temporary support outcomes and the optimal timing of deployment. The fourth and final theme is an update on cardiac surgery, with a discussion of the diverse aspects of concomitant valvular surgery and the optimal approach to procedural treatment for coronary artery disease. The themes selected for this 15th special article are only a few of the diverse advances in the specialties during 2022. These highlights will inform the reader of key updates on a variety of topics, leading to the improvement of perioperative outcomes for patients with cardiothoracic and vascular disease.

YEASTRACT+: a portal for the exploitation of global transcription regulation and metabolic model data in yeast biotechnology and pathogenesis.

YEASTRACT+ (http://yeastract-plus.org/) is a tool for the analysis, prediction and modelling of transcription regulatory data at the gene and genomic levels in yeasts. It incorporates three integrated databases: YEASTRACT (http://yeastract-plus.org/yeastract/), PathoYeastract (http://yeastract-plus.org/pathoyeastract/) and NCYeastract (http://yeastract-plus.org/ncyeastract/), focused on Saccharomyces cerevisiae, pathogenic yeasts of the Candida genus, and non-conventional yeasts of biotechnological relevance. In this release, YEASTRACT+ offers upgraded information on transcription regulation for the ten previously incorporated yeast species, while extending the database to another pathogenic yeast, Candida auris. Since the last release of YEASTRACT+ (January 2020), a fourth database has been integrated. CommunityYeastract (http://yeastract-plus.org/community/) offers a platform for the creation, use, and future update of YEASTRACT-like databases for any yeast of the users' choice. CommunityYeastract currently provides information for two Saccharomyces boulardii strains, Rhodotorula toruloides NP11 oleaginous yeast, and Schizosaccharomyces pombe 972h-. In addition, YEASTRACT+ portal currently gathers 304 547 documented regulatory associations between transcription factors (TF) and target genes and 480 DNA binding sites, considering 2771 TFs from 11 yeast species. A new set of tools, currently implemented for S. cerevisiae and C. albicans, is further offered, combining regulatory information with genome-scale metabolic models to provide predictions on the most promising transcription factors to be exploited in cell factory optimisation or to be used as novel drug targets. The expansion of these new tools to the remaining YEASTRACT+ species is ongoing.

Multiple genome analysis of Candida glabrata clinical isolates renders new insights into genetic diversity and drug resistance determinants.

The emergence of drug resistance significantly hampers the treatment of human infections, including those caused by fungal pathogens such as Candida species. Candida glabrata ranks as the second most common cause of candidiasis worldwide, supported by rapid acquisition of resistance to azole and echinocandin antifungals frequently prompted by single nucleotide polymorphisms (SNPs) in resistance associated genes, such as PDR1 (azole resistance) or FKS1/2 (echinocandin resistance). To determine the frequency of polymorphisms and genome rearrangements as the possible genetic basis of C. glabrata drug resistance, we assessed genomic variation across 94 globally distributed isolates with distinct resistance phenotypes, whose sequence is deposited in GenBank. The genomes of three additional clinical isolates were sequenced, in this study, including two azole resistant strains that did not display Gain-Of-Function (GOF) mutations in the transcription factor encoding gene PDR1. Genomic variations in susceptible isolates were used to screen out variants arising from genome diversity and to identify variants exclusive to resistant isolates. More than half of the azole or echinocandin resistant isolates do not possess exclusive polymorphisms in PDR1 or FKS1/2, respectively, providing evidence of alternative genetic basis of antifungal resistance. We also identified copy number variations consistently affecting a subset of chromosomes. Overall, our analysis of the genomic and phenotypic variation across isolates allowed to pinpoint, in a genome-wide scale, genetic changes enriched specifically in antifungal resistant strains, which provides a first step to identify additional determinants of antifungal resistance. Specifically, regarding the newly sequenced strains, a set of mutations/genes are proposed to underlie the observed unconventional azole resistance phenotype.

Genomic evolution towards azole resistance in Candida glabrata clinical isolates unveils the importance of CgHxt4/6/7 in azole accumulation.

The increasing prevalence of candidosis caused by Candida glabrata is related to its ability to acquire azole resistance. Although azole resistance mechanisms are well known, the mechanisms for azole import into fungal cells have remained obscure. In this work, we have characterized two hexose transporters in C. glabrata and further investigate their role as potential azole importers. Three azole susceptible C. glabrata clinical isolates were evolved towards azole resistance and the acquired resistance phenotype was found to be independent of CgPDR1 or CgERG11 mutations. Through whole-genome sequencing, CgHXT4/6/7 was found to be mutated in the three evolved strains, when compared to their susceptible parents. CgHxt4/6/7 and the 96% identical CgHxt6/7 were found to confer azole susceptibility and increase azole accumulation in C. glabrata cells, strikingly rescuing the susceptibility phenotype imposed by CgPDR1 deletion, while the identified loss-of-function mutation in CgHXT4/6/7, leads to increased azole resistance. In silico docking analysis shows that azoles display a strong predicted affinity for the glucose binding site of CgHxt4/6/7. Altogether, we hypothesize that hexose transporters, such as CgHxt4/6/7 and CgHxt6/7, may constitute a family of azole importers, involved in clinical drug resistance in fungal pathogens, and constituting promising targets for improved antifungal therapy.

Insights into the Antimicrobial Activities and Metabolomes of Aquimarina (Flavobacteriaceae, Bacteroidetes) Species from the Rare Marine Biosphere.

Two novel natural products, the polyketide cuniculene and the peptide antibiotic aquimarin, were recently discovered from the marine bacterial genus Aquimarina. However, the diversity of the secondary metabolite biosynthetic gene clusters (SM-BGCs) in Aquimarina genomes indicates a far greater biosynthetic potential. In this study, nine representative Aquimarina strains were tested for antimicrobial activity against diverse human-pathogenic and marine microorganisms and subjected to metabolomic and genomic profiling. We found an inhibitory activity of most Aquimarina strains against Candida glabrata and marine Vibrio and Alphaproteobacteria species. Aquimarina sp. Aq135 and Aquimarina muelleri crude extracts showed particularly promising antimicrobial activities, amongst others against methicillin-resistant Staphylococcus aureus. The metabolomic and functional genomic profiles of Aquimarina spp. followed similar patterns and were shaped by phylogeny. SM-BGC and metabolomics networks suggest the presence of novel polyketides and peptides, including cyclic depsipeptide-related compounds. Moreover, exploration of the 'Sponge Microbiome Project' dataset revealed that Aquimarina spp. possess low-abundance distributions worldwide across multiple marine biotopes. Our study emphasizes the relevance of this member of the microbial rare biosphere as a promising source of novel natural products. We predict that future metabologenomics studies of Aquimarina species will expand the spectrum of known secondary metabolites and bioactivities from marine ecosystems.

DNA Markers for Detection and Genotyping of Xanthomonas euroxanthea.

Xanthomonas euroxanthea is a bacterial species encompassing both pathogenic and non-pathogenic strains and is frequently found colonizing the same host plants as X. arboricola. This presents the need to develop a detection and genotyping assay able to track these bacteria in microbial consortia with other xanthomonads. Eight X. euroxanthea-specific DNA markers (XEA1-XEA8) were selected by comparative genomics and validated in silico regarding their specificity and consistency using BLASTn, synteny analysis, CG content, codon usage (CAI/eCAI values) and genomic proximity to plasticity determinants. In silico, the selected eight DNA markers were found to be specific and conserved across the genomes of 11 X. euroxanthea strains, and in particular, five DNA markers (XEA4, XEA5, XEA6, XEA7 and XEA8) were unfailingly found in these genomes. A multiplex of PCR targeting markers XEA1 (819 bp), XEA8 (648 bp) and XEA5 (295 bp) was shown to successfully detect X. euroxanthea down to 1 ng of DNA (per PCR reaction). The topology of trees generated with the concatenated sequences of three markers (XEA5, XEA6 and XEA8) and four housekeeping genes (gyrB, rpoD, fyuA and acnB) underlined the equal discriminatory power of these features and thus the suitability of the DNA markers to discriminate X. euroxanthea lineages. Overall, this study displays a DNA-marker-based method for the detection and genotyping of X. euroxanthea strains, contributing to monitoring for its presence in X. arboricola-colonizing habitats. The present study proposes a workflow for the selection of species-specific detection markers. Prospectively, this assay could contribute to unveil alternative host species of Xanthomonas euroxanthea; and improve the control of phytopathogenic strains.

Prediction of Gene and Genomic Regulation in Candida Species, Using the PathoYeastract Database: A Comparative Genomics Approach.

The ability of living organisms to survive changing environmental conditions is dependent on the implementation of gene expression programs underlying adaptation and fitness. Transcriptional networks can be exceptionally complex: a single transcription factor (TF) may regulate hundreds of genes, and multiple TFs may regulate a single gene-depending on the environmental conditions. Moreover, the same TF may act as an activator or repressor in distinct conditions. In turn, the activity of regulators themselves may be dependent on other TFs, as well as posttranscriptional and posttranslational regulation. These traits greatly contribute to the intricate networks governing gene expression programs.In this chapter, a step-by-step guide of how to use PathoYeastract, one of several interconnecting databases within the YEASTRACT+ portal, to predict gene and genomic regulation in Candida spp. is provided. PathoYeastract contains a set of analysis tools to study regulatory associations in human pathogenic yeasts, enabling: (1) the prediction and ranking of TFs that contribute to the regulation of individual genes; (2) the prediction of the genes regulated by a given TF; and (3) the prediction and ranking of TFs that regulate a genome-wide transcriptional response. These capabilities are illustrated, respectively, with the analysis of: (1) the TF network controlling the C. glabrata QDR2 gene; (2) the regulon controlled by the C. glabrata TF Rpn4; and (3) the regulatory network controlling the C. glabrata transcriptome-wide changes induced upon exposure to the antifungal drug fluconazole. The newest potentialities of this information system are explored, including cross-species network comparison. The results are discussed considering the performed queries and integrated with the current knowledge on the biological data for each case-study.

Erg25 Controls Host-Cholesterol Uptake Mediated by Aus1p-Associated Sterol-Rich Membrane Domains in Candida glabrata.

The uptake of cholesterol from the host is closely linked to the proliferation of pathogenic fungi and protozoa during infection. For some pathogenic fungi, cholesterol uptake is an important strategy for decreasing susceptibility to antifungals that inhibit ergosterol biosynthesis. In this study, we show that Candida glabrata ERG25, which encodes an enzyme that demethylates 4,4-dimethylzymosterol, is required for cholesterol uptake from host serum. Based on the screening of C. glabrata conditional knockdown mutants for each gene involved in ergosterol biosynthesis, ERG25 knockdown was found to decrease lethality of infected mice. ERG25 knockdown impairs the plasma membrane localization of the sterol importer Aus1p, suggesting that the accumulated 4,4-dimethylzymosterol destabilizes the lipid domain with which Aus1p functionally associates. ERG25 knockdown further influences the structure of the membrane compartment of Can1p (MCC)/eisosomes (ergosterol-rich lipid domains), but not the localization of the membrane proteins Pma1p and Hxt1p, which localize to sterol-poor domains. In the sterol-rich lipid domain, Aus1p-contining domain was mostly independent of MCC/eisosomes, and the nature of these domains was also different: Ausp1-contining domain was a dynamic network-like domain, whereas the MCC/eisosomes was a static dot-like domain. However, deletion of MCC/eisosomes was observed to influence the localization of Aus1p after Aus1p was transported from the endoplasmic reticulum (ER) through the Golgi apparatus to the plasma membrane. These findings suggest that ERG25 plays a key role in stabilizing sterol-rich lipid domains, constituting a promising candidate target for antifungal therapy.

A Genome-Scale Metabolic Model for the Human Pathogen Candida Parapsilosis and Early Identification of Putative Novel Antifungal Drug Targets.

Candida parapsilosis is an emerging human pathogen whose incidence is rising worldwide, while an increasing number of clinical isolates display resistance to first-line antifungals, demanding alternative therapeutics. Genome-Scale Metabolic Models (GSMMs) have emerged as a powerful in silico tool for understanding pathogenesis due to their systems view of metabolism, but also to their drug target predictive capacity. This study presents the construction of the first validated GSMM for C. parapsilosis-iDC1003-comprising 1003 genes, 1804 reactions, and 1278 metabolites across four compartments and an intercompartment. In silico growth parameters, as well as predicted utilisation of several metabolites as sole carbon or nitrogen sources, were experimentally validated. Finally, iDC1003 was exploited as a platform for predicting 147 essential enzymes in mimicked host conditions, in which 56 are also predicted to be essential in C. albicans and C. glabrata. These promising drug targets include, besides those already used as targets for clinical antifungals, several others that seem to be entirely new and worthy of further scrutiny. The obtained results strengthen the notion that GSMMs are promising platforms for drug target discovery and guide the design of novel antifungal therapies.