A biomimetic ion channel shortens the QT interval of type 2 long QT syndrome through efficient transmembrane transport of potassium ions.

Potassium ion transport across myocardial cell membrane is essential for type 2 long QT syndrome (LQT2). However, the dysfunction of potassium ion transport due to genetic mutations limits the therapeutic effect in treating LQT2. Biomimetic ion channels that selectively and efficiently transport potassium ions across the cellular membranes are promising for the treatment of LQT2. To corroborate this, we synthesized a series of foldamer-based ion channels with different side chains, and found a biomimetic ion channel of K+ (BICK) with the highest transport activity among them. The selected BICK can restore potassium ion transport and increase transmembrane potassium ion current, thus shortening phase 3 of action potential (AP) repolarization and QT interval in LQT2. Moreover, BICK does not affect heart rate and cardiac rhythm in treating LQT2 model induced by E4031 in isolated heart as well as in guinea pigs. By restoring ion transmembrane transport tactic, biomimetic ion channels, such as BICK, will show great potential in treating diseases related to ion transport blockade. STATEMENT OF SIGNIFICANCE: Type 2 long QT syndrome (LQT2) is a disease caused by K+ transport disorder, which can cause malignant arrhythmia and even death. There is currently no radical cure, so it is critical to explore ways to improve K+ transmembrane transport. In this study, we report that a small-molecule biomimetic ion channel BICK can efficiently simulate natural K+ channel proteins on the cardiomyocyte and cure E4031-induced LQT2 in guinea pig by restoring K+ transport function for the first time. This study found that the potassium transmembrane transport by BICK significantly reduced the QT interval, which provides a conceptually new strategy for the treatment of LQT2 disease.

Antibacterial Activity and Mechanism of Action of Osthole against Listeria monocytogenes.

The purpose of this study was to investigate the antibacterial activity and mechanism of action of osthole against Listeria monocytogenes. The antibacterial activity of osthole was evaluated by determining the minimum inhibitory concentration (MIC) and growth curve. Cell morphology, membrane permeability, membrane integrity, bacterial physiology, and metabolism were explored using different methods to elucidate the mechanism of action of osthole. It was shown that the MIC of osthole against L. monocytogenes was 62.5 µg/mL and it inhibited the growth of L. monocytogenes effectively in a concentration-dependent manner. Scanning electron microscopy (SEM) images demonstrated morphology changes of L. monocytogenes, including rough surface, cell shrinkage, and rupture. It was found that extracellular conductivity and macromolecule content were increased significantly in the presence of osthole, indicating the disruption of cell membrane integrity and permeability. Laser confocal microscopy results supported the conclusion that osthole caused severe damage to the cell membrane. It was also noticed that osthole depleted intracellular adenosine triphosphate (ATP), inhibited Na+-K+-ATPase and Ca2+-Mg2+-ATPase activity, and promoted the accumulation of intracellular reactive oxygen species (ROS), leading to cell death. This study suggests that osthole is a promising antibacterial agent candidate against L. monocytogenes, and it shows potential in the prevention and control of foodborne pathogens.

Extraction of tannin, colour and aroma compounds in pinot noir wines as affected by clone selection and whole bunch addition.

Our previous study revealed stem inclusion fermentation reduced anthocyanin, and increased tannin and aroma compounds responsible for green notes. This study further investigated the effect of clone selection and whole bunch fermentation on Pinot noir wine composition, with focus on tannin composition. Three treatments were conducted using two clones (AM10/5 and UCD5) in 2021 and 2022: 100% destemmed (DS), 30% whole bunch (WB30), and 60% whole bunch (WB60). WB60 increased stem and skin derived tannins but reduced seed derived tannin proportion in wines. Clone selection had an impact on tannin composition and an even greater impact on tannin concentration, colour, and aroma compounds. AM10/5 produced wines with higher tannin, polymeric pigments and darker colour. AM10/5 wines also had higher concentration of phenylethyl alcohol, but lower concentrations of 3-isobutyl-2-methoxypyrazine and ethyl esters, indicating more floral but less fruity and green notes.

Printed Self-Healing Stretchable Electronics for Bio-signal Monitoring and Intelligent Packaging.

Integrating self-healing capabilities into printed stretchable electronic devices is important for improving performance and extending device life. However, achieving printed self-healing stretchable electronic devices with excellent device-level healing ability and stretchability while maintaining outstanding electrical performance remains challenging. Herein, a series of printed device-level self-healing stretchable electronic devices is achieved by depositing liquid metal/silver fractal dendrites/polystyrene-block-polyisoprene-block-polystyrene (LM/Ag FDs/SIS) conductive inks onto a self-healing thermoplastic polyurethane (TPU) film via screen printing method. Owing to the fluidic properties of the LM and the interfacial hydrogen bonding and disulfide bonds of TPU, the as-obtained stretchable electronic devices maintain good electronic properties under strain and exhibit device-level self-healing properties without external stimulation. Printed self-healing stretchable electrodes possess high electrical conductivity (1.6 × 105 S m-1), excellent electromechanical properties, and dynamic stability, with only a 2.5-fold increase in resistance at 200% strain, even after a complete cut and re-healing treatment. The printed self-healing capacitive stretchable strain sensor shows good linearity (R2 ≈0.9994) in a wide sensing range (0%-200%) and is successfully applied to bio-signal detection. Furthermore, the printed self-healing electronic smart label is designed and can be used for real-time environmental monitoring, which exhibits promising potential for practical application in food preservation packaging.

Multiple functions of the nonstructural protein 3D in picornavirus infection.

3D polymerase, also known as RNA-dependent RNA polymerase, is encoded by all known picornaviruses, and their structures are highly conserved. In the process of picornavirus replication, 3D polymerase facilitates the assembly of replication complexes and directly catalyzes the synthesis of viral RNA. The nuclear localization signal carried by picornavirus 3D polymerase, combined with its ability to interact with other viral proteins, viral RNA and cellular proteins, indicate that its noncatalytic role is equally important in viral infections. Recent studies have shown that 3D polymerase has multiple effects on host cell biological functions, including inducing cell cycle arrest, regulating host cell translation, inducing autophagy, evading immune responses, and triggering inflammasome formation. Thus, 3D polymerase would be a very valuable target for the development of antiviral therapies. This review summarizes current studies on the structure of 3D polymerase and its regulation of host cell responses, thereby improving the understanding of picornavirus-mediated pathogenesis caused by 3D polymerase.

Vulnerable brain regions in adolescent major depressive disorder: A resting-state functional magnetic resonance imaging activation likelihood estimation meta-analysis.

BACKGROUND: Adolescent major depressive disorder (MDD) is a significant mental health concern that often leads to recurrent depression in adulthood. Resting-state functional magnetic resonance imaging (rs-fMRI) offers unique insights into the neural mechanisms underlying this condition. However, despite previous research, the specific vulnerable brain regions affected in adolescent MDD patients have not been fully elucidated. AIM: To identify consistent vulnerable brain regions in adolescent MDD patients using rs-fMRI and activation likelihood estimation (ALE) meta-analysis. METHODS: We performed a comprehensive literature search through July 12, 2023, for studies investigating brain functional changes in adolescent MDD patients. We utilized regional homogeneity (ReHo), amplitude of low-frequency fluctuations (ALFF) and fractional ALFF (fALFF) analyses. We compared the regions of aberrant spontaneous neural activity in adolescents with MDD vs healthy controls (HCs) using ALE. RESULTS: Ten studies (369 adolescent MDD patients and 313 HCs) were included. Combining the ReHo and ALFF/fALFF data, the results revealed that the activity in the right cuneus and left precuneus was lower in the adolescent MDD patients than in the HCs (voxel size: 648 mm3, P < 0.05), and no brain region exhibited increased activity. Based on the ALFF data, we found decreased activity in the right cuneus and left precuneus in adolescent MDD patients (voxel size: 736 mm3, P < 0.05), with no regions exhibiting increased activity. CONCLUSION: Through ALE meta-analysis, we consistently identified the right cuneus and left precuneus as vulnerable brain regions in adolescent MDD patients, increasing our understanding of the neuropathology of affected adolescents.

Iron efflux by IetA enhances ß-lactam aztreonam resistance and is linked to oxidative stress through cellular respiration in Riemerella anatipestifer.

BACKGROUND: Riemerella anatipestifer encodes an iron acquisition system, but whether it encodes the iron efflux pump and its role in antibiotic resistance are largely unknown. OBJECTIVES: To screen and identify an iron efflux gene in R. anatipestifer and determine whether and how the iron efflux gene is involved in antibiotic resistance. METHODS: In this study, gene knockout, streptonigrin susceptibility assay and inductively coupled plasma mass spectrometry were used to screen for the iron efflux gene ietA. The MIC measurements, scanning electron microscopy and reactive oxygen species (ROS) detection were used to verify the role of IetA in aztreonam resistance and its mechanism. Mortality and colonization assay were used to investigate the role of IetA in virulence. RESULTS: The deletion mutant ΔietA showed heightened susceptibility to streptonigrin, and prominent intracellular iron accumulation was observed in ΔfurΔietA under excess iron conditions. Additionally, ΔietA exhibited increased sensitivity to H2O2-produced oxidative stress. Under aerobic conditions with abundant iron, ΔietA displayed increased susceptibility to the ß-lactam antibiotic aztreonam due to heightened ROS production. However, the killing efficacy of aztreonam was diminished in both WT and ΔietA under anaerobic or iron restriction conditions. Further experiments demonstrated that the efficiency of aztreonam against ΔietA was dependent on respiratory complexes Ⅰ and Ⅱ. Finally, in a duckling model, ΔietA had reduced virulence compared with the WT. CONCLUSION: Iron efflux is critical to alleviate oxidative stress damage and ß-lactam aztreonam killing in R. anatipestifer, which is linked by cellular respiration.

Influence of Grape Flesh on the Retention and Composition of Polyphenols from Skins and Seeds.

This study examined the impact of grape flesh polysaccharide, protein, and amino acid contents on polyphenol retention from skins and seeds in Pinot noir (Vitis vinifera) and cold-hardy interspecific cultivars Marquette and Frontenac (Vitis spp.). After isolating grape tissues (skin, seed, and flesh), they were soaked either individually or combined with other tissues in a wine-like solution for up to 7 days. Findings revealed that flesh significantly reduces the concentration of condensed tannin, and mono- and diglucoside forms of anthocyanins in the supernatants, due to its rich content in polysaccharides and proteins. Frontenac skin and flesh tissues were the main sources of soluble proteins, amino acids, and soluble polysaccharides. Surprisingly, Marquette exhibited a higher retention of skin tannin than Pinot noir, likely due to its smaller tannin molecular mass, and a potential competitive effect with anthocyanins for the binding sites of flesh.

Tensile Strain-Mediated Bimetallene Nanozyme for Enhanced Photothermal Tumor Catalytic Therapy.

Nanozymes have demonstrated significant potential in combating malignant tumor proliferation through catalytic therapy. However, the therapeutic effect is often limited by insufficient catalytic performance. In this study, we propose the utilization of strain engineering in metallenes to fully expose the active regions due to their ultrathin nature. Here, we present the first report on a novel tensile strain-mediated local amorphous RhRu (la-RhRu) bimetallene with exceptional intrinsic photothermal effect and photo-enhanced multiple enzyme-like activities. Through geometric phase analysis, electron diffraction profile, and X-ray diffraction, it is revealed that crystalline-amorphous heterophase boundaries can generate approximately 2 % tensile strain in the bimetallene. The ultrathin structure and in-plane strain of the bimetallene induce an amplified strain effect. Both experimental and theoretical evidence support the notion that tensile strain promotes multiple enzyme-like activities. Functioning as a tumor microenvironment (TME)-responsive nanozyme, la-RhRu exhibits remarkable therapeutic efficacy both in vitro and in vivo. This work highlights the tremendous potential of atomic-scale tensile strain engineering strategy in enhancing tumor catalytic therapy.

Matrine and icariin can inhibit bovine viral diarrhoea virus replication by promoting type I interferon response in vitro.

Introduction: Bovine viral diarrhoea virus (BVDV) can cause diarrhoea (BVD) in an animal herd, leading to heavy economic losses. There are limited drugs available for treating and controlling BVD. This research aims to investigate the antiviral and immunoregulatory effects of two traditional Chinese herb extracts against BVDV infection. The extracts are matrine and icariin, which have been proved to have immunostimulant and antiviral effects. Material and Methods: A cell counting kit-8 assay was used to analyse the toxicity of matrine and icariin to Madin-Darby bovine kidney (MDBK) cells. The model of MDBK cells infected with BVDV was utilised to uncover the antiviral mechanism of matrine and icariin, which along with their immunoregulatory ability was evaluated by quantitative reverse-transcription PCR and ELISA. Results: The results showed that matrine and icariin can significantly inhibit the gene expression level of the BVDV 5' untranslated region through various pathways. Both matrine and icariin can statistically upregulate the gene expression level of interferon alpha, interferon beta (IFN-ß), toll-like receptor 3, retinoic acid-inducible gene I and interferon regulatory factor 3, and raise the concentration of IFN-ß after BVDV infection. Conclusion: This study proves that both matrine and icariin have inhibitory effects on BVDV replication by activating IFN production and the IFN signalling pathway. The finding is promising and should open up the possibility of larger-scale in vitro research followed by in vivo experiments evaluating matrine and icariin as therapeutic agents in BVD cases.

Image rectangling network based on reparameterized transformer and assisted learning.

Stitched images can offer a broader field of view, but their boundaries can be irregular and unpleasant. To address this issue, current methods for rectangling images start by distorting local grids multiple times to obtain rectangular images with regular boundaries. However, these methods can result in content distortion and missing boundary information. We have developed an image rectangling solution using the reparameterized transformer structure, focusing on single distortion. Additionally, we have designed an assisted learning network to aid in the process of the image rectangling network. To improve the network's parallel efficiency, we have introduced a local thin-plate spline Transform strategy to achieve efficient local deformation. Ultimately, the proposed method achieves state-of-the-art performance in stitched image rectangling with a low number of parameters while maintaining high content fidelity. The code is available at https://github.com/MelodYanglc/TransRectangling .

Artificial intelligence-based MRI radiomics and radiogenomics in glioma.

The specific genetic subtypes that gliomas exhibit result in variable clinical courses and the need to involve multidisciplinary teams of neurologists, epileptologists, neurooncologists and neurosurgeons. Currently, the diagnosis of gliomas pivots mainly around the preliminary radiological findings and the subsequent definitive surgical diagnosis (via surgical sampling). Radiomics and radiogenomics present a potential to precisely diagnose and predict survival and treatment responses, via morphological, textural, and functional features derived from MRI data, as well as genomic data. In spite of their advantages, it is still lacking standardized processes of feature extraction and analysis methodology among different research groups, which have made external validations infeasible. Radiomics and radiogenomics can be used to better understand the genomic basis of gliomas, such as tumor spatial heterogeneity, treatment response, molecular classifications and tumor microenvironment immune infiltration. These novel techniques have also been used to predict histological features, grade or even overall survival in gliomas. In this review, workflows of radiomics and radiogenomics are elucidated, with recent research on machine learning or artificial intelligence in glioma.

FDG PET/CT Findings of a Small Cell Carcinoma of the Lung Presented as Diffuse Dermatomyositis.

ABSTRACT: Patients with dermatomyositis are prone to have occult malignancy. A previously healthy 68-year-old man with dermatomyositis underwent FDG PET/CT to detect possible malignancy of unknown origin. The images showed not only diffuse increased activity in the muscle, which was related to the known dermatomyositis, but also intense activity in the anterior chest with foci of abnormal activity throughout the body. Pathology examination confirmed small cell carcinoma of the lung with widespread metastases.

NS1: a promising novel target antigen with strong immunogenicity and protective efficacy for avian flavivirus vaccine development.

Tembusu virus (TMUV), an avian pathogenic flavivirus, has emerged as a significant threat to the duck industry in Southeast Asia, causing substantial economic losses. Due to the antibody-dependent enhancement (ADE) effect of TMUV subneutralizing antibodies, there is a pressing need to further develop new TMUV vaccine target antigens that ensure both safety and efficacy. Here, the TMUV non-structural protein 1 (NS1) as a target for development of effective anti-TMUV vaccines was unveiled. The amino acid sequences of TMUV NS1 exhibit a high degree of conservation across different strains (92.63-100%). To investigate the potential of TMUV NS1 as a vaccine target, the TMUV NS1-based plasmids were constructed and identified the C-terminal 30 amino acids residues of TMUV E (EC30) as an effective signal peptide for promoting NS1 expression and secretion. Subsequently, the plasmid pVAX1-EC30-NS1 was employed to immunize ducks, resulting in specific anti-NS1 IgG responses being stimulated, while without inducing anti-TMUV neutralizing antibodies. Furthermore, the cellular immune responses triggered by the TMUV NS1 were evaluated, observing a notable increase in lymphocyte proliferation at 4 wk and 6 wk postinjection with the pVAX1-EC30-NS1. Additionally, there was a significant up-regulation of NS1-specific Il-4 and Ifnγ levels at these time points. Following this, ducks from different groups were challenged with TMUV, and remarkably, those immunized with the NS1 vaccine displayed significantly lower viral copies both at 3 d postinfection (dpi) and 7 dpi (P < 0.05) compared to ducks immunized with the control vector. Notably, the NS1 demonstrated remarkable protection against TMUV challenge without causing severe gross lesions. Collectively, these findings highlighted the impressive immunogenicity and protectivity of the TMUV NS1. Consequently, NS1 holds great promise as a novel antigen target for the development of efficient and safe TMUV vaccines.

An alpha-herpesvirus employs host HEXIM1 to promote viral transcription.

Although it is widely accepted that herpesviruses utilize host RNA polymerase II (RNAPII) to transcribe viral genes, the mechanism of utilization varies significantly among herpesviruses. With the exception of herpes simplex virus 1 (HSV-1) in alpha-herpesviruses, the mechanism by which RNAPII transcribes viral genes in the remaining alpha-herpesviruses has not been reported. In this study, we investigated the transcriptional mechanism of an avian alpha-herpesvirus, Anatid herpesvirus 1 (AnHV-1). We discovered for the first time that hexamethylene-bis-acetamide-inducing protein 1 (HEXIM1), a major inhibitor of positive elongation factor B (P-TEFb), was significantly upregulated during AnHV-1 infection, and its expression was dynamically regulated throughout the progression of the disease. However, the expression level of HEXIM1 remained stable before and after HSV-1 infection. Excessive HEXIM1 assists AnHV-1 in progeny virus production, gene expression, and RNA polymerase II recruitment by promoting the formation of more inactive P-TEFb and the loss of RNAPII S2 phosphorylation. Conversely, the expression of some host survival-related genes, such as SOX8, CDK1, MYC, and ID2, was suppressed by HEXIM1 overexpression. Further investigation revealed that the C-terminus of the AnHV-1 US1 gene is responsible for the upregulation of HEXIM1 by activating its promoter but not by interacting with P-TEFb, which is the mechanism adopted by its homologs, HSV-1 ICP22. Additionally, the virus proliferation deficiency caused by US1 deletion during the early infection stage could be partially rescued by HEXIM1 overexpression, suggesting that HEXIM1 is responsible for AnHV-1 gaining transcription advantages when competing with cells. Taken together, this study revealed a novel HEXIM1-dependent AnHV-1 transcription mechanism, which has not been previously reported in herpesvirus or even DNA virus studies.IMPORTANCEHexamethylene-bis-acetamide-inducing protein 1 (HEXIM1) has been identified as an inhibitor of positive transcriptional elongation factor b associated with cancer, AIDS, myocardial hypertrophy, and inflammation. Surprisingly, no previous reports have explored the role of HEXIM1 in herpesvirus transcription. This study reveals a mechanism distinct from the currently known herpesvirus utilization of RNA polymerase II, highlighting the dependence on high HEXIM1 expression, which may be a previously unrecognized facet of the host shutoff manifested by many DNA viruses. Moreover, this discovery expands the significance of HEXIM1 in pathogen infection. It raises intriguing questions about whether other herpesviruses employ similar mechanisms to manipulate HEXIM1 and if this molecular target can be exploited to limit productive replication. Thus, this discovery not only contributes to our understanding of herpesvirus infection regulation but also holds implications for broader research on other herpesviruses, even DNA viruses.

Duck plague virus Us3 regulates the expression of pUL48.

Duck plague (DP) is one of the contagious diseases caused by Duck plague virus (DPV), which is a serious threat to the development of duck farming. Us3 is a PKA-like protein kinase in alphaherpesvirus, which can regulate the biological functions of many viral proteins, but whether Us3 regulates pUL48 protein has not been reported. In this paper, Western Blot, qRT-PCR, dual luciferase reporter system and Co-IP were used to investigate the relationship between pUL48 and Us3. The results showed that: 1) pUL48 interacted with Us3 at 138-256aa through its DBD region. 2) Us3 enhanced the protein expression of pUL48 in a dose-dependent manner. 3) Us3 promoted the mRNA level of pUL48 by activating its promoter activity. 4) Us3 inhibited the transcriptional activation function of pUL48. The results can provide scientific data for perfecting and supplementing the function of alpha herpesvirus Us3 and pUL48.

CCCP inhibits DPV infection in DEF cells by attenuating DPV manipulated ROS, apoptosis, and mitochondrial stability.

Duck plague virus (DPV) is extremely infectious and lethal, so antiviral drugs are urgently needed. Our previous study shows that DPV infection with duck embryo fibroblast (DEF) induces reactive oxygen species (ROS) changes and promotes apoptosis. In this study, we tested the antiviral effect of the carbonyl cyanide m-chlorophenyl hydrazone (CCCP), a common mitochondrial autophagy inducer. Our results demonstrated a dose-dependent anti-DPV effect of CCCP, CCCP-treatment blocked the intercellular transmission of DPV after infection, and we also proved that CCCP could have an antiviral effect up to 48 hpi. The addition of CCCP reversed the DPV-induced ROS changes, CCCP can inhibit virus-induced apoptosis; meanwhile, CCCP can affect mitochondrial fusion and activate mitophagy to inhibit DPV. In conclusion, CCCP can be an effective antiviral candidate against DPV.

Efficient photocatalytic degradation of antibiotics by binary heterojunction complex boron subphthalocyanine bromide/bismuth oxychloride.

The development of stable and efficient heterojunction photocatalysts for wastewater environmental purification exhibits a significant challenge. Herien, a promising binary heterojunction complex comprising boron subphthalocyanine bromide/bismuth oxychloride (SubPc-Br/BiOCl) was successfully synthesized using the hydrothermal method, which involved the self-assembled of SubPc-Br on the surface of BiOCl via intermolecular π-π stacking interactions to compose an electron-transporting layer. The photocatalytic efficiency of SubPc-Br/BiOCl for the degradation of tetracycline and the minocycline exhibited a substantial improvement of 29.14% and 53.72%, respectively, compared to the original BiOCl. Experimental characterization and theoretical calculations elucidated that the enhanced photocatalytic performance of the SubPc-Br/BiOCl composite photocatalysts stemmed from the S-scheme electron transport mechanism at the interface between BiOCl and SubPc-Br supramolecules, which broadened the visible light absorption range, increased the carrier molecular efficiency, and accelerated the carriers. Furthermore, molecular dynamic (MD) simulations provided insights into the action trajectories of the two semiconductors, revealing that the presence of SubPc-Br enhances the water and organic pollutant adsorption capabilities of the BiOCl surface within the supramolecular array system. In conclusion, the synthesis and analysis of the binary heterojunction complex SubPc-Br/BiOCl yield valuable insights into the efficient photocatalytic degradation of antibiotics, holding great promise for diverse environmental applications.

Systematic characterization of p53-regulated long non-coding RNAs across human cancers reveals remarkable heterogeneity among different tumor types.

The p53 tumor suppressor protein, a sequence specific DNA binding transcription factor, regulates the expression of a large number of genes, in response to various forms of cellular stress. While the protein coding target genes of p53 have been well studied, less is known about its role in regulating long non-coding genes and their functional relevance to cancer. Here we report the genome-wide identification of a large set (>1000) of long non-coding RNAs(lncRNAs) that are putative p53 targets in a colon cancer cell line and in human patient datasets from five different common types of cancer. These lncRNAs have not been annotated by other studies of normal unstressed systems. In the colon cancer cell line a high proportion of these lncRNAs are uniquely induced by different chemotherapeutic agents that activate p53, while others are induced by more than one agent tested. Further, subsets of these lncRNAs independently predict overall and disease-free survival of patients across the five different common cancer types. Interestingly, both genetic alterations and patient survival associated with different lncRNAs are unique to each cancer tested, indicating extraordinary tissue-specific variability in the p53 non-coding response. The newly identified non-coding p53 target genes have allowed us to construct a classifier for tumor diagnosis and prognosis. Implications: Our results not only identify myriad p53-regulated lncRNAs, they also reveal marked drug-induced, as well as tissue- and tumor-specific heterogeneity in these putative p53 targets and our findings have enabled the construction of robust classifiers for diagnosis and prognosis.