Mitochondria complex I deficiency in Candida albicans arrests the cell cycle at S phase through suppressive TOR and PKA pathways.

How mutations in mitochondrial electron transport chain (ETC) proteins impact the cell cycle of Candida albicans was investigated in this study. Using genetic null mutants targeting ETC complexes I (CI), III (CIII), and IV (CIV), the cell cycle stages (G0/G1, S phase, and G2/M) were analyzed via fluorescence-activated cell sorting (FACS). Four CI null mutants exhibited distinct alterations, including extended S phase, shortened G2/M population, and a reduction in cells size exceeding 10 µM. Conversely, CIII mutants showed an increased population in G1/G0 phase. Among four CI mutants, ndh51Δ/Δ and goa1Δ/Δ displayed aberrant cell cycle patterns correlated with previously reported cAMP/PKA downregulation. Specifically, nuo1Δ/Δ and nuo2Δ/Δ mutants exhibited increased transcription of RIM15, a central hub linking cell cycle with nutrient-dependent TOR1 and cAMP/PKA pathways and Snf1 aging pathway. These findings suggest that suppression of TOR1 and cAMP/PKA pathways or enhanced Snf1 disrupts cell cycle progression, influencing cell longevity and growth among CI mutants. Overall, our study highlights the intricate interplay between mitochondrial ETC, cell cycle, and signaling pathways.

The mitochondrial complex I proteins of Candida albicans moderate phagocytosis and the production of pro-inflammatory cytokines in murine macrophages and dendritic cells.

Loss of respiratory functions impairs Candida albicans colonization of host tissues and virulence in a murine model of candidiasis. Furthermore, it is known that respiratory inhibitors decrease mannan synthesis and glucan exposure and thereby promotes phagocytosis. To understand the impact of respiratory proteins of C. albicans on host innate immunity, we characterized cell wall defects in three mitochondrial complex I (CI) null mutants (nuo1Δ, nuo2Δ and ndh51Δ) and in one CI regulator mutant (goa1Δ), and we studied the corresponding effects of these mutants on phagocytosis, neutrophil killing and cytokine production by dendritic cells (DCs). We find that reductions of phosphopeptidomannan (PPM) in goa1Δ, nuo1Δ and phospholipomannan (PLM) in nuo2Δ lead to reductions of IL-2, IL-4, and IL-10 but increase of TNF-α in infected DCs. While PPM loss is a consequence of a reduced phospho-Cek1/2 MAPK that failed to promote phagocytosis and IL-22 production in goa1Δ and nuo1Δ, a 30% glucan reduction and a defective Mek1 MAPK response in ndh51Δ lead to only minor changes in phagocytosis and cytokine production. Glucan exposure and PLM abundance seem to remain sufficient to opsonize neutrophil killing perhaps via humoral immunity. The diversity of immune phenotypes in these mutants possessing divergent cell wall defects is further supported by their transcriptional profiles in each infected murine macrophage scenario. Since metabolic processes, oxidative stress-induced senescence, and apoptosis are differently affected in these scenarios, we speculate that during the early stages of infection, host immune cells coordinate their bioactivities based upon a mixture of signals generated during host-fungi interactions.

Salivary and serum IgA and IgG responses to SARS-CoV-2-spike protein following SARS-CoV-2 infection and after immunization with COVID-19 vaccines.

Background: Secretory immunoglobulin A (sIgA) plays an important role in antiviral protective immunity. Although salivary testing has been used for many viral infections, including severe acute respiratory syndrome (SARS) and Middle East Respiratory Syndrome (MERS), its use has not yet been well established with the SARS coronavirus 2 (SARS-CoV-2). Quantification of salivary IgA and IgG antibodies can elucidate mucosal and systemic immune responses after natural infection or vaccination. Here, we report the development and validation of a rapid enzyme-linked immunosorbent assay (ELISA) for anti-SARS-CoV-2 salivary IgA and serum IgG antibodies, and present quantitative results for immunized subjects both prior to or following COVID-19 infections. Objective: Total and serum SARS-CoV-2 spike-specific IgG responses were compared with salivary spike-specific IgA and IgG responses in samples obtained from patients recently infected with SARS-CoV-2 and from subjects recently immunized with COVID-19 vaccines. Methods: A total of 52 paired saliva and serum samples were collected from 26 study participants: 7 subjects after COVID-19 infection and 19 subjects who were uninfected. The ELISA results from these samples were compared with five prepandemic control serum samples. Total IgG and SARS-CoV-2 spike-specific IgG in the serum samples from the subjects who were infected and vaccinated were also measured in a commercial laboratory with an enzyme immunoassay. Results: A wide variation in antibody responses was seen in salivary and serum samples measured by both methods. Three groups of serum total and IgG spike-specific SARS-CoV-2 antibody responses were observed: (1) low, (2) intermediate, and (3) high antibody responders. A correlational analysis of salivary IgA (sIgA) responses with serum IgG concentrations showed a statistical correlation in the low and intermediate antibody responder groups but not in the high group (which we believe was a result of saturation). Conclusion: These preliminary findings suggest measuring salivary and serum IgG and IgA merit further investigation as markers of current or recent SARS-CoV-2 infections.

Genetic Screens of an Anti-Candida Natural Product Using the Heterozygous Saccharomyces cerevisiae Mutant Library.

Candida albicans is one of the most common fungal pathogens in humans. Due to the development of resistance to antifungal drugs, today there is a need for finding new antifungal agents with new pharmacological targets for a more efficient management of C. albicans infections. Drug repositioning or drug repurposing has been exploited to develop new antifungal approaches. Natural products may be more easily developed because they have been a useful source of active antimicrobials. Additionally, new antifungals are needed to combat drug-resistant infections caused by fungi such as by Candida species. Once compounds are identified, determining the mode of action (MOA) of natural products is a key objective. Genetic screens utilizing the Saccharomyces cerevisiae heterozygous mutant library provides a direct link between a phenotypic screen (easy read-out) and the identity of the gene target. Screens using mutant libraries can identify chemical-genetic interactions and genes or pathways affected by compounds to decipher the mechanism of action. Herein, we describe a genetic screen of an anti-Candida natural product.

Assessing Complex I (CI) Mitochondrial Subunit Protein Functions in Candida albicans.

Mitochondria of Candida species play critical roles in cell metabolism and pathogenesis. Greater emphasis in specific mitochondria activities of this fungus have been revealed through studies that defined fungal or Candida-specific subunit proteins of ETC Complexes I, III, and IV (CI, CIII, and CIV). Functional activities of these subunits have been characterized through the construction of single-gene null mutants. Activities common to mitochondria of most eukaryotes include their importance in metabolism, ATP synthesis, oxidative phosphorylation, oxygen consumption, and redox potential. An important difference among specific subunits compared to eukaryotic species is the role of CI fungal-specific subunit proteins in activities specific to Candida albicans, such as cell wall synthesis, especially cell wall mannan and ß-glucan synthesis. We have associated cell wall synthesis with a signal transduction pathway that includes a Chk1p fungal-specific pathway. Recently, based upon the specificity of CI subunit specificities, a suggestion is the development of novel antifungals that target mitochondrial activity.

Measurements of Treg Cell Induction by Candida albicans DNA Using Flow Cytometry.

Microbiota and their metabolites in the human gut regulate a variety of immune cells including regulatory T cells (Treg cells) and cytokine production. The T helper 17 (Th17)/Treg ratio biomarker (Th17/Treg), for example, has been linked to the development and progression of certain inflammatory diseases, insulin resistance, and systemic lupus erythematosus (SLE). Candida albicans is an opportunistic fungal pathogen that also colonizes the gut. T cell reactivity through T helper cells play critical roles in fungal clearance by the host through the secretion of proinflammatory cytokines. While these cytokines are mainly produced by the Th1 and Th17 subsets of T cells, another subset of T cells, the Treg cells, are also induced by antigenic ligands from pathogens that inhibit the responses of other effector T cells during the inflammation. The antigenic ligands for Treg induction have been found to include microbial cell wall polysaccharides (PSA), metabolites like short chain fatty acids (SCFA), or even microbial DNA.

Deletion of Non-histidine Domains of Histidine Kinase CHK1 Diminishes the Infectivity of Candida albicans in an Oral Mucosal Model.

Objectives: The histidine kinase (HK) CHK1 and other protein kinases in Candida albicans are key players in the development of hyphae. This study is designed to determine the functional roles of the S_Tkc domain (protein kinase) and the GAF domain of C. albicans CHK1 in hyphal formation and mucosal invasion. Methods: The domain mutants CHK25 (ΔS_Tkc CHK1/Δchk1) and CHK26 (ΔS_TkcΔgaf CHK1/Δchk1) were first constructed by the his1-URA3-his1 method and confirmed by sequencing and Southern blots. A mouse tongue infection model was used to evaluate the hyphal invasion and fungal loads in each domain mutant, full-gene deletion mutant CHK21 (chk1Δ/chk1Δ), re-constituted strain CHK23 (chk1Δ/CHK1), and wild type (WT) from day 1 to day 5. The degree of invasion and damage to the oral mucosa of mice in each strain-infected group was evaluated in vivo and compared with germ tube rate and hyphal formation in vitro. Result: When compared with severe mucosal damage and massive hyphal formation in WT- or CHK23-infected mouse tongues, the deletion of S_Tkc domain (CHK25) caused mild mucosal damage, and fungal invasion was eliminated as we observed in full-gene mutant CHK21. However, the deletion of S_Tkc and GAF (CHK26) partially restored the hyphal invasion and mucosal tissue damage that were exhibited in WT and CHK23. Regardless of the in vivo results, the decreased hyphal formation and germ tube in vitro were less apparent and quite similar between CHK25 and CHK26, especially at the late stage of the log phase where CHK26 was closer to WT and CHK23. However, growth defect and hyphal impairment of both domain mutants were similar to CHK21 in the early stages. Conclusion: Our data suggest that both protein kinase (S_Tkc) and GAF domains in C. albicans CHK1 are required for hyphal invasiveness in mucosal tissue. The appropriate initiation of cell growth and hyphal formation at the lag phase is likely mediated by these two functional domains of CHK1 to maintain in vivo infectivity of C. albicans.

Similarities and Differences: A Comparative Review of the Molecular Mechanisms and Effectors of NAFLD and AFLD.

Non-alcoholic fatty liver disease (NAFLD) and alcoholic fatty liver disease (AFLD) are the most prevalent metabolic liver diseases globally. Due to the complex pathogenic mechanisms of NAFLD and AFLD, no specific drugs were approved at present. Lipid accumulation, oxidative stress, insulin resistance, inflammation, and dietary habits are all closely related to the pathogenesis of NAFLD and AFLD. However, the mechanism that promotes disease progression has not been fully elucidated. Meanwhile, the gut microbiota and their metabolites also play an important role in the pathogenesis and development of NAFLD and AFLD. This article comparatively reviewed the shared and specific signaling pathways, clinical trials, and potential intervention effectors of NAFLD and AFLD, revealing their similarities and differences. By comparing the shared and specific molecular regulatory mechanisms, this paper provides mutual reference strategies for preventing and treating NAFLD, AFLD, and related metabolic diseases. Furthermore, it provides enlightenment for discovering novel therapies of safe and effective drugs targeting the metabolic liver disease.

Berberine reverses multidrug resistance in Candida albicans by hijacking the drug efflux pump Mdr1p.

Clinical use of antimicrobials faces great challenges from the emergence of multidrug-resistant pathogens. The overexpression of drug efflux pumps is one of the major contributors to multidrug resistance (MDR). Reversing the function of drug efflux pumps is a promising approach to overcome MDR. In the life-threatening fungal pathogen Candida albicans, the major facilitator superfamily (MFS) transporter Mdr1p can excrete many structurally unrelated antifungals, leading to MDR. Here we report a counterintuitive case of reversing MDR in C. albicans by using a natural product berberine to hijack the overexpressed Mdr1p for its own importation. Moreover, we illustrate that the imported berberine accumulates in mitochondria and compromises the mitochondrial function by impairing mitochondrial membrane potential and mitochondrial Complex I. This results in the selective elimination of Mdr1p overexpressed C. albicans cells. Furthermore, we show that berberine treatment can prolong the mean survival time of mice with blood-borne dissemination of Mdr1p overexpressed multidrug-resistant candidiasis. This study provides a potential direction of novel anti-MDR drug discovery by screening for multidrug efflux pump converters.

Mitochondrial Complex I Core Protein Regulates cAMP Signaling via Phosphodiesterase Pde2 and NAD Homeostasis in Candida albicans.

The cyclic adenosine 3',5'-monophosphate (cAMP)/protein kinase A (PKA) pathway of Candida albicans responds to nutrient availability to coordinate a series of cellular processes for its replication and survival. The elevation of cAMP for PKA signaling must be both transitory and tightly regulated. Otherwise, any abnormal cAMP/PKA pathway would disrupt metabolic potential and ergosterol synthesis and promote a stress response. One possible mechanism for controlling cAMP levels is direct induction of the phosphodiesterase PDE2 gene by cAMP itself. Our earlier studies have shown that most single-gene-deletion mutants of the mitochondrial electron transport chain (ETC) complex I (CI) are hypersensitive to fluconazole. To understand the fluconazole hypersensitivity observed in these mutants, we focused upon the cAMP/PKA-mediated ergosterol synthesis in CI mutants. Two groups of the ETC mutants were used in this study. Group I includes CI mutants. Group II is composed of CIII and CIV mutants; group II mutants are known to have greater respiratory loss. All mutants are not identical in cAMP/PKA-mediated ergosterol response. We found that ergosterol levels are decreased by 47.3% in the ndh51Δ (CI core subunit mutant) and by 23.5% in goa1Δ (CI regulator mutant). Both mutants exhibited a greater reduction of cAMP and excessive trehalose production compared with other mutants. Despite the normal cAMP level, ergosterol content decreased by 33.0% in the CIII mutant qce1Δ as well, thereby displaying a cAMP/PKA-independent ergosterol response. While the two CI mutants have some unique cAMP/PKA-mediated ergosterol responses, we found that the degree of cAMP reduction correlates linearly with a decrease in total nicotinamide adenine dinucleotide (NAD) levels in all mutants, particularly in the seven CI mutants. A mechanism study demonstrates that overactive PDE2 and cPDE activity must be the cause of the suppressive cAMP-mediated ergosterol response in the ndh51Δ and goa1Δ. While the purpose of this study is to understand the impact of ETC proteins on pathogenesis-associated cellular events, our results reveal the importance of Ndh51p in the regulation of the cAMP/PKA pathway through Pde2p inhibition in normal physiological environments. As a direct link between Ndh51p and Pde2p remains elusive, we suggest that Ndh51p participates in NAD homeostasis that might regulate Pde2p activity for the optimal cAMP pathway state.

Treg-inducing capacity of genomic DNA of Bifidobacterium longum subsp. infantis.

Background: Allergic and autoimmune diseases comprise a group of inflammatory disorders caused by aberrant immune responses in which CD25+ forkhead box P3-positive regulatory T cells (Treg) cells that normally suppress inflammatory events are often poorly functioning. This has stimulated an intensive investigative effort to find ways of increasing Tregs as a method of therapy for these conditions. Commensal microbiota known to have health benefits in humans include the lactic acid-producing, probiotic bacteria B. longum subsp. infantis and Lactobacillus rhamnosus. Mechanistically, several mechanisms have been proposed to explain how probiotics may favorably affect host immunity, including the induction of Tregs. Analysis of emerging data from several laboratories, including our own, suggest that DNA methylation may be an important determinant of immune reactivity responsible for Treg induction. Although methylated CpG moieties in normal mammalian DNA are both noninflammatory and lack immunogenicity, unmethylated CpGs, found largely in microbial DNA, are immunostimulatory and display proinflammatory properties. Objective: We hypothesize that microbiota with more DNA methylation may potentiate Treg induction to a greater degree than microbiota with a lower content of methylation. The purpose of the present study was to test this hypothesis by studying the methylation status of whole genomic DNA (gDNA) and the Treg-inducing capacity of purified gDNA in each of the probiotic bacteria B. longum subsp. infantis and L. rhamnosus, and a pathogenic Escherichia coli strain B. Results: We showed that gDNA from B. longum subsp. infantis is a potent Treg inducer that displays a dose-dependent response pattern at a dose threshold of 20 µg of gDNA. No similar Treg-inducing responses were observed with the gDNA from L. rhamnosus or E. coli. We identified a unique CpG methylated motif in the gDNA sequencing of B. longum subsp. infantis which was not found in L. rhamnosus or E. coli strain B. Conclusion: Although the literature indicates that both B. longum subsp. infantis and L. rhamnosus strains contribute to health, our data suggest that they do so by different mechanisms. Further, because of its small molecular size, low cost, ease of synthesis, and unique Treg-inducing feature, this methylated CpG oligodeoxynucleotide (ODN) from B. longum would offer many attractive features for an ideal novel therapeutic vaccine candidate for the treatment of immunologic diseases, such as the allergic and autoimmune disorders, in which Treg populations are diminished.

The antifungal pipeline: the need is established. Are there new compounds?

Our review summarizes and compares the temporal development (eras) of antifungal drug discovery as well as antibacterial ventures. The innovation gap that occurred in antibacterial discovery from 1960 to 2000 was likely due to tailoring of existing compounds to have better activity than predecessors. Antifungal discovery also faced innovation gaps. The semi-synthetic antibiotic era was followed closely by the resistance era and the heightened need for new compounds and targets. With the immense contribution of comparative genomics, antifungal targets became part of the discovery focus. These targets by definition are absolutely required to be fungal- or even lineage (clade) specific. Importantly, targets need to be essential for growth and/or have important roles in disease and pathogenesis. Two types of antifungals are discussed that are mostly in the FDA phase I-III clinical trials. New antifungals are either modified to increase bioavailability and stability for instance, or are new compounds that inhibit new targets. One of the important developments in incentivizing new antifungal discovery has been the prolific number of publications of global and country-specific incidence. International efforts that champion global antimicrobial drug discovery are discussed. Still, interventions are needed. The current pipeline of antifungals and alternatives to antifungals are discussed including vaccines.

Unique, Diverged, and Conserved Mitochondrial Functions Influencing Candida albicans Respiration.

Candida albicans is an opportunistic fungal pathogen of major clinical concern. The virulence of this pathogen is intimately intertwined with its metabolism. Mitochondria, which have a central metabolic role, have undergone many lineage-specific adaptations in association with their eukaryotic host. A screen for lineage-specific genes identified seven such genes specific to the CTG clade of fungi, of which C. albicans is a member. Each is required for respiratory growth and is integral to expression of complex I, III, or IV of the electron transport chain. Two genes, NUO3 and NUO4, encode supernumerary subunits of complex I, whereas NUE1 and NUE2 have nonstructural roles in expression of complex I. Similarly, the other three genes have nonstructural roles in expression of complex III (QCE1) or complex IV (COE1 and COE2). In addition to these novel additions, an alternative functional assignment was found for the mitochondrial protein encoded by MNE1MNE1 was required for complex I expression in C. albicans, whereas the distantly related Saccharomyces cerevisiae ortholog participates in expression of complex III. Phenotypic analysis of deletion mutants showed that fermentative metabolism is unable to support optimal growth rates or yields of C. albicans However, yeast-hypha morphogenesis, an important virulence attribute, did not require respiratory metabolism under hypoxic conditions. The inability to respire also resulted in hypersensitivity to the antifungal fluconazole and in attenuated virulence in a Galleria mellonella infection model. The results show that lineage-specific adaptations have occurred in C. albicans mitochondria and highlight the significance of respiratory metabolism in the pathobiology of C. albicansIMPORTANCECandida albicans is an opportunistic fungal pathogen of major clinical concern. The virulence of this pathogen is intimately intertwined with its metabolic behavior, and mitochondria have a central role in that metabolism. Mitochondria have undergone many evolutionary changes, which include lineage-specific adaptations in association with their eukaryotic host. Seven lineage-specific genes required for electron transport chain function were identified in the CTG clade of fungi, of which C. albicans is a member. Additionally, examination of several highly diverged orthologs encoding mitochondrial proteins demonstrated functional reassignment for one of these. Deficits imparted by deletion of these genes revealed the critical role of respiration in virulence attributes of the fungus and highlight important evolutionary adaptations in C. albicans metabolism.

Interplay of interferon-gamma and macrophage polarization during Talaromyces marneffei infection.

Talaromyces marneffei is an increasingly destructive dimorphic fungal pathogen in clinical settings that can cause lethal Talaromycosis. The activation of macrophages is known to be important for host defenses against T. marneffei, and these macrophages are known to be activated in two ways (polarization), known as M1 and M2. We investigated the plasticity of these polarizations, in order to understand if cross-conversion of macrophages may be possible even after they have been programmed. We conducted in vitro experiments using a murine macrophage cell line to investigate the ability of T. marneffei to activate these polarizations. The pre-polarized (M0) macrophage subsets were challenged with LPS as a control, and the sets of M1 markers (iNOS and CD86) and M2 markers (Arg-1 and CD206) were assessed for a possible cross-conversion among M1, M2 and M0 (unstimulated) populations. We found that either conidia or yeast forms of T. marneffei initiate the repression of Arg-1 in M2 cells with no change in the M1 subtype marker molecule iNOS. However, an additional IFN-γ stimulus caused the three macrophage groups to fully exhibit an LPS-induced M2 suppression and a shift to M1 from M0 and M2. We conclude that the conversion of macrophages is required for maintenance of sufficient iNOS production against this organism in the host. The cytokine environment is the key factor that manipulates the plasticity changes among macrophage subtypes. Furthermore, IFN-γ is a crucial host defense factor against pathogenic T. marneffei that has significant therapeutic potential to promote an M1 polarization phenotype.

Vitiligo and Hashimoto's thyroiditis: Autoimmune diseases linked by clinical presentation, biochemical commonality, and autoimmune/oxidative stress-mediated toxicity pathogenesis.

Vitiligo (VL) is a chronic autoimmune pigmentation disorder characterized by destruction of melanocytes. The condition is associated with several other autoimmune diseases, but autoimmune thyroid diseases, especially Hashimoto's thyroiditis (HT), is the most prevalent organ-specific autoimmune disease with a co-morbidity up to 34%. Among the many hypotheses that have been proposed for the pathogenesis of both diseases, autoimmunity and oxidative stress-mediated toxicity in melanocytes or thyrocytes, respectively, have been the most widely accepted - with autoimmunity being the presumed consequence of oxidative stress-mediated toxicity. However, the predominant etiologic basis for impairment of redox balance has rarely been studied. The two autoimmune diseases are not only linked by a concordance of clinical presentations and an autoimmune/oxidative stress-mediated toxicity pathogenesis but also by an apparent biochemical commonality. The target molecules produced in the thyroid and skin, i.e., thyroxine and melanin, respectively, are derived from the same primordial parent molecule, tyrosine. On the basis of these similarities between Hashimoto's thyroiditis and vitiligo, specifically with respect to the activation of oxidative stress, we propose a novel hypothesis accounting for the destruction of melanocytes or thyrocytes in VL and AT. We suggest a new therapeutic regimen of quinone derivatives to combat ROS-induced autoimmunity resulting from this common biochemical etiologic error.

Requirements for Transparency and Communicability of Regulatory Science.

This article presents the results of a study attempting to provide examples that implement transparency and communicability elements of Ethical Rules Principle of Best Available Regulatory Science (BARS) and Metrics for Evaluation of Regulatory Science Claims (MERSC). It starts with an overview of regulatory science and briefly summarizes principles of BARS and key pillars of MERSC. Subsequently, the BARS/MERSC system is used to evaluate the linear nonthreshold (LNT) process used in cancer assessments and the similar process used for evaluating in particulate matter (PM) exposure. The study identifies 3 parts in dose-response curves, where the first part is reproducible science and the second part includes uncertainties and often requires the application of precautionary principle. The primary reason for disagreements on LNT and PM is a lack of recognition that the third part is based on desire of regulators to be protective, a policy decision process. Two PM epidemiological examples are included in this study to demonstrate the point. The regulatory process would benefit from recognizing the distinction between science and policy and excluding policy from regulatory science. Furthermore, the society would greatly benefit from increased transparency in the regulatory process and compliance with the Jeffersonian communication principle.

Methodologies for in vitro and in vivo evaluation of efficacy of antifungal and antibiofilm agents and surface coatings against fungal biofilms.

Unlike superficial fungal infections of the skin and nails, which are the most common fungal diseases in humans, invasive fungal infections carry high morbidity and mortality, particularly those associated with biofilm formation on indwelling medical devices. Therapeutic management of these complex diseases is often complicated by the rise in resistance to the commonly used antifungal agents. Therefore, the availability of accurate susceptibility testing methods for determining antifungal resistance, as well as discovery of novel antifungal and antibiofilm agents, are key priorities in medical mycology research. To direct advancements in this field, here we present an overview of the methods currently available for determining (i) the susceptibility or resistance of fungal isolates or biofilms to antifungal or antibiofilm compounds and compound combinations; (ii) the in vivo efficacy of antifungal and antibiofilm compounds and compound combinations; and (iii) the in vitro and in vivo performance of anti-infective coatings and materials to prevent fungal biofilm-based infections.

Regulating Ionizing Radiation Based on Metrics for Evaluation of Regulatory Science Claims.

This article attempts to reconcile differences within the relevant scientific community on the effect of exposure to low levels of ionizing radiation notably the applicability of linear nonthreshold (LNT) process at exposures below a certain limit. This article applies an updated version of Metrics for Evaluation of Regulatory Science Claims (MERSC) derived form Best Available Regulatory Science (BARS) to the arguments provided by the proponents and opponents of LNT. Based on BARS/MERSC, 3 categories of effects of exposure to ionizing radiation are identified. One category (designated as S) consists of reproducible and undisputed adverse effects. A second category (designated as U) consists of areas where the scientific evidence for potential adverse effects includes uncertainties. The scientific evidence in the U category leads to a threshold. In contrast, the scientific foundation of the third category (designated as P) is questionable, as the scientific evidence indicates that adverse effects of the exposure at this level are not only questionable but may be helpful. This article claims that the third area is the domain of policy makers including regulators. This article describes Jeffersonian Principle that categorizes the affected community into specialists, knowledgeable nonspecialists, and the general public. Based on Jeffersonian Principle, the relevant scientific information, particularly the U and P areas, must be translated into a language that at a minimum is understandable to the knowledgeable group. Once this process is completed, the policy makers including regulators may select exposure limits based on their judgment.

A mitochondrial proteomics view of complex I deficiency in Candida albicans.

Proteomic analyses were carried out on isolated mitochondrial samples of C. albicans from gene-deleted mutants (nuo1Δ, nuo2Δ and goa1Δ) as well as the parental strain in order to better understand the contribution of these three fungal-specific mitochondrial ETC complex I (CI) subunits to cellular activities. Herein, we identify 2333 putative proteins from four strains, in which a total of 663 proteins (28.5%) are putatively located in mitochondria. Comparison of protein abundances between mutants and the parental strain reveal 146 differentially-expressed proteins, of which 78 are decreased and 68 are increased in at least one mutant. The common changes across the three mutants include the down-regulation of nuclear-encoded CI subunit proteins as well as phospholipid, ergosterol and cell wall mannan synthesis, and up-regulated proteins in CIV and the alternative oxidase (AOX2). As for gene-specific functions, we find that NUO1 participates in nucleotide synthesis and ribosomal biogenesis; NUO2 is involved in vesicle trafficking; and GOA1 appears to regulate membrane transporter proteins, ROS removal, and substrates trafficking between peroxisomes and mitochondria. The proteomic view of general as well as mutant-specific proteins further extends our understanding of the functional roles of non-mammalian CI-specific subunit proteins in cell processes. Particularly intriguing is the confirmation of a regulatory role for GOA1 on ETC function, a protein found almost exclusively in Candida species. SIGNIFICANCE: Fungal mitochondria are critical for fungal pathogenesis. The absence of any of the three fungal specific CI subunits in mitochondria causes an avirulence phenotype of C. albicans in a murine model of invasive disease. As model yeast (Saccharomyces cerevisiae) lacks a CI and is rarely a pathogen of humans, C. albicans is a better choice for establishing a link between mitochondrial CI and pathogenesis. Apart from the general effects of CI mutants on respiration, previous phenotyping of these mutants were quite similar to each other or to CI conservative subunit. By comparison to transcriptional data, the proteomic data obtained in this study indicate that biosynthetic events in each mutant such as cell wall and cell membrane phospholipids and ergosterol are generally decreased in both transcriptomal and translational levels. However, in the case of mitochondrial function, glycolysis/gluconeogenesis, and ROS scavengers, often gene changes are opposite that of proteomic data in mutants. We hypothesize that the loss of energy production in mutants is compensated by increases in protein levels of glycolysis, gluconeogenesis, and anti-ROS scavengers that at least extend mutant survival.