Synthesis, In Silico Study, and In Vitro Antifungal Activity of New 5-(1,3-Diphenyl-1H-Pyrazol-4-yl)-4-Tosyl-4,5-Dihydrooxazoles.

The increase in multi-drug resistant Candida strains has caused a sharp rise in life-threatening fungal infections in immunosuppressed patients, including those with SARS-CoV-2. Novel antifungal drugs are needed to combat multi-drug-resistant yeasts. This study aimed to synthesize a new series of 2-oxazolines and evaluate the ligands in vitro for the inhibition of six Candida species and in silico for affinity to the CYP51 enzymes (obtained with molecular modeling and protein homology) of the same species. The 5-(1,3-diphenyl-1H-pyrazol-4-yl)-4-tosyl-4,5-dihydrooxazoles 6a-j were synthesized using the Van Leusen reaction between 1,3-diphenyl-4-formylpyrazoles 4a-j and TosMIC 5 in the presence of K2CO3 or KOH without heating, resulting in short reaction times, high compound purity, and high yields. The docking studies revealed good affinity for the active site of the CYP51 enzymes of the Candida species in the following order: 6a-j > 4a-j > fluconazole (the reference drug). The in vitro testing of the compounds against the Candida species showed lower MIC values for 6a-j than 4a-j, and for 4a-j than fluconazole, thus correlating well with the in silico findings. According to growth rescue assays, 6a-j and 4a-j (like fluconazole) inhibit ergosterol synthesis. The in silico toxicity assessment evidenced the safety of compounds 6a-j, which merit further research as possible antifungal drugs.

Plant growth-promoting and heavy metal-resistant Priestia and Bacillus strains associated with pioneer plants from mine tailings.

Open mine tailings dams are extreme artificial environments containing sizeable potentially toxic elements (PTEs), including heavy metals (HMs), transition metals, and metalloids. Furthermore, these tailings have nutritional deficiencies, including assimilable phosphorus sources, organic carbon, and combined nitrogen, preventing plant colonization. Bacteria, that colonize these environments, have mechanisms to tolerate the selective pressures of PTEs. In this work, several Priestia megaterium (formerly Bacillus megaterium), Bacillus mojavensis, and Bacillus subtilis strains were isolated from bulk tailings, anthills, rhizosphere, and endosphere of pioneer plants from abandoned mine tailings in Zacatecas, Mexico. Bacillus spp. tolerated moderate HMs concentrations, produced siderophores and indole-3-acetic acid (IAA), solubilized phosphates, and reduced acetylene in the presence of HMs. The strains harbored different PIB-type ATPase genes encoding for efflux pumps and Cation Diffusion Facilitator (CDF) genes. Moreover, nifH and nifD nitrogenase genes were detected in P. megaterium and B. mojavensis genomic DNA. They showed similarity with sequences of the beta-Proteobacteria species, which may represent likely horizontal transfer events. These Bacillus species precede the colonization of mine tailings by plants. Their phenotypic and genotypic features could be essential in the natural recovery of the sites by reducing the oxidative stress of HMs, fixing nitrogen, solubilizing phosphate, and accumulating organic carbon. These traits of the strains reflect the adaptations of Bacillus species to the mine tailings environment and could contribute to the success of phytoremediation efforts.

Inhibitors of 3-Hydroxy-3-methylglutaryl Coenzyme A Reductase Decrease the Growth, Ergosterol Synthesis and Generation of petite Mutants in Candida glabrata and Candida albicans.

Candida glabrata and Candida albicans, the most frequently isolated candidiasis species in the world, have developed mechanisms of resistance to treatment with azoles. Among the clinically used antifungal drugs are statins and other compounds that inhibit 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), resulting in decreased growth and ergosterol levels in yeasts. Ergosterol is a key element for the formation of the yeast cell membrane. However, statins often cause DNA damage to yeast cells, facilitating mutation and drug resistance. The aim of the current contribution was to synthesize seven series of compounds as inhibitors of the HMGR enzyme of Candida ssp., and to evaluate their effect on cellular growth, ergosterol synthesis and generation of petite mutants of C. glabrata and C. albicans. Compared to the reference drugs (fluconazole and simvastatin), some HMGR inhibitors caused lower growth and ergosterol synthesis in the yeast species and generated fewer petite mutants. Moreover, heterologous expression was achieved in Pichia pastoris, and compounds 1a, 1b, 6g and 7a inhibited the activity of recombinant CgHMGR and showed better binding energy values than for α-asarone and simvastatin. Thus, we believe these are good candidates for future antifungal drug development.

Pyrrole-Based Enaminones as Building Blocks for the Synthesis of Indolizines and Pyrrolo[1,2-a]pyrazines Showing Potent Antifungal Activity.

As a new approach, pyrrolo[1,2-a]pyrazines were synthesized through the cyclization of 2-formylpyrrole-based enaminones in the presence of ammonium acetate. The enaminones were prepared with a straightforward method, reacting the corresponding alkyl 2-(2-formyl-1H-pyrrol-1-yl)acetates, 2-(2-formyl-1H-pyrrol-1-yl)acetonitrile, and 2-(2-formyl-1H-pyrrol-1-yl)acetophenones with DMFDMA. Analogous enaminones elaborated from alkyl (E)-3-(1H-pyrrol-2-yl)acrylates were treated with a Lewis acid to afford indolizines. The antifungal activity of the series of substituted pyrroles, pyrrole-based enaminones, pyrrolo[1,2-a]pyrazines, and indolizines was evaluated on six Candida spp., including two multidrug-resistant ones. Compared to the reference drugs, most test compounds produced a more robust antifungal effect. Docking analysis suggests that the inhibition of yeast growth was probably mediated by the interaction of the compounds with the catalytic site of HMGR of the Candida species.

Intracellular Aminopeptidase Activity Determination from the Fungus Sporisorium reilianum: Purification and Biochemical Characterization of psrAPEi Enzyme.

The aims of this study were to, first, determine the intracellular aminopeptidase activity (APEi) and second, purify and biochemically characterize one intracellular aminopeptidase enzyme from the phytopathogen fungus Sporisorium reilianum (psrAPEi), the causal agent of head smut in corn. The fungus produced APEi activity in all media cultures evaluated. The psrAPEi was purified by a procedure that involved ammonium sulfate fractionation and four chromatographic steps using an FPLC system (Fast Protein Liquid Chromatography). Results showed an estimated molecular mass of 52.2 kDa. Enzymatic activity was optimal at pH 7.0 and 35 °C and was inhibited by EDTA-Na2, 1,10-phenanthroline, bestatin, and PMSF. This aminopeptidase showed a preference for leucine, arginine, and lysine at the N-position. The Km and Vmax values were 3.72 µM and 188.0 µmol/min, respectively, for L-lysyl-4-nitroanilide. This is the first study to report on intracellular aminopeptidase activity in S. reilianum and the purification and characterization of an intracellular metallo-serine-aminopeptidase (psrAPEi).

Molecular Recognition of Citroflavonoids Naringin and Naringenin at the Active Site of the HMG-CoA Reductase and DNA Topoisomerase Type II Enzymes of Candida spp. and Ustilago maydis.

Two agents from natural sources, citroflavonoids naringin and naringenin, can target enzymes in pathogenic yeasts responsible for hospital infections and crop failure. The aim of this study was to examine the molecular recognition site for naringin and naringenin on the HMGR and TOPOII enzymes of eleven Candida species and one phytopathogen, U. maydis, and evaluate yeast susceptibility to these flavonoids. The HMGR and TOPOII enzymes were analyzed in silico. The alignment of the two enzymes in the twelve pathogenic organisms with the corresponding enzyme of Homo sapiens revealed highly conserved amino acid sequences. Modeling studies of the enzymes indicated highly conserved structures. According to molecular docking simulations, both citroflavonoids recognized the amino acid residues of the active site of the enzymes. Binding energy values were higher for naringin (-10.75 and -9.38 kcal/mol, respectively) than simvastatin on HMGR (-9.9) and curcumin on TOPOII (-8.37). The appraisal of twenty-nine virtual mutations provided evidence of probable mechanisms of resistance (high binding energy) or susceptibility (low energy) to the drugs and emphasized the role of key residues. An in vitro susceptibility evaluation of the twelve yeasts demonstrated that the two flavonoids have similar or better MIC values than those reported for the reference compounds, obtaining the lowest with Candida dubliniensis (2.5 µg/ml) and U. maydis (5 µg/ml). Based on the present findings, naringin and naringenin could possibly be effective for treating diseases caused by pathogenic yeasts of the Candida species and U. maydis, presumably by inhibition of their HMGR and TOPOII enzymes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12088-021-00980-0.

Inhibitors of DNA topoisomerases I and II applied to Candida dubliniensis reduce growth, viability, the generation of petite mutants and toxicity, while acting synergistically with fluconazole.

The increasing resistance of Candida species to azoles emphasizes the urgent need for new antifungal agents with novel mechanisms of action. The aim of this study was to examine the effect of three DNA topoisomerase inhibitors of plant origin (camptothecin, etoposide and curcumin) on the growth of Candida dubliniensis. The phylogenetic analysis showed a close relationship between the topoisomerase enzymes of C. dubliniensis and Candida albicans. The alignment of the amino acid sequences of topoisomerase I and II of yeasts and humans evidenced conserved domains. The docking study revealed affinity of the test compounds for the active site of topoisomerase I and II in C. dubliniensis. Curcumin and camptothecin demonstrated a stronger in vitro antifungal effect than the reference drugs (fluconazole and itraconazole). Significant synergistic activity between the topoisomerase inhibitors and fluconazole at the highest concentration (750 µM) was observed. Fluconazole induced the petite phenotype to a greater degree than the topoisomerase inhibitors, indicating a tendency to generate resistance. Lower toxicity was found for such inhibitors versus reference drugs on Galleria mellonella larva. The topoisomerase inhibitors exhibited promising antifungal activity, and the DNA topoisomerase enzymes of C. dubliniensis proved to be an excellent model for evaluating new antifungal compounds.

Candida pseudoglaebosa and Kodamaea ohmeri are capable of degrading alkanes in the presence of heavy metals.

The aim of this study was to examine four strains of two yeast species in relation to their capability for assimilating alkanes in the presence of heavy metals (HMs). The four strains tested were Candida pseudoglaebosa ENCB-7 and Kodamaea ohmeri ENCB-8R, ENCB-23, and ENCB-VIK. Determination was made of the expression of CYP52 genes involved in alkane hydroxylation. When exposed to Cu2+ , Zn2+ , Pb2+ , Cd2+ , and As3+ at pH 3 and 5, all four strains could assimilate several n-alkanes having at least six carbon atoms. The three K. ohmeri strains could also utilize branched alkanes, cycloalkanes, and n-octanol as sole carbon sources. Kinetic assays demonstrated greater biomass production and specific growth of the yeasts exposed to long-chain n-alkanes. Fragments of paralogous CYP52 genes of C. pseudoglaebosa ENCB-7 and K. ohmeri ENCB-23 were amplified, sequenced, and phylogenetically evaluated. Reverse-transcription polymerase chain reaction revealed that n-nonane and n-decane induced to CpCYP52-G3, CpCYP52-G9, and CpCYP52-G10. KoCYP52-G3 was induced with n-decane and n-octanol. Also, CpCYP52-G3 and CpCYP52-G9 were induced by glucose. In conclusion, C. pseudoglaebosa and K. ohmeri were able to degrade several alkanes in the presence of HMs and under acidic conditions. These yeasts harbor paralogous alkane-induced CYP52 genes, which display different profiles of transcriptional expression.

Candida pseudoglaebosa and Kodamaea ohmeri are capable of degrading alkanes in the presence of heavy metals.

The aim of this study was to examine four strains of two yeast species in relation to their capability for assimilating alkanes in the presence of heavy metals (HMs). The four strains tested were Candida pseudoglaebosa ENCB-7 and Kodamaea ohmeri ENCB-8R, ENCB-23, and ENCB-VIK. Determination was made of the expression of CYP52 genes involved in alkane hydroxylation. When exposed to Cu2+ , Zn2+ , Pb2+ , Cd2+ , and As3+ at pH 3 and 5, all four strains could assimilate several n-alkanes having at least six carbon atoms. The three K. ohmeri strains could also utilize branched alkanes, cycloalkanes, and n-octanol as sole carbon sources. Kinetic assays demonstrated greater biomass production and specific growth of the yeasts exposed to long-chain n-alkanes. Fragments of paralogous CYP52 genes of C. pseudoglaebosa ENCB-7 and K. ohmeri ENCB-23 were amplified, sequenced, and phylogenetically evaluated. Reverse-transcription polymerase chain reaction revealed that n-nonane and n-decane induced to CpCYP52-G3, CpCYP52-G9, and CpCYP52-G10. KoCYP52-G3 was induced with n-decane and n-octanol. Also, CpCYP52-G3 and CpCYP52-G9 were induced by glucose. In conclusion, C. pseudoglaebosa and K. ohmeri were able to degrade several alkanes in the presence of HMs and under acidic conditions. These yeasts harbor paralogous alkane-induced CYP52 genes, which display different profiles of transcriptional expression.

Activity and expression of Candida glabrata vacuolar proteases in autophagy-like conditions.

Candida glabrata is an emerging opportunistic pathogen that has intrinsic resistance to azoles. During infection or while living as a commensal, it encounters nutritional stresses such as deficiency of carbon or nitrogen sources. Herein, we investigate the expression and activity of PrA, Ape1, Ape3 and CpY vacuolar proteases during these stressful nutrimental conditions. Our findings demonstrate a differential activity profile depending on the addition or lack of carbon, nitrogen or both. Of the four proteases tested, PrA and Ape3 showed a higher activity in the absence of nitrogen. Steady-state RNA levels for all the proteases were also differentially expressed although not always correlated with its activity, suggesting multiple levels of regulation. Microscopy observations of C. glabrata cells subjected to the different conditions showed an increase in the vacuolar volume. Moreover, the presence of ATG8-PE and an increased expression of ATG8 were observed in the yeast under the tested conditions suggesting that C. glabrata is in autophagy stage. Taken together, our results showed that PrA, Ape1, Ape3 and CpY have varying activities and expression depending on whether nitrogen or carbon is added to the media, and that these vacuolar proteases might have a role in the autophagy process.

Cyanotrophic and arsenic oxidizing activities of Pseudomonas mendocina P6115 isolated from mine tailings containing high cyanide concentration.

Mine tailings and wastewater generate man-made environments with several selective pressures, including the presence of heavy metals, arsenic and high cyanide concentrations, but severe nutritional limitations. Some oligotrophic and pioneer bacteria can colonise and grow in mine wastes containing a low concentration of organic matter and combined nitrogen sources. In this study, Pseudomonas mendocina P6115 was isolated from mine tailings in Durango, Mexico, and identified through a phylogenetic approach of 16S rRNA, gyrB, rpoB, and rpoD genes. Cell growth, cyanide consumption, and ammonia production kinetics in a medium with cyanide as sole nitrogen source showed that at the beginning, the strain grew assimilating cyanide, when cyanide was removed, ammonium was produced and accumulated in the culture medium. However, no clear stoichiometric relationship between both nitrogen sources was observed. Also, cyanide complexes were assimilated as nitrogen sources. Other phenotypic tasks that contribute to the strain's adaptation to a mine tailing environment included siderophores production in media with moderate amounts of heavy metals, arsenite and arsenate tolerance, and the capacity of oxidizing arsenite. P. mendocina P6115 harbours cioA/cioB and aoxB genes encoding for a cyanide-insensitive oxidase and an arsenite oxidase, respectively. This is the first report where P. mendocina is described as a cyanotrophic and arsenic oxidizing species. Genotypic and phenotypic tasks of P. mendocina P6115 autochthonous from mine wastes are potentially relevant for biological treatment of residues contaminated with cyanide and arsenic.

Inhibition of dengue virus infection by small interfering RNAs that target highly conserved sequences in the NS4B or NS5 coding regions.

Dengue fever is one of the most common viral infections in the world. Although a vaccine against dengue virus (DENV) has been approved in several countries, this disease is still considered a public health priority worldwide. The ability of three small interfering RNAs (FG-siRNAs) targeting conserved sequences in the NS4B and NS5 regions of the DENV genome to inhibit DENV replication was tested in vitro in both Vero and C6/36 cells. The FG-siRNAs were effective against DENV-1, -3, and -4, but not DENV-2. A fourth siRNA specifically targeting the NS5 region of the DENV-2 genome (SG-siRNA) was designed and tested against two different DENV-2 strains, showing high levels of inhibition in both mammalian and insect cells.

Heterologous expression and characterization of the aspartic endoprotease Pep4um from Ustilago maydis, a homolog of the human Chatepsin D, an important breast cancer therapeutic target.

The pep4um gene (um04926) of Ustilago maydis encodes a protein related to either vacuolar or lysosomal aspartic proteases. Bioinformatic analysis of the Pep4um protein revealed that it is a soluble protein with a signal peptide suggesting that it likely passes through the secretory pathway, and it has two probable self-activation sites, which are similar to those in Saccharomyces cerevisiae PrA. Moreover, the active site of the Pep4um has the two characteristic aspartic acid residues of aspartyl proteases. The pep4um gene was cloned, expressed in Pichia pastoris and a 54 kDa recombinant protein was observed. Pep4um-rec was confirmed to be an aspartic protease by specifically inhibiting its enzymatic activity with pepstatin A. Pep4um-rec enzymatic activity on acidic hemoglobin was optimal at pH 4.0 and at 40 °C. To the best of our knowledge this is the first report about the heterologous expression of an aspartic protease from a basidiomycete. An in-depth in silico analysis suggests that Pep4um is homolog of the human cathepsin D protein. Thus, the Pep4um-rec protein may be used to test inhibitors of human cathepsin D, an important breast cancer therapeutic target.

Dibutyltin(IV) Complexes Derived from L-DOPA: Synthesis, Molecular Docking, Cytotoxic and Antifungal Activity.

A series of organotin(IV) complexes was herein prepared and characterized. A one-pot synthetic strategy afforded reasonable to high yields, depending on the nature of the ligand. All new complexes were fully characterized by spectroscopic techniques, consisting of IR, MS and NMR (1H, 13C and 119Sn). The in vitro cytotoxicity tests demonstrated that the organotin complexes produced a greater inhibition, versus cisplatin (the positive control), of the growth of six human cancer cell lines: U-251 (glioblastoma), K-562 (chronic myelogenous leukemia), HCT-15 (colorectal), MCF-7 (breast), MDA-MB-231 (breast) and SKLU-1 (non-small cell lung). The potency of this cytotoxic activity depended on the nature of the substituent bonded to the aromatic ring. All complexes exhibited excellent IC50 values. The test compounds were also screened in vitro for their antifungal effect against Candida glabrata and Candida albicans, showing minimum inhibitory concentration (MIC) values lower than those obtained for fluconazole. A brine shrimp bioassay was performed to examine the toxic properties. Molecular docking studies demonstrated that the organotin(IV) complexes bind at the active site of topoisomerase I in a similar manner to topotecan, sharing affinity for certain amino acid side chains (Ile535, Arg364 and Asp533), as well as for similar DNA regions (DA113, DC112 and DT10).

Synthesis, Molecular Docking, and Antimycotic Evaluation of Some 3-Acyl Imidazo[1,2-a]pyrimidines.

A series of 3-benzoyl imidazo[1,2-a]pyrimidines, obtained from N-heteroarylformamidines in good yields, was tested in silico and in vitro for binding and inhibition of seven Candida species (Candida albicans (ATCC 10231), Candida dubliniensis (CD36), Candida glabrata (CBS138), Candida guilliermondii (ATCC 6260), Candida kefyr, Candida krusei (ATCC 6358) and Candida tropicalis (MYA-3404)). To predict binding mode and energy, each compound was docked in the active site of the lanosterol 14α-demethylase enzyme (CYP51), essential for fungal growth of Candida species. Antimycotic activity was evaluated as the 50% minimum inhibitory concentration (MIC50) for the test compounds and two reference drugs, ketoconazole and fluconazole. All test compounds had a better binding energy (range: -6.11 to -9.43 kcal/mol) than that found for the reference drugs (range: 48.93 to -6.16 kcal/mol). In general, the test compounds showed greater inhibitory activity of yeast growth than the reference drugs. Compounds 4j and 4f were the most active, indicating an important role in biological activity for the benzene ring with electron-withdrawing substituents. These compounds show the best MIC50 against C. guilliermondii and C. glabrata, respectively. The current findings suggest that the 3-benzoyl imidazo[1,2-a]pyrimidine derivatives, herein synthesized by an accessible methodology, are potential antifungal drugs.

Polymorphism in the regulatory regions of genes CgYPS1 and CgYPS7 encoding yapsins in Candida glabrata is associated with changes in expression levels.

Candida glabrata is an opportunistic fungus infecting mainly immunocompromised people. Its adherence capacity and exoenzymes contribute to damaging host cells. In particular, the yapsins are a family of aspartyl proteases involved in maturation of proteins and cell wall function, and yapsins 1 and 7, respectively encoded by genes CgYPS1 and CgYPS7, are potential virulence factors. In this study, the polymorphism of regulatory regions and the expression profiles of both genes were compared in C. glabrata clinical strains. The sequence analysis of regulatory regions revealed that the distribution of transcription factor binding sites (TFBSs) was similar, although some TFBSs were not universally distributed. The quantita-tive expression of CgYPS1 and CgYPS7 genes of different C. glabrata strains in rich and poor media was estimated by RT-qPCR. The primary sequences of genes CgYPS1 and CgYPS7 of C. glabrata strains were highly conserved among different strains, but the regulatory regions were polymorphic, harboring different TFBS arrays, and showing differential expression profiles.

Ammonia-Oligotrophic and Diazotrophic Heavy Metal-Resistant Serratia liquefaciens Strains from Pioneer Plants and Mine Tailings.

Mine tailings are man-made environments characterized by low levels of organic carbon and assimilable nitrogen, as well as moderate concentrations of heavy metals. For the introduction of nitrogen into these environments, a key role is played by ammonia-oligotrophic/diazotrophic heavy metal-resistant guilds. In mine tailings from Zacatecas, Mexico, Serratia liquefaciens was the dominant heterotrophic culturable species isolated in N-free media from bulk mine tailings as well as the rhizosphere, roots, and aerial parts of pioneer plants. S. liquefaciens strains proved to be a meta-population with high intraspecific genetic diversity and a potential to respond to these extreme conditions. The phenotypic and genotypic features of these strains reveal the potential adaptation of S. liquefaciens to oligotrophic and nitrogen-limited mine tailings with high concentrations of heavy metals. These features include ammonia-oligotrophic growth, nitrogen fixation, siderophore and indoleacetic acid production, phosphate solubilization, biofilm formation, moderate tolerance to heavy metals under conditions of diverse nitrogen availability, and the presence of zntA, amtB, and nifH genes. The acetylene reduction assay suggests low nitrogen-fixing activity. The nifH gene was harbored in a plasmid of ∼60 kb and probably was acquired by a horizontal gene transfer event from Klebsiella variicola.

Synthesis and highly potent hypolipidemic activity of alpha-asarone- and fibrate-based 2-acyl and 2-alkyl phenols as HMG-CoA reductase inhibitors.

In the search for new potential hypolipidemic agents, the present study focused on the synthesis of 2-acyl phenols (6a-c and 7a-c) and their saturated side-chain alkyl phenols (4a-c and 5a-c), and on the evaluation of their hypolipidemic activity using a murine Tyloxapol-induced hyperlipidemic protocol. The whole series of compounds 4-7 greatly and significantly reduced elevated serum levels of total cholesterol, LDL-cholesterol, and triglycerides, with series 6 and 7 showing the greatest potency ever found in our laboratory. At the minimum dose (25mg/kg/day), the latter compounds lowered cholesterol by 68-81%, LDL by 72-86%, and triglycerides by 59-80%. This represents a comparable performance than that shown by simvastatin. Experimental evidence and docking studies suggest that the activity of these derivatives is associated with the inhibition of HMG-CoA reductase.

Comparative Genomics of the First Resistant Candida auris Strain Isolated in Mexico: Phylogenomic and Pan-Genomic Analysis and Mutations Associated with Antifungal Resistance.

Candida auris is an emerging multidrug-resistant and opportunistic pathogenic yeast. Whole-genome sequencing analysis has defined five major clades, each from a distinct geographic region. The current study aimed to examine the genome of the C. auris 20-1498 strain, which is the first isolate of this fungus identified in Mexico. Based on whole-genome sequencing, the draft genome was found to contain 70 contigs. It had a total genome size of 12.86 Mbp, an N50 value of 1.6 Mbp, and an average guanine-cytosine (GC) content of 45.5%. Genome annotation revealed a total of 5432 genes encoding 5515 proteins. According to the genomic analysis, the C. auris 20-1498 strain belongs to clade IV (containing strains endemic to South America). Of the two genes (ERG11 and FKS1) associated with drug resistance in C. auris, a mutation was detected in K143R, a gene located in a mutation hotspot of ERG11 (lanosterol 14-α-demethylase), an antifungal drug target. The focus on whole-genome sequencing and the identification of mutations linked to the drug resistance of fungi could lead to the discovery of new therapeutic targets and new antifungal compounds.

Interactions between the AraC/XylS-like transcriptional activator InvF of Salmonella Typhimurium, the RNA polymerase alpha subunit and the chaperone SicA.

Salmonella enterica serovar Typhimurium causes gastroenteritis and systemic infections in humans. For this bacterium the expression of a type III secretion system (T3SS) and effector proteins encoded in the Salmonella pathogenicity island-1 (SPI-1), is keystone for the virulence of this bacterium. Expression of these is controlled by a regulatory cascade starting with the transcriptional regulators HilD, HilC and RtsA that induce the expression of HilA, which then activates expression of the regulator InvF, a transcriptional regulator of the AraC/XylS family. InvF needs to interact with the chaperone SicA to activate transcription of SPI-1 genes including sicA, sopB, sptP, sopE, sopE2, and STM1239. InvF very likely acts as a classical activator; however, whether InvF interacts with the RNA polymerase alpha subunit RpoA has not been determined. Results from this study confirm the interaction between InvF with SicA and reveal that both proteins interact with the RNAP alpha subunit. Thus, our study further supports that the InvF/SicA complex acts as a classical activator. Additionally, we showed for the first time an interaction between a chaperone of T3SS effectors (SicA) and the RNAP.