Microarray of Candida albicans treated with sublethal concentrations of the antifungal Jagaricin
ABSTRACT: Candida albicans were treated with a sublethal concentration of the antifungal Jagaricin for either a short time (30 min) or until an OD of 0.5 (indicating log growth) was reached. Controls were grown without any antifungal to determine cellular reactions to the compound.
Project description:We performed a gene expression analysis of C. albicans SC5314 planktonic cells exposed to the antifungal peptide ApoEdpL-W. Exponentially-growing C. albicans SC5314 cells in SD at 30°C medium were exposed to 2.5 µM ApoEdpL-W and samples were collected after 10 and 30 min. for transcript profiling
Project description:Azole resistance and varying degrees of cross-resistance to other members of the azole family in clinical isolates have been documented, which has necessitated additional and prolonged use of the antifungal agents available. 2-Amino-Nonyl-6-Methoxyl-Tetralin Muriate (10b), a novel chemical structural aminotetralin derivate, is synthesized as an antifungal agent and exibited strong antifungal activity. To further investigated the action mechanism, we used microarray analysis to investigate the genes expression profiles of C. albicans cells treated or untreated with 10b and found 957 genes were differentially expressed. Of them,457 showed a decrease in expression and 500 showed an increase in expression. 33 down-regulated genes were involved in glycolysis (e.g., PFK1, CDC19 and HXK2), fermentation (e.g., PDC11, ALD5 and ADH1) and respiratory electron transport chain (e.g., CBP3, COR1 and QCR8). 30 differentially expressed genes were found to relate to biofilm formation, filamentous or hyphal growth. It was noticed that striking up-regulation of SFL1 and marked down-regulation of YWP1 directly related to prevent C. albicans from changing its morphology from the yeast form to the hyphal. Two genes related to specifically hydrolyzing beta-1, 3 glucan (e.g., XOG1) and chitin (e.g., CHT1) were significantly increased. 40 overexpressed genes and 15 down-regulated genes were related to the lipid metabolic process. Of them, Eight were directly linked to ergosterol biosynthesis, including ERG2, ERG6 and ERG11. 99 genes related to translation were down-regulated following exposure to 10b, which account for 21.66% in down-regulated genes. This suggested that translation might be lower in SC5314 cells exposed to 10b than in control. Total RNA from the control SC5314 cells and 10b-treated SC5314 cells were used to generate target cDNA, and then hybridized to 8k Candida albicans Genome Array Genechips, representing about 7925 characterized Candida albicans genes. Two independent experiments were conducted. Reference strain was control SC5314 cells and test strain was SC5314 cells treated with 10b.
Project description:Biomaterial infections are an increasingly alarming problem, and because of their intrinsic recalcitrance to conventional therapy, a new class of antifungal drugs must be explored. 10b, a 2-aminotetralin derivate, was synthesized as a novel chemical structural antifungal agent and exibited strong anti-biofilm activity. To further investigate the action mechanism, we used microarray analysis to investigate the genes expression profiles of C. albicans biofilms treated or untreated with 10b and found 150 genes were differentially expressed. Of them, 69 showed a decrease in expression and 81 showed an increase in expression -10 differentially expressed genes related to biofilm formation, Filamentous or hypha growth. A gene related to specifically hydrolyzing β-1, 3 glucan was significantly increased. 10 down-regulated genes were involved in glycolysis, fermentation and active oxygen scavenging. 15 overexpressed genes were related to the lipid metabolic process. Of them, 13 genes were directly linked to ergosterol biosynthesis including ERG2, ERG6 and ERG11. 10 genes related to translation were over-expressed. Among them, 2 genes involved in negative regulation of transcription were significantly up-regulated. Total RNA from the control SC5314 biofilms and 10b-treated SC5314 biofilms were used to generate target cDNA, and then hybridized to 8k Candida albicans Genome Array Genechips, representing about 7925 characterized Candida albicans genes. Two independent experiments were conducted. Reference strain was control SC5314 biofilms and test strain was SC5314 biofilms treated with 10b.
Project description:Following antifungal treatment, Candida albicans, and other human pathogenic fungi can undergo microevolution, which leads to the emergence of drug resistance. However, the capacity for microevolutionary adaptation of fungi goes beyond the development of drug resistance. Here we used an experimental microevolution approach to show that one of the central pathogenicity mechanisms of C. albicans, the yeast-to-hyphae transition, can be subject to experimental evolution. The C. albicans cph1Δ/efg1Δ mutant is non-filamentous, as central signalling pathways linking environmental cues to hypha formation are disrupted. We subjected this mutant to constant selection pressure in the hostile environment of the macrophage phagosome. In a comparatively short time-frame, the mutant evolved the ability to escape macrophages by filamentation. To investigate the transcriptional response underlying the yeast-to-filament transition in the evolved strain, we applied RNA-Seq technology. Furthermore, RNA-Seq data were used to identify SNPs, which are specific for the evolved strain. For both strains, the cph1Δ/efg1Δ mutant and the Evo-strain, two conditions, one promotes yeast growth the other filamentous growth, were investigated. For each condition three biological replicates were analysed.
Project description:Invasion of host tissue by the human fungal pathogen, Candida albicans is an important step during many forms of candidosis. However, not all C. albicans strains possess the same invasive and virulence properties. It is known for example that the two clinical isolates SC5314 and ATCC10231 differ in their ability to invade into host tissue and to cause infections. Strain SC5314 is invasive whereas strain ATCC10231 is non-invasive and strongly attenuated in virulence as compared to SC5314. In this study we compare the in vitro transcriptional profiles and the genotypic profiles of these two widely used laboratory strains in order to determine the principal biological and genetic properties which may govern the different potential for invasiveness and virulence. Keywords: transcriptional profiling, comparative genomic hybridisation, invasive vs. non-invasive C. albicans strain Genomic DNA from C. albicans strains SC5314 and ATCC10231 hybridisations were done in duplicate including one dye swap. Total RNA from C. albicans strains SC5314 and ATCC10231 strains were compared in triplicate including one biological replicate and one dye swap.
Project description:Recent studies have shown that the transcriptional landscape of the pleiomorphic fungus Candida albicans is highly dependent upon growth conditions. Here using a dual RNA-seq approach we identified 299 C. albicans and 72 Streptococcus gordonii genes that were either up- or down-regulated specifically as a result of co-culturing these human oral cavity microorganisms. Seventy five C. albicans genes involved in responses to chemical stimuli, regulation, homeostasis, protein modification and cell cycle were statistically (P ≤0.05) upregulated, while 36 genes mainly involved in transport and translation were down-regulated. Upregulation of filamentation-associated TEC1 and FGR42 genes, and of ALS1 adhesin gene, concurred with previous evidence that the C. albicans yeast to hypha transition is promoted by S. gordonii. Increased expression of genes required for arginine biosynthesis in C. albicans was potentially indicative of a novel oxidative stress response. The transcriptional response of S. gordonii to C. albicans was less dramatic, with only eight S. gordonii genes significantly (P ≤0.05) up-regulated ≥ twofold (glpK, rplO, celB, rplN, rplB, rpsE, ciaR, and gat). The expression patterns suggest that signals from S. gordonii cause a positive filamentation response in C. albicans, while S. gordonii appears to be transcriptionally less influenced by C. albicans. Five Samples; Sample 1 - Candida albicans cells grown in hypha inducing conditions for two hours; Sample 2 - Candida albicans cells grown in hypha-inducing conditions for two hours before co-culture with Streptococcus gordonii cells for one hour in a 2:1 rato; Sample 3 - Candida albicans cells grown in hypha-inducing conditions for two hours before culture in Streptococcus gordonii media for one hour; Sample 4 - Candida albicans cells grown in hypha inducing conditions for two hours, filtered to remove Candida albicans cells and media added to Streptococcus gordonii cells for one hour; Sample 5 - Streptococcus gordonii cells alone for one hour. All samples extracted and sequenced in biological triplicate using Illumina HiSeq2500. Samples 1, 2 and 3 aligned to the reference genome for Candida albicans and Samples 2, 4 and 5 aligned to the reference genome for Streptococcus gordonii.
Project description:Fungal infections are a serious health problem in the clinic especially in the immunocompromised patient. Disease ranges from widespread superficial vulvovaginal infections to life-threatening systemic candidiasis. Especially for systemic mycoses only a limited arsenal of antimycotica are available, including azoles, polyenes, echinocandines and amphothericin B. Due to emerging resistance to standard therapy and significant side effects for some antimycotica there is a medical need for new antifungals in the clinic and general practice. In order to expand the arsenal of compounds with antifungal activities we screened compound libraries, including combinatorial libraries as well as more than 30 000 pure compounds derived from organic synthesis for antimycotic activity. In total more than 100 000 compounds were screened using an innovative AS (activity-selectivity) assay analyzing both the antifungal activity and the compatability with human cells at the same time. One promising hit, a Benzimidazol-2-yl-alkylamine derivative, was developed in a series of lead compounds showing potent antifungal activity. ((1S)-1-[1-(3-chlorobenzyl)-1H-benzimidazol-2-yl]-2-methylpropyl-amine) (EMC120B12) showed the highest antifungal activity and best compatability with human cells in several cell culture models and against a number of different yeasts and clinical isolates. Transcriptional profiling indicates that the newly discovered compound is a potential inhibitor of the ergosterol-pathway. In total, three biological replicates were performed. All experiments were performed as dye swaps. Thus, in total six arrays have been hybridzed. Hybridization experiments included an untreated reference sample and a sample of cells treated with ((1S)-1-[1-(3-chlorobenzyl)-1H-benzimidazol-2-yl]-2-methylpropyl-amine) (EMC120B12). The array included one technical replicate of each probe.
Project description:Fungal infections are a serious health problem in clinics especially in the immune-compromised patient. Disease ranges from widespread superficial infections like vulvovaginal infections to life-threatening systemic candidiasis. Especially for systemic mycoses only a limited arsenal of antifungals is available. The most commonly used classes of antifungal compounds used include azoles, polyenes and echinocandines. Due to emerging resistance to standard therapy and significant side effects and high costs for several antifungals.,there is a medical need for new antifungals in the clinic and general practice. In order to expand the arsenal of compounds with antifungal activities we previously screened a compound library, using a new type of activity-selectivity (AS) assay analysing both the antifungal activity and the compatibility with human cells at the same time. One compound, ((S)-2-(1-aminoisobutyl)-1-(3-chlorobenzyl) benzimidazole (EMC120B12)), showed high antifungal activity against several species of pathogenic yeasts including C. glabrata and C. krusei, species which are highly refractory to antifungals, especially to the commonly used azoles. Here we could show by transcriptional profiling and sterol analysis that the target of this new antifungal compound is the ergosterol pathway. The effects of EMC120B12 on sterol biosynthesis mimic those of fluconazole, strongly indicating that EMC120B12 also targets ERG11 like the azols. But not only the marker sterol 14 methylergosta 8,24(28) dien 3β,6α diol accumulated in C. krusei under EMC120B12 treatment, but also hitherto unknown related sterols. The novel sterols have a 3β,6α diol structure. Furthermore, this is the first time that a benzimidazole structure has been shown to result in a block of the sterol pathway by accumulating marker sterols connected to ERG11 inactivation. In total, three biological replicates were performed. All experiments were performed as dye swaps. Thus, in total 18 arrays have been hybridzed. Hybridization experiments included an untreated reference sample and a sample of cells treated with either ((1S)-1-[1-(3-chlorobenzyl)-1H-benzimidazol-2-yl]-2-methylpropyl-amine) (EMC120B12), Fluconazole or Nocodazole. The array included one technical replicate of each probe.
Project description:Perforin is a major effector molecule of human natural killer (NK) cells. It can induce delayed growth of Candida albicans hyphae. Here, the fungal transcriptome was analyzed after a co-incubation with 500ng/µl Perforin.
Project description:Global transcriptional profiling revealed that IL-17A induced artificial nutrient starvation conditions in Candida albicans, resulting in a downregulated target of rapamycin (TOR) signaling pathway and in increased autophagic responses and intracellular cAMP. We used microarray to detail the global programme of gene expression underlying IL17A sensing by Candida albicans at different time points (T0_0h T1_4h, T2_24h) and identified distinct classes of up-regulated and down regulated genes.