Hypha essential genes in Candida albicans pathogenesis of oral lichen planus: an in-vitro study

Background Hypha essential genes (HEGs) of Candida Albicans have been emerging into scholar’s attention, little known about their functions in oral lichen planus (OLP) with an uncovered etiology. This research aimed to observe necessary genes in biphasic C. albicans from OLP and study their relevance in pathogenesis, so as to evaluate possible roles of morphologic switching in etiology of OLP. Methods Samples were collected from OLP lesions of patients, mycelia were cultured and total RNA was extracted then subjected to reverse transcription-PCR and real-time PCR. Results HWP1 and HGC1 were significantly expressed in hyphae phase and weakly detected in yeast phase, while there was no significant difference of EFG1, ALS3, and ECE1 between in yeast and mycelia. Conclusion HGC1 and HWP1 were confirmed to be hypha essential genes, with HGC1 for hypha morphogenesis and HWP1 for adhesion invasion in pathogenesis of C. albicans in OLP. ALS3, ECE1 and EFG1 played minor roles in hyphae maintenance and adhesion for hyphae. These might be deemed as hints for the etiology of OLP and indicate HGC1 and HWP1 to be a priority of potential drug target.

Dimorphism in Neopseudocercosporella capsellae, an Emerging Pathogen Causing White Leaf Spot Disease of Brassicas

White leaf spot pathogen: Neopseudocercosporella capsellae causes significant damage to many economically important Brassicaceae crops, including oilseed rape through foliar, stem, and pod lesions under cool and wet conditions. A lack of information on critical aspects of the pathogen’s life cycle limits the development of effective control measures. The presence of single-celled spores along with multi-celled conidia on cotyledons inoculated with multi-celled conidia suggested that the multi-celled conidia were able to form single-celled spores on the host surface. This study was designed to demonstrate N. capsellae morphological plasticity, which allows the shift between a yeast-like single-celled phase and the multi-celled hyphal phase. Separate experiments were designed to illustrate the pathogen’s morphological transformation to single-celled yeast phase from multi-celled hyphae or multi-celled macroconidia in-vitro and in-planta. Results confirmed the ability of N. capsellae to switch between two morphologies (septate hyphae and single-celled yeast phase) on a range of artificial culture media (in-vitro) or in-planta on the host surface before infection occurs. The hyphae-to-yeast transformation occurred through the production of two morphologically distinguishable blastospore (blastoconidia) types (meso-blastospores and micro-blastospores), and arthrospores (arthroconidia).

A modified culture medium and hyphae isolation method can increase quality of the RNA extracted from mycelia of a dimorphic fungal species

The capability of RNA isolation with good efficiency and high quality is essential for a downstream application such as RNA sequencing. It requires successful cell culturing and an effective RNA isolation method. Although effective methods are available, production of the homogenous mycelia and extraction of good-quality mycelial RNA from true invasive hyphae, which penetrated into the agar plates, are difficult. To overcome these problems, the aim of this study was to develop technical modifications which allow production of homogenous mycelial biomass without extra stimuli agents and improve quality of the RNA extracted from the fungal hyphae. Our alternative culture medium was suitable for production both yeast-phase cells and hyphae of the Schizosaccharomyces japonicus and other dimorphic species, such as the Candida albicans, Saccharomyces cerevisiae, and Jaminaea angkorensis. To improve quality of the mycelial RNA, we developed an isolation procedure of the hyphal tip, which eliminated the unnecessary vacuoles-containing parts of the hyphae. To increase RNA quantity, we used glass beads in the RNA extraction protocol to achieve stronger breaking of the mycelial walls. All these modifications can also be useful for researchers working with other dimorphic fungi and can contribute to the higher comparability of the transcriptional data coming from yeast-phase cells and hyphae or even from different species.

Dietary Inclusion of Blood Plasma with Yeast (Saccharomyces cerevisiae) Supplementation Enhanced the Growth Performance, Nutrient Digestibility, Lactobacillus Count, and Reduced Gas Emissions in Weaning Pigs

This experiment was performed to examine the hypothesis that blood plasma (BP) with yeast (Saccharomyces cerevisiae) supplement in the diet of weaning pigs could provoke the growth performance, nutrient digestibility, fecal microbial, and reduce harmful gas excretion. A total of one hundred and eighty healthy piglets were taken and assigned (complete random blocks) to three dietary treatments as: Phase 1: Treatment (TRT) 1-6% BP; TRT 2-3% BP + 3% yeast; TRT 3-6% yeast. Phase 2: TRT 1-3%; BP., TRT 2-1.5% BP + 1.5% yeast; TRT 3- 3% yeast. Phase 3: TRT 1- Control (CON) (Basal diet); TRT 2- CON; TRT 3- CON for six- weeks. Each treatment had twelve replicates and five (three gilts and two barrows) pigs per pen. Dietary inclusion of BP with yeast supplementation significantly increased the body weight of piglets during phase 2 (p = 0.003) and phase 3 (p = 0.032). In addition, TRT2 group piglets had a significant improvement in average daily gain at the end of each phase and overall (p = 0.047, 0.025, 0.018 and 0.012, respectively). At phase 3, TRT2 group piglets showed a significant improvement on nutrient digestibility of dry matter (p = 0.012) and nitrogen (p = 0.040). The fecal microbiota of TRT2 group piglets showed a tendency to increase the number of Lactobacillus counts at phase 1 (p = 0.07) and phase 2 (p = 0.06) as well as, a significant improvement at phase 3 (p = 0.021). In addition, TRT2 group piglets had trend to decrease NH3 (p = 0.074) and H2S (p = 0.069) during phase 2, and significantly reduced NH3 (p = 0.038) and H2S (p = 0.046) at phase 3. However, the fecal score of piglets remains unaffected during the entire trial. At the end of phase 1 piglets’ IgG (p = 0.008) was significantly increased with the inclusion of BP with yeast supplementation. Based on the positive effects on body weight, average daily gain, nutrient digestibility, Lactobacillus count, and reduced gas emission, we suggest that dietary supplement with BP and yeast in the diet of weaned piglet could serve as an excellent alternative to antibiotics growth promoters.

The Trojan Horse Model in Paracoccidioides: A Fantastic Pathway to Survive Infecting Human Cells

Paracoccidioidomycosis (PCM) is the most relevant systemic endemic mycosis limited to Latin American countries. The etiological agents are thermally dimorphic species of the genus Paracoccidioides. Infection occurs via respiratory tract by inhalation of propagules from the environmental (saprophytic) phase. In the lung alveoli the fungus converts to the characteristic yeast phase (parasitic) where interact with extracellular matrix proteins, epithelial cells, and the host cellular immunity. The response involves phagocytic cells recognition but intracellular Paracoccidioides have demonstrated the ability to survive and also multiply inside the neutrophils, macrophages, giant cells, and dendritic cells. Persistence of Paracoccidioides as facultative intracellular pathogen is important in terms of the fungal load but also regarding to the possibility to disseminate penetrating other tissues even protected by the phagocytes. This strategy to invade other organs via transmigration of infected phagocytes is called Trojan horse mechanism and it was also described for other fungi and considered a factor of pathogenicity. This mini review comprises a literature revision of the spectrum of tools and mechanisms displayed by Paracoccidioides to overcame phagocytosis, discusses the Trojan horse model and the immunological context in proven models or the possibility that Paracoccidioides apply this tool for dissemination to other tissues.

Investigating the Smuts: Common Cues, Signaling Pathways, and the Role of MAT in Dimorphic Switching and Pathogenesis

The corn smut fungus Ustilago maydis serves as a model species for studying fungal dimorphism and its role in phytopathogenic development. The pathogen has two growth phases: a saprobic yeast phase and a pathogenic filamentous phase. Dimorphic transition of U. maydis involves complex processes of signal perception, mating, and cellular reprogramming. Recent advances in improvement of reference genomes, high-throughput sequencing and molecular genetics studies have been expanding research in this field. However, the biology of other non-model species is frequently overlooked. This leads to uncertainty regarding how much of what is known in U. maydis is applicable to other dimorphic fungi. In this review, we will discuss dimorphic fungi in the aspects of physiology, reproductive biology, genomics, and molecular genetics. We also perform comparative analyses between U. maydis and other fungi in Ustilaginomycotina, the subphylum to which U. maydis belongs. We find that lipid/hydrophobicity is a potential common cue for dimorphic transition in plant-associated dimorphic fungi. However, genomic profiles alone are not adequate to explain dimorphism across different fungi.

The Ndr/LATS Kinase Cbk1 Regulates a Specific Subset of Ace2 Functions and Suppresses the Hypha-to-Yeast Transition in Candida albicans

ABSTRACT The regulation of Ace2 and morphogenesis (RAM) pathway is an important regulatory network in the human fungal pathogen Candida albicans. The RAM pathway’s two most well-studied components, the NDR/Lats kinase Cbk1 and its putative substrate, the transcription factor Ace2, have a wide range of phenotypes and functions. It is not clear, however, which of these functions are specifically due to the phosphorylation of Ace2 by Cbk1. To address this question, we first compared the transcriptional profiles of CBK1 and ACE2 deletion mutants. This analysis indicates that, of the large number of genes whose expression is affected by deletion of CBK1 and ACE2, only 5.5% of those genes are concordantly regulated. Our data also suggest that Ace2 directly or indirectly represses a large set of genes during hyphal morphogenesis. Second, we generated strains containing ACE2 alleles with alanine mutations at the Cbk1 phosphorylation sites. Phenotypic and transcriptional analysis of these ace2 mutants indicates that, as in Saccharomyces cerevisiae, Cbk1 regulation is important for daughter cell localization of Ace2 and cell separation during yeast-phase growth. In contrast, Cbk1 phosphorylation of Ace2 plays a minor role in C. albicans yeast-to-hypha transition. We have, however, discovered a new function for the Cbk1-Ace2 axis. Specifically, Cbk1 phosphorylation of Ace2 prevents the hypha-to-yeast transition. To our knowledge, this is one of the first regulators of the C. albicans hypha-to-yeast transition to be described. Finally, we present an integrated model for the role of Cbk1 in the regulation of hyphal morphogenesis in C. albicans. IMPORTANCE The regulation of Ace2 and morphogenesis (RAM) pathway is a key regulatory network that plays a role in many aspects of C. albicans pathobiology. In addition to characterizing the transcriptional effects of this pathway, we discovered that Cbk1 and Ace2, a key RAM pathway regulator-effector pair, mediate a specific set of the overall functions of the RAM pathway. We have also discovered a new function for the Cbk1-Ace2 axis: suppression of the hypha-to-yeast transition. Very few regulators of this transition have been described, and our data indicate that maintenance of hyphal morphogenesis requires suppression of yeast phase growth by Cbk1-regulated Ace2.

The Ndr/LATS kinase Cbk1 regulates a specific subset of Ace2 functions and suppresses the hyphae-to-yeast transition in Candida albicans

The Regulation of Ace2 and Morphogenesis (RAM) pathway is an important regulatory network in the human fungal pathogen Candida albicans. The RAM pathway’s two most well-studied components, the NDR/Lats kinase Cbk1 and its putative substrate, the transcription factor Ace2, have a wide range of phenotypes and functions. It is not clear, however, which of these functions are specifically due to the phosphorylation of Ace2 by Cbk1. To address this question, we first compared the transcriptional profiles of CBK1 and ACE2 deletion mutants. This analysis indicates that, of the large number of genes whose expression is affected by deletion of CBK1 and ACE2, only 5.5% of those genes are concordantly regulated. Our data also suggest that Ace2 directly or indirectly represses a large set of genes during hyphal morphogenesis. Second, we generated strains containing ACE2 alleles with alanine mutations at the Cbk1 phosphorylation sites. Phenotypic and transcriptional analysis of these ace2 mutants indicates that, as in Saccharomyces cerevisiae, Cbk1 regulation is important for daughter cell localization of Ace2 and cell separation during yeast phase growth. In contrast, Cbk1 phosphorylation of Ace2 plays a minor role in C. albicans yeast-to-hyphae transition. We have, however, discovered a new function for the Cbk1-Ace2 axis. Specifically, Cbk1 phosphorylation of Ace2 prevents the hyphae-to-yeast transition. To our knowledge, this is one of the first regulators of the C. albicans hyphae-to-yeast transition to be described. Finally, we present an integrated model for the role of Cbk1 in the regulation of hyphal morphogenesis in C. albicans.ImportanceRegulation of Ace2 and Morphogenesis (RAM) pathway is a key regulatory network that plays a role in many aspects of C. albicans pathobiology. In addition to characterizing the transcriptional effects of this pathway, we discovered that Cbk1 and Ace2, a key RAM pathway regulator-effector pair, mediate a specific set of the overall functions of the RAM pathway. We have also discovered a new function for the Cbk1-Ace2 axis; suppression of the hyphae-to-yeast transition. Very few regulators of this transition have been described and our data indicate that maintenance of hyphal morphogenesis requires suppression of yeast phase growth by Cbk1-regulated Ace2.

Expression of Talaromyces marneffei acuM and acuK Genes in Gluconeogenic Substrates and Various Iron Concentrations

Talaromyces marneffei is an opportunistic, dimorphic fungal pathogen that causes a disseminated infection in people with a weakened immunological status. The ability of this fungus to acquire nutrients inside the harsh environment of the macrophage phagosome is presumed to contribute to its pathogenicity. The transcription factors AcuM and AcuK are known to regulate gluconeogenesis and iron acquisition in Aspergillus fumigatus. This study demonstrated that they are also involved in both of these processes in the dimorphic fungus T. marneffei. Expression of acuM and acuK genes was determined by real time-polymerase chain reaction (RT-PCR) on the cells grown in media containing gluconeogenic substrates and various iron concentrations. We found that the acuM and acuK transcript levels were sequentially reduced when growing the fungus in increasing amounts of iron. The acuM transcript was upregulated in the gluconeogenic condition, while the acuK transcript showed upregulation only in the acetate medium in the yeast phase. These results suggest the involvement of acuM and acuK in gluconeogenesis and iron homeostasis in T. marneffei.