N-acetylglucosamine-mediated morphological transition in Candida albicans and Candida tropicalis.

Morphological transitions in Candida species are key factors in facilitating invasion and adapting to environmental changes. N-acetylglucosamine (GlcNAc) is a monosaccharide signalling molecule that can regulate morphological transitions in Candida albicans and Candida tropicalis. Interestingly, although the uptake and metabolic pathways of GlcNAc and GlcNAc-mediated white-to-opaque cell switching are similar between the two Candida species, GlcNAc induces hyphal development in C. albicans, whereas it suppresses hyphal development in C. tropicalis. These findings indicate that the characteristics of C. albicans and C. tropicalis in response to GlcNAc are remarkably different. Here, we compare the conserved and divergent GlcNAc-mediated signalling pathways and catabolism between the two Candida species. Deletion of NGT1, a GlcNAc transportation gene, inhibited hyphal formation in C. albicans but promoted hyphal development in C. tropicalis. To further understand these opposite effects on filamentous growth in response to GlcNAc in the two Candida species, the cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) signalling pathways in both C. albicans and C. tropicalis were compared. Interestingly, GlcNAc activated the cAMP/PKA signalling pathway of the two Candida species, suggesting that the hyphal development-regulated circuit is remarkably diverse between the two species. Indeed, the Ndt80-like gene REP1, which is critical for regulating GlcNAc catabolism, exhibits distinct roles in the hyphal development of C. albicans and C. tropicalis. These data suggest possible reasons for the divergent hyphal growth response in C. albicans and C. tropicalis upon GlcNAc induction.

Candida tropicalis RON1 is required for hyphal formation, biofilm development, and virulence but is dispensable for N-acetylglucosamine catabolism.

NDT80-like family genes are highly conserved across a large group of fungi, but the functions of each Ndt80 protein are diverse and have evolved differently among yeasts and pathogens. The unique NDT80 gene in budding yeast is required for sexual reproduction, whereas three NDT80-like genes, namely, NDT80, REP1, and RON1, found in Candida albicans exhibit distinct functions. Notably, it was suggested that REP1, rather than RON1, is required for N-acetylglucosamine (GlcNAc) catabolism. Although Candida tropicalis, a widely dispersed fungal pathogen in tropical and subtropical areas, is closely related to Candida albicans, its phenotypic, pathogenic and environmental adaptation characteristics are remarkably divergent. In this study, we focused on the Ron1 transcription factor in C. tropicalis. Protein alignment showed that C. tropicalis Ron1 (CtRon1) shares 39.7% identity with C. albicans Ron1 (CaRon1). Compared to the wild-type strain, the C. tropicalis ron1Δ strains exhibited normal growth in different carbon sources and had similar expression levels of several GlcNAc catabolic genes during GlcNAc treatment. In contrast, C. tropicalis REP1 is responsible for GlcNAc catabolism and is involved in GlcNAc catabolic gene expressions, similar to C. albicans Rep1. However, REP1 deletion strains in C. tropicalis promote hyphal development in GlcNAc with low glucose content. Interestingly, CtRON1, but not CaRON1, deletion mutants exhibited significantly impaired hyphal growth and biofilm formation. As expected, CtRON1 was required for full virulence. Together, the results of this study showed divergent functions of CtRon1 compared to CaRon1; CtRon1 plays a key role in yeast-hyphal dimorphism, biofilm formation and virulence. LAY ABSTRACT: In this study, we identified the role of RON1, an NDT80-like gene, in Candida tropicalis. Unlike the gene in Candida albicans, our studies showed that RON1 is a key regulator of hyphal formation, biofilm development and virulence but is dispensable for N-acetylglucosamine catabolism in C. tropicalis.

Time-resolved RNA-seq analysis to unravel the in vivo competence induction by Streptococcus pneumoniae during pneumonia-derived sepsis.

Competence development in Streptococcus pneumoniae (pneumococcus) is tightly intertwined with virulence. In addition to genes encoding genetic transformation machinery, the competence regulon also regulates the expression of allolytic factors, bacteriocins, and cytotoxins. Pneumococcal competence system has been extensively interrogated in vitro where the short transient competent state upregulates the expression of three distinct phases of "early," "late," and "delayed" genes. Recently, we have demonstrated that the pneumococcal competent state develops naturally in mouse models of pneumonia-derived sepsis. To unravel the underlying adaptive mechanisms driving the development of the competent state, we conducted a time-resolved transcriptomic analysis guided by the spatiotemporal live in vivo imaging system of competence induction during pneumonia-derived sepsis. Mouse lungs infected by the serotype 2 strain D39 expressing a competent state-specific reporter gene (D39-ssbB-luc) were subjected to RNA sequencing guided by monitoring the competence development at 0, 12, 24, and, at the moribund state, >40 hours post-infection (hpi). Transcriptomic analysis revealed that the competence-specific gene expression patterns in vivo were distinct from those under in vitro conditions. There was significant upregulation of early, late, and some delayed phase competence-specific genes as early as 12 hpi, suggesting that the pneumococcal competence regulon is important for adaptation to the lung environment. Additionally, members of the histidine triad (pht) gene family were sharply upregulated at 12 hpi followed by a steep decline throughout the rest of the infection cycle, suggesting that Pht proteins participate in the early adaptation to the lung environment. Further analysis revealed that Pht proteins execute a metal ion-dependent regulatory role in competence induction.IMPORTANCEThe induction of pneumococcal competence for genetic transformation has been extensively studied in vitro but poorly understood during lung infection. We utilized a combination of live imaging and RNA sequencing to monitor the development of a competent state during acute pneumonia. Upregulation of competence-specific genes was observed as early as 12 hour post-infection, suggesting that the pneumococcal competence regulon plays an important role in adapting pneumococcus to the stressful lung environment. Among others, we report novel finding that the pneumococcal histidine triad (pht) family of genes participates in the adaptation to the lung environment and regulates pneumococcal competence induction.

Amphiphilic Dendrimer as Potent Antibacterial against Drug-Resistant Bacteria in Mouse Models of Human Infectious Diseases.

Modern medicine continues to struggle against antibiotic-resistant bacterial pathogens. Among the pathogens of critical concerns are the multidrug-resistant (MDR) Pseudomonas aeruginosa, Staphylococcus aureus, and Klebsiella pneumoniae. These pathogens are major causes of nosocomial infections among immunocompromised individuals, involving major organs such as lung, skin, spleen, kidney, liver, and bloodstream. Therefore, novel approaches are direly needed. Recently, we developed an amphiphilic dendrimer DDC18-8A exhibiting high antibacterial and antibiofilm efficacy in vitro. DDC18-8A is composed of a long hydrophobic alkyl chain and a small hydrophilic poly(amidoamine) dendron bearing amine terminals, exerting its antibacterial activity by attaching and inserting itself into bacterial membranes to trigger cell lysis. Here, we examined the pharmacokinetics and in vivo toxicity as well as the antibacterial efficacy of DDC18-8A in mouse models of human infectious diseases. Remarkably, DDC18-8A significantly reduced the bacterial burden in mouse models of acute pneumonia and bacteremia by P. aeruginosa, methicillin-resistant S. aureus (MRSA), and carbapenem-resistant K. pneumoniae and neutropenic soft tissue infection by P. aeruginosa and MRSA. Most importantly, DDC18-8A outperformed pathogen-specific antibiotics against all three pathogens by achieving a similar bacterial clearance at 10-fold lower therapeutic concentrations. In addition, it showed superior stability and biodistribution in vivo, with excellent safety profiles yet without any observable abnormalities in histopathological analysis of major organs, blood serum biochemistry, and hematology. Collectively, we provide strong evidence that DDC18-8A is a promising alternative to the currently prescribed antibiotics in addressing challenges associated with nosocomial infections by MDR pathogens.

Convergent and divergent roles of the glucose-responsive kinase SNF4 in Candida tropicalis.

The sucrose non-fermenting 1 (SNF1) complex is a heterotrimeric protein kinase complex that is an ortholog of the mammalian AMPK complex and is evolutionally conserved in most eukaryotes. This complex contains a catalytic subunit (Snf1), a regulatory subunit (Snf4) and a scaffolding subunit (Sip1/Sip2/Gal73) in budding yeast. Although the function of AMPK has been well studied in Saccharomyces cerevisiae and Candida albicans, the role of AMPK in Candida tropicalis has never been investigated. In this study, we focused on SNF4 in C. tropicalis as this fungus cannot produce a snf1Δ mutant. We demonstrated that C. tropicalis SNF4 shares similar roles in glucose derepression and is necessary for cell wall integrity and virulence. The expression of both SNF1 and SNF4 was significantly induced when glucose was limited. Furthermore, snf4Δ strains exhibited high sensitivity to many surface-perturbing agents because the strains contained lower levels of glucan, chitin and mannan. Interestingly, in contrast to C. albicans sak1Δ and snf4Δ, C. tropicalis snf4Δ exhibited phenotypes for cell aggregation and pseudohypha production. These data indicate that SNF4 performs convergent and divergent roles in C. tropicalis and possibly other unknown roles in the C. tropicalis SNF1-SNF4 AMPK pathway.