Control of β-glucan exposure by the endo-1,3-glucanase Eng1 in Candida albicans modulates virulence

Candida albicans is a major opportunistic pathogen of humans. It can grow as morphologically distinct yeast, pseudohyphae and hyphae, and the ability to switch reversibly among different forms is critical for its virulence. The relationship between morphogenesis and innate immune recognition is not quite clear. Dectin-1 is a major C-type lectin receptor that recognizes β-glucan in the fungal cell wall. C. albicans β-glucan is usually masked by the outer mannan layer of the cell wall. Whether and how β-glucan masking is differentially regulated during hyphal morphogenesis is not fully understood. Here we show that the endo-1,3-glucanase Eng1 is differentially expressed in yeast, and together with Yeast Wall Protein 1 (Ywp1), regulates β-glucan exposure and Dectin-1-dependent immune activation of macrophage by yeast cells. ENG1 deletion results in enhanced Dectin-1 binding at the septa of yeast cells; while eng1 ywp1 yeast cells show strong overall Dectin-1 binding similar to hyphae of wild-type and eng1 mutants. Correlatively, hyphae of wild-type and eng1 induced similar levels of cytokines in macrophage. ENG1 expression and Eng1-mediated β-glucan trimming are also regulated by antifungal drugs, lactate and N-acetylglucosamine. Deletion of ENG1 modulates virulence in the mouse model of hematogenously disseminated candidiasis in a Dectin-1-dependent manner. The eng1 mutant exhibited attenuated lethality in male mice, but enhanced lethality in female mice, which was associated with a stronger renal immune response and lower fungal burden. Thus, Eng1-regulated β-glucan exposure in yeast cells modulates the balance between immune protection and immunopathogenesis during disseminated candidiasis.

IL-23 signaling prevents ferroptosis-driven renal immunopathology during candidiasis

During infection the host relies on pattern-recognition receptors to sense invading fungal pathogens to launch immune defense mechanisms. While fungal recognition and immune effector responses are organ and cell type specific, during disseminated candidiasis myeloid cells exacerbate collateral tissue damage. However, the complex interplay between protective antifungal immunity and immunopathology remains incompletely understood. The β-glucan receptor ephrin type-A 2 receptor (EphA2) is required to initiate mucosal inflammatory responses during oral Candida infection. Here we report that Epha2 promotes renal immunopathology during disseminated candidiasis. EphA2 deficiency leads to reduced renal inflammation and injury. Comprehensive analyses reveal that EphA2 limits IL-23 secretion in dendritic cells, while IL-23 signaling prevents ferroptotic myeloid cell death during infection. Further, ferroptosis aggravates inflammation during infection, while at the same time reducing the fungal killing capacity of macrophages. Thus, we identify ferroptotic cell death as a critical pathway of Candida-mediated renal immunopathology that opens a new avenue to tackle Candida infection and inflammation.

Disseminated Candidiasis and Candidemia Caused by Candida palmioleophila in a Green Sea Turtle (Chelonia mydas)

A sub-adult green sea turtle (Chelonia mydas) was rescued and treated for carapace and plastron shell fractures. The turtle was kept dry-docked for the first 2 months with a placement of a long-term jugular central venous catheter (CVC). Pain management, aggressive antibiotic and anthelmintic therapy, fluid therapy, force feeding, and wound debridement were provided to manage the shell fractures and control bacteremia. Human albumin was administered to treat severe hypoalbuminemia. On day 59, small budding yeasts were noted on the blood smears. Candidemia was confirmed by blood culture, as the yeasts were identified as Candida palmioleophila by the molecular multi-locus identification method. The CVC was removed, and the patient was treated with itraconazole. Although the carapace and plastron wounds had epithelized by 5.5 months after the rescue, the turtle died unexpectedly by 7.5 months. The postmortem examination revealed numerous necrogranulomas with intralesional yeasts, morphologically compatible with Candida spp., in joints, bones, brain, and lungs, suggestive of disseminated candidiasis. We describe a rare case of candidemia in the veterinary field. To our knowledge, this is the first report of candidiasis caused by C. palmioleophila in a reptile. The present results should improve veterinary medical care and, therefore, enhance the conservation of endangered sea turtle species.

DectiSomes: Glycan Targeting of Liposomal Drugs Improves the Treatment of Disseminated Candidiasis

Candida albicans causes life-threatening disseminated candidiasis. Individuals at greatest risk have weakened immune systems. An outer cell wall, exopolysaccharide matrix, and biofilm rich in oligoglucans and oligomannans help Candida spp. evade host defenses. Even after antifungal treatment, the one-year mortality rate exceeds 25%. Undoubtedly, there is room to improve drug performance. The mammalian C-type lectin pathogen receptors Dectin-1 and Dectin-2 bind to fungal oligoglucans and oligomannans, respectively. We previously coated amphotericin B-loaded liposomes, AmB-LLs, pegylated analogs of AmBisome, with the ligand binding domains of these two Dectins. DectiSomes, DEC1-AmB-LLs and DEC2-AmB-LLs, showed two distinct patterns of binding to the exopolysaccharide matrix surrounding C. albicans hyphae grown in vitro. Here we showed that DectiSomes were preferentially associated with fungal colonies in the kidneys. In a neutropenic mouse model of candidiasis, DEC1-AmB-LLs and DEC2-AmB-LLs delivering only one dose of 0.2 mg/kg AmB reduced the kidney fungal burden several fold relative to AmB-LLs. DEC1-AmB-LLs and DEC2-AmB-LLs increased the percent of surviving mice 2.5-fold and 8.3-fold, respectively, relative to AmB-LLs. Dectin-2 targeting of anidulafungin loaded liposomes, DEC2-AFG-LLs, and of commercial AmBisome, DEC2-AmBisome, reduced fungal burden in the kidneys several fold over their untargeted counterparts. The data herein suggest that targeting of a variety of antifungal drugs to fungal glycans may achieve lower safer effective doses and improve drug efficacy against a variety of invasive fungal infections.

DectiSomes: Glycan Targeting of Liposomal Drugs Improves the Treatment of Disseminated Candidiasis

Candida albicans causes life-threatening disseminated candidiasis. Individuals at greatest risk have weakened immune systems. An outer cell wall, exopolysaccharide matrix, and biofilm rich in oligoglucans and oligomannans help Candida albicans. evade host defenses. Even after antifungal drug treatment the one-year mortality rate exceeds 25%. Undoubtedly there is room to improve antifungal drug performance. The mammalian C-type lectin pathogen receptors Dectin-1 and Dectin-2 bind to fungal oligoglucans and oligomannans, respectively. We previously coated amphotericin B-loaded liposomes, AmB-LLs, pegylated analogs of AmBisome, with the ligand binding domains of these two Dectins. DectiSomes, DEC1-AmB-LLs and DEC2-AmB-LLs, showed two distinct patterns of binding to the exopolysaccharide matrix surrounding C. albicans hyphae grown in vitro, while untargeted AmB-LLs did not bind. DectiSomes were preferentially associated with fungal colonies in the kidneys. In a neutropenic mouse model of candidiasis, DEC1-AmB-LLs and DEC2-AmB-LLs delivering only one dose of 0.2 mg/kg AmB significantly reduced the kidney fungal burden several fold relative to AmB-LLs, based on either colony forming units (P=0.013 to 8.8×10-5) or quantitative PCR of fungal rRNA ITS (P=5.5×10-5 to 3.0×10-10). DEC1-AmB-LLs and DEC2-AmB-LLs significantly increased the percent of surviving mice relative to AmB-LLs. Dectin-2 targeted anidulafungin loaded liposomes and AmBisomes, DEC2-AFG-LLs and DEC2-AmBisome reduced fungal burden in the kidneys several fold over their untargeted counterparts (P=7.8×10-5 and 0.0020, respectively). The data herein suggest that targeting of a variety of antifungal drugs to fungal glycans may achieve lower safer effective doses and improve drug efficacy against a variety of invasive fungal infections.

#34: Characteristics of Pediatric Patients with Candidemia and Risk Factors for Disseminated Candidiasis

Background Candida species are the most common cause of fungal infection in hospitalized patients. Patients with candidemia are at risk of developing disseminated infection. Studies in adults have described risk factors for disseminated candidiasis in patients with candidemia, but there are fewer such studies in pediatric populations. The objective of the study is to examine the prevalence of disseminated candidiasis in pediatric patients with candidemia and to compare patients with disseminated candidiasis to those with candidemia alone. Methods All patients with blood cultures positive for a Candida species from 2007–2018 were identified from the UM Clinical Microbiology Laboratory. Retrospective chart review was conducted to gather demographic, clinical, and microbiologic data. Disseminated candidiasis was defined as ocular infection; endocarditis; renal, hepatic, or splenic involvement; peritonitis; or CNS infection. Patients with disseminated candidiasis were compared to those with candidemia alone using odds ratios. Results There were 81 episodes of candidemia during the study period, representing 61 unique patients. The most common comorbidity was intestinal failure (43%), and the presence of a central line (99%) and use of TPN (73%) were both highly prevalent. The most frequently identified Candida species was C. albicans, followed by C. parapsilosis. There were 14 patients (17%) with disseminated candidiasis. The most common site for dissemination was the abdomen (5 patients). Immunosuppressed patients were significantly more likely to have disseminated candidiasis (OR 4.1, 95% CI 1.1–15.3), while those with intestinal failure were significantly less likely to develop disseminated candidiasis (OR 0.07, 95% CI 0.01–0.6). Patients with disseminated candidiasis were significantly more likely to require ICU admission (OR 5.8, 95% CI 1.2–28) and were significantly more likely to die within three months of infection (OR 9.3, 95% CI 2.3–37.6) than those with candidemia alone. Conclusions The prevalence of disseminated candidiasis in patients with candidemia was 17%. Immunosuppression was positively associated and intestinal failure was negatively associated with the development of disseminated candidiasis. Patients with disseminated candidiasis had worse outcomes (including ICU admission and death) than those with candidemia alone. Our data suggest that immunosuppressed patients with candidemia should be thoroughly evaluated for signs of disseminated candidiasis.