A fungal metabolic regulator underlies infectious synergism during Candida albicans - Staphylococcus aureus intra-abdominal co-infection.

Candida albicans and Staphylococcus aureus are two commonly associated pathogens that cause nosocomial infections with high morbidity and mortality. Our prior and current work using a murine model of polymicrobial intra-abdominal infection (IAI) uncovered synergistic lethality that was driven by Candida -induced upregulation of functional S. aureus ⍺-toxin leading to polymicrobial sepsis and organ damage. In order to determine the candidal effector(s) mediating enhanced virulence, an unbiased screen of C. albicans transcription factor mutants was undertaken and revealed that zcf13 Δ/Δ failed to drive augmented ⍺-toxin or lethal synergism during co-infection. Using a combination of transcriptional and phenotypic profiling approaches, ZCF13 was shown to regulate genes involved in pentose metabolism, including RBK1 and HGT7 that contribute to fungal ribose catabolism and uptake, respectively. Subsequent experiments revealed that ribose inhibited the staphylococcal agr quorum sensing system and concomitantly repressed toxicity. Unlike wild-type C. albicans , zcf13 Δ/Δ was unable to effectively utilize ribose during co-culture or co-infection leading to exogenous ribose accumulation and agr repression. Forced expression of RBK1 and HGT7 in the zcf13 Δ/Δ mutant fully restored pathogenicity during co-infection. Collectively, our results detail the interwoven complexities of cross-kingdom interactions and highlight how intermicrobial metabolism impacts polymicrobial disease pathogenesis with devastating consequences for the host.

Candida-induced granulocytic myeloid-derived suppressor cells are protective against polymicrobial sepsis.

IMPORTANCE: Polymicrobial intra-abdominal infections are serious clinical infections that can lead to life-threatening sepsis, which is difficult to treat in part due to the complex and dynamic inflammatory responses involved. Our prior studies demonstrated that immunization with low-virulence Candida species can provide strong protection against lethal polymicrobial sepsis challenge in mice. This long-lived protection was found to be mediated by trained Gr-1+ polymorphonuclear leukocytes with features resembling myeloid-derived suppressor cells (MDSCs). Here we definitively characterize these cells as MDSCs and demonstrate that their mechanism of protection involves the abrogation of lethal inflammation, in part through the action of the anti-inflammatory cytokine interleukin (IL)-10. These studies highlight the role of MDSCs and IL-10 in controlling acute lethal inflammation and give support for the utility of trained tolerogenic immune responses in the clinical treatment of sepsis.

Protection against lethal sepsis following immunization with Candida species varies by isolate and inversely correlates with bone marrow tissue damage.

Protection against lethal Candida albicans (Ca)/Staphylococcus aureus (Sa) intra-abdominal infection (IAI)-mediated sepsis can be achieved by a novel form of trained innate immunity (TII) involving Gr-1+ myeloid-derived suppressor cells (MDSCs) that are induced by inoculation (immunization) with low virulence Candida species [i.e., Candida dubliniensis (Cd)] that infiltrate the bone marrow (BM). In contrast, more virulent Candida species (i.e., C. albicans), even at sub-lethal inocula, fail to induce similar levels of protection. The purpose of the present study was to test the hypothesis that the level of TII-mediated protection induced by Ca strains inversely correlates with damage in the BM as a reflection of virulence. Mice were immunized by intraperitoneal inoculation with several parental and mutant strains of C. albicans deficient in virulence factors (hyphal formation and candidalysin production), followed by an intraperitoneal Ca/Sa challenge 14 d later and monitored for sepsis and mortality. Whole femur bones were collected 24 h and 13 d after immunization and assessed for BM tissue/cellular damage via ferroptosis and histology. While immunization with standard but not sub-lethal inocula of most wild-type C. albicans strains resulted in considerable mortality, protection against lethal Ca/Sa IAI challenge varied by strain was usually less than that for C. dubliniensis, with no differences observed between parental and corresponding mutants. Finally, levels of protection afforded by the Ca strains were inversely correlated with BM tissue damage (R 2 = -0.773). TII-mediated protection against lethal Ca/Sa sepsis induced by Candida strain immunization inversely correlates with BM tissue/cellular damage as a reflection of localized virulence.

Effect of MMR Vaccination to Mitigate Severe Sequelae Associated With COVID-19: Challenges and Lessons Learned.

Mortality in COVID-19 cases was strongly associated with progressive lung inflammation and eventual sepsis. There is mounting evidence that live attenuated vaccines commonly administered during childhood, also provide beneficial non-specific immune effects, including reduced mortality and hospitalization due to unrelated infections. It has been proposed that live attenuated vaccine-associated non-specific effects are a result of inducing trained innate immunity to function more effectively against broader infections. In support of this, our laboratory has reported that immunization with a live attenuated fungal strain induces a novel form of trained innate immunity which provides protection against various inducers of sepsis in mice via myeloid-derived suppressor cells. Accordingly, we initiated a randomized control clinical trial with the live attenuated Measles, Mumps, Rubella (MMR) vaccine in healthcare workers in the greater New Orleans area aimed at preventing/reducing severe lung inflammation/sepsis associated with COVID-19 (ClinicalTrials.gov Identifier: NCT04475081). Included was an outcome to evaluate the myeloid-derived suppressor cell populations in blood between those administered the MMR vaccine vs placebo. The unanticipated emergency approval of several COVID-19 vaccines in the midst of the MMR clinical trials eliminated the ability to examine effects of the MMR vaccine on COVID-19-related health status. Unfortunately, we were also unable to show any impact of the MMR vaccine on peripheral blood myeloid-derived suppressor cells due to several inherent limitations (low percentages of blood leukocytes, small sample size), that also included a collaboration with a similar trial (CROWN CORONATION; ClinicalTrials.gov Identifier: NCT04333732) in St. Louis, MO. In contrast, monitoring the COVID-19 vaccine response in trial participants revealed that high COVID-19 antibody titers occurred more often in those who received the MMR vaccine vs placebo. While the trial was largely inconclusive, lessons learned from addressing several trial-associated challenges may aid future studies that test the non-specific beneficial immune effects of live attenuated vaccines.

Efficacy of Candida dubliniensis and Fungal ß-Glucans in Inducing Trained Innate Immune Protection Against Inducers of Sepsis.

Fungal-bacterial intra-abdominal infections (IAI) can lead to sepsis with significant morbidity and mortality. We have established a murine model of Candida albicans (Ca) and Staphylococcus aureus (Sa) IAI that results in acute lethal sepsis. Prior intraperitoneal or intravenous inoculation with low virulence Candida dubliniensis (Cd) confers high level protection against lethal Ca/Sa IAI and sepsis. Protection via Cd immunization is associated with decreased pro-inflammatory cytokines and mediated by Gr-1+ putative myeloid-derived suppressor cells (MDSCs) representing a novel form of trained innate immunity (TII). The objective of these studies was to determine the extent of Cd-mediated TII against sepsis of broad origin and explore the potential of fungal cell wall components as abiotic immunogen alternatives to induce TII, including zymosan depleted of TLR2 activity (d-zymosan), or purified preparations of ß-glucan. Immunized mice were challenged 14 days post-immunization with a lethal array of live or abiotic inducers of sepsis, including Ca/Sa, Ca/Escherichia coli (Ca/Ec), LPS or untreated zymosan. Results showed that live Cd immunization was protective against sepsis induced by Ca/Ec and zymosan, but not LPS. Similar to protection against Ca/Sa, survival was dependent on Gr-1+ cells with no role for macrophages. Among the fungal cell wall compounds as immunogens, immunization with d-zymosan and an alkali-treated form of ß-glucan also resulted in significant protection against sepsis induced by Ca/Sa or Ca/Ec, but not LPS sepsis. Again, there was a strong dependence on Gr-1+ cells for protection with one exception, an added role for macrophages in the case of protection induced by alkali-treated ß-glucan. Overall, these results demonstrate that immunization with Cd as well as abiotic fungal cell components are capable of Gr-1+ cell-mediated trained innate immune protection against sepsis of broad microbial origin. In addition, abiotic ß-glucans represent potential alternatives to live Cd for protection against lethal polymicrobial sepsis.

Trained Innate Immunity Induced by Vaccination with Low-Virulence Candida Species Mediates Protection against Several Forms of Fungal Sepsis via Ly6G+ Gr-1+ Leukocytes.

We recently discovered a novel form of trained innate immunity (TII) induced by low-virulence Candida species (i.e., Candida dubliniensis) that protects against lethal fungal/bacterial infection. Mice vaccinated by intraperitoneal (i.p.) inoculation are protected against lethal sepsis following Candida albicans/Staphylococcus aureus (Ca/Sa) intra-abdominal infection (IAI) or Ca bloodstream infection (BSI). The protection against IAI is mediated by long-lived Gr-1+ leukocytes as putative myeloid-derived suppressor cells (MDSCs) and not by prototypical trained macrophages. This study aimed to determine if a similar TII mechanism (Gr-1+ cell-mediated suppression of sepsis) is protective against BSI and whether this TII can also be induced following intravenous (i.v.) vaccination. For this, mice were vaccinated with low-virulence Candida strains (i.p. or i.v.), followed by lethal challenge (Ca/Sa i.p. or Ca i.v.) 14 days later, and observed for sepsis (hypothermia, sepsis scoring, and serum cytokines), organ fungal burden, and mortality. Similar parameters were monitored following depletion of macrophages or Gr-1+ leukocytes during lethal challenge. The results showed that mice vaccinated i.p. or i.v. were protected against lethal Ca/Sa IAI or Ca BSI. In all cases, protection was mediated by Ly6G+ Gr-1+ putative granulocytic MDSCs (G-MDSCs), with no role for macrophages, and correlated with reduced sepsis parameters. Protection also correlated with reduced fungal burden in spleen and brain but not liver or kidney. These results suggest that Ly6G+ G-MDSC-mediated TII is induced by either the i.p. and i.v. route of inoculation and protects against IAI or BSI forms of systemic candidiasis, with survival correlating with amelioration of sepsis and reduced organ-specific fungal burden. IMPORTANCE Trained innate immunity (TII) is induced following immunization with live attenuated microbes and represents a clinically important strategy to enhance innate defenses. TII was initially demonstrated following intravenous inoculation with low-virulence Candida albicans, with protection against a subsequent lethal C. albicans intravenous bloodstream infection (BSI) mediated by monocytes with enhanced cytokine responses. We expanded this by describing a novel form of TII induced by intraperitoneal inoculation with low-virulence Candida that protects against lethal sepsis induced by polymicrobial intra-abdominal infection (IAI) via Gr-1+ leukocytes as putative myeloid-derived suppressor cells (MDSCs). In this study, we addressed these two scenarios and confirmed an exclusive role for Ly6G+ Gr-1+ leukocytes in mediating TII against either IAI or BSI via either route of inoculation, with protection associated with suppression of sepsis. These studies highlight the previously unrecognized importance of Ly6G+ MDSCs as central mediators of a novel form of TII termed trained tolerogenic immunity.

A Contemporary Warming/Restraining Device for Efficient Tail Vein Injections in a Murine Fungal Sepsis Model.

In rodent models, tail vein injections are important methods for intravenous administration of experimental agents. Tail vein injections typically involve warming of the animal to promote vasodilation, which aids in both the identification of the blood vessels and positioning of the needle into the vessel lumen while securely restraining the animal. Although tail vein injections are common procedures in many protocols and are not considered highly technical if performed correctly, accurate and consistent injections are crucial to obtain reproducible results and minimize variability. Conventional methods for inducing vasodilation prior to tail vein injections generally depend on the use of a heat source such as a heat lamp, electrical/rechargeable heat pads, or pre-heated water at 37 °C. Despite being readily accessible in a standard laboratory setting, these tools evidently suffer from poor/limited thermo-regulatory capacity. Similarly, although various forms of restraining devices are commercially available, they must be used carefully to avoid trauma to the animals. These limitations of the current methods create unnecessary variables in experiments or result in varying outcomes between experiments and/or laboratories. In this article, we demonstrate an improved protocol using an innovative device that combines an independent, thermally regulated, warming device with an adjustable restraining unit into one system for efficient streamlined tail vein injection. The example we use is an intravenous model of fungal bloodstream infection that results in sepsis. The warming apparatus consists of a heat-reflective acrylic box installed with an adjustable automatic thermostat to maintain the internal temperature at a pre-set threshold. Likewise, the width and height of the cone restraining apparatus can be adjusted to safely accommodate various rodent sizes. With the advanced and versatile features of the device, the technique shown here could become a useful tool across a range of research areas involving rodent models that employ tail vein injections.

Analysis of Measles-Mumps-Rubella (MMR) Titers of Recovered COVID-19 Patients.

The measles-mumps-rubella (MMR) vaccine has been theorized to provide protection against coronavirus disease 2019 (COVID-19). Our aim was to determine whether any MMR IgG titers are inversely correlated with severity in recovered COVID-19 patients previously vaccinated with MMR II. We divided 80 subjects into two groups, comparing MMR titers to recent COVID-19 severity levels. The MMR II group consisted of 50 subjects who would primarily have MMR antibodies from the MMR II vaccine, and a comparison group of 30 subjects consisted of those who would primarily have MMR antibodies from sources other than MMR II, including prior measles, mumps, and/or rubella illnesses. There was a significant inverse correlation (rs = -0.71, P < 0.001) between mumps virus titers (mumps titers) and COVID-19 severity within the MMR II group. There were no significant correlations between mumps titers and severity in the comparison group, between mumps titers and age in the MMR II group, or between severity and measles or rubella titers in either group. Within the MMR II group, mumps titers of 134 to 300 arbitrary units (AU)/ml (n = 8) were found only in those who were functionally immune or asymptomatic; all with mild symptoms had mumps titers below 134 AU/ml (n = 17); all with moderate symptoms had mumps titers below 75 AU/ml (n = 11); all who had been hospitalized and had required oxygen had mumps titers below 32 AU/ml (n = 5). Our results demonstrate that there is a significant inverse correlation between mumps titers from MMR II and COVID-19 severity.IMPORTANCE COVID-19 has presented various paradoxes that, if understood better, may provide clues to controlling the pandemic, even before a COVID-19 vaccine is widely available. First, young children are largely spared from severe disease. Second, numerous countries have COVID-19 death rates that are as low as 1% of the death rates of other countries. Third, many people, despite prolonged close contact with someone who is COVID-19 positive, never test positive themselves. Fourth, nearly half of people who test positive for COVID-19 are asymptomatic. Some researchers have theorized that the measles-mumps-rubella (MMR) vaccine may be responsible for these disparities. The significance of our study is that it showed that mumps titers related to the MMR II vaccine are significantly and inversely correlated with the severity of COVID-19-related symptoms, supporting the theorized association between the MMR vaccine and COVID-19 severity.

Could an Unrelated Live Attenuated Vaccine Serve as a Preventive Measure To Dampen Septic Inflammation Associated with COVID-19 Infection?

We propose the concept that administration of an unrelated live attenuated vaccine, such as MMR (measles, mumps, rubella), could serve as a preventive measure against the worst sequelae of coronavirus disease 2019 (COVID-19). There is mounting evidence that live attenuated vaccines provide nonspecific protection against lethal infections unrelated to the target pathogen of the vaccine by inducing "trained" nonspecific innate immune cells for improved host responses against subsequent infections. Mortality in COVID-19 cases is strongly associated with progressive lung inflammation and eventual sepsis. Vaccination with MMR in immunocompetent individuals has no contraindications and may be especially effective for health care workers who can easily be exposed to COVID-19. Following the lead of other countries conducting clinical trials with the live attenuated Mycobacterium bovis BCG (BCG) vaccine under a similar concept, a clinical trial with MMR in high-risk populations may provide a "low-risk-high-reward" preventive measure in saving lives during this unprecedented COVID-19 pandemic.

Applying the Host-Microbe Damage Response Framework to Candida Pathogenesis: Current and Prospective Strategies to Reduce Damage.

Disease is a complex outcome that can occur as a result of pathogen-mediated damage, host-mediated damage or both. This has led to the revolutionary concept of the damage response framework (DRF) that defines microbial virulence as a function of host immunity. The DRF outlines six scenarios (classes) of host damage or beneficial outcomes, depending on the microbe and the strength of the immune response. Candida albicans is uniquely adapted to its human host and can exist as either a commensal, colonizing various anatomical sites without causing notable damage, or as a pathogen, with the ability to cause a diverse array of diseases, ranging from mucosal to invasive systemic infections that result in varying levels of microbe-mediated and/or host-mediated damage. We recently categorized six different forms of candidiasis (oropharyngeal, hematogenous, intra-abdominal, gastrointestinal, denture stomatitis, and vulvovaginitis) into independent DRF classes, supporting a contemporary view of unique mechanisms of pathogenesis for these Candida infections. In this review, we summarize the evidence for the pathogenesis of these various forms of candidiasis in the context of the DRF with the further intent to provide insights into strategies to achieve a level of host response or outcome otherwise, that limits host damage.

Candida albicans Impacts Staphylococcus aureus Alpha-Toxin Production via Extracellular Alkalinization.

Candida albicans and Staphylococcus aureus are common causes of nosocomial infections with severe morbidity and mortality. Murine polymicrobial intra-abdominal infection (IAI) with C. albicans and S. aureus results in acute mortality dependent on the secreted cytolytic effector alpha-toxin. Here, we confirmed that alpha-toxin is elevated during polymicrobial growth compared to monomicrobial growth in vitro Therefore, this study sought to unravel the mechanism by which C. albicans drives enhanced staphylococcal alpha-toxin production. Using a combination of functional and genetic approaches, we determined that an intact agr quorum sensing regulon is necessary for enhanced alpha-toxin production during coculture and that a secreted candidal factor likely is not implicated in elevating agr activation. As the agr system is pH sensitive, we observed that C. albicans raises the pH during polymicrobial growth and that this correlates with increased agr activity and alpha-toxin production. Modulation of the pH could predictably attenuate or activate agr activity during coculture. By using a C. albicans mutant deficient in alkalinization (stp2Δ/Δ), we confirmed that modulation of the extracellular pH by C. albicans can drive agr expression and toxin production. Additionally, the use of various Candida species (C. glabrata, C. dubliniensis, C. tropicalis, C. parapsilosis, and C. krusei) demonstrated that those capable of raising the extracellular pH correlated with elevated agr activity and alpha-toxin production during coculture. Overall, we demonstrate that alkalinization of the extracellular pH by the Candida species leads to sustained activation of the staphylococcal agr system.IMPORTANCECandida albicans and Staphylococcus aureus are commonly coisolated from central venous catheters and deep-seated infections, including intra-abdominal sepsis. Thus, they represent a significant cause of nosocomial morbidity and mortality. Yet how these organisms behave in the context of polymicrobial growth remains poorly understood. In this work, we set out to determine the mechanism by which activation of the staphylococcal agr quorum sensing system and production of its major virulence effector alpha-toxin is enhanced during coculture with C. albicans Surprisingly, we likely ruled out that a secreted candidal factor drives this process. Instead, we demonstrated that alkalinization of the extracellular milieu by C. albicans and other Candida species correlated with elevated agr activity. Thus, we propose a mechanism where modulation of the extracellular pH by fungal opportunists can indirectly alter virulence of a bacterial pathogen. Uncovering molecular events that drive interkingdom pathogenicity mechanisms may enhance surveillance and treatment for devastating polymicrobial infections.

Special Issue: Fungal-Bacterial Interactions-Current Knowledge and Future Perspectives.

We would like to thank all the contributors to the Special Issue on Fungal-Bacterial Interactions-Current Knowledge and Future Perspectives [...].

Candida albicans Augments Staphylococcus aureus Virulence by Engaging the Staphylococcal agr Quorum Sensing System.

Candida albicans and Staphylococcus aureus are among the most prevalent nosocomial pathogens that are responsible for severe morbidity and mortality, even with appropriate treatment. Using a murine model of polymicrobial intra-abdominal infection (IAI), we have previously shown that coinfection with these pathogens results in synergistic lethality that is partially dependent on exacerbated prostaglandin signaling, while monomicrobial infection is nonlethal. Therefore, the objective of this study was to identify staphylococcal virulence determinants that drive lethal synergism during polymicrobial IAI. Using the toxigenic S. aureus strain JE2, we observed that coinfection with C. albicans led to a striking 80 to 100% mortality rate within 20 h postinoculation (p.i.) while monomicrobial infections were nonlethal. Use of a green fluorescent protein (GFP)-P3 promoter S. aureus reporter strain revealed enhanced activation of the staphylococcal agr quorum sensing system during in vitro polymicrobial versus monomicrobial growth. Analyses by quantitative real-time PCR (qPCR), Western blot, and toxin functional assays confirmed enhanced agr-associated gene transcription and increases in secreted alpha- and delta-toxins. C. albicans-mediated elevated toxin production and hemolytic activity were determined to be agrA dependent, and genetic knockout and complementation of hla identified alpha-toxin as the key staphylococcal virulence factor driving lethal synergism. Analysis of mono- and polymicrobial infections 8 h p.i. demonstrated equivalent bacterial burdens in the peritoneal cavity but significantly elevated levels of alpha-toxin (3-fold) and the eicosanoid prostaglandin E2 (PGE2) (4-fold) during coinfection. Importantly, prophylactic passive immunization using the monoclonal anti-alpha-toxin antibody MEDI4893* led to significantly improved survival rates compared to those following treatment with isotype control antibody. Collectively, these results define alpha-toxin as an essential virulence determinant during C. albicans-S. aureus IAI and describe a novel mechanism by which a human-pathogenic fungus can augment the virulence of a highly pathogenic bacterium in vivoIMPORTANCE Relatively little is known about the complex interactions and signaling events that occur between microbes and even less so about how microbial "cross talk" shapes human health and disease. Candida albicans (a fungus) and Staphylococcus aureus (a bacterium) are formidable human nosocomial pathogens, causing severe morbidity and mortality. Moreover, they are frequently coisolated from central venous catheters and deep-seated infections, including intra-abdominal sepsis. In this work, we have shown that coinfection with C. albicans and S. aureus is highly lethal, leading to >80% mortality by day 1 postinfection, whereas monoinfection with C. albicans or S. aureus does not cause mortality. This infectious synergism is dependent on the expression of staphylococcal alpha-toxin, and secretion of this potent virulence factor is actually augmented by C. albicans via an agr-dependent mechanism. Moreover, prophylactic neutralization of alpha-toxin with a monoclonal antibody is sufficient to elicit protection during coinfection. Therefore, we have demonstrated that a pathogenic fungus can enhance virulence determinants of a bacterium in vivo with devastating consequences to the host. These results have important implications in the surveillance and treatment of polymicrobial disease and highlight the dynamic intersection of environment, pathogens, and host.

Candida/Staphylococcal Polymicrobial Intra-Abdominal Infection: Pathogenesis and Perspectives for a Novel Form of Trained Innate Immunity.

Polymicrobial sepsis is difficult to diagnose and treat and causes significant morbidity and mortality, especially when fungi are involved. In vitro, synergism between Candida albicans and various bacterial species has been described for many years. Our laboratory has developed a murine model of polymicrobial intra-abdominal infection with Candida albicans and Staphylococcus aureus, demonstrating that polymicrobial infections cause high levels of mortality, while monoinfections do not. By contrast, closely related Candida dubliniensis does not cause synergistic lethality and rather provides protection against lethal polymicrobial infection. This protection is thought to be driven by a novel form of trained innate immunity mediated by myeloid-derived suppressor cells (MDSCs), which we are proposing to call "trained tolerogenic immunity". MDSC accumulation has been described in patients with sepsis, as well as in in vivo sepsis models. However, clinically, MDSCs are considered detrimental in sepsis, while their role in in vivo models differs depending on the sepsis model and timing. In this review, we will discuss the role of MDSCs in sepsis and infection and summarize our perspectives on their development and function in the spectrum of trained innate immune protection against fungal-bacterial sepsis.

Spectrum of Trained Innate Immunity Induced by Low-Virulence Candida Species against Lethal Polymicrobial Intra-abdominal Infection.

Polymicrobial intra-abdominal infections (IAI) are clinically prevalent and cause significant morbidity and mortality, especially those involving fungi. Our laboratory developed a mouse model of polymicrobial IAI and demonstrated that coinfection with Candida albicans and Staphylococcus aureus (C. albicans/S. aureus) results in 80 to 90% mortality in 48 to 72 h due to robust local and systemic inflammation. Surprisingly, inoculation with Candida dubliniensis and S. aureus resulted in minimal mortality, and rechallenge of mice with lethal C. albicans/S. aureus conferred >90% protection up to 60 days postinoculation. Protection was mediated by Gr-1+ polymorphonuclear leukocytes, indicating a novel form of trained innate immunity (TII). The purpose of this study was to determine the microbial requirements and spectrum of innate-mediated protection. In addition to Candida dubliniensis, several other low-virulence Candida species (C. glabrata, C. auris, and C. albicansefg1Δ/Δ cph1Δ/Δ) and Saccharomyces cerevisiae conferred significant protection with or without S. aureus For C. dubliniensis-mediated protection, hyphal formation was not required, with protection conferred as early as 7 days after primary challenge but not at 120 days, and also following multiple lethal C. albicans/S. aureus rechallenges. This protection also extended to a lethal intravenous (i.v.) C. albicans challenge but had no effect in the C. albicans vaginitis model. Finally, studies revealed the ability of the low-virulence Candida species that conferred protection to invade the bone marrow by 24 h post-primary challenge, with a positive correlation between femoral bone marrow fungal infiltration at 48 h and protection upon rechallenge. These results support and further extend the characterization of this novel TII in protection against lethal fungal-bacterial IAI and sepsis.

Candida glabrata Has No Enhancing Role in the Pathogenesis of Candida-Associated Denture Stomatitis in a Rat Model.

Denture stomatitis (DS) is a condition characterized by inflammation of the oral mucosa in direct contact with dentures and affects a significant number of otherwise healthy denture wearers. Candida-associated DS is predominantly caused by Candida albicans, a dimorphic fungus that readily colonizes and forms biofilms on denture materials. Previous studies showed a requirement for Candida biofilm formation on both palate and dentures in infection and identified fungal morphogenic transcription factors, Efg1 and Bcr1, as key players in DS pathogenesis. While both C. albicans and Candida glabrata are frequently coisolated in mucosal candidiasis, a pathogenic role for C. glabrata in DS remains unknown. Using an established rat model of DS, we sought to determine whether C. glabrata alone or coinoculation with C. albicans establishes colonization and causes palatal tissue damage and inflammation. Rats fitted with custom dentures were inoculated with C. albicans and/or C. glabrata and monitored over a 4-week period for fungal burden (denture/palate), changes in body weight, and tissue damage via lactate dehydrogenase (LDH) release as well as palatal staining by hematoxylin and eosin (H&E) and immunohistochemistry for myeloperoxidase (MPO) as measures of inflammation. C. glabrata colonized the denture/palate similarly to C. albicans In contrast to C. albicans, colonization by C. glabrata resulted in minimal changes in body weight, palatal LDH release, and MPO expression. Coinoculation with both species had no obvious modulation of C. albicans-mediated pathogenic effects. These data suggest that C. glabrata readily establishes colonization on denture and palate but has no apparent role for inducing/enhancing C. albicans pathogenesis in DS.IMPORTANCE Many denture wearers suffer from Candida-associated denture stomatitis (DS), a fungal infection of the hard palate in contact with dentures. Biofilm formation by Candida albicans on denture/palate surfaces is considered a central process in the infection onset. Although Candida glabrata is frequently coisolated with C. albicans, its role in DS pathogenesis is unknown. We show here, using a contemporary rat model that employed a patented intraoral denture system, that C. glabrata established stable colonization on the denture/palate. However, in contrast to C. albicans inoculated rats, rats inoculated with C. glabrata exhibited minimal changes in weight gain or palatal tissue damage. Likewise, coinoculation with the two Candida species resulted in no exacerbation of C. albicans-induced DS pathology. Together, our findings indicate that C. glabrata has no inducing/enhancing role in DS pathogenesis.

Current patient perspectives of vulvovaginal candidiasis: incidence, symptoms, management and post-treatment outcomes.

BACKGROUND: Vulvovaginal candidiasis (VVC) is a common infection affecting women worldwide. Reports of patterns/risk factors/trends for episodic/recurrent VVC (RVVC) are largely outdated. The purpose of this study was to obtain current patient perspectives of several aspects of VVC/RVVC. METHODS: Business cards containing on-line survey information were distributed to healthy volunteers and patients seeking standard, elective, or referral gynecologic care in university-affiliated Obstetrics/Gynecology clinics. The internet-based questionnaire was completed by 284 non-pregnant women (78% Caucasian, 14% African American, 8% Asian). RESULTS: The majority of the participants (78%) indicated a history of VVC with 34% defined as having RVVC. The most common signs/symptoms experienced were itching, burning and redness with similar ranking of symptoms among VVC and RVVC patients. Among risk factors, antibiotic use ranked highest followed by intercourse, humid weather and use of feminine hygiene products. A high number of respondents noted 'no known cause' (idiopathic episodes) that was surprisingly similar among women with a history of either VVC or RVVC. VVC/RVVC episodes reported were primarily physician-diagnosed (73%) with the remainder mostly reporting self-diagnosis and treating with over-the-counter (OTC) medications. Most physician-diagnosed attacks utilized a combination of pelvic examination and laboratory tests followed by prescribed antifungals. Physician-treated cases achieved a higher level of symptom relief (84%) compared to those who self-medicated (57%). The majority of women with RVVC (71%) required continual or long-term antifungal medication as maintenance therapy to control symptoms. CONCLUSIONS: Current patient perspectives closely reflect historically documented estimates of VVC/RVVC prevalence and trends regarding symptomatology, disease management and post-treatment outcomes.

Identification of Specific Components of the Eicosanoid Biosynthetic and Signaling Pathway Involved in Pathological Inflammation during Intra-abdominal Infection with Candida albicans and Staphylococcus aureus.

Polymicrobial intra-abdominal infections (IAIs) are a significant cause of morbidity and mortality, particularly when fungal pathogens are involved. Our experimental murine model of IAI involving intraperitoneal inoculation of Candida albicans and Staphylococcus aureus results in synergistic lethality (∼80%) due to exacerbated inflammation. Monomicrobial infection results in no mortality, despite a microbial burden and dissemination similar to those in a coinfection. In the coinfection model, the immunomodulatory eicosanoid prostaglandin E2 (PGE2) was determined to be necessary and sufficient to induce mortality, implicating PGE2 as the central mediator of the amplified inflammatory response. The aim of this study was to identify key components of the PGE2 biosynthetic and signaling pathway involved in the inflammatory response and explore whether these can be targeted to prevent or reduce mortality. Using selective pharmacological inhibitors of cyclooxygenases (COX) or PGE2 receptor antagonists in the C. albicans-S. aureus IAI mouse model, we found that inhibition of COX and/or blocking of PGE2 receptor 1 (EP1) or PGE2 receptor 3 (EP3) signaling reduced proinflammatory cytokine production, promoted interleukin-10 production, reduced cellular damage in the peritoneal cavity, and, most importantly, significantly improved survival. The greatest effect on survival was obtained by the simultaneous inhibition of COX-1 activity and EP1 and EP3 receptor signaling. Importantly, early inhibition of PGE2 pathways dramatically improved the survival of fluconazole-treated mice compared with that achieved with fluconazole treatment alone. These findings indicate that COX-1 and the EP1 and EP3 receptors mediate the downstream pathological effects of PGE2 during polymicrobial IAI and may serve as effective therapeutic targets.