Salivary metals, age, and gender correlate with cultivable oral Candida carriage levels.

Background: Little is known about the normal range of metal levels in unstimulated saliva, nor whether these might impact Candida carriage in healthy individuals. Both are important in determining which populations are at risk for candidiasis, as the availability of metal ions can influence the growth and pathogenesis of Candida albicans. Objective: We quantified salivary metals of healthy individuals to determine the correlation with C. albicans oral colonization. Design: Unstimulated whole saliva was collected from healthy adults and plated to determine fungal carriage, and metal content was measured using ICP-mass spectrometry (ICP-MS). Results: Zinc was most abundant, followed by iron, copper, manganese, and nickel. Cultivable oral Candida carriage was found in 48% of people. Total protein levels did not differ in salivas from donors with or without carriage. However, innate fungicidal activity was increased in donors with carriage; correlations between levels of several metals were stronger in salivas with fungal carriage, indicating a shift in the oral environment. Concentrations of copper and manganese, as well as age and gender, were significantly predictive of carriage levels in a multiple regression model. Conclusions: Salivary copper and manganese content along with age and gender could be used as a predictive metric for individuals that are more susceptible to Candida overgrowth.

Fluconazole-Resistant Candida auris Is Susceptible to Salivary Histatin 5 Killing and to Intrinsic Host Defenses.

Candida auris is a newly identified species causing invasive candidemia and candidiasis. It has broad multidrug resistance (MDR) not observed for other pathogenic Candida species. Histatin 5 (Hst 5) is a well-studied salivary cationic peptide with significant antifungal activity against Candida albicans and is an attractive candidate for treating MDR fungi, since antimicrobial peptides induce minimal drug resistance. We investigated the susceptibility of C. auris to Hst 5 and neutrophils, two first-line innate defenses in the human host. The majority of C. auris clinical isolates, including fluconazole-resistant strains, were highly sensitive to Hst 5: 55 to 90% of cells were killed by use of 7.5 µM Hst 5. Hst 5 was translocated to the cytosol and vacuole in C. auris cells; such translocation is required for the killing of C. albicans by Hst 5. The inverse relationship between fluconazole resistance and Hst 5 killing suggests different cellular targets for Hst 5 than for fluconazole. C. auris showed higher tolerance to oxidative stress than C. albicans, and higher survival within neutrophils, which correlated with resistance to oxidative stress in vitro Thus, resistance to reactive oxygen species (ROS) is likely one, though not the only, important factor in the killing of C. auris by neutrophils. Hst 5 has broad and potent candidacidal activity, enabling it to combat MDR C. auris strains effectively.

Patient Demographic and Clinician Factors in Antibiotic Prescribing for Upper Respiratory Tract Infections in the Australian Capital Territory from 2006-2015.

INTRODUCTION: National antibiotic stewardship programs aim to mitigate rising antimicrobial resistance and associated healthcare costs by promoting safe and appropriate antibiotic prescribing. AIM: This study aimed to analyse patient and clinician demographic factors that may influence antibiotic prescribing for Upper Respiratory Tract Infections (URTIs). Trends in antibiotic prescribing patterns were also analysed over the study period. MATERIALS AND METHODS: This retrospective cross-sectional study analysed data from the Australian National University Medical School Clinical Audit Project database, comprising data collected by students during patient encounters over a two week period each April-May between 2006 and 2015 (excluding 2013). Data was collected via standardised survey in multiple healthcare settings and locations in the Australian Capital Territory (ACT) and Southeast New South Wales. (NSW) URTI diagnosis and symptomatology were defined using the International Classification of Disease (ICD-10) and International Classification of Primary Care, version 2 PLUS (ICPC-2+) criteria. RESULTS: URTI accounted for 5.6% (n=698) of total patient encounters (n=12,468), and of these, 42.7% (n=289) were prescribed an antibiotic intervention. Antibiotics were significantly more likely to be prescribed in the hospital setting (44.2%; n=237) compared to community GP (32.1%; n=52; p<0.05) and for patients presenting with localised symptoms (65.9%; n=109) compared to generalised symptoms (33.7%; n=122; p<0.01). No significant association was observed for age, rurality, patient gender, clinical gender or Indigenous status. The most frequently prescribed antibiotic was penicillin (67.8%; n=196). Over the decade of study, antibiotic prescribing for URTIs showed decreasing trend both overall (R2=0.204) and with respect to all demographic factors assessed. CONCLUSION: This study supports the effectiveness to-date of antibiotic stewardship programs in Australia. While continued efforts are required to further mitigate antibiotic resistance, this study suggests target areas may include improving clinician resistance to patient demand for antibiotics and increasing confidence in prescribing for special populations such as Indigenous peoples and the extremes of age.

An acquired mechanism of antifungal drug resistance simultaneously enables Candida albicans to escape from intrinsic host defenses.

The opportunistic fungal pathogen Candida albicans frequently produces genetically altered variants to adapt to environmental changes and new host niches in the course of its life-long association with the human host. Gain-of-function mutations in zinc cluster transcription factors, which result in the constitutive upregulation of their target genes, are a common cause of acquired resistance to the widely used antifungal drug fluconazole, especially during long-term therapy of oropharyngeal candidiasis. In this study, we investigated if C. albicans also can develop resistance to the antimicrobial peptide histatin 5, which is secreted in the saliva of humans to protect the oral mucosa from pathogenic microbes. As histatin 5 has been shown to be transported out of C. albicans cells by the Flu1 efflux pump, we screened a library of C. albicans strains that contain artificially activated forms of all zinc cluster transcription factors of this fungus for increased FLU1 expression. We found that a hyperactive Mrr1, which confers fluconazole resistance by upregulating the multidrug efflux pump MDR1 and other genes, also causes FLU1 overexpression. Similarly to the artificially activated Mrr1, naturally occurring gain-of-function mutations in this transcription factor also caused FLU1 upregulation and increased histatin 5 resistance. Surprisingly, however, Mrr1-mediated histatin 5 resistance was mainly caused by the upregulation of MDR1 instead of FLU1, revealing a previously unrecognized function of the Mdr1 efflux pump. Fluconazole-resistant clinical C. albicans isolates with different Mrr1 gain-of-function mutations were less efficiently killed by histatin 5, and this phenotype was reverted when MRR1 was deleted. Therefore, antimycotic therapy can promote the evolution of strains that, as a consequence of drug resistance mutations, simultaneously have acquired increased resistance against an innate host defense mechanism and are thereby better adapted to certain host niches.

Candida albicans Shed Msb2 and Host Mucins Affect the Candidacidal Activity of Salivary Hst 5.

Salivary Histatin 5 (Hst 5) is an antimicrobial peptide that exhibits potent antifungal activity towards Candida albicans, the causative agent of oral candidiasis. However, it exhibits limited activity in vivo, largely due to inactivation by salivary components of both host and pathogen origin. Proteins secreted by C. albicans during infection such as secreted aspartyl proteases (Saps) and shed mucin Msb2 can reduce Hst 5 activity; and human salivary mucins, while suggested to protect Hst 5 from proteolytic degradation, can entrap peptides into mucin gels, thereby reducing bioavailability. We show here that Sap6 that is secreted during hyphal growth reduces Hst 5 activity, most likely a result of proteolytic degradation of Hst 5 since this effect is abrogated with heat inactivated Sap 6. We further show that just like C. albicans shedding Msb2, mammalian mucins, fetuin and porcine gut mucin (that is related to salivary mucins), also reduce Hst 5 activity. However, we identify mucin-like protein-induced changes in C. albicans cell morphology and aggregation patterns, suggesting that the effect of such proteins on Hst 5 cannot be interpreted independently of their effect on yeast cells.