Fluconazole Resistance Research Articles

A multidimensional assessment of in-host fitness costs of drug resistance in the opportunistic fungal pathogen Candida glabrata.

Drug-resistant microbes typically carry mutations in genes involved in critical cellular functions and may therefore be less fit under drug-free conditions than susceptible strains. Candida glabrata is a prevalent opportunistic yeast pathogen with a high rate of fluconazole resistance (FLZR), echinocandin resistance (ECR), and multidrug resistance (MDR) relative to other Candida. However, the fitness of C. glabrata MDR isolates, particularly in the host, is poorly characterized, and studies of FLZR isolate fitness have produced contradictory findings. Two important host niches for C. glabrata are macrophages, in which it survives and proliferates, and the gut. Herein we used a collection of clinical and lab-derived C. glabrata isolates to show that FLZR C. glabrata isolates are less fit inside macrophages than susceptible isolates and that this fitness cost is reversed by acquiring ECR mutations. Interestingly, dual-RNAseq revealed that macrophages infected with drug-resistant isolates mount an inflammatory response whereas intracellular drug-resistant cells downregulate processes required for in-host adaptation. Furthermore, drug-resistant isolates were outcompeted by their susceptible counterparts during gut colonization and in infected kidneys, while showing comparable fitness in the spleen. Collectively, our study shows that macrophage-rich organs, such as the spleen, favor the retention of MDR isolates of C. glabrata.

Batzelladine D, a Marine Natural Product, Reverses the Fluconazole Resistance Phenotype Mediated by Transmembrane Transporters in Candida albicans and Interferes with Its Biofilm: An In Vitro and In Silico Study

Numerous Candida species are responsible for fungal infections; however, Candida albicans stands out among the others. Treatment with fluconazole is often ineffective due to the resistance phenotype mediated by transmembrane transporters and/or biofilm formation, mechanisms of resistance commonly found in C. albicans strains. A previous study by our group demonstrated that batzelladine D can inhibit the Pdr5p transporter in Saccharomyces cerevisiae. In the present study, our aim was to investigate the efficacy of batzelladine D in inhibiting the main efflux pumps of Candida albicans, CaCdr1p and CaCdr2p, as well as to evaluate the effect of the compound on C. albicans biofilm. Assays were conducted using a clinical isolate of Candida albicans expressing both transporters. Additionally, to allow the study of each transporter, S. cerevisiae mutant strains overexpressing CaCdr1p or CaCdr2p were used. Batzelladine D was able to reverse the fluconazole resistance phenotype by acting on both transporters. The compound synergistically improved the effect of fluconazole against the clinical isolate when tested in the Caenorhabditis elegans animal model. Moreover, the compound disrupted the preformed biofilm. Based on the obtained data, the continuation of batzelladine D studies as a potential new antifungal agent and/or chemosensitizer in Candida albicans infections can be suggested.

Detection of Mutated erg11 and fks1 Genes among Resistant Candida Species Isolated in Pregnant Women in Mbarara, Uganda

Background: Vulvovaginal candidiasis is estimated to range between 35-60% among pregnant women worldwide. The emergency of anti-fungal resistance in Candida species against azoles and caspofungin is a rising concern because there is a limited range of choices of antifungals to be used in pregnant women with low toxicity. In Uganda, the burden of vulvovaginal candidiasis is estimated to be 48%. However, there is limited data regarding in vitro phenotypic and genotypic anti-fungal susceptibility patterns among candida species isolated from pregnant women. Thus, this study aimed to screen for the two mutated famous erg11 and fks1 genes that lead to anti-fungal resistance among clinical candida isolates. Methods: A cross-sectional study involving 90 Candida species isolates previously collected from a larger study carried out from Mbarara regional referral hospital. Phenotypic susceptibility methods (Kirby-Bauer and minimum inhibitory concentration) while Polymerase chain reaction method and gel electrophoresis were used for detection of the amplified mutated ERG11 and FKS1 genes. Mean and chi-square tests were used to evaluate the associations of resistance patterns between resistant and susceptible isolates. Results: Out of the 90 Candida isolates recovered, 56% were Candida albicans, C.glabrata were 31.11% (28/90), C. parapsilosis and C.famata accounted for 4.44% (4/90) each, C. krusei accounted for 3.33% (3/90) while C. tropicalis accounted for 1.11% (1/90). All the Candida isolates were susceptible to caspofungin while fluconazole resistance was 34.4%. The FKS1 mutated gene was not detected in randomly selected caspofungin susceptible isolates. The ERG11 mutated gene was detected in 80.6% of the fluconazole-resistant isolates and 87.5% of the isolates with intermediate activity towards fluconazole. Conclusion: This study provides evidence that mutated erg11 gene causes reduced fluconazole drug susceptibility (p-value 0.001). Susceptible dose dependence should not be ignored as it may be associated with ERG11 gene mutation leading to resistance to fluconazole.

Multicentre Study of Candida parapsilosis Blood Isolates in Türkiye Highlights an Increasing Rate of Fluconazole Resistance and Emergence of Echinocandin and Multidrug Resistance.

Worldwide emergence of clonal outbreaks caused by fluconazole-resistant (FLCR) and the recent emergence of echinocandin- and multidrug-resistant (ECR and MDR) Candida parapsilosis isolates pose serious threats to modern clinics. Conducting large-scale epidemiological studies aimed at determining the genetic composition and antifungal resistance rates is necessary to devise antifungal stewardship and infection control strategies at international, national and local levels. Despite being severely hit by outbreaks due to FLCR C. parapsilosis isolates, such knowledge at the national level is lacking in Türkiye. Herein, we conducted a prospective multicentre study involving five major clinical centres in Türkiye to determine antifungal resistance rates, underlying mechanisms and genetic composition of all isolates. In total, 341 isolates were collected from 265 patients including clinical information. Antifungal susceptibility testing against common antifungals was performed in addition to sequencing of ERG11 and FKS1. Last, isolates were genotyped with short tandem repeat (STR) genotyping to investigate potential nosocomial transmission. The FLCR rate was 26.7% (91/341), out of which 75.8% (69/91) harboured the ERG11Y132F mutation. Patients infected with FLCR isolates had a higher mortality rate compared to their susceptible counterparts (49% for FLCR vs. 42% for susceptible). ECR rate was 2.1% (7/341) and isolates carried FKS1F652L/R658G/W1370R mutations. Concerningly, four ECR isolates were MDR. FLCR isolates grouped in distinct clusters without evidence of inter-hospital transmission, whereas large clusters containing susceptible isolates from all centres were noted. Overall, the increasing prevalence of FLCR C. parapsilosis at national level and the emergence of ECR and MDR isolates pose serious clinical challenges in Türkiye. Therefore, conducting large-scale epidemiological studies are critical to determine the trend of antifungal resistance and to tailor pertinent antifungal stewardship and infection control strategies to effectively curb the spread of drug-resistant C. parapsilosis.

Molecular and Microbiological Characterization of Candida auris Strains Isolated from Colonized and Infected Patients in a New York City Tertiary Care Center

Abstract Candida auris is a multidrug resistant pathogen that causes healthcare associated invasive infections and outbreaks with high mortality rates, often involving intensive care units (ICU). High rates of C. auris colonization among patients in ICUs are associated with increased risk of subsequent C. auris hospital-acquired infections. In this study, microbiological characteristics, and antifungal susceptibility patterns of C. auris isolates collected between January and December 2023 were investigated. Whole-genome sequencing was used to examine the genetic variation and molecular epidemiological characteristics. During the study period, a total of 20 patients were found to be colonized with C. auris by three site skin swab and eight of these patients subsequently developed invasive infection with C. auris. Among patients with both colonization and infection, all invasive isolates were closely related genetically to colonizing isolates and belonged to clade I. Candidemia was the most common infection identified in 4/8 (50%) patients. All isolates were resistant to fluconazole and thirteen isolates from six patients were resistant to amphotericin B. Paired isolates from patients with both colonization and invasive infection showed the same antifungal resistance patterns. All isolates had the ERG11 p.K143R mutation known to cause fluconazole resistance. Three isolates from one patient with a FKS1 hotspot region 3 p.M690I mutation were resistant to caspofungin and two isolates from one patient with a FKS1 hotspot region 2 p.R1354H mutation were resistant to caspofungin, anidulafungin and micafungin. Our study demonstrates that colonization with C. auris is associated with increased risk of infection by the same strain amongst critical ill patients. C. auris screening and implementation of adequate infection control measures are urgently needed to prevent the dissemination of C. auris infections.

Temporal transcriptional response of Candida glabrata during macrophage infection reveals a multifaceted transcriptional regulator CgXbp1 important for macrophage response and fluconazole resistance.

Candida glabrata can thrive inside macrophages and tolerate high levels of azole antifungals. These innate abilities render infections by this human pathogen a clinical challenge. How C. glabrata reacts inside macrophages and what is the molecular basis of its drug tolerance are not well understood. Here, we mapped genome-wide RNA polymerase II (RNAPII) occupancy in C. glabrata to delineate its transcriptional responses during macrophage infection in high temporal resolution. RNAPII profiles revealed dynamic C. glabrata responses to macrophages with genes of specialized pathways activated chronologically at different times of infection. We identified an uncharacterized transcription factor (CgXbp1) important for the chronological macrophage response, survival in macrophages, and virulence. Genome-wide mapping of CgXbp1 direct targets further revealed its multi-faceted functions, regulating not only virulence-related genes but also genes associated with drug resistance. Finally, we showed that CgXbp1 indeed also affects fluconazole resistance. Overall, this work presents a powerful approach for examining host-pathogen interaction and uncovers a novel transcription factor important for C. glabrata's survival in macrophages and drug tolerance.

Elucidation of the mechanisms of fluconazole resistance and repurposing treatment options against urinary Candida spp. isolated from hospitalized patients in Alexandria, Egypt

BackgroundThe incidence of fungal urinary tract infections (UTIs) has dramatically increased in the past decades, with Candida arising as the predominant etiological agent. Managing these infections poses a serious challenge to clinicians, especially with the emergence of fluconazole-resistant (FLC-R) Candida species. In this study, we aimed to determine the mechanisms of fluconazole resistance in urinary Candida spp. isolated from hospitalized patients in Alexandria, Egypt, assess the correlation between fluconazole resistance and virulence, and explore potential treatment options for UTIs caused by FLC-R Candida strains.ResultsFluconazole susceptibility testing of 34 urinary Candida isolates indicated that 76.5% were FLC-R, with a higher prevalence of resistance recorded in non-albicans Candida spp. (88.9%) than in Candida albicans (62.5%). The calculated Spearman’s correlation coefficients implied significant positive correlations between fluconazole minimum inhibitory concentrations and both biofilm formation and phospholipase production. Real-time PCR results revealed that most FLC-R isolates (60%) significantly overexpressed at least one efflux pump gene, while 42.3% significantly upregulated the ERG11 gene. The most prevalent mutation detected upon ERG11 sequencing was G464S, which is conclusively linked to fluconazole resistance. The five repurposed agents: amikacin, colistin, dexamethasone, ketorolac, and sulfamethoxazole demonstrated variable fluconazole-sensitizing activities in vitro, with amikacin, dexamethasone, and colistin being the most effective. However, the fluconazole/colistin combination produced a notable reduction (49.1%) in bladder bioburden, a 50% decrease in the inflammatory response, and tripled the median survival span relative to the untreated murine models.ConclusionsThe fluconazole/colistin combination offers a promising treatment option for UTIs caused by FLC-R Candida, providing an alternative to the high-cost, tedious process of novel antifungal drug discovery in the battle against antifungal resistance.

Microbiological Profile and Antimicrobial Susceptibility Pattern of Uropathogens Isolated From Pregnant Women Attending a Tertiary Care Hospital in Central India.

Urinary tract infections (UTIs) are more common in females than males, and during pregnancy, frequency is increased due to hormonal changes affecting the anatomy and physiology. If not treated on time, complications may develop in both the mother and fetus. This study aimed to analyze the microbiological (bacterial and yeast) profile of the isolated uropathogens and their antimicrobial susceptibility. Aseptically collected urine specimens were processed by the standard loop method. Identification of uropathogens was done by standard microbiological tests, and antimicrobial susceptibility was performed as per the Clinical and Laboratory Standards Institute (CLSI) guidelines. Out of 400 study participants, UTI was detected in 26.75% (107/400) of cases. Overall, Escherichia coli (42.99%, 46/107) was the most frequently isolated uropathogen followed by Candida species (20.56%, 22/107), whereas, among gram-positive bacteria, Enterococcus faecalis (7.47%, 8/107) was the commonest. Among Enterobacterales, extended-spectrum beta-lactamase (ESBL) and AmpC beta-lactamase (AmpC) production was observed in 19.35% (12/62, eight E. coli and four Klebsiella) and 6.45% (4/62, all E. coli) isolates, respectively. However, all the gram-negative organisms showed 100% (n=70) sensitivity to carbapenems. Among staphylococci, 50% (2/4) of isolates were methicillin-resistant, and high-level aminoglycoside resistance (HLAR) was observed in 75% (6/8) of Enterococcus faecalis isolates, but all were sensitive to vancomycin, linezolid, and nitrofurantoin. Among Candida species, non-albicans Candida (NAC) species (59%, 13/22) outweigh the number of Candida albicans (41%, 9/22), with Candida tropicalis (22.72%, 5/22) being the commonest NAC. All the Candida species were susceptible to voriconazole, whereas fluconazole resistance was observed in 31.81% (7/22) of Candida isolates, with a higher percentage in NAC species (22.72%, 5/22) than in Candida albicans (9.09%, 2/22). UTIs in pregnancy are caused by a spectrum of both gram-negative bacilli and gram-positive cocci. Moreover, there is a rise of both Candida albicans and NAC species as uropathogens, with the emergence of fluconazole resistance. There is an increased prevalence of asymptomatic bacteriuria in pregnancy, which necessitates the screening of pregnant females for UTI using culture and antimicrobial susceptibility testing.

Similarities and distinctions in the activation of the Candida glabrata Pdr1 regulatory pathway by azole and non-azole drugs.

Incidences of fluconazole (FLC) resistance among Candida glabrata clinical isolates is a growing issue in clinics. The pleiotropic drug response (PDR) network in C. glabrata confers azole resistance and is defined primarily by the Zn2Cys6 zinc cluster-containing transcription factor Pdr1 and target genes such as CDR1, that encodes an ATP-binding cassette transporter protein thought to act as a FLC efflux pump. Mutations in the PDR1 gene that render the transcription factor hyperactive are the most common cause of fluconazole resistance among clinical isolates. The phenothiazine class drug fluphenazine and a molecular derivative, CWHM-974, which both exhibit antifungal properties, have been shown to induce the expression of Cdr1 in Candida spp. We have used a firefly luciferase reporter gene driven by the CDR1 promoter to demonstrate two distinct patterns of CDR1 promoter activation kinetics: gradual promoter activation kinetics that occur in response to ergosterol limitations imposed by exposure to azole and polyene class antifungals and a robust and rapid CDR1 induction occurring in response to the stress imposed by fluphenazines. We can attribute these different patterns of CDR1 induction as proceeding through the promoter region of this gene since this is the only segment of the gene included in the luciferase reporter construct. Genetic analysis indicates that the signaling pathways responsible for phenothiazine and azole induction of CDR1 overlap but are not identical. The short time course of phenothiazine induction suggests that these compounds may act more directly on the Pdr1 protein to stimulate its activity.

Evaluation of Etest and MICRONAUT-AM Assay for Antifungal Susceptibility Testing of Candida auris: Underestimation of Fluconazole Resistance by MICRONAUT-AM and Overestimation of Amphotericin B Resistance by Etest.

Multidrug-resistant Candida auris has recently caused major outbreaks in healthcare facilities. Rapid and accurate antifungal susceptibility testing (AST) of C. auris is crucial for proper management of invasive infections. The Commercial Sensititre Yeast One and Vitek 2 methods underestimate or overestimate the resistance of C. auris to fluconazole and amphotericin B (AMB). This study evaluated the AST results of C. auris against fluconazole and AMB by gradient-MIC-strip (Etest) and broth microdilution-based MICRONAUT-AM-EUCAST (MCN-AM) assays. Clinical C. auris isolates (n = 121) identified by phenotypic and molecular methods were tested. Essential agreement (EA, ±1 two-fold dilution) between the two methods and categorical agreement (CA) based on the Centers for Disease Control and Prevention's (CDC's) tentative resistance breakpoints were determined. Fluconazole resistance-associated mutations were detected by PCR-sequencing of ERG11. All isolates identified as C. auris belonged to South Asian clade I and contained the ERG11 Y132F or K143R mutation. The Etest-MCN-AM EA was poor (33%) for fluconazole and moderate (76%) for AMB. The CA for fluconazole was higher (94.2%, 7 discrepancies) than for AMB (91.7%, 10 discrepancies). Discrepancies were reduced when an MCN-AM upper-limit value of 4 µg/mL for fluconazole-susceptible C. auris and an Etest upper-limit value of 8 µg/mL for the wild type for AMB were used. Our data show that resistance to fluconazole was underestimated by MCN-AM, while resistance to AMB was overestimated by Etest when using the CDC's tentative resistance breakpoints of ≥32 µg/mL for fluconazole and ≥2 µg/mL for AMB. Method-specific resistance breakpoints should be devised for accurate AST of clinical C. auris isolates for proper patient management.

Increasing rate of non-Candida albicans yeasts and fluconazole resistance in yeast isolates from women with recurrent vulvovaginal candidiasis in Leeds, United Kingdom

ObjectivesAzoles have been the mainstay of recurrent vulvovaginal candidiasis (RVVC) for many years. Because of a recent anecdotal increase in non-Candida albicans yeasts (NCAY) and azole-resistant C. albicans cases, their...

Step-wise evolution of azole resistance through copy number variation followed by KSR1 loss of heterozygosity in Candida albicans.

Antimicrobial drug resistance poses a global health threat, requiring a deeper understanding of the evolutionary processes that lead to its emergence in pathogens. Complex evolutionary dynamics involve multiple mutations that can result in cooperative or competitive (clonal interference) effects. Candida albicans, a major fungal pathogen, displays high rates of copy number variation (CNV) and loss of heterozygosity (LOH). CNV and LOH events involve large numbers of genes and could synergize during evolutionary adaptation. Understanding the contributions of CNV and LOH to antifungal drug adaptation is challenging, especially in the context of whole-population genome sequencing. Here, we document the sequential evolution of fluconazole tolerance and then resistance in a C. albicans isolate involving an initial CNV on chromosome 4, followed by an LOH on chromosome R that involves KSR1. Similar LOH events involving KSR1, which encodes a reductase in the sphingolipid biosynthesis pathway, were also detected in independently evolved fluconazole resistant isolates. We dissect the specific KSR1 codons that affect fluconazole resistance and tolerance. The combination of the chromosome 4 CNV and KSR1 LOH results in a >500-fold decrease in azole susceptibility relative to the progenitor, illustrating a compelling example of rapid, yet step-wise, interplay between CNV and LOH in drug resistance evolution.

Molecular Evaluation of the mRNA Expression of the ERG11, ERG3, CgCDR1, and CgSNQ2 Genes Linked to Fluconazole Resistance in Candida glabrata in a Colombian Population.

The study of Candida glabrata genes associated with fluconazole resistance, from a molecular perspective, increases the understanding of the phenomenon with a view to its clinical applicability. We sought to establish the predictive molecular profile of fluconazole resistance in Candida glabrata by analyzing the ERG11, ERG3, CgCDR1, and CgSNQ2 genes. Expression was quantified using RT-qPCR. Metrics were obtained through molecular docking and Fisher discriminant functions. Additionally, a predictive classification was made against the susceptibility of C. glabrata to fluconazole. The relative expression of the ERG3, CgCDR1, and CgSNQ2 genes was higher in the fluconazole-resistant strains than in the fluconazole-susceptible, dose-dependent strains. The gene with the highest relative expression in the fluconazole-exposed strains was CgCDR1, and in both the resistant and susceptible, dose-dependent strains exposed to fluconazole, this was also the case. The molecular docking model generated a median number of contacts between fluconazole and ERG11 that was lower than the median number of contacts between fluconazole and ERG3, -CgCDR1, and -CgSNQ2. The predicted classification through the multivariate model for fluconazole susceptibility achieved an accuracy of 73.5%. The resistant strains had significant expression levels of genes encoding efflux pumps and the ERG3 gene. Molecular analysis makes the identification of a low affinity between fluconazole and its pharmacological target possible, which may explain the lower intrinsic susceptibility of the fungus to fluconazole.

Overlapping coactivator function is required for transcriptional activation by the Candida glabrata Pdr1 transcription factor.

Azole resistance in the pathogenic yeast Candida glabrata is a serious clinical complication and increasing in frequency. The majority of resistant organisms have been found to contain a substitution mutation in the Zn2Cys6 zinc cluster-containing transcription factor Pdr1. These mutations typically lead to this factor driving high, constitutive expression of target genes like the ATP-binding cassette transporter-encoding gene CDR1. Overexpression of Cdr1 is required for the observed elevated fluconazole resistance exhibited by strains containing one of these hyperactive PDR1 alleles. While the identity of hyperactive PDR1 alleles has been extensively documented, the mechanisms underlying how these gain-of-function (GOF) forms of Pdr1 lead to elevated target gene transcription are not well understood. We have used a tandem affinity purification-tagged form of Pdr1 to identify coactivator proteins that biochemically purify with the wild-type and 2 different GOF forms of Pdr1. Three coactivator proteins were found to associate with Pdr1: the SWI/SNF complex Snf2 chromatin remodeling protein and 2 different components of the SAGA complex, Spt7 and Ngg1. We found that deletion mutants lacking either SNF2 or SPT7 exhibited growth defects, even in the absence of fluconazole challenge. To overcome these issues, we employed a conditional degradation system to acutely deplete these coactivators and determined that loss of either coactivator complex, SWI/SNF or SAGA, caused defects in Pdr1-dependent transcription. A double degron strain that could be depleted for both SWI/SNF and SAGA exhibited a profound defect in PDR1 autoregulation, revealing that these complexes work together to ensure high-level Pdr1-dependent gene transcription.

Long-term stability of acquired drug resistance and resistance associated mutations in the fungal pathogen Nakaseomyces glabratus (Candida glabrata).

The limited number of available antifungal drugs and the increasing number of fungal isolates that show drug or multidrug resistance pose a serious medical threat. Several yeast pathogens, such as Nakaseomyces glabratus (Candida glabrata), show a remarkable ability to develop drug resistance during treatment through the acquisition of genetic mutations. However, how stable this resistance and the underlying mutations are in non-selective conditions remains poorly characterized. The stability of acquired drug resistance has fundamental implications for our understanding of the appearance and spread of drug-resistant outbreaks and for defining efficient strategies to combat them. Here, we used an in vitro evolution approach to assess the stability under optimal growth conditions of resistance phenotypes and resistance-associated mutations that were previously acquired under exposure to antifungals. Our results reveal a remarkable stability of the resistant phenotype and the underlying mutations in a significant number of evolved populations, which conserved their phenotype for at least two months in the absence of drug-selective pressure. We observed a higher stability of anidulafungin resistance over fluconazole resistance, and of resistance-conferring point mutations as compared with aneuploidies. In addition, we detected accumulation of novel mutations in previously altered resistance-associated genes in non-selective conditions, which suggest a possible compensatory role. We conclude that acquired resistance, particularly to anidulafungin, is a long-lasting phenotype, which has important implications for the persistence and propagation of drug-resistant clinical outbreaks.

Candida albicans in the oral cavities of pets: biofilm formation, putative virulence, antifungal resistance profiles and classification of the isolates.

The study aimed to investigate Candida albicans presence, antifungal resistance, biofilm formation, putative virulence genes, and molecular characterization in oral samples of dogs and cats. A total of 239 oral samples were collected from cats and dogs of various breeds and ages at Erciyes University, Faculty of Veterinary Medicine Clinics, between May 2017 and April 2018. Among 216 isolates obtained, 15 (6.95%) were identified as C. albicans, while 8 (3.7%) were non-albicans Candida species. Antifungal susceptibility testing revealed sensitivities to caspofungin, fluconazole, and flucytosine in varying proportions. Molecular analysis indicated the presence of fluconazole and caspofungin resistance genes in all C. albicans isolates. Additionally, virulence genes ALS1, HWP1, and HSP90 showed variable presence. Biofilm formation varied among isolates, with 46.7% strong, 33.3% moderate, and 20% weak producers. PCA analysis categorized isolates into two main clusters, with some dog isolates grouped separately. The findings underscore the significance of oral care and protective measures in pets due to C. albicans prevalence, biofilm formation, virulence factors, and antifungal resistance in their oral cavity, thereby aiding clinical diagnosis and treatment in veterinary medicine.

Creation of a Multi-Year Pediatric Candidemia Antibiogram in Georgia Identifies Changing Epidemiology and Resistance Trends

Background: Invasive candidiasis, including candidemia, is a significant cause of morbidity and mortality in medically complex and immunocompromised children. Understanding the epidemiology and antifungal susceptibility patterns of Candida infections could help guide empiric antifungal therapy. Methods: This fungal antibiogram was created at a large quaternary children’s health system in Georgia. Blood isolates positive for Candida spp. from 2019 through 2023 were included. The number and percentage of isolates for each Candida spp was recorded by year and then as the combined 5-year total. The Clinical and Laboratory Standards Institute (CLSI) antifungal interpretative criteria were used, and we only included one unique Candida spp isolate per patient. Due to the limited number of isolates, the combined 5 years of isolates were used to create the fungal antibiogram. Data are shown as percent susceptible using CLSI interpretative criteria and number of isolates. Results: Between 2019 and 2023 there were 124 unique blood isolates of Candida spp identified. The most common isolates were C. albicans (33%), C. parapsilosis (27%), C. glabrata (14%) and C. tropicalis (11%). Over the 5 years of the study, the percentage of C. albicans isolates decreased from 47% to 21%. The change in epidemiology was not driven by a single Candida species but varied from year to year. For C. albicans, susceptibility was 100% for fluconazole and micafungin. For C. parapsilosis, susceptibility to fluconazole and micafungin was 97% and 94%, respectively. Fluconazole susceptibility was lowest for C. glabrata (88%) and C. krusei (0%). Using CLSI epidemiological cutoff values (ECV) to evaluate the amphotericin B results, none of the isolates had results greater than the CLSI ECVs. Comparing 2019 and 2023, the percentage of Candida blood isolates resistant to fluconazole increased from 5% to 18.5%. Conclusion: C. albicans was the most frequently identified cause of candidemia in children, but there was a gradual increase in fungemia caused by other Candida spp. over the past 5 years including Candida with fluconazole resistance. Overall, our findings demonstrate high susceptibility rates to fluconazole and echinocandins in Candida spp. blood isolates. Further research is needed to identify risk factors for antifungal resistant candidemia in pediatric patients.Disclosure: Mark Gonzalez: Honoria for a one time consulation with NaviDx consulting in May of 2022. Honoria from the American Society for Microbiology for writing of a chapter in the Clinical Microbiology Procedures Handbook.

Clinical and microbiological characteristics of a hospital outbreak of Candida auris in a referral hospital in Lima, Peru.

Candida auris, a multidrug-resistant fungal pathogen, has received considerable attention owing to its recent surge, especially in South America, which coincides with the ongoing global COVID-19 pandemic. Understanding the clinical and microbiological characteristics of outbreaks is crucial for their effective management and control. This retrospective observational study aimed to characterize a C. auris outbreak at a Peruvian referral hospital between January 2021 and July 2023. Data were collected from hospitalized patients with positive C. auris culture results. Microbiological data and antifungal susceptibility test results were analysed. Additionally, infection prevention and control measures have been described. Statistical analysis was used to compare the characteristics between the infected and colonized patients. Thirty-three patients were identified, mostly male (66.7%), with a median age of 53 years. Among them, 18 (54.5%) were colonized, and 15 (45.5%) were infected. Fungemia was the predominant presentation (80%), with notable cases of fungemia in tuberculosis patients with long-stay devices for parenteral anti-tuberculosis therapy. Seventy-five percent of the isolates exhibited fluconazole resistance. Echinocandins were the primary treatment, preventing fungemia recurrence within 30 days. Infected patients had significantly longer hospital stays than colonized patients (100 vs. 45 days; p = .023). Hospital mortality rates were 46.7% and 25% in the infected and fungemia patients, respectively. Simultaneous outbreaks of multidrug-resistant bacteria were documented. This study underscores the severity of a C. auris outbreak at a referral hospital in Peru, highlighting its significant impact on patient outcomes and healthcare resources. The high prevalence of fluconazole-resistant isolates, leading to prolonged hospital stay and high mortality rates, particularly in cases of fungemia, underscores the critical need for effective infection prevention and control strategies.

Resistance to Antifungals in Non-albicans Candida Species Isolates in the Southeast Region

Background: Antimicrobial resistance is a growing problem in Candida spp., leading to treatment challenges and increased morbidity and mortality. The World Health Organization (WHO) fungal priority pathogens list classifies C. glabrata, C. tropicalis, and C. parapsilosis as high priority and leading causes of candidemia with high fluconazole resistance. In the US, these organisms are the most frequently isolated non-albicans Candida species. In 2016, the Antibiotic Resistance Laboratory Network (ARLN) was created to monitor resistance threats, including in Candida spp. This study describes the proportion of resistance in C. glabrata, C. parapsilosis, and C. tropicalis isolates sent to the Southeast ARLN from 2017 to 2023. Methods: This study evaluated C. glabrata, C. parapsilosis, and C. tropicalis submitted to the Southeast ARLN from Alabama, Florida, Georgia, Louisiana, Mississippi, and Tennessee from February 2017- September 2023. Species identification was confirmed by Bruker Biotyper matrix assisted laser desorption-ionization time of flight (MALDI-TOF). Antifungal susceptibility testing (AFST) was performed using TREK frozen broth microdilution panels. Minimum inhibitory concentration values from the clinical instrument were used to determine susceptibility based on Clinical and Laboratory Standards Institute (CLSI) standard interpretations from the 2020 CLSI M60 guidelines. Data were extracted from the laboratory information management system. Analyses were conducted using SAS v9.4. Results: AFST testing was performed on 660 C. glabrata, 500 C. parapsilosis, and 233 C. tropicalis isolates from within the Southeast region. The predominant specimen sources by species were blood 25.30% C. glabrata; other/not specified 27.80% C. parapsilosis; and lower respiratory 36.91% C. tropicalis. Resistance to fluconazole is as follows: C. glabrata, 12.88%; C. parapsilosis, 3.41%; C. tropicalis, 36.64%. Resistance to voriconazole is as follows: C. parapsilosis, 1.00%; C. tropicalis 30.04%. Resistance to at least one echinocandin (Anidulafungin, Capsofungin, Micafungin) is as follows: C. glabrata, 1.67%; C. parapsilosis, 0.60%; C. tropicalis, 0.43%. Overall, there was a decreasing trend in resistance to fluconazole, and voriconazole in all three species between 2017 and 2023. Conclusions: Antifungal resistance in non-albicans Candida species represents an emerging public health threat, however, within the Southeast region, ARLN data has shown a decreasing trend of azole resistance. This may be due in part to changes in reporting requirements and submission criteria from within the region. Nevertheless, C. tropicalis showed high resistance to azoles within the Southeast region. These Candida species should be monitored to inform clinical decision making and identify resistance patterns in other US regions due to their increase in resistance worldwide.

Increase in candidemia cases and emergence of fluconazole-resistant Candida parapsilosis and C. auris isolates in a tertiary care academic hospital during the COVID-19 pandemic, Greece, 2020 to 2023.

BackgroundThe COVID-19 pandemic and the emergence of Candida auris have changed the epidemiological landscape of candidaemia worldwide.AimWe compared the epidemiological trends of candidaemia in a Greek tertiary academic hospital before (2009-2018) and during the early COVID-19 (2020-2021) and late COVID-19/early post-pandemic (2022-2023) era.MethodsIncidence rates, species distribution, antifungal susceptibility profile and antifungal consumption were recorded, and one-way ANOVA or Fisher's exact test performed. Species were identified by MALDI-ToF MS, and in vitro susceptibility determined with CLSI M27-Ed4 for C. auris and the EUCAST-E.DEF 7.3.2 for other Candida spp.ResultsIn total, 370 candidaemia episodes were recorded during the COVID-19 pandemic. Infection incidence (2.0 episodes/10,000 hospital bed days before, 3.9 during the early and 5.1 during the late COVID-19 era, p<0.0001), C. auris (0%, 9% and 33%, p<0.0001) and fluconazole-resistant C. parapsilosis species complex (SC) (20%, 24% and 33%, p=0.06) infections increased over time, with the latter not associated with increase in fluconazole/voriconazole consumption. A significant increase over time was observed in fluconazole-resistant isolates regardless of species (8%, 17% and 41%, p<0.0001). Resistance to amphotericin B or echinocandins was not recorded, with the exception of a single pan-echinocandin-resistant C. auris strain.ConclusionCandidaemia incidence nearly tripled during the COVID-19 era, with C. auris among the major causative agents and increasing fluconazole resistance in C. parapsilosis SC. Almost half of Candida isolates were fluconazole-resistant, underscoring the need for increased awareness and strict implementation of infection control measures.