Antifungal Resistance Trends of Candida auris Clinical Isolates, New York-New Jersey, 2016-2020

About 55% of U.S. Candida auris clinical cases were reported from New York and New Jersey from 2016 through 2020. Nearly all New York-New Jersey clinical isolates (99.8%) were fluconazole resistant, and 50% were amphotericin B resistant. Echinocandin resistance increased from 0% to 4% and pan-resistance increased from 0 to <1% for New York C. auris clinical isolates but not for New Jersey, highlighting the regional differences.

Bioactive triterpenoids from Solanum torvum fruits with antifungal, resistance modulatory and anti-biofilm formation activities against fluconazole-resistant candida albicans strains

Vulvovaginal candidiasis (VVC) is the second most common vaginal infection that affects women of reproductive age. Its increased occurrence and associated treatment cost coupled to the rise in resistance of the causative pathogen to current antifungal therapies has necessitated the need for the discovery and development of novel effective antifungal agents for the treatment of the disease. We report in this study the anti-Candida albicans activity of Solanum torvum 70% ethanol fruit extract (STF), fractions and some isolated compounds against four (4) fluconazole-resistant strains of C. albicans. We further report on the effect of the isolated compounds on the antifungal activity of fluconazole and voriconazole in the resistant isolates as well as their inhibitory effect on C. albicans biofilm formation. STF was fractionated using n-hexane, chloroform (CHCl3) and ethyl acetate (EtOAc) to obtain four respective major fractions, which were then evaluated for anti-C. albicans activity using the microbroth dilution method. The whole extract and fractions recorded MICs that ranged from 0.25 to 16.00 mg/mL. From the most active fraction, STF- CHCl3 (MIC = 0.25–1.00 mg/mL), four (4) known compounds were isolated as Betulinic acid, 3-oxo-friedelan-20α-oic acid, Sitosterol-3-β-D-glucopyranoside and Oleanolic acid. The compounds demonstrated considerably higher antifungal activity (0.016 to 0.512 mg/mL) than the extract and fractions and caused a concentration-dependent anti-biofilm formation activity. They also increased the sensitivity of the C. albicans isolates to fluconazole. This is the first report of 3-oxo-friedelan-20α-oic acid in the plant as well as the first report of betulinic acid, sitosterol-3-β-D-glucopyranoside and oleanolic acid from the fruits of S. torvum. The present study has demonstrated the anti-C. albicans activity of the constituents of S. torvum ethanol fruit extract and also shown that the constituents possess anti-biofilm formation and resistance modulatory activities against fluconazole-resistant clinical C. albicans isolates.

Azole uptake in Candida auris is strongly correlated with drug resistance

Analyses of fluconazole uptake in clinical isolates of C. auris, with wide ranging drug resistance profiles, has revealed interesting differences within the species as well as major distinctions from other yeast species. We previously proposed that prevention of drug uptake is a potential mechanism of drug resistance and our C. auris data provide further support for this. We developed an assay using radio-labeled fluconazole to measure intracellular azole accumulation in fungal cells. The assay is performed under glucose-replete conditions to inhibit ATP-dependent efflux. A comparative study measuring fluconazole uptake in 63 C. auris isolates as well as a panel of other species such as C. albicans, S. cerevisiae, C. glabrata, C. krusie, C. lusitanea, C. tropicalis, and C. dublinienses revealed striking C. auris phenotypes that we have not seen in other fungal species. There is a strong correlation between fluconazole resistance and reduced drug uptake in C. auris. Fluconazole-resistant C. auris isolates had reduced levels of intracellular fluconazole accumulation compared to susceptible isolates. Drug-resistant C. auris isolates had the lowest drug accumulation of any of the yeast species tested. Fluconazole-susceptible C. auris isolates had dramatically increased fluconazole accumulation compared to the resistant isolates as well as when compared to other yeast species. Of the 63 C. auris isolates, 28 of 32 (∼88%) resistant isolates had extremely low fluconazole uptake, whereas 15 of 18 (∼83%) susceptible isolates had high fluconazole uptake. This association between reduced drug uptake and resistance could be a C. auris-distinctive mechanism of drug resistance.

Candida glabrata MSH2 deletion increases antifungal tolerance in vitro and during intra-abdominal candidiasis (IAC), it does not impact pathogenesis of peritonitis or intra-abdominal abscesses

Background: IAC is the second most common type of invasive Candidiasis, but its pathogenesis is poorly understood. We have shown that Candida albicans DNA damage response genes are strongly induced within intra-abdominal abscesses. Deletion of C. glabrata MSH2, A DNA mismatch repair (MMR) gene, results in a mutator phenotype that facilitates multidrug resistance in vitro and in mouse gastrointestinal tracts. Our goal was to determine if CGMSH2 Contributed to pathogenesis or resistance to the new antifungal rezafungin during IAC. Methods: We createdΔMSH2 in BG2 using SAT-Flipper, and tested virulence and rezafungin responses in a mouse model of IAC. Results: ΔMSH2 displayed no growth defects at 30°C in liquid (YPD, Ypglycerol) or solid media (YPD+0.02% MMS, 1MM H2O2, 1M NACL, 20 UG/ML CW, 250 UG/ML OR 0.02% SDS). ΔMSH2 longevity in YPD was comparable to BG2. Caspofungin-, Rezafungin- and Fluconazole-resistant mutants arose 24-, 16- and 3-fold more often, respectively, for ΔMSH2 than BG2 (108-106 CFU overnight in YPD, selected on 8XMIC-Containing plates). However, respective minimum inhibitory concentrations (MICS) were not different, nor were rezafungin time-kills.ΔMSH2 was comparable to BG2 in peritonitis and abscess burdens in mouse IAC.ΔMSH2 demonstrated significantly greater caspofungin- and fluconazole-tolerance than BG2 in abscesses. Rezafungin reduced peritonitis and abscess burdens ofΔMSH2,BG2 ANDFKS mutant strains to similar extents. Conclusions: CgMSH2 deletionincreased the frequency of spontaneously-arising echinocandin- and fluconazole-resistant colonies in vitro and tolerance in intra-abdominal abscesses, but it did not attenuate virulence or rezafungin responses during IAC.

Susceptibility of Fluconazole-Resistant Candida albicans to Thyme Essential Oil

Candida spp. is the most common microbial pathogen in fungal infections. There has been a tremendous increase in cases of candidiasis, especially among critically ill non-neutropenic patients. Candida albicans’ isolates were procured from the Prince Sultan Military Hospital, Riyadh, KSA. The isolates were characterized for their identification using CHROMagar, carbohydrate metabolism, germ tube formation, and RAPD-PCR techniques. The essential oil of Thymus vulgaris was obtained by hydro-distillation and characterized to decipher the major bioactive phytoconstituents. The antifungal activity of the thyme essential oil (TEO) was evaluated against fluconazole-resistant C. albicans isolates. The major phytocomponents identified by GC/MS were thymol (68.1%) followed by γ-terpinene (8.9%), cymol (7.7%), caryophyllene (1.1%), linalool (1.4%). The TEO successfully reduced the growth of C. albicans isolates. At very low doses, the TEO proved to be fungi static and fungicidal. TEO also effectively inhibited the germ tube formation and budging of fungal pathogens. The time kill assays have shown that TEO was more effective against drug resistant clinical isolates than fluconazole. This study provides an array of experimental evidence regarding the therapeutic efficacy of TEO against the drug-resistant clinical isolates of C. albicans. The findings may be used in the development of a new antifungal agent accordingly.

317. Invasive Fungal Infections in Critically-ill Patients with COVID-19 in Mexico City

Background Invasive fungal infections (IFI) are emergent complications in SARS-CoV-2 infection. We aimed to describe the epidemiology, characteristics and outcome of IFI during the pandemic. Methods Between March 2020 and April 2021, patients admitted to the Intensive Care Unit (ICU) in a COVID-19 center in Mexico City who developed IFI were included. COVID-19 associated pulmonary aspergillosis (CAPA) was defined according to the ECMM/ISHAM criteria. Demographic and clinical data were obtained from the electronic medical record. Descriptive analysis was made. The study was approved by the Institutional Review Board. Results Sixty-seven (67/743, 9%) patients with COVID-19 developed IFI during ICU stay, of which 37 (55%) had CAPA, 24 (36%) had Invasive Candidiasis (IC), 3 Cryptococcosis and 3 pulmonary Mucormycosis. The median age was 57.5 (IQR 48-68) and 46 (69%) were male. Thirty-six (54%) had obesity and 20 (30%) type 2 diabetes. Sixty-two received COVID-19 directed therapy: 48/67 (72%) steroids, 4/67 (6%) tocilizumab and 8/67 (12%) were included in clinical trials. Among 24 patients with IC, 13 (54%) were fluconazole-resistant C. parapsilosis, 11 (46%) C. albicans and 2 C. glabrata. Twenty-two received antifungal treatment, 20 with echinocandins and 2 fluconazole. Among 37 CAPA, 8 (22%) were probable and 29 (78%) possible. Serum galactomannan was positive in 8 (22%), 33 respiratory cultures grew Aspergillus (31 tracheal aspirates and 2 bronchoalveolar lavage). Aspergillus fumigatus was the most frequent isolate in 18/33 (55%). Chest CT showed ground glass opacities in 21 (57%). Most received voriconazole (26/37, 70%). The median time from ICU admission to IFI was 9.5 (IQR 3-14) days. The median ICU and hospital stay length were 30 days (IQR 16-41) and 40 days (IQR 23-49), respectively. In-hospital mortality was 48%. The incidence rate of IC was higher early in the pandemic, due to Infection Control breaches, while higher CAPA incidence may have occurred later due to ventilation system gaps (Figure 1). Bi-monthly Invasive Fungal Infection incidence rate/100 ICU admissions. Conclusion We found 9% incidence of IFIs in critically-ill COVID-19 patients with high mortality. The majority received steroids, had obesity and had a prolonged hospital stay. Most had possible CAPA. An outbreak of fluconazole-resistant C. parapsilosis was found. Disclosures All Authors: No reported disclosures

263. Antibody Spill-Over vs. True Coccidioidal Meningitis in a Patient with HIV/AIDS and Disseminated Coccidioidomycosis

Background The diagnosis of coccidioidal meningitis merits life-long antifungal therapy given high rates of disease recurrence. Accurate diagnosis is important. Antibody spill-over into cerebrospinal fluid (CSF) can happen when serum titers are high. We present a case of antibody spill-over vs. true fluconazole-resistant coccidioidal meningitis. Methods A 49-year-old man presented with 6 months of intermittent fever, myalgias, decreased appetite, vomiting, diarrhea, unsteadiness and 60-pound weight loss. He was recently diagnosed with HIV and a prior lymph node biopsy had grown Coccidioides immitis (C. i) for which he was given fluconazole 100 mg twice daily. Figure 1. Timeline of Coccidioides immitis lab results in relation to treatment regimen. CF: Complement fixation, EIA: Enzyme immunoassay for Coccidioides antigen, CSF: Cerebrospinal fluid, c/mL: copies/mL, R: resistant, S: susceptible. Results Vitals revealed a temperature of 102°F. He was cachectic and a 0.5 cm right supraclavicular lymph node was palpable. No meningeal signs were appreciated. CD4 count was 50/µL (18%), HIV-1 viral load 2,969,945 copies/mL. Computed tomography (CT) of the abdomen/pelvis suggested lung and spleen involvement. Serum C. i enzyme immunoassay (EIA) was 1.38 ng/mL, immunodiffusion (ID) was positive and complement fixation (CF) titer was 1:256. C. i was isolated from expectorated sputum. CSF cell count was normal, but ID was positive and CF titer was 1:2 however, lab reported concern for spill-over due to high serum IgG titers. He left against medical advice with fluconazole 400 mg daily. He was hospitalized a month later for failure to thrive. MRI head revealed enlarged lateral and third ventricles with increased periventricular hyperintensity concerning for coccidioidal meningitis. Repeat serum studies were stable. CSF revealed CF 1:4 and C. i antigen by EIA 1.31ng/mL, distinguishing between spill-over and meningitis. Susceptibility results showed resistance to fluconazole and amphotericin B with minimum inhibitory concentrations (MICs) of 50 and 4 respectively, posaconazole susceptibility (MIC &lt; 1) and itraconazole borderline (MIC 3.7). Despite amphotericin B resistance, it was used for bridge to posaconazole. ART was initiated after concern for immune reconstitution had resolved. Conclusion This case highlights the difficulty in making an accurate diagnosis of coccidioidal meningitis. It also describes a fluconazole-resistant C. i isolate in the setting of prolonged low-dose fluconazole therapy. Disclosures All Authors: No reported disclosures