Caspofungin Research Articles

The Influence of Aspergillus fumigatus Fatty Acid Oxygenases PpoA and PpoC on Caspofungin Susceptibility

Aspergillus fumigatus can cause invasive pulmonary aspergillosis (IPA). Fungicidal azoles and fungistatic caspofungin (CAS) are the first- and second-line therapies, respectively, used to treat IPA. Treatment of A. fumigatus with CAS or micafungin induces the production of the oxylipin 5,8-diHODE by the fungal oxygenase PpoA. For this article, we investigated the influence of ppo genes, which encode the fatty acid oxygenases responsible for oxylipin biosynthesis, on CAS tolerance. The influence of PpoA and PpoC on CAS tolerance is mediated by MpkA phosphorylation and protein kinase A (PKA) activity. RNAseq transcriptional profiling and the label-free quantitative proteomics of the ppoA and ppoC mutants showed that differentially expressed genes and proteins are related to secondary metabolites and carbohydrate metabolism. We also characterized two clinical isolates, CM7555 and IFM61407, which decrease and increase susceptibility to CAS, respectively. CM7555 does not exhibit increased oxylipin production in the presence of CAS but oxylipin induction upon CAS exposure is increased in IFM61407, suggesting that oxylipins are not the only mechanism involved in CAS tolerance in these isolates. Upon CAS exposure, CM7555 has higher MpkA phosphorylation and PKA activity than IFM61407. Our results reveal the different aspects and genetic determinants involved in A. fumigatus CAS tolerance.

Synergistic Antifungal Effect and In Vivo Toxicity of a Monoterpene Isoespintanol Obtained from Oxandra xylopioides Diels.

Candida sp. infections are a threat to global health, with high morbidity and mortality rates due to drug resistance, especially in immunocompromised people. For this reason, the search for new alternatives is urgent, and in recent years, a combined therapy with natural compounds has been proposed. Considering the biological potential of isoespintanol (ISO) and continuing its study, the objective of this research was to assess the effect of ISO in combination with the antifungals fluconazole (FLZ), amphotericin B (AFB) and caspofungin (CASP) against clinical isolates of C. tropicalis and to evaluate the cytotoxic effect of this compound in the acute phase (days 0 and 14) and chronic phase (days 0, 14, 28, 42, 56, 70 and 84) in female mice (Mus musculus) of the Balb/c lineage. The results show that ISO can potentiate the effect of FLZ, AFB and CASP, showing synergism with these antifungals. An evaluation of the mice via direct observation showed no behavioral changes or variations in weight during treatment; furthermore, an analysis of the cytokines IFN-γ and TNF in plasma, peritoneal cavity lavage (PCL) and bronchoalveolar lavage (BAL) indicated that there was no inflammation process. In addition, histopathological studies of the lungs, liver and kidneys showed no signs of toxicity caused by ISO. This was consistent with an analysis of oxaloacetic transaminases (GOT) and pyruvic transaminases (GPT), which remained in the standard range. These findings indicate that ISO does not have a cytotoxic effect at the doses evaluated, placing it as a monoterpene of interest in the search for compounds with pharmacological potential.

Development of ROS-sensitive capofungin hydrogel by crosslinking chitosan with four-arm polyethylene glycol derivative for treatment of vulvovaginal candidiasis

Both exogenous and endogenous reactive oxygen species (ROS) in vulvovaginal candidiasis (VVC) play pivotal roles in promoting the hyphal formation of Candida albicans (CA), which suggests that clearing ROS could inhibit CA hyphae formation. A ROS-sensitive hydrogel (CAS@4Arm-PB/CS) was formulated by using a novel four-arm polyethylene glycol (4Arm-PEG) derivative (4Arm-PB) as a crosslinking agent, chitosan (CS) as the hydrogel matrix, and caspofungin (CAS) as the antifungal drug against CA. The ROS-sensitivity, disintegration mechanism, crosslinking action, swelling degree, microstructure, modulus, and rheological properties of 4Arm-PB were characterized. According to the results, 5.0 % 4Arm-PB could quickly and efficiently cross-link 0.5 mg/mL of CS. The ROS-sensitivity of 4Arm-PB was 10–50 μM, indicating a strong ROS sensitivity. The in vitro and in vivo anti-CA results indicated that CAS@4Arm-PB/CS not only cleared endogenous and exogenous ROS and inhibited the formation of CA hyphae and biofilm but also contributed beneficially to the treatment of VVC mice caused by CA infection, implying a certain safety aspect and an in vivo applicability. This research introduces a novel functional crosslinking agent for CS hydrogel formulation, presenting a new avenue for hydrogel-based drug delivery systems and therapeutic strategies for VVC treatment.

Two outbreaks and sporadic occurrences of Candida auris from one hospital in China: an epidemiological, genomic retrospective study.

To investigate the clinical relevance, origin, transmission, and resistance of Candida auris (C. auris) isolates from two outbreaks and sporadic occurrences from one hospital in China. A total of 135 C. auris isolates were collected. Clinical characteristics were obtained and antifungal susceptibility testing (AFST) was performed using the method of broth microdilution. Phylogenetic tree, WGS analysis, and single nucleotide polymorphisms (SNPs) were used to determine the origin, transmission, and resistance mechanisms. A total of 31 patients (91.2%, 31/34) received invasive medical procedures and 13 patients (38.2%, 13/34) had antifungal agents before C. auris infection/colonization, except one patient whose clinical information was missing. Only 4 cases of C. auris candidemia were observed. 18 patients died, 13 patients recovered, and the outcomes of 3 patients were not available. A total of 35 C. auris isolates, which were successfully cultivated and the first isolated or harbored specific drug-resistant phenotype from each patient, were selected to be sequenced and further analyzed. C. auris isolates presented low genetic variability and belonged to clade I, possibly originating from BJ004-H7 in Beijing. All 35 isolates were resistant to Fluconazole (FCZ) and amphotericin B (AMB), and 3 isolates were resistant to caspofungin (CAS). Mutations in ERG11 and FKS1 were linked to reduced azole and echinocandin susceptibility, respectively. Two outbreaks of highly clonal, multidrug-resistant C. auris isolates within the medical facility were reported. The intensive performance of disinfection measures helped block in-hospital transmission. Understanding the epidemiology, drug resistance and management of C. auris will be helpful for implementing effective infection control and treatment strategies.

Verapamil enhances the activity of Caspofungin against Cryptococcus neoformans， coinciding with inhibited Ca2+/CN pathway and damage to cell wall integrity

Given the challenges posed by toxicity and drug resistance in the treatment of cryptococcal infections, we sought to explore the antifungal potential of verapamil (VER), a calcium channel blocker, against Cryptococcus neoformans (C. neoformans), and its potential synergy with antifungals, specifically caspofungin (CAS). In vitro and in vivo (Galleria mellonella) models were employed to assess VER's antifungal activity and its interaction with CAS. Mechanisms underlying the synergism were explored through analysis of cell wall integrity, membrane permeability, and gene expression related to the calcineurin pathway. Additionally, the influence of Ca2+ on chitin deacetylase activity was investigated. VER exhibited a pronounced antifungal effect on C. neoformans and synergized with CAS, enhancing antifungal efficacy in Galleria mellonella. VER reduced chitosan content and disrupted cell wall integrity, evidenced by melanin leakage and fluorescence staining. VER+CAS modified membrane permeability, triggering intracellular ROS accumulation and mitochondrial membrane potential alterations. VER mitigated CAS-induced calcium fluctuations and downregulated calcineurin pathway genes. Furthermore, it was found that the enzyme activity of chitin deacetylase of C. neoformans is significantly influenced by the presence of Ca2+, suggesting that the use of VER may affect this activity. The synergistic antifungal effect of VER and CAS represents a promising therapeutic strategy for cryptococcal infections. The multifaceted mechanisms, including disruption of cell wall integrity and modulation of membrane permeability, and regulation of intracellular calcium signaling pathways, offer new insights into antifungal drug development.

Brilacidin, a novel antifungal agent against Cryptococcus neoformans.

Invasive fungal infections have a high mortality rate causing more deaths annually than tuberculosis or malaria. Cryptococcosis, one of the most prevalent fungal diseases, is generally characterized by meningitis and is mainly caused by two closely related species of basidiomycetous yeasts, Cryptococcus neoformans and Cryptococcus gattii. There are few therapeutic options for treating cryptococcosis, and searching for new antifungal agents against this disease is very important. Here, we present brilacidin (BRI) as a potential antifungal agent against C. neoformans. BRI is a small molecule host defense peptide mimetic that has previously exhibited broad-spectrum immunomodulatory/anti-inflammatory activity against bacteria and viruses. BRI alone was shown to inhibit the growth of C. neoformans, acting as a fungicidal drug, but surprisingly also potentiated the activity of caspofungin (CAS) against this species. We investigated the mechanism of action of BRI and BRI + CAS against C. neoformans. We propose BRI as a new antifungal agent against cryptococcosis.

Investigation of antifungal susceptibility of Aspergillus species isolated from systemic clinical specimens by different methods

PurposeDue to the potential for Aspergillus species to cause lethal infections and the rising rates of antifungal resistance, the significance of antifungal susceptibility tests has increased. We aimed to assess the sensitivities of Aspergillus species to amphotericin B (AMB), voriconazole (VOR), itraconazole (ITZ), and caspofungin (CAS) using disk diffusion (DD) and gradient diffusion (GD) methods and compare them with broth microdilution (BMD) as the reference susceptibility method. MethodsThe study involved 62 Aspergillus fumigatus, 28 Aspergillus flavus, and 16 Aspergillus terreus isolates, totaling 106 Aspergillus isolates. BMD and DD methods were performed in accordance with CLSI M38-A2 and CLSI M51-A documents, respectively. The GD method utilized nonsupplemented Mueller Hinton agar (MHA) as the medium. ResultsIn the BMD method, the lowest minimal inhibitory concentration (MIC)90 or minimal effective concentration (MEC)90 values were observed for VOR and CAS (0.5 μg/mL and 0.06 μg/mL, respectively). AMB and ITZ MIC90 values were both 2 μg/mL. In our comparison of the GD method with the BMD method at ±2 dilution, we observed essential agreement rates of 91.6%, 99.1%, 100%, and 38.6% for AMB, VOR, ITZ, and CAS, respectively. When comparing DD and BMD methods, we found categorical agreement rates of 65.1%, 99.1%, 77.3%, and 100% for AMB, VOR, ITZ, and CAS, respectively. For GD and BMD methods, these rates were 79.2%, 99.1%, 87.8%, and 100%. ConclusionsGiven the high essential and categorical agreement rates, we posit that the GD method is a viable alternative to the BMD method for AMB, ITZ and VOR but not for CAS. In addition, the use of nonsupplemented MHA in the GD method proves advantageous due to its cost-effectiveness and widespread availability compared to other growth media.

Effect of Conformational Variability on the Drug Resistance of Candida auris ERG11p and FKS1.

Candida auris infection has been recognized as an urgent threat to antifungal drug resistance, and the Eagle effect of C. auris FKS1 (1,3-β-d-glucan synthase) wild-type isolates has also been noted. The Eagle effect, namely, where higher concentrations of antifungals reduce fungicidal activity relative to lower concentrations, is a confounding factor of apparent antifungal resistance, but the detailed mechanism remains unclear. Here, we present the conformational variability of mutation sites for ERG11p (lanosterol 14α-demethylase) and FKS1 from deep neural network-based prediction along with the reported X-ray crystallographic and cryo-electron microscopy (cryo-EM) structures of antifungals. The sequence variability maps provide valuable insights into the inconsistent correlation between azole resistance and the mysterious Eagle effect with the dispersion of minimal inhibitory concentration (MIC) for echinocandin resistance. The conformational variability prediction supports the hypothesis that mutations K143R of clade I, VF125AL of clade III, and Y132F of clade IV for C. auris ERG11p make the corresponding site variable and that an increased population of invisible variable conformations potentially contributes to triazole resistance. In contrast, the predicted rigid conformation by the S639F mutation of hot spot region 1 (HS1) for FKS1 suggests that caspofungin (CAS) is involved in an uncompetitive inhibition, and a decreased population of the CAS-bound state of FKS1 with Rho1 leads to drug resistance. The predicted variable HS1 region for FKS1 WT isolates and the rigid one for FKS1 S639F mutants support the in vivo drug response and the in vitro MIC dispersion. A plausible mechanism of the Eagle effect is hereby proposed, namely, that a high concentration of CAS with a high membrane affinity reduces the population of the CAS-bound state of FKS1 with Rho1, as well as accompanying events such as aggregation or association depending on the conformational variability of HS1.

SAR study of N′-(Salicylidene)heteroarenecarbohydrazides as promising antifungal agents

Clinically available antifungal drugs have therapeutic limitations due to toxicity, narrow spectrum of activity, and intrinsic or acquired drug resistance. Thus, there is an urgent need for new broad-spectrum antifungal agents with low toxicity and a novel mechanism of action. In this context, we have successfully identified several highly promising lead compounds, i.e., aromatic N′-(salicylidene)carbohydrazides, exhibiting excellent antifungal activities against Cryptococcus neoformans, Candida albicans, Aspergillus fumigatus and several other fungi both in vitro and in vivo. Building upon these highly promising results, 71 novel N′-(salicylidene)heteroarenecarbohydrazides 5 were designed, synthesized and their antifungal activities examined against fungi. Based on the SAR study, four highly promising lead compounds, i.e., 5.6a, 5.6b, 5.7b and 5.13a were identified, which exhibited excellent potency against C. neoformans, C. albicans and A. fumigatus, and displayed impressive time-kill profiles against C. neoformans with exceptionally high selectivity indices (SI ≥ 500). These four lead compounds also showed synergy with clinical antifungal drugs, fluconazole, caspofungin (CS) and amphotericin B against C. neoformans. For the SAR study, we also employed quantitative structure–activity relationship (QSAR) analysis by taking advantage of the accumulated data on a large number of aromatic and heteroaromatic N′-(salicylidene)carbohydrazides, which successfully led to rational design and selection of promising compounds for chemical synthesis and biological evaluation.

Caspofungin enhances the potency of rifampin against Gram-negative bacteria.

Developing antibiotic adjuvants is an effective strategy to combat antimicrobial resistance (AMR). The envelope of Gram-negative bacteria (GNB) is a barrier to prevent the entry of antibiotics, making it an attractive target for novel antibiotic and adjuvant development. In this study, we identified Caspofungin acetate (CAS) as an antibiotic adjuvant against GNB in the repurposing screen of 3,158 FDA-approved drugs. Checkerboard assays suggested that CAS could enhance the antimicrobial activity of rifampin or colistin against various GNB strains in vitro, Moreover, Galleria mellonella larvae infection model also indicated that CAS significantly potentiated the efficacy of rifampin against multidrug-resistant Escherichia coli 72 strain in vivo. Most importantly, resistance development assay showed that CAS was less susceptible to accelerating the resistance development of drug-sensitive strain E. coli MG1655. Functional studies and RNA-seq analysis confirmed that the mechanisms by which CAS enhanced the antimicrobial activities of antibiotics were involved in permeabilizing the bacterial cell envelope, disrupting proton motive force and inhibiting bacterial biofilm formation. Additionally, it has been found that PgaC is the CAS target and enzymatic assay has confirmed the inhibition activity. Our results illustrate the feasibility of CAS as an antibiotic adjuvant against GNB, which is an alternative strategy of anti-infection.

Method Development and Validation of Indigenously Isolated Caspofungin Acetate and its related Substances using RP-HPLC Method

Caspofungin is prepared from a precursor PneumocandinB0 derived from a fungus Glarea lozoyensis. Prepared crude caspofungin acetate has many impurities, namely, A, B, D, E and PneumocandinB0, and these impurities vary slightly in chemical or structural composition and need to be eliminated by purification. The present study involves the preparation of caspofungin acetate from PneumocandinB0, followed by purification and method development for detecting caspofungin and its related substances.”We used a high-tech liquid chromatograph system, which included components like the Agilent 1260 infinity quaternary pump module, a sophisticated MWD detector, and the Empower 3 data handling system. To separate and analyze substances, we employed an analytical column made of stainless steel. This column was 150 mm long and had an internal diameter of 3.0 mm. It was filled with octadecyl silane chemically bonded to porous silica particles, each with a tiny diameter of 3 micrometers. The test method underwent validation to ensure it could accurately identify specific elements, detect minimal quantities, establish the lowest measurable amounts, maintain linearity, demonstrate precision, achieve accuracy, and suitably handle sample solutions and the system.” and the developed test method is suitable and can be introduced into the routine use for the detection of caspofungin acetate and its related substances.

2115. Rezafungin Activity against Invasive Candidiasis Isolates Globally: Results from the 2022 Rezafungin Surveillance Program

Abstract Background Rezafungin (RZF) is a new echinocandin (ECH) approved by the US FDA to treat candidemia and invasive candidiasis (IC). Fluconazole (FLC) resistance (R) is a raising concern to treat IC, and ECHs are often used as first-line therapy. We evaluated the in vitro activity of RZF, caspofungin (CSF), micafungin (MCF), anidulafungin (ANF), and azoles against a global collection of 500 Candida isolates causing IC. Methods C. albicans (CA; 233 isolates; 46.6%), C. glabrata (CG; 102; 20.4%), C. parapsilosis (CP; 82; 16.4%), C. tropicalis (CT; 41; 8.2%), C. krusei (CK; 18; 3.6%), C. dubliniensis (CD; 15; 3.0%), and C. auris (CARS; 9; 1.8%) isolates were collected (1/patient) in 2022 from 52 medical centers located in Europe (EU; n=203; 19 centers), North America (NA; n=167; 14 centers), Asia-Pacific (AP; n=63; 8 centers), and Latin America (LA, n=67; 7 centers), identified by MALDI-TOF MS and/or sequencing and tested by CLSI broth microdilution. CLSI breakpoints (BP) were applied where available. CARS CDC tentative BPs were applied. Results RFZ exhibited activity against CA (Table) inhibiting 99.1% of the isolates overall, and 98.6%, 99.0%, 100%, and 100% of isolates from NA, EU, AP, and LA. Only 1 CA isolate (from US) displayed a FLC MIC value within the susceptible-dose dependent (SDD) category. CSP, ANF, and MCF showed S rates of 99.1% against CA. All but 1 CG isolate was susceptible (S) to RZF (99.0%S). CG S rates to CSP, ANF, MCF, and FLC were 98.0%, 98.0%, 97.1%, and 96.1% (SDD), respectively. All CP isolates were S to RZF, CSF, and MCF. However, only 84.1% of CP isolates were S to FLC. RZF and other ECHs inhibited all FLC-R isolates (10 total, 9 EU and 1 US). All CT and CK isolates were S to RZF and other ECHs. All CT were also S to azoles, and 94.4% of CK were S to VRC. Only RZF BPs are available by CLSI against CD (86.7%S). All CARS isolates (9 total; 4 EU, 3 NA, and 2 LA) were FLC-R; but 77.8% and 88.9% were S to RZF and other ECHs, respectively. Conclusion RZF demonstrated potent in vitro activity against invasive candidiasis isolates regardless of the region, and remained active against FLC-R CP and most of the FLC-R CARS isolates. Based on MIC50 and MIC90 results, RZF had similar activity to the other ECHs overall. Disclosures Cecilia G. Carvalhaes, MD, PhD, AbbVie: Grant/Research Support|bioMerieux: Grant/Research Support|Cipla: Grant/Research Support|CorMedix: Grant/Research Support|Melinta: Grant/Research Support|Pfizer: Grant/Research Support Paul Rhomberg, BS, MT(ASCP), bioMerieux: Grant/Research Support|Melinta: Grant/Research Support|Pfizer: Grant/Research Support Abby Klauer, BS, Melinta: Grant/Research Support Mariana Castanheira, PhD, AbbVie: Grant/Research Support|Basilea: Grant/Research Support|bioMerieux: Grant/Research Support|Cipla: Grant/Research Support|CorMedix: Grant/Research Support|Entasis: Grant/Research Support|Melinta: Grant/Research Support|Paratek: Grant/Research Support|Pfizer: Grant/Research Support|Shionogi: Grant/Research Support

1947. Activity of Rezafungin against Echinocandin–Non-wild type C. glabrata Clinical Isolates from the Rezafungin Surveillance Program (2014–2021)

Abstract Background Fluconazole (FLC) resistance (R) is common in C. glabrata (CGLA). Echinocandins (ECHs) are often used as first-line therapy. R to ECHs has been associated with FKS1 and FKS2 gene alterations. Rezafungin (RZF), a new ECH approved by the US FDA to treat candidemia and invasive candidiasis, was evaluated against a collection of ECH–non-wild type (NWT) CGLA isolates. Methods A total of 1045 CGLA collected (1/patient) in 2014–2021 from 58 medical centers located in North America (NA; n=459; 20 centers), Europe (EU; n=396; 23 centers), Asia-Pacific (AP; n=130; 10 centers), and Latin America (LA, n=60; 5 centers) were identified by MALDI-TOF and/or sequencing and tested by CLSI broth microdilution. CLSI breakpoints (BP) and epidemiological cut-off values were applied. ECH NWT isolates were submitted to FKS analysis by whole genome sequencing. Results RZF showed similar activity to other ECHs against CGLA (Table), inhibiting 98.5% at ≤0.5 mg/L. Anidulafungin (ANF), caspofungin (CSF), and micafungin (MCF) susceptibility (S) rates were 96.2%, 97.5%, and 97.3%, respectively. ECH-NWT CGLA were detected in 42 isolates (4.0% overall): NA showed the highest rate of ECH NWT isolates (29; 6.3%), followed by EU (10; 2.5%), AP (2; 1.5%), and LA (1; 1.7%). The RZF S rate was 61.9% of ECH-NWT CGLA, while the S rate to ANF, CSF, and MCF was 21.4%, 40.5%, and 33.3%, respectively. Alteration in FKS genes were observed in 31 CGLA (73.8% of ECH NWT; 3.0% overall). RZF was active against >50% of these isolates. RFZ was also active against >90% of ECH-NWT CGLA carrying wildtype FKS genes. Equivalent activity was noted to other ECHs against ECH-NWT isolates. However, ECHs were 4 to 8-fold more active against ECH-NWT CGLA that did not display alterations in FKS genes than those with FKS alterations. FKS2 hot spot(HS)-1 alterations were observed in 22 isolates (12 displayed S663F), while FKS1-HS1 alterations were noted in 9 isolates (7 displayed S629P). FLC-R was observed in 6.9% of CGLA overall and 16.7% of the ECH-NWT isolates. Conclusion RZF demonstrated potent in vitro activity against CGLA and remained active against most isolates of CGLA displaying an ECH-NWT phenotype with or without FKS alterations. Disclosures Cecilia G. Carvalhaes, MD, PhD, AbbVie: Grant/Research Support|bioMerieux: Grant/Research Support|Cipla: Grant/Research Support|CorMedix: Grant/Research Support|Melinta: Grant/Research Support|Pfizer: Grant/Research Support Paul Rhomberg, BS, MT(ASCP), bioMerieux: Grant/Research Support|Melinta: Grant/Research Support|Pfizer: Grant/Research Support Abby Klauer, BS, Melinta: Grant/Research Support Lalitagauri M. Deshpande, PhD, Melinta: Grant/Research Support|Paratek: Grant/Research Support Mariana Castanheira, PhD, AbbVie: Grant/Research Support|Basilea: Grant/Research Support|bioMerieux: Grant/Research Support|Cipla: Grant/Research Support|CorMedix: Grant/Research Support|Entasis: Grant/Research Support|Melinta: Grant/Research Support|Paratek: Grant/Research Support|Pfizer: Grant/Research Support|Shionogi: Grant/Research Support

Antifungal susceptibility of Aspergillusgenus determined by the Etest® method: eleven years of experience at the Instituto Médico La Floresta. Caracas, Venezuela.

Abstract. This research aimed to determine the susceptibility of Aspergil-lus spp. to four antifungal agents using the Etest® method in several clinical samples (respiratory samples, soft tissue, otic tissue, and ocular tissue, among others) from a private health center in Venezuela. Thirty-three strains were evaluated: 11 Aspergillus section Flavi, eight Aspergillus section Fumigati, six Aspergillus section Nigri, four Aspergillus section Terrei, and four Aspergillus spp. A 0.5 McFarland standard suspension of a 5-day culture of each Aspergillusstrain was prepared on Potato Dextrose agar and then inoculated on Sabouraud agar plates with 2% glucose. Voriconazole (VCZ), amphotericin B (AMB), caspo-fungin (CAS), and posaconazole (PCZ) were tested. Minimal inhibitory concen-trations (MIC) in μg/mL were determined after 24 and 48 hours of incubation at 35 °C and th range (R), geometric mean (GM), MIC50, and MIC90 were calcu-lated. The results for the 33 Aspergillus spp. tested after 24 h were the follow-ing: VCZ (R = 0.031- 16; GM = 0.145; MIC50 = 0.125 and MIC90 = 0.5), AMB (R = 0.031-16; GM = 0.644; MIC50 = 0.5 and MIC90 = 8), CAS (R = 0.031-16; GM = 0.1076; MIC50 = 0.063 and MIC90 = 1), PCZ (R =0.031 - 0.5; GM = 0.0755; MIC50 = 0.063 and MIC90 = 0.25). This investigation allowed assessing the an-tifungal susceptibility profiles of Aspergillus spp. isolated from clinical samples by the Etest® method, which is practical, reproducible and easy to perform in microbiology laboratories.

Antifungal susceptibility of clinically isolated invasive Candida tropicalis in East China from 2017 to 2021

Objective: To explore the epidemiological characteristics of sample distribution and antifungal susceptibilities of clinically invasive C. tropicalis isolates from 2017 to 2021 in East China. Methods: Using a retrospective analysis, the East China Invasive Fungal Infection Group (ECIFIG) collected C. tropicalis clinically isolated from 32 hospitals in East China between January 2017 and December 2021. The identification results of the strains were reviewed using mass spectrometry by the central laboratory of the Shanghai East Hospital. The minimum inhibitory concentrations (MICs) of the strains against fluconazole (FLU), voriconazole (VOR), itraconazole (ITR), Posaconazole (POS), isavuconazole (ISA), anidulafungin (ANI), caspofungin (CAS), micafungin (MICA) and 5-fluorocytosine (FCT) were tested by the ThermoFisher CMC1JHY colorimetric microdilution method. The MIC of amphotericin B (AMB) was tested by the broth microdilution method. The MIC results were analyzed based on the clinical breakpoints and epidemiological cutoff values (ECV) published by the Clinical and Laboratory Standards Institute (CLSI) M27 Ed3 and M57 Ed4 documents. Data analysis was conducted using the Kruskal-Wallis test and paired t-test. Results: In total, 305 C. tropicalis isolates were collected. There were 38.0% (116/305) strains isolated from blood, 11.5% (35/305) ascites, 8.9% (27/305) catheter and 8.9% (27/305) drainage fluid. The resistance rate of C. tropicalis to FLU was 32.5%, to VOR was 28.5%, and the cross-resistance rate to FLU and VOR was 28.5%. The wild-type proportions for ITR and POS were 79.3% and 29.2% respectively. There was no significant difference in resistance rates, MIC50, and MIC90 of FLU and VOR, or in the wild-type rates of ITR and POS over five years. More than 95.0% of the isolates were susceptible to echinocandins. However, one strain was identified as being multi-drug resistant. In azole antifungals, voriconazole, itraconazole, posaconazole, and isavuconazole have similar GM MIC values. The GM MIC of fluconazole is significantly higher than that of itraconazole (t=9.95, P<0.05), posaconazole (t=9.99, P<0.05), and voriconazole (t=10.01, P<0.05), Meanwhile, among echinocandins, the GM MIC of ANI was comparable to that of CAS (t=1.17, P>0.05), both of which were significantly higher than MICA (t=11.56, P<0.05; t=4.15, P<0.05). Conclusion: The clinical isolates of C. tropicalis in East China from 2017 to 2021 were relatively susceptible to echinocandins. However, there was consistently high resistance to fluconazole and voriconazole. More intensive efforts should be made on the monitoring of drug resistance in C. tropicalis.

Inter-laboratory variability of caspofungin MICs for Nakaseomyces glabrata isolates – an Irish tertiary hospital experience

Background. Nakaseomyces glabrata, formerly Candida glabrata, is an opportunistic yeast and emerging cause of human infections. The use of broth microdilution (BMD) methodologies for caspofungin (CSP) antifungal susceptibility testing (AFST) against N. glabrata is reported to be prone to high inter-laboratory variation. We aimed to compare CSP MICs of N. glabrata isolates from our institution with those obtained by the Reference Laboratory for the same isolates. Methods. All clinically significant N. glabrata isolates from 2019 to 2021 inclusive were reviewed. AFST was performed locally using the VITEK2 system with the AST-YS08 card, while E-tests were performed at the Mycology Reference Laboratory (MRL), and agreement between these two methods was evaluated – categorical and essential. Results. Forty-one isolates were reviewed during the study period – 30 from blood cultures, seven from intra-operative theatre specimens and four from sterile site drain fluids. Despite an essential agreement of 100 % within ±2 log2 dilutions, marked discrepancies were noted in interpretative breakpoints between assays with 17 Minor and 16 Major category errors. Categorical agreement was 19.5 %, with the VITEK2 over-estimating resistance. A Mann–Whitney U-test assessed the relationship of MICs across the AFST modalities, and a statistically significant difference was noted, P&lt;0.01, with a higher mean rank for VITKEK2 outputs. Conclusion. While the VITEK2 system is highly applicable, its performance for CSP AFST is unreliable and potentially results in the mis-classification of susceptible isolates as highlighted in our study. The use of VITEK2 AST-YS08 micafungin as a sentinel echinocandin should be explored and/or the evaluation of CSP-specific E-tests as utilized by the MRL. These methods appear more consistent and less prone to the variation seen with BMD for CSP.

Clinical and Microbiological Characteristics of Candidemia Cases in Saudi Arabia.

Candidemia and antifungal resistance are major healthcare challenges. The aim of this study is to describe the frequency of candidemia cases, distribution of Candida spp., and the associated risk factors for mortality in an academic institution in Saudi Arabia over an 18-month period. We also evaluated the susceptibility patterns of Candida blood isolates. Candidemia cases were collected from King Fahad Hospital of the University over the period between July 1st, 2020 through December 31st, 2021. They were prospectively reviewed for the preceding risk factors and antifungal (AF) susceptibility, testing results to fluconazole (FL), voriconazole (VO), itraconazole (IT), posaconazole (PO), caspofungin (CASP), anidulafungin (AND), micafungin (MYC), flucytosine (FLC) and amphotericin B (AMPB) using a broth microdilution kit (Sensititre™ YeastOne). A total of 48 candidemia isolates were included that were isolated from 43 patients. The median age of cases was 62 ± 23.3 years (60.4% males and 83% ICU patients). Independent risk factors for mortality at 30 days in candidemia patients were age, COVID-19 co-infection, and use of tocilizumab. The most commonly isolated species were C. glabrata and C. parapsilosis (22.9% each) followed by C. albicans (18.75%). AF resistance for ≥1 antifungal was detected in 39.3% of 33 cases tested, with no cross-resistance identified. Resistance rates for each AF were as follows: FL (18%), VO (6%), IT (6%), PO (9%) and AMPB (3%). No resistance was seen for echinocandins apart from one C. krusei strain showing an intermediate result for CASP. The study showed an overall high rate of non-albicans Candida, with the predominance of C. parapsilosis and C. glabrata, representing a therapeutic challenge. AF resistance rate was high which emphasizes the importance of continuing surveillance and providing accurate and reliable tools in the laboratories for rapid speciation and susceptibility testing.

Non disseminative nano-strategy against in vivo Staphylococcus aureus biofilms

Staphylococcus aureus is considered a high priority pathogen by the World Health Organization due to its high prevalence and the potential to form biofilms. Currently, the available treatments for S. aureus biofilm-associated infections do not target the extracellular polymeric substances (EPS) matrix. This matrix is a physical barrier to bactericidal agents, contributing to the increase of antimicrobial tolerance. The present work proposes the development of lipid nanoparticles encapsulating caspofungin (CAS) as a matrix-disruptive nanosystem. The nanoparticles were functionalized with D-amino acids to target the matrix. In a multi-target nano-strategy against S. aureus biofilms, CAS-loaded nanoparticles were combined with a moxifloxacin-loaded nanosystem, as an adjuvant to promote the EPS matrix disruption. In vitro and in vivo studies showed biofilm reduction after combining the two nanosystems. Besides, the combinatory therapy showed no signs of bacterial dissemination into vital organs of mice, while dissemination was observed for the treatment with the free compounds. Additionally, the in vivo biodistribution of the two nanosystems revealed their potential to reach and accumulate in the biofilm region, after intraperitoneal administration. Thus, this nano-strategy based on the encapsulation of matrix-disruptive and antibacterial agents is a promising approach to fight S. aureus biofilms.

P27 Rezafungin treatment of candidaemia and invasive candidiasis: outcomes stratified by baseline renal function—analysis of the Phase 2 + Phase 3 trials

Abstract Background Rezafungin (RZF) once weekly (QWk) is a next-generation echinocandin in development for treatment of candidaemia and invasive candidiasis (IC) and prevention of invasive fungal disease caused by Candida, Aspergillus, and Pneumocystis spp. in BMT. RZF QWk was compared with caspofungin (CAS) QD in two double-blind, randomized, controlled trials of treatment of candidaemia and/or IC: STRIVE (Phase 2, NCT02734862) and ReSTORE (NCT03667690). Objectives Trial data (Phase 2 + Phase 3) were analysed to evaluate outcomes stratified by renal function at baseline: CrCl ≥ 60 mL/min (normal/mild impairment [Norm/Mild]) and &amp;lt;60 mL/min (moderate/severe impairment [Mod/Sev]). Methods Outcomes were evaluated for differences between CrCl categories and between treatment groups: RZF QWk 400 mg on Wk 1 then 200 mg versus CAS QD 70 mg on Day (D)1 then 50 mg, for ≥14 days (≤4 Wks) w/optional oral fluconazole stepdown for CAS. Results D30 all-cause mortality (ACM)—Mod/Sev: RZF, 13% (7/54); CAS, 30.5% (18/59); Norm/Mild: RZF, 22.7% (17/75); CAS, 10.8% (9/83). Mycological eradication (ME) at D5—Mod/Sev: RZF, 75.9% (41/54); CAS, 61.0% (36/59); Norm/Mild: RZF, 74.7% (56/75); CAS, 66.3% (55/83). ME at D14—Mod/Sev: RZF, 75.9% (41/54); CAS, 57.6% (34/59); Norm/Mild: RZF, 69.3% (52/75); CAS, 74.7% (62/83). ≥1 treatment-emergent AE—Mod/Sev: RZF, 93.2% (55/59); CAS, 88.9% (56/63); Norm/Mild: RZF, 88.9% (72/81); CAS, 76.7% (69/90). Conclusions RZF efficacy was comparable across CrCl categories, with higher ME and lower D30 ACM in the Mod/Sev group. Further analyses are needed to evaluate the observed differences between treatment groups.

P28 Rezafungin efficacy and safety in immunocompromised patients: sub-analyses of the Phase 3 trial in treatment of candidaemia and invasive candidiasis

Abstract Background Rezafungin (RZF) once weekly (QWk) is a next-generation echinocandin in development for treatment of candidaemia and invasive candidiasis (IC) and for prevention of invasive fungal disease caused by Candida, Aspergillus, and Pneumocystis spp. in BMT. Objectives The Phase 3 ReSTORE treatment trial (NCT03667690) demonstrated RZF QWk noninferiority to caspofungin (CAS) QD. This sub-analysis evaluated efficacy and safety outcomes in patients identified as immunocompromised (immuno-c) during the trial. Methods As previously described, adults with confirmed candidaemia and/or IC were randomized to RZF QWk (400 mg Wk1 then 200 mg QWk) or CAS QD for ≥14 days (≤4 weeks) with optional oral fluconazole stepdown for CAS. In this sub-analysis, immuno-c patients were those with prior and/or concomitant use of immunosuppressants (e.g. calcineurin inhibitors and corticosteroids) and/or medical history ongoing at screening of neutropenia, BMT, SOT, lymphoma or leukaemia. Results Of 187 patients (mITT population), 90 were immuno-c as defined. Results are for those with data available for analyses: Day 14 global cure—Immuno-c: RZF, 51.1% (23/45); CAS, 55.6% (25/45); –No immuno-c: RZF, 66.7% (32/48); CAS 65.3% (32/49). Day 5 mycological eradication—Immuno-c: RZF, 64.4% (29/45); CAS, 51.1% (23/45); no immuno-c: RZF, 72.9% (35/48); CAS, 71.4% (35/49). Among immuno-c, more had ≥1 AE (96.9%) and ≥1 SAE (64.6%) versus no immuno-c (79.0% and 45.0% respectively). Conclusions In this sub-analysis of ReSTORE, immune-c status reduced efficacy rates overall but did not change efficacy rate differences between RZF and CAS.