scholarly journals EUCAST Susceptibility Testing of Isavuconazole: MIC Data for Contemporary Clinical Mold and Yeast Isolates

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
Karin Meinike Jørgensen ◽  
Karen Marie Thyssen Astvad ◽  
Rasmus Krøger Hare ◽  
Maiken Cavling Arendrup
Keyword(s):  
Susceptibility Testing ◽  
Pearson Correlation ◽  
Acquired Resistance ◽  
Cross Resistance ◽  
Wild Type ◽  
Content Type ◽  
Wild Type Phenotype ◽  
Mold And Yeast ◽  
Gene Alterations

ABSTRACTIsavuconazole is the newest medical azole. We investigated EUCAST MICs for isavuconazole and seven comparators against 1,498 contemporary isolates (2016 to 2017). EUCAST susceptibility testing was performed. Isavuconazole MICs >2 dilution steps above the modal MIC were regarded as non-wild type for species without EUCAST epidemiological cutoff values (ECOFFs).CYP51Asequencing was performed when relevant. Pearson correlation analysis was adopted for comparing activity.Aspergillusaccounted for 90% of mold andCandidaaccounted for 97% of yeast isolates. Thirty (9.3%)Aspergillusfumigatusisolates were classified as resistant, and 10 (3.1%) were classified as non-wild type. Thirteen (4%) were cross-resistant to other mold-active azoles. Target gene alterations were found in 10 (76.9%) isolates, including 4 (30.8%) of environmental origin (TR34/L98H [n = 3] and Trip343/L98H [n = 1]). SixAspergillusterreusisolates were resistant, including two (17%) with MICs of >2 mg/liter and M217I alterations. Modal MICs/MIC50s (milligrams per liter) againstCandidaspp. were ≤0.004/≤0.004 forC. albicansandC. dubliniensis, 0.008/0.008 forC. tropicalis, 0.016/0.016 forC. parapsilosis, 0.06/0.06 forC. glabrata, and 0.125/0.125 forC. krusei. A non-wild-type phenotype was observed for 6.6% of isolates (C. glabrata[11.8%] andC. tropicalis[12.3%], specifically). All of these isolates were nonsusceptible/non-wild type to fluconazole (96.1%) or voriconazole (86.2%). Low MICs were found for several other species, exceptScedosporium apiospermumandFusarium. The best correlation was found between isavuconazole and voriconazole overall but forA. terreusand Mucorales to itraconazole and posaconazole, respectively. Isavuconazole displayed broadin vitroactivity. Acquired resistance was infrequent except inA. terreus,C. glabrata, andC. tropicalisand, when present, was associated with cross-resistance to other azoles. Revising the EUCAST breakpoints forA. fumigatus(defining an MIC of 2 mg/liter as intermediate [“I”]) would minimize major errors.

scholarly journals EUCAST Determination of Olorofim (F901318) Susceptibility of Mold Species, Method Validation, and MICs

2018 ◽  
Vol 62 (8) ◽  
Author(s):  
Karin Meinike Jørgensen ◽  
Karen M. T. Astvad ◽  
Rasmus Krøger Hare ◽  
Maiken Cavling Arendrup
Keyword(s):  
Fusarium Solani ◽  
Susceptibility Testing ◽  
Wild Type ◽  
Preparation Methods ◽  
Content Type ◽  
Microtiter Plates ◽  
Upper Limits ◽  
Limited Variation

ABSTRACT Olorofim is a novel antifungal agent with in vitro activity against Aspergillus and some other molds. Here, we addressed technical aspects for EUCAST olorofim testing and generated contemporary MIC data. EUCAST E.Def 9.3.1 testing was performed comparing two plate preparation methods (serial dilution in medium [serial plates] versus predilution in DMSO [ISO plates]), two lots of olorofim, visual (visual-MIC) versus spectrophotometer (spec-MIC) reading, and four polystyrene plates using 34 to 53 Aspergillus isolates from five genera. Subsequently, olorofim MICs were compared to itraconazole, voriconazole, posaconazole, and amphotericin B MICs for 298 clinical mold isolates (2016 to 2017). Wild-type upper limits (WT-UL) were determined following EUCAST principles for epidemiologic cutoff value (ECOFF) setting. Olorofim median MICs comparing serial plates and ISO plates were identical (25/36 [69%]) or one dilution apart (11/36 [31%]). Interperson agreement for visual-MICs was 92% to 94%/100% for ≤1/≤2 dilutions, respectively. The visual-MIC values across tested microtiter plates and olorofim lots revealed only discrete differences (≤1 dilution lower for treated plates). No single spec-MIC criterion was applicable to all species. Olorofim MICs were low against 275 Aspergillus species isolates (modal MIC, 0.06 mg/liter; MIC range, < 0.004 to 0.25 mg/liter) and three dermatophytes (MICs 0.03 to 0.06 mg/liter). MICs against Fusarium were diverse, with full inhibition of F. proliferatum (MIC, 0.016), 50% growth inhibition of Fusarium solani at 1 to 2 mg/liter, and no inhibition of F. dimerum. Olorofim displayed potent in vitro activity against most mold isolates and was associated with limited variation in EUCAST susceptibility testing.

scholarly journals Implications of the EUCAST Trailing Phenomenon inCandida tropicalisfor theIn VivoSusceptibility in Invertebrate and Murine Models

2018 ◽  
Vol 62 (12) ◽  
Author(s):  
K. M. T. Astvad ◽  
D. Sanglard ◽  
E. Delarze ◽  
R. K. Hare ◽  
M. C. Arendrup
Keyword(s):  
Susceptibility Testing ◽  
Galleria Mellonella ◽  
Growth Control ◽  
Resistant Isolate ◽  
Murine Models ◽  
Wild Type ◽  
Content Type ◽  
Gradual Loss

ABSTRACTCandida tropicalisisolates often display reduced but persistent growth (trailing) over a broad fluconazole concentration range during EUCAST susceptibility testing. Whereas weak trailing (<25% of the positive growth control) is common and found not to impair fluconazole efficacy, we investigated if more pronounced trailing impacted treatment efficacy. Fluconazole efficacy against two weakly (≤25% growth), two moderately (26% to 50% growth), and one heavily (>70% growth) trailing resistant isolate and one resistant (100% growth) isolate were investigatedin vitroandin vivo(in aGalleria mellonellasurvival model and two nonlethal murine models).CDR1expression levels andERG11sequences were characterized. The survival in fluconazole-treatedG. mellonellawas inversely correlated with the degree of trailing (71% to 9% survival in treatment groups). In mice, resistant and heavily trailing isolates responded poorly to fluconazole treatment.CDR1expression was significantly higher in trailing and resistant isolates than in wild-type isolates (1.4-fold to 10-fold higher). All isolates exhibitedERG11wild-type alleles. Heavily trailing isolates were less responsive to fluconazole in allin vivomodels, indicating an impact on fluconazole efficacy.CDR1upregulation may have contributed to the observed differences. Moderately trailing isolates responded less well to fluconazole in larvae only. This confirms clinical data suggesting fluconazole is effective against infections with such isolates in less severely ill patients and supports the current 50% growth endpoint for susceptibility testing. However, it is still unclear if the gradual loss of efficacy observed for moderately trailing isolates in the larva model may be a reason for concern in selected vulnerable patient populations.

scholarly journals APX001A In Vitro Activity against Contemporary Blood Isolates and Candida auris Determined by the EUCAST Reference Method

2018 ◽  
Vol 62 (10) ◽  
Author(s):  
Maiken Cavling Arendrup ◽  
Anuradha Chowdhary ◽  
Karen M. T. Astvad ◽  
Karin Meinike Jørgensen
Keyword(s):  
Susceptibility Testing ◽  
Yeast Species ◽  
Reference Method ◽  
Common Species ◽  
Drug Candidate ◽  
Its Sequencing ◽  
Wild Type ◽  
Candida Auris ◽  
Content Type

ABSTRACT APX001A is the active moiety of the first-in-class drug candidate APX001. So far, most susceptibility testing studies have examined ≤30 isolates/species, and only one used the EUCAST method. Here, we investigated the in vitro activity of APX001A and five comparators against 540 candidemia and 122 C. auris isolates. Isolates (17 Candida and 3 yeast species) were identified using CHROMagar, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF) and, when needed, internal transcribed space (ITS) sequencing. EUCAST E.Def 7.3.1 susceptibility testing included APX001A, amphotericin B, anidulafungin, micafungin, fluconazole, and voriconazole. Wild-type upper limits (WT-UL) were established following the EUCAST principles for epidemiological cutoff value setting for APX001A, allowing classification as wild type (WT) or non-WT. APX001A MIC50 values (mg/liter) were as follows: Candida albicans, Candida dubliniensis, and Candida tropicalis, 0.004 to 0.008; Candida parapsilosis and Candida auris, 0.016; Candida glabrata, 0.06; and Candida krusei, >0.5. APX001A MICs against the rare species varied from ≤0.0005 (C. pelliculosa) to >0.5 (Candida norvegensis). APX001A was equally or more active in vitro than the comparators against all species except C. krusei and C. norvegensis. Four isolates were APX001A non-WT; all were fluconazole resistant. A correlation was observed between APX001A and fluconazole MICs across all species except Candida guilliermondii and C. auris, and when comparing high and low fluconazole MIC isolates of C. albicans, C. dubliniensis, C. glabrata, C. tropicalis, and C. auris. APX001A showed promising in vitro activity against most Candida and other yeast species, including C. auris, compared to five comparators. WT-UL were suggested for the common species, and a new and unexplained correlation to fluconazole susceptibility was observed.

scholarly journals Effects of KPC Variant and Porin Genotype on the In Vitro Activity of Meropenem-Vaborbactam against Carbapenem-Resistant Enterobacteriaceae

2019 ◽  
Vol 63 (3) ◽  
Author(s):  
William R. Wilson ◽  
Ellen G. Kline ◽  
Chelsea E. Jones ◽  
Kristin T. Morder ◽  
Roberta T. Mettus ◽  
...  
Keyword(s):  
Susceptibility Testing ◽  
Disk Diffusion ◽  
Broth Microdilution ◽  
Wild Type ◽  
Content Type ◽  
E Coli ◽  
Carbapenem Resistant ◽  
Strip Testing ◽  
Carbapenem Resistant Enterobacteriaceae

ABSTRACT Meropenem-vaborbactam is a new agent with the potential to treat carbapenem-resistant Enterobacteriaceae (CRE) infections. We describe the in vitro activity of meropenem-vaborbactam against representative CRE genotypes and laboratory-engineered Escherichia coli isolates harboring mutant blaKPC genes associated with ceftazidime-avibactam resistance. We also compared disk diffusion and gradient strip testing methods to standard broth microdilution methods. Against 120 CRE isolates, median ceftazidime-avibactam and meropenem-vaborbactam MICs were 1 and 0.03 µg/ml, respectively. Ninety-eight percent (117/120) of isolates were susceptible to meropenem-vaborbactam (MICs ≤ 4 µg/ml). Against Klebsiella pneumoniae isolates harboring mutant blaKPC, the addition of vaborbactam lowered the meropenem MICs in 78% of isolates (14/18); 100% were susceptible to meropenem-vaborbactam. Median meropenem-vaborbactam MICs were higher against K. pneumoniae carbapenemase (KPC)-producing K. pneumoniae isolates with mutant ompK36 porin genes (n = 26) than against those with wild-type ompK36 porin genes (n = 54) (0.25 versus 0.03 µg/ml; P < 0.0001). Against E. coli TOP10 isolates with plasmid constructs containing wild-type blaKPC or mutant blaKPC, the addition of vaborbactam at 8 µg/ml lowered the meropenem MICs 2- to 512-fold, resulting in meropenem-vaborbactam MICs of 0.03 µg/ml. The rates of categorical agreement with broth microdilution for disk diffusion or gradient strips ranged from 90 to 95%. Essential agreement rates were higher for research-use-only (RUO) gradient strips manufactured by bioMérieux (82%) than for those manufactured by Liofilchem (48%) (P < 0.0001). Taken together, our data highlight the potent in vitro activity of meropenem-vaborbactam against CRE, including isolates resistant to ceftazidime-avibactam. Vaborbactam inhibited both wild-type and variant KPC enzymes. On the other hand, KPC-producing K. pneumoniae isolates with ompK36 mutations displayed higher meropenem-vaborbactam MICs than isolates with wild-type ompK36. The results of susceptibility testing with RUO bioMérieux gradient strips most closely aligned with those of broth microdilution methods.

scholarly journals Echinocandin-Induced Microevolution of Candida parapsilosis Influences Virulence and Abiotic Stress Tolerance

mSphere ◽  
2018 ◽  
Vol 3 (6) ◽  
Author(s):  
Csaba Papp ◽  
Katica Kocsis ◽  
Renáta Tóth ◽  
László Bodai ◽  
Jesse R. Willis ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Amino Acid ◽  
Candida Parapsilosis ◽  
Hot Spot ◽  
Abiotic Stress Tolerance ◽  
Acquired Resistance ◽  
Cross Resistance ◽  
First Line ◽  
Content Type

ABSTRACT Candida species are a major cause of life-threatening bloodstream infections worldwide. Although Candida albicans is responsible for the vast majority of infections, the clinical relevance of other Candida species has also emerged over the last twenty years. This shift might be due in part to changes in clinical guidelines, as echinocandins became the first line of therapeutics for the treatment. Candida parapsilosis is an emerging non-albicans Candida species that exhibits lower susceptibility levels to these drugs. Candida species frequently display resistance to echinocandins, and the mechanism for this is well-known in C. albicans and Candida glabrata, where it is mediated by amino acid substitutions at defined locations of the β-1,3-glucan synthase, Fks1p. In C. parapsilosis isolates, Fks1p harbors an intrinsic amino acid change at position 660 of the hot spot 1 (HS1) region, which is thought to be responsible for the high MIC values. Less is known about acquired substitutions in this species. In this study, we used directed evolution experiments to generate C. parapsilosis strains with acquired resistance to caspofungin, anidulafungin, and micafungin. We showed that cross-resistance was dependent on the type of echinocandin used to generate the evolved strains. During their characterization, all mutant strains showed attenuated virulence in vivo and also displayed alterations in the exposure of inner cell wall components. The evolved strains harbored 251 amino acid changes, including three in the HS1, HS2, and HS3 regions of Fks1p. Altogether, our results demonstrate a direct connection between acquired antifungal resistance and virulence of C. parapsilosis. IMPORTANCE Candida parapsilosis is an opportunistic fungal pathogen with the ability to cause infections in immunocompromised patients. Echinocandins are the currently recommended first line of treatment for all Candida species. Resistance of Candida albicans to this drug type is well characterized. C. parapsilosis strains have the lowest in vitro susceptibility to echinocandins; however, patients with such infections typically respond well to echinocandin therapy. There is little knowledge of acquired resistance in C. parapsilosis and its consequences on other characteristics such as virulence properties. In this study, we aimed to dissect how acquired echinocandin resistance influences the pathogenicity of C. parapsilosis and to develop explanations for why echinocandins are clinically effective in the setting of acquired resistance.

scholarly journals In VitroActivity of ASP2397 against Aspergillus Isolates with or without Acquired Azole Resistance Mechanisms

2015 ◽  
Vol 60 (1) ◽  
pp. 532-536 ◽  
Author(s):  
Maiken Cavling Arendrup ◽  
Rasmus Hare Jensen ◽  
Manuel Cuenca-Estrella
Keyword(s):  
Amphotericin B ◽  
Target Gene ◽  
Antifungal Agents ◽  
Susceptibility Testing ◽  
Resistance Mechanisms ◽  
Azole Resistance ◽  
Wild Type ◽  
Resistance Mutations ◽  
Content Type

ABSTRACTASP2397 is a new compound with a novel and as-yet-unknown target different from that of licensed antifungal agents. It has activity againstAspergillusandCandida glabrata. We compared itsin vitroactivity against wild-type and azole-resistantA. fumigatusandA. terreusisolates with that of amphotericin B, itraconazole, posaconazole, and voriconazole. Thirty-four isolates, including 4 wild-typeA. fumigatusisolates, 24A. fumigatusisolates with alterations in CYP51A TR/L98H (5 isolates), M220 (9 isolates), G54 (9 isolates), and HapE (1 isolate), andA. terreusisolates (2 wild-type isolates and 1 isolate with an M217I CYP51A alteration), were analyzed. EUCAST E.Def 9.2 and CLSI M38-A2 MIC susceptibility testing was performed. ASP2397 MIC50values (in milligrams per liter, with MIC ranges in parentheses) determined by EUCAST and CLSI were 0.5 (0.25 to 1) and 0.25 (0.06 to 0.25) againstA. fumigatusCYP51A wild-type isolates and were similarly 0.5 (0.125 to >4) and 0.125 (0.06 to >4) against azole-resistantA. fumigatusisolates, respectively. These values were comparable to those for amphotericin B, which were 0.25 (0.125 to 0.5) and 0.25 (0.125 to 0.25) against wild-type isolates and 0.25 (0.125 to 1) and 0.25 (0.125 to 1) against isolates with azole resistance mechanisms, respectively. In contrast, MICs for the azole compounds were elevated and highest for itraconazole: >4 (1 to >4) and 4 (0.5 to >4) against isolates with azole resistance mechanisms compared to 0.125 (0.125 to 0.25) and 0.125 (0.06 to 0.25) against wild-type isolates, respectively. ASP2397 was active againstA. terreusCYP51A wild-type isolates (MIC 0.5 to 1), whereas MICs of both azole and ASP2397 were elevated for the mutant isolate. ASP2397 displayedin vitroactivity againstA. fumigatusandA. terreusisolates which was independent of the presence or absence of azole target gene resistance mutations inA. fumigatus. The findings are promising at a time when azole-resistantA. fumigatusis emerging globally.

scholarly journals In Vitro Antifungal Susceptibility Testing of Candida Isolates with the EUCAST Methodology, a New Method for ECOFF Determination

2017 ◽  
Vol 61 (4) ◽  
Author(s):  
J. Meletiadis ◽  
I. Curfs-Breuker ◽  
J. F. Meis ◽  
J. W. Mouton
Keyword(s):  
Susceptibility Testing ◽  
Antifungal Susceptibility ◽  
Antifungal Drugs ◽  
Second Derivative ◽  
Wild Type ◽  
Content Type ◽  
Microdilution Method ◽  
In Vitro Antifungal Susceptibility ◽  
Resistance Rates

ABSTRACT The in vitro susceptibilities of 1,099 molecularly identified clinical Candida isolates against 8 antifungal drugs were determined using the EUCAST microdilution method. A new simple, objective, and mathematically solid method for determining epidemiological cutoff values (ECOFFs) was developed by derivatizing the MIC distribution and determining the derivatized ECOFF (dECOFF) as the highest MIC with the maximum second derivative. The dECOFFs were similar (95% agreement within 1 dilution) to the EUCAST ECOFFs. Overall, low non-wild-type/resistance rates were found. The highest rates were found for azoles with C. parapsilosis (2.7 to 9.8%), C. albicans (7%), and C. glabrata (1.7 to 2.3%) and for echinocandins with C. krusei (3.3%), C. albicans (1%), and C. tropicalis (1.7%).

scholarly journals In VitroActivity of a Novel Broad-Spectrum Antifungal, E1210, Tested against Aspergillus spp. Determined by CLSI and EUCAST Broth Microdilution Methods

2011 ◽  
Vol 55 (11) ◽  
pp. 5155-5158 ◽  
Author(s):  
Michael A. Pfaller ◽  
Frederick Duncanson ◽  
Shawn A. Messer ◽  
Gary J. Moet ◽  
Ronald N. Jones ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Broad Spectrum ◽  
Antifungal Agents ◽  
Susceptibility Testing ◽  
Hyphal Growth ◽  
Wild Type ◽  
Minimum Effective Concentration ◽  
Content Type ◽  
Excellent Activity

ABSTRACTE1210 is a first-in-class broad-spectrum antifungal that suppresses hyphal growth by inhibiting fungal glycophosphatidylinositol (GPI) biosynthesis. In the present study, we extend these findings by examining the activity of E1210 and comparator antifungal agents againstAspergillusspp. by using the methods of the Clinical and Laboratory Standards Institute (CLSI) and the European Committee for Antimicrobial Susceptibility Testing (EUCAST) to test wild-type (WT) as well as amphotericin B (AMB)-resistant (-R) and azole-R strains (as determined by CLSI methods). Seventy-eight clinical isolates ofAspergilluswere tested including 20 isolates ofAspergillus flavusspecies complex (SC), 22 ofA. fumigatusSC, 13 ofA. nigerSC, and 23 ofA. terreusSC. The collection included 15 AMB-R (MIC, ≥2 μg/ml) isolates ofA. terreusSC and 10 itraconazole-R (MIC, ≥4 μg/ml) isolates ofA. fumigatusSC (7 isolates),A. nigerSC (2 isolates), andA. terreusSC (1 isolate). Comparator antifungal agents included anidulafungin, caspofungin, amphotericin B, itraconazole, posaconzole, and voriconazole. Both CLSI and EUCAST methods were highly concordant for E1210 and all comparators. The essential agreement (EA; ±2 log2dilution steps) was 100% for all comparisons with the exception of posaconazole versusA. terreusSC (EA = 91.3%). The minimum effective concentration (MEC)/MIC90values (μg/ml) for E1210, anidulafungin, caspofungin, itraconazole, posaconazole, and voriconazole, respectively, were as follows for each species: forA. flavusSC, 0.03, ≤0.008, 0.12, 1, 1, and 1; forA. fumigatusSC, 0.06, 0.015, 0.12, >8, 1, and 4; forA. nigerSC, 0.015, 0.03, 0.12, 4, 1, and 2; and forA. terreusSC, 0.06, 0.015, 0.12, 1, 0.5, and 1. E1210 was very active against AMB-R strains ofA. terreusSC (MEC range, 0.015 to 0.06 μg/ml) and itraconazole-R strains ofA. fumigatusSC (MEC range, 0.03 to 0.12 μg/ml),A. nigerSC (MEC, 0.008 μg/ml), andA. terreusSC (MEC, 0.015 μg/ml). In conclusion, E1210 was a very potent and broad-spectrum antifungal agent regardless ofin vitromethod applied, with excellent activity against AMB-R and itraconazole-R strains ofAspergillusspp.

scholarly journals Rezafungin In Vitro Activity against Contemporary Nordic Clinical Candida Isolates and Candida auris Determined by the EUCAST Reference Method

2020 ◽  
Vol 64 (4) ◽  
Author(s):  
Marie Helleberg ◽  
Karin Meinike Jørgensen ◽  
Rasmus Krøger Hare ◽  
Raluca Datcu ◽  
Anuradha Chowdhary ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Target Genes ◽  
Specific Activity ◽  
Significant Proportion ◽  
Reference Method ◽  
Cross Resistance ◽  
Wild Type ◽  
Candida Auris ◽  
Content Type

ABSTRACT Rezafungin (formerly CD101) is a novel echinocandin in clinical development. EUCAST epidemiological cutoff values (ECOFFs) have not yet been established. We determined the in vitro activity of rezafungin and comparators against 1,293 Nordic yeast isolates and 122 Indian Candida auris isolates and established single-center wild-type upper limits (WT-UL). The isolates (19 Candida spp. and 13 other yeast species) were identified using Chromagar; matrix-assisted laser desorption ionization–time of flight (MALDI-TOF); and, when needed, internal transcribed spacer sequencing. EUCAST E.Def 7.3.1 susceptibility testing included rezafungin, anidulafungin, micafungin, amphotericin B, and fluconazole. WT-UL were established following EUCAST principles for visual and statistical ECOFF setting. fks target genes were sequenced for rezafungin non-wild-type isolates. EUCAST clinical breakpoints for fungi version 9.0 were adopted for susceptibility classification. Rezafungin had species-specific activity similar to that of anidulafungin and micafungin. On a milligram-per-liter basis, rezafungin was overall less active than anidulafungin and micafungin but equally or more active than fluconazole and amphotericin B against the most common Candida species, except C. parapsilosis. We identified 37 (3.1%) rezafungin non-wild-type isolates of C. albicans (1.9%), C. glabrata (3.0%), C. tropicalis (2.7%), C. dubliniensis (2.9%), C. krusei (1.2%), and C. auris (14.8%). Alterations in Fks hot spots were found in 26/26 Nordic and 8/18 non-wild-type C. auris isolates. Rezafungin displayed broad in vitro activity against Candida spp., including C. auris. Adopting WT-UL established here, few Nordic strains, but a significant proportion of C. auris isolates, had elevated MICs with mutations in fks target genes that conferred echinocandin cross-resistance. fks1 mutations raised rezafungin MICs notably less than anidulafungin and micafungin MICs in C. auris.

scholarly journals Mutations in fbiD (Rv2983) as a Novel Determinant of Resistance to Pretomanid and Delamanid in Mycobacterium tuberculosis

2020 ◽  
Vol 65 (1) ◽  
pp. e01948-20
Author(s):  
Dalin Rifat ◽  
Si-Yang Li ◽  
Thomas Ioerger ◽  
Keshav Shah ◽  
Jean-Philippe Lanoix ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Clinical Evidence ◽  
Clinical Samples ◽  
Loss Of Function ◽  
Wild Type ◽  
Content Type ◽  
Solid Media ◽  
High Level ◽  
Level Resistance

ABSTRACTThe nitroimidazole prodrugs delamanid and pretomanid comprise one of only two new antimicrobial classes approved to treat tuberculosis (TB) in 50 years. Prior in vitro studies suggest a relatively low barrier to nitroimidazole resistance in Mycobacterium tuberculosis, but clinical evidence is limited to date. We selected pretomanid-resistant M. tuberculosis mutants in two mouse models of TB using a range of pretomanid doses. The frequency of spontaneous resistance was approximately 10−5 CFU. Whole-genome sequencing of 161 resistant isolates from 47 mice revealed 99 unique mutations, of which 91% occurred in 1 of 5 genes previously associated with nitroimidazole activation and resistance, namely, fbiC (56%), fbiA (15%), ddn (12%), fgd (4%), and fbiB (4%). Nearly all mutations were unique to a single mouse and not previously identified. The remaining 9% of resistant mutants harbored mutations in Rv2983 (fbiD), a gene not previously associated with nitroimidazole resistance but recently shown to be a guanylyltransferase necessary for cofactor F420 synthesis. Most mutants exhibited high-level resistance to pretomanid and delamanid, although Rv2983 and fbiB mutants exhibited high-level pretomanid resistance but relatively small changes in delamanid susceptibility. Complementing an Rv2983 mutant with wild-type Rv2983 restored susceptibility to pretomanid and delamanid. By quantifying intracellular F420 and its precursor Fo in overexpressing and loss-of-function mutants, we provide further evidence that Rv2983 is necessary for F420 biosynthesis. Finally, Rv2983 mutants and other F420H2-deficient mutants displayed hypersusceptibility to some antibiotics and to concentrations of malachite green found in solid media used to isolate and propagate mycobacteria from clinical samples.

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