scholarly journals Identification of Azole Resistance Markers in Clinical Isolates ofCandida tropicalisUsing cDNA-AFLP Method

2015 ◽  
Vol 30 (3) ◽  
pp. 266-272 ◽  
Author(s):  
Ali Kanani ◽  
Farideh Zaini ◽  
Parivash Kordbacheh ◽  
Mehraban Falahati ◽  
Sassan Rezaie ◽  
...  
Keyword(s):  
Clinical Isolates ◽  
Azole Resistance ◽  
Resistance Markers

Azole resistance among clinical isolates of Aspergillus fumigatus in Lima-Peru

2019 ◽  
Vol 58 (1) ◽  
pp. 54-60 ◽  
Author(s):  
Beatriz Bustamante ◽  
Luis Ricardo Illescas ◽  
Andrés Posadas ◽  
Pablo E Campos
Keyword(s):  
Aspergillus Fumigatus ◽  
Latin American ◽  
Pcr Amplification ◽  
Sensu Stricto ◽  
Clinical Isolates ◽  
Azole Resistance ◽  
Molecular Testing ◽  
Naive Patient ◽  
In Vitro Susceptibility

Abstract Azole resistance among Aspergillus fumigatus isolates, which is mainly related to mutations in the cyp51A gene, is a concern because it is rising, worldwide disseminated, and associated with treatment failure and death. Data on azole resistance of aspergillus from Latin American countries is very scarce and do not exist for Peru. Two hundred and seven Aspergillus clinical isolates collected prospectively underwent mycology and molecular testing for specie identification, and 143 isolates were confirmed as A. fumigatus sensu stricto (AFSS). All AFSS were tested for in vitro azole susceptibility, and resistant isolates underwent PCR amplification and sequencing of the whole cyp51A gene and its promoter. The in vitro susceptibility showed a minimal inhibitory concentration (MIC) range, MIC50 and MIC90 of 0.125 to >16, 0.25, and 0.5 μg/ml for itraconazole; 0.25 to 2, 0.5, and 0.5 μg/ml for voriconazole; and 0.003 to 1, 0.06, and 0.125 μg/ml for posaconazole. Three isolates (2%) showed resistance to itraconazole and exhibited different mutations of the cyp51A gene. One isolate harbored the mutation M220K, while a second one exhibited the G54 mutation plus a modification in the cyp51A gene promoter. The third isolate, from an azole naive patient, presented an integration of a 34-bp tandem repeat (TR34) in the promoter region of the gene and a substitution of leucine 98 by histidine (L98H). The three source patients had a diagnosis or suspicion of chronic pulmonary aspergillosis.

scholarly journals Role of ATP-Binding-Cassette Transporter Genes in High-Frequency Acquisition of Resistance to Azole Antifungals in Candida glabrata

2001 ◽  
Vol 45 (4) ◽  
pp. 1174-1183 ◽  
Author(s):  
Dominique Sanglard ◽  
Francoise Ischer ◽  
Jacques Bille
Keyword(s):  
Abc Transporter ◽  
High Frequency ◽  
Candida Glabrata ◽  
Antifungal Agents ◽  
Clinical Isolates ◽  
Azole Resistance ◽  
Atp Binding ◽  
Transporter Gene ◽  
Atp Binding Cassette

ABSTRACT Candida glabrata has been often isolated from AIDS patients with oropharyngeal candidiasis treated with azole antifungal agents, especially fluconazole. We recently showed that the ATP-binding-cassette (ABC) transporter gene CgCDR1 was upregulated in C. glabrata clinical isolates resistant to azole antifungal agents (D. Sanglard, F. Ischer, D. Calabrese, P. A. Majcherczyk, and J. Bille, Antimicrob. Agents Chemother. 43:2753–2765, 1999). Deletion of CgCDR1 in C. glabrata rendered the null mutant hypersusceptible to azole derivatives and showed the importance of this gene in mediating azole resistance. We observed that wild-type C. glabrata exposed to fluconazole in a medium containing the drug at 50 μg/ml developed resistance to this agent and other azoles at a surprisingly high frequency (2 × 10−4 to 4 × 10−4). We show here that this high-frequency azole resistance (HFAR) acquired in vitro was due, at least in part, to the upregulation ofCgCDR1. The CgCDR1 deletion mutant DSY1041 could still develop HFAR but in a medium containing fluconazole at 5 μg/ml. In the HFAR strain derived from DSY1041, a distinct ABC transporter gene similar to CgCDR1, calledCgCDR2, was upregulated. This gene was slightly expressed in clinical isolates but was upregulated in strains with the HFAR phenotype. Deletion of both CgCDR1 and CgCDR2suppressed the development of HFAR in a medium containing fluconazole at 5 μg/ml, showing that both genes are important mediators of resistance to azole derivatives in C. glabrata. We also show here that the HFAR phenomenon was linked to the loss of mitochondria in C. glabrata. Mitochondrial loss could be obtained by treatment with ethidium bromide and resulted in acquisition of resistance to azole derivatives without previous exposure to these agents. Azole resistance obtained in vitro by HFAR or by agents stimulating mitochondrial loss was at least linked to the upregulation of both CgCDR1 and CgCDR2.

Effect of antifungals on itraconazole resistant Candida glabrata

2010 ◽  
Vol 5 (3) ◽  
pp. 318-323 ◽  
Author(s):  
Soňa Kucharíková ◽  
Patrick Dijck ◽  
Magdaléna Lisalová ◽  
Helena Bujdáková
Keyword(s):  
Amphotericin B ◽  
Biofilm Formation ◽  
Candida Glabrata ◽  
Antifungal Agents ◽  
Inhibitory Concentration ◽  
Clinical Isolates ◽  
Azole Resistance ◽  
Low Concentrations ◽  
Reduction Assay ◽  
Very High

AbstractIn the last decade, infections caused by Candida glabrata have become more serious, particularly due to its decreased susceptibility to azole derivatives and its ability to form biofilm. Here we studied the resistance profile of 42 C. glabrata clinical isolates to different azoles, amphotericin B and echinocandins. This work was also focused on the ability to form biofilm which plays a role in the development of antifungal resistance. The minimal inhibitory concentration testing to antifungal agents was performed according to the CLSI (Clinical and Laboratory Standards Institute) M27-A3 protocol. Quantification of biofilm was done by XTT reduction assay. All C. glabrata clinical isolates were resistant to itraconazole and sixteen also showed resistance to fluconazole. All isolates remained susceptible to voriconazole. Amphotericin B was efficient in a concentration range of 0.125–1 mg/L. The most effective antifungal agents were micafungin and caspofungin with the MIC100 values of ≤0.0313–0.125 mg/L. Low concentrations of these agents reduced biofilm formation as well. Our results show that resistance of different C. glabrata strains is azole specific and therefore a single azole resistance cannot be assumed to indicate general azole resistance. Echinocandins proved to have very high efficacy against clinical C. glabrata strains including those with ability to form biofilm.

scholarly journals Multiplecyp51A-Based Mechanisms Identified in Azole-Resistant Isolates of Aspergillus fumigatus from China

2015 ◽  
Vol 59 (7) ◽  
pp. 4321-4325 ◽  
Author(s):  
Musang Liu ◽  
Rong Zeng ◽  
Lili Zhang ◽  
Dongmei Li ◽  
Guixia Lv ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Clinical Isolates ◽  
Resistant Isolate ◽  
Azole Resistance ◽  
Content Type ◽  
Resistant Strains

ABSTRACTSeventy-twoA. fumigatusclinical isolates from China were investigated for azole resistance based on mutations ofcyp51A. We identified four azole-resistant strains, among which we found three strains highly resistant to itraconazole, two of which exhibit the TR34/L98H/S297T/F495I mutation, while one carries only the TR34/L98H mutation. To our knowledge, the latter has not been found previously in China. The fourth multiazole-resistant isolate (with only moderate itraconazole resistance) carries a new G432A mutation.

scholarly journals Genome-Wide Expression Profile Analysis Reveals Coordinately Regulated Genes Associated with Stepwise Acquisition of Azole Resistance in Candida albicans Clinical Isolates

2003 ◽  
Vol 47 (4) ◽  
pp. 1220-1227 ◽  
Author(s):  
P. David Rogers ◽  
Katherine S. Barker
Keyword(s):  
Candida Albicans ◽  
Expression Profile ◽  
Profile Analysis ◽  
Clinical Isolates ◽  
Azole Resistance ◽  
Down Regulation ◽  
Human Fungal Pathogen ◽  
Expression Profile Analysis ◽  
Genome Wide ◽  
First Time

ABSTRACT Candida albicans is an opportunistic human fungal pathogen and a causative agent of oropharyngeal candidiasis (OPC), the most frequent opportunistic infection among patients with AIDS. Fluconazole and other azole antifungal agents have proven effective in the management of OPC; however, with increased use of these agents treatment failures have occurred. Such failures have been associated with the emergence of azole-resistant strains of C. albicans. In the present study we examined changes in the genome-wide gene expression profile of a series of C. albicans clinical isolates representing the stepwise acquisition of azole resistance. In addition to genes previously associated with azole resistance, we identified many genes whose differential expression was for the first time associated with this phenotype. Furthermore, the expression of these genes was correlated with that of the known resistance genes CDR1, CDR2, and CaMDR1. Genes coordinately regulated with the up-regulation of CDR1 and CDR2 included the up-regulation of GPX1 and RTA3 and the down-regulation of EBP1. Genes coordinately regulated with the up-regulation of CaMDR1 included the up-regulation of IFD1, IFD4, IFD5, IFD7, GRP2, DPP1, CRD2, and INO1 and the down-regulation of FET34, OPI3, and IPF1222. Several of these appeared to be coordinately regulated with both the CDR genes and CaMDR1. Many of these genes are involved in the oxidative stress response, suggesting that reduced susceptibility to oxidative damage may contribute to azole resistance. Further evaluation of the role these genes and their respective gene products play in azole antifungal resistance is warranted.

scholarly journals Analytical and Clinical Evaluation of the PathoNostics AsperGenius Assay for Detection of Invasive Aspergillosis and Resistance to Azole Antifungal Drugs during Testing of Serum Samples

2015 ◽  
Vol 53 (7) ◽  
pp. 2115-2121 ◽  
Author(s):  
P. Lewis White ◽  
Raquel B. Posso ◽  
Rosemary A. Barnes
Keyword(s):  
Invasive Aspergillosis ◽  
Clinical Performance ◽  
Antifungal Drugs ◽  
Azole Resistance ◽  
Serum Samples ◽  
Content Type ◽  
Management Application ◽  
Noninvasive Samples ◽  
Resistance Markers ◽  
Small Cohort

The commercially developed PathoNostics AsperGenius species assay is a multiplex real-time PCR capable of detecting aspergillosis and genetic markers associated with azole resistance. The assay is validated for testing bronchoalveolar lavage fluids, replacing the requirement for culture and benefiting patient management. Application of this assay to less invasive, easily obtainable samples (e.g., serum) might be advantageous. The aim of this study was to determine the analytical and clinical performance of the AsperGenius species and resistance assays for testing serum samples. For the analytical evaluations, serum samples were spiked with various concentrations ofAspergillusgenomic DNA for extraction, following international recommendations. For the clinical study, 124 DNA extracts from 14 proven/probable invasive aspergillosis (IA) cases, 2 possible IA cases, and 33 controls were tested. The resistance assay was performed onAspergillus fumigatusPCR-positive samples when a sufficient fungal burden was evident. The limits of detection of the species and resistance assays forA. fumigatusDNA were 10 and ≥75 genomes/sample, respectively. Nonreproducible detection at lower burdens was achievable for all markers. With a positivity threshold of 39 cycles, the sensitivity and specificity of the species assay were 78.6% and 100%, respectively. For 7 IA cases, at least one genetic region potentially associated with azole resistance was successfully amplified, although no resistance markers were detected in this small cohort. The AsperGenius assay provides good clinical performance with the added ability to detect azole resistance directly from noninvasive samples. While the available burden will limit application, it remains a significant advancement in the diagnosis and management of aspergillosis.

scholarly journals Resistance Mechanisms and Clinical Features of Fluconazole-Nonsusceptible Candida tropicalis Isolates Compared with Fluconazole-Less-Susceptible Isolates

2016 ◽  
Vol 60 (6) ◽  
pp. 3653-3661 ◽  
Author(s):  
Min Ji Choi ◽  
Eun Jeong Won ◽  
Jong Hee Shin ◽  
Soo Hyun Kim ◽  
Wee-Gyo Lee ◽  
...  
Keyword(s):  
Amino Acid ◽  
Clinical Features ◽  
Candida Tropicalis ◽  
Resistance Mechanisms ◽  
Therapeutic Failure ◽  
Clinical Isolates ◽  
Azole Resistance ◽  
Amino Acid Substitutions ◽  
Content Type ◽  
Expression Levels

We investigated the azole resistance mechanisms and clinical features of fluconazole-nonsusceptible (FNS) isolates ofCandida tropicalisrecovered from Korean surveillance cultures in comparison with fluconazole-less-susceptible (FLS) isolates. Thirty-five clinical isolates ofC. tropicalis, comprising 9 FNS (fluconazole MIC, 4 to 64 μg/ml), 12 FLS (MIC, 1 to 2 μg/ml), and 14 control (MIC, 0.125 to 0.5 μg/ml) isolates, were assessed.CDR1,MDR1, andERG11expression was quantified, and theERG11andUPC2genes were sequenced. Clinical features of 16 patients with FNS or FLS bloodstream isolates were analyzed. Both FNS and FLS isolates had >10-fold higher mean expression levels ofCDR1,MDR1, andERG11genes than control isolates (Pvalues of <0.02 for all). When FNS and FLS isolates were compared, FNS isolates had 3.4-fold higher meanERG11expression levels than FLS isolates (P= 0.004), but there were no differences in those ofCDR1orMDR1. Of all 35 isolates, 4 (2 FNS and 2 FLS) and 28 (8 FNS, 11 FLS, and 9 control) isolates exhibited amino acid substitutions in Erg11p and Upc2p, respectively. Both FNS and FLS bloodstream isolates were associated with azole therapeutic failure (3/4 versus 4/7) or uncleared fungemia (4/6 versus 4/10), but FNS isolates were identified more frequently from patients with previous azole exposure (6/6 versus 3/10;P= 0.011) and immunosuppression (6/6 versus 3/10;P= 0.011). These results reveal that the majority of FNSC. tropicalisisolates show overexpression ofCDR1,MDR1, andERG11genes, and fungemia develops after azole exposure in patients with immunosuppression.

scholarly journals Candida albicans Clinical Isolates from a Southwest Brazilian Tertiary Hospital Exhibit MFS-mediated Azole Resistance Profile

2019 ◽  
Vol 91 (3) ◽  
Author(s):  
ANA CAROLINA C. PINTO ◽  
DEBORA A.S. ROCHA ◽  
DANIEL C. DE MORAES ◽  
MARIA L. JUNQUEIRA ◽  
ANTONIO FERREIRA-PEREIRA
Keyword(s):  
Candida Albicans ◽  
Tertiary Hospital ◽  
Clinical Isolates ◽  
Azole Resistance ◽  
Resistance Profile
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