scholarly journals Distinct Patterns of Gene Expression Associated with Development of Fluconazole Resistance in Serial Candida albicansIsolates from Human Immunodeficiency Virus-Infected Patients with Oropharyngeal Candidiasis

1998 ◽  
Vol 42 (11) ◽  
pp. 2932-2937 ◽  
Author(s):  
Jose L. Lopez-Ribot ◽  
Robert K. McAtee ◽  
Linda N. Lee ◽  
William R. Kirkpatrick ◽  
Theodore C. White ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Immunodeficiency Virus ◽  
Sequence Data ◽  
Point Mutations ◽  
Oropharyngeal Candidiasis ◽  
Fluconazole Resistance ◽  
Expression Of Genes ◽  
Immunodeficiency Virus ◽  
Major Facilitator ◽  
Abc Transporter Genes

ABSTRACT Resistance to fluconazole is becoming an increasing problem in the management of oropharyngeal candidiasis in human immunodeficiency virus-infected patients. Strains obtained from five patients developed decreased fluconazole susceptibility over time. DNA strain typing confirmed the high degree of relatedness among isolates from one patient and the variability among isolates from different patients. Expression of genes involved in development of fluconazole resistance was monitored in each isolate using probes specific for ERG11 (lanosterol 14α-demethylase), MDR1 (a major facilitator), andCDR (ATP-binding cassette or ABC transporter) genes. Increased expression of CDR genes was detected in the series of isolates from two patients. Isolates from one of the two patients also demonstrated increased ERG11 expression, whereas isolates from the other patient did not. Increased levels ofMDR1 mRNA correlated with increased resistance in sequential isolates from another patient. Initial overexpression ofMDR1 with subsequent overexpression of CDRgenes and a final isolate again overexpressing MDR1 were detected in serial isolates from another patient. In another patient, overexpression of these genes was not detected despite an eightfold increase in fluconazole MIC. In this patient, sequence data of theERG11 gene revealed no point mutations associated with decreased susceptibility. Five different patterns of gene expression were observed in isolates recovered from five patients who developed resistance. Therefore, these experiments demonstrate that a variety of mechanisms or combinations of mechanisms are associated with the development of fluconazole drug resistance. Additional studies are needed to estimate the frequency and clinical impact of these mechanisms of resistance.

scholarly journals Increased mRNA levels of ERG16, CDR, and MDR1 correlate with increases in azole resistance in Candida albicans isolates from a patient infected with human immunodeficiency virus.

1997 ◽  
Vol 41 (7) ◽  
pp. 1482-1487 ◽  
Author(s):  
T C White
Keyword(s):  
Human Immunodeficiency Virus ◽  
Candida Albicans ◽  
Abc Transporter ◽  
Genetic Alterations ◽  
Antifungal Drugs ◽  
Cross Resistance ◽  
Mrna Levels ◽  
Fluconazole Resistance ◽  
Immunodeficiency Virus ◽  
Major Facilitator

Resistance to antifungal drugs, specifically azoles such as fluconazole, in the opportunistic yeast Candida albicans has become an increasing problem in human immunodeficiency virus (HIV)-infected individuals. The molecular mechanisms responsible for this resistance have only recently become apparent and can include alterations in the target enzyme of the azole drugs (lanosterol 14alpha demethylase [14DM]), or in various efflux pumps from both the ABC transporter and major facilitator gene families. To determine which of these possible mechanisms was associated with the development of drug resistance in a particular case, mRNA levels have been studied in a series of 17 clinical isolates taken from a single HIV-infected patient over 2 years, during which time the levels of fluconazole resistance of the strain increased over 200-fold. Using Northern blot analysis of steady-state levels of total RNA from these isolates, we observed increased mRNA levels of ERG16 (the 14DM-encoding gene), CDR1 (an ABC transporter), and MDR1 (a major facilitator) in this series. The timing of the increase in mRNA levels of each of these genes correlated with increases in fluconazole resistance of the isolates. Increased mRNA levels were not observed for three other ABC transporters, two other genes in the ergosterol biosynthetic pathway, or the NADPH-cytochrome P-450 oxidoreductase gene that transfers electrons from NADPH to 14DM. Increases in mRNA levels of ERG16 and CDR1 correlated with increased cross-resistance to ketoconazole and itraconazole but not to amphotericin B. A compilation of the genetic alterations identified in this series suggests that resistance develops gradually and is the sum of several different changes, all of which contribute to the final resistant phenotype.

scholarly journals Low Levels of Antigenic Variability in Fluconazole-Susceptible and -Resistant Candida albicansIsolates from Human Immunodeficiency Virus-Infected Patients with Oropharyngeal Candidiasis

1999 ◽  
Vol 6 (5) ◽  
pp. 665-670 ◽  
Author(s):  
Jose L. Lopez-Ribot ◽  
Robert K. McAtee ◽  
William R. Kirkpatrick ◽  
Roberto La Valle ◽  
Thomas F. Patterson
Keyword(s):  
Human Immunodeficiency Virus ◽  
Cell Wall ◽  
Yeast Cells ◽  
National Committee ◽  
Oropharyngeal Candidiasis ◽  
Fluconazole Resistance ◽  
Homogeneous Population ◽  
Antigenic Variability ◽  
Immunodeficiency Virus ◽  
Different Strains

ABSTRACT Three serial isolates of Candida albicans were obtained by direct swab or by oral saline rinses from each of five human immunodeficiency virus-infected patients with recurrent oropharyngeal candidiasis. Genotyping techniques confirmed the presence of a persistent strain in multiple episodes from the same patient, which was different from the strains isolated from other patients. Fluconazole susceptibility was determined by both an agar dilution method and the National Committee for Clinical Laboratory Standards macrobroth procedure. In four of these patients the strains developed fluconazole resistance, and in one patient the strain remained susceptible. The different isolates were propagated as yeast cells on a synthetic medium, and their cell wall proteinaceous components were extracted by treatment with β-mercaptoethanol. Protein and mannoprotein components present in the extracts were analyzed by electrophoresis, immunoblotting, and lectin-blotting techniques. The analysis showed a similar composition, with only minor qualitative and quantitative differences in the polypeptidic and antigenic patterns associated with the cell wall extracts from serial isolates from the same patient, as well as those from different strains isolated from different patients. Use of monospecific antibodies generated against two immunodominant antigens during candidiasis (enolase and the 58-kDa fibrinogen-binding mannoprotein) demonstrated their expression in all isolates tested. Overall, the antigenic makeup of C. albicans strains remained constant during the course of infection and was not affected by development of fluconazole resistance. In contrast to previous reports, the low degree of antigenic variability observed in this study may be due to the fact that the isolates were obtained from a highly homogeneous population of patients and to the uniformity in techniques used for the isolation, storage, and culture of the different strains, as well as extraction methodologies.

scholarly journals Molecular Mechanisms of Fluconazole Resistance in Candida dubliniensis Isolates from Human Immunodeficiency Virus-Infected Patients with Oropharyngeal Candidiasis

2002 ◽  
Vol 46 (6) ◽  
pp. 1695-1703 ◽  
Author(s):  
Sofia Perea ◽  
José L. López-Ribot ◽  
Brian L. Wickes ◽  
William R. Kirkpatrick ◽  
Olga P. Dib ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Molecular Mechanisms ◽  
Systemic Disease ◽  
Antifungal Susceptibility ◽  
Point Mutations ◽  
Candida Dubliniensis ◽  
Clinical Isolates ◽  
Oropharyngeal Candidiasis ◽  
Mechanisms Of Resistance ◽  
Immunodeficiency Virus

ABSTRACT Candida dubliniensis is a newly identified species of Candida that is phenotypically similar to but genetically distinct from C. albicans. This organism has been recovered with increasing frequency from the oral cavities of human immunodeficiency virus (HIV)-infected and AIDS patients and has been implicated as a causative agent of oral candidiasis and systemic disease. In the present study we characterized the molecular mechanisms of resistance to fluconazole (FLC) in C. dubliniensis clinical isolates from two different HIV-infected patients with oropharyngeal candidiasis. Isolates were identified to the species level by phenotypic and genotypic tests. DNA-typing techniques were used to assess strain identity. Antifungal susceptibility testing was performed by NCCLS techniques. Northern blotting analysis was used to monitor the expression of genes encoding lanosterol demethylase (ERG11) and efflux transporters (CDR and MDR1) in matched sets of C. dubliniensis-susceptible and -resistant isolates by using probes generated from their homologous C. albicans sequences. In addition, ERG11 genes were amplified by PCR, and their nucleotide sequences were determined in order to detect point mutations with a possible effect in the affinity for azoles. Decreasing susceptibilities to FLC were detected in C. dubliniensis isolates recovered from both patients during the course of treatment. FLC-resistant C. dubliniensis isolates from one patient demonstrated combined upregulation of the MDR1, CDR1, and ERG11 genes. Among the isolates from the second patient, all isolates showing decreased susceptibility to FLC demonstrated upregulation of MDR1, whereas the levels of mRNA for the ERG11 genes remained constant and the expression of CDR genes was negligible. Fourteen point mutations were found in the ERG11 genes of the isolates with decreased susceptibility to FLC. These data demonstrate that the development of azole resistance in C. dublinensis clinical isolates from HIV-infected patients treated with FLC is mediated by multiple molecular mechanisms of resistance, similar to the observations found in the case of C. albicans.

scholarly journals Detection and Significance of Fluconazole Resistance in Oropharyngeal Candidiasis in Human Immunodeficiency Virus-Infected Patients

1996 ◽  
Vol 174 (4) ◽  
pp. 821-827 ◽  
Author(s):  
S. G. Revankar ◽  
W. R. Kirkpatrick ◽  
R. K. McAtee ◽  
O. P. Dib ◽  
A. W. Fothergill ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Oropharyngeal Candidiasis ◽  
Fluconazole Resistance ◽  
Immunodeficiency Virus

scholarly journals Comparison of Predicted Scaffold-Compatible Sequence Variation in the Triple-Hairpin Structure of Human Immunodeficiency Virus Type 1 gp41 with Patient Data

2002 ◽  
Vol 76 (15) ◽  
pp. 7595-7606 ◽  
Author(s):  
Nathalie Boutonnet ◽  
Wouter Janssens ◽  
Carlo Boutton ◽  
Jean-Luc Verschelde ◽  
Leo Heyndrickx ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Sequence Variation ◽  
Single Point ◽  
Point Mutations ◽  
Hairpin Structure ◽  
Single Amino Acid ◽  
Immunodeficiency Virus

ABSTRACT It has been proposed that the ectodomain of human immunodeficiency virus type 1 (HIV-1) gp41 (e-gp41), involved in HIV entry into the target cell, exists in at least two conformations, a pre-hairpin intermediate and a fusion-active hairpin structure. To obtain more information on the structure-sequence relationship in e-gp41, we performed in silico a full single-amino-acid substitution analysis, resulting in a Fold Compatible Database (FCD) for each conformation. The FCD contains for each residue position in a given protein a list of values assessing the energetic compatibility (ECO) of each of the 20 natural amino acids at that position. Our results suggest that FCD predictions are in good agreement with the sequence variation observed for well-validated e-gp41 sequences. The data show that at a minECO threshold value of 5 kcal/mol, about 90% of the observed patient sequence variation is encompassed by the FCD predictions. Some inconsistent FCD predictions at N-helix positions packing against residues of the C helix suggest that packing of both peptides may involve some flexibility and may be attributed to an altered orientation of the C-helical domain versus the N-helical region. The permissiveness of sequence variation in the C helices is in agreement with FCD predictions. Comparison of N-core and triple-hairpin FCDs suggests that the N helices may impose more constraints on sequence variation than the C helices. Although the observed sequences of e-gp41 contain many multiple mutations, our method, which is based on single-point mutations, can predict the natural sequence variability of e-gp41 very well.

scholarly journals A genetic-algorithm approach to simulating human immunodeficiency virus evolution reveals the strong impact of multiply infected cells and recombination

2005 ◽  
Vol 86 (11) ◽  
pp. 3109-3118 ◽  
Author(s):  
Gennady Bocharov ◽  
Neville J. Ford ◽  
John Edwards ◽  
Tanja Breinig ◽  
Simon Wain-Hobson ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Copy Number ◽  
Hamming Distance ◽  
Point Mutations ◽  
Strong Impact ◽  
Hiv Replication ◽  
Immunodeficiency Virus ◽  
Genetic Algorithm Approach ◽  
Infected Cells

It has been previously shown that the majority of human immunodeficiency virus type 1 (HIV-1)-infected splenocytes can harbour multiple, divergent proviruses with a copy number ranging from one to eight. This implies that, besides point mutations, recombination should be considered as an important mechanism in the evolution of HIV within an infected host. To explore in detail the possible contributions of multi-infection and recombination to HIV evolution, the effects of major microscopic parameters of HIV replication (i.e. the point-mutation rate, the crossover number, the recombination rate and the provirus copy number) on macroscopic characteristics (such as the Hamming distance and the abundance of n-point mutants) have been simulated in silico. Simulations predict that multiple provirus copies per infected cell and recombination act in synergy to speed up the development of sequence diversity. Point mutations can be fixed for some time without fitness selection. The time needed for the selection of multiple mutations with increased fitness is highly variable, supporting the view that stochastic processes may contribute substantially to the kinetics of HIV variation in vivo.

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