scholarly journals Antifungal Susceptibility Profiles and Drug Resistance Mechanisms of Clinical Lomentospora prolificans Isolates

2020 ◽  
Vol 64 (11) ◽  
Author(s):  
Yongqin Wu ◽  
Nina Grossman ◽  
Marissa Totten ◽  
Warda Memon ◽  
Anna Fitzgerald ◽  
...  
Keyword(s):  
Amino Acid ◽  
Tandem Repeats ◽  
Hot Spot ◽  
Antifungal Susceptibility ◽  
Antifungal Drugs ◽  
Upstream Region ◽  
Amino Acid Residues ◽  
Content Type ◽  
Echinocandin Resistance

ABSTRACT Lomentospora prolificans is an opportunistic fungal pathogen with low susceptibility to current antifungal drugs. Here, we tested the in vitro susceptibility of 8 drugs against 42 clinical L. prolificans isolates. All isolates showed high MICs to voriconazole (MIC90>16 μg/ml), itraconazole (MIC90>16 μg/ml), posaconazole (MIC90>16 μg/ml), isavuconazole (MIC90>16 μg/ml), amphotericin B (MIC90>16 μg/ml), and terbinafine (MIC90>64 μg/ml) and high minimum effective concentrations (MECs) to micafungin (MEC90>8 μg/ml), with the exception of miltefosine showing an MIC90 value of 4 μg/ml. We examined six different in vitro drug combinations and found that the combination of voriconazole and terbinafine achieved the most synergistic effort against L. prolificans. We then annotated the L. prolificans whole genome and located its Cyp51 and Fks1 genes. We completely sequenced the two genes to determine if any mutation would be related to azole and echinocandin resistance in L. prolificans. We found no amino acid changes in Cyp51 protein and no tandem repeats in the 5′ upstream region of the Cyp51 gene. However, we identified three intrinsic amino acid residues (G138S, M220I, and T289A) in the Cyp51 protein that were linked to azole resistance. Likewise, two intrinsic amino acid residues (F639Y, W695F) that have reported to confer echinocandin resistance were found in Fks1 hot spot regions. In addition, three new amino acid alterations (D440A, S634R, and H1245R) were found outside Fks1 hot spot regions, and their contributions to echinocandin resistance need future investigation. Overall, our findings support the notion that L. prolificans is intrinsically resistant to azoles and echinocandins.

scholarly journals Understanding Echinocandin Resistance in the Emerging Pathogen Candida auris

2018 ◽  
Vol 62 (6) ◽  
Author(s):  
Milena Kordalewska ◽  
Annie Lee ◽  
Steven Park ◽  
Indira Berrio ◽  
Anuradha Chowdhary ◽  
...  
Keyword(s):  
Hot Spot ◽  
Antifungal Susceptibility ◽  
Antifungal Drugs ◽  
Clinical Isolates ◽  
Growth Effect ◽  
Candida Auris ◽  
Content Type ◽  
Broth Microdilution Method ◽  
Eagle Effect

ABSTRACT Candida auris has simultaneously emerged on five continents as a fungal pathogen causing nosocomial outbreaks. The challenges in the treatment of C. auris infections are the variable antifungal susceptibility profiles among clinical isolates and the development of resistance to single or multiple classes of available antifungal drugs. Here, the in vitro susceptibility to echinocandin antifungal drugs was determined and FKS1 sequencing was performed on 106 C. auris clinical isolates. Four isolates were identified to be resistant to all tested echinocandins (MIC ≥ 4 mg/liter) and harbored an S639F mutation in FKS1 hot spot region 1. All remaining isolates were FKS1 wild type (WT) and echinocandin susceptible, with micafungin being the most potent echinocandin (MIC 50 = 0.125 mg/liter). Antifungal susceptibility testing with caspofungin was challenging due to the fact that all FKS1 WT isolates exhibited an Eagle effect (also known as the paradoxical growth effect), which occurred at various intensities. To assess whether the Eagle effect resulted in pharmacodynamic resistance, 8 representative isolates were evaluated for their in vivo drug response in a murine model of invasive candidiasis. All isolates were susceptible to caspofungin at a human therapeutic dose, except for those harboring the S639F mutation. The data suggest that only isolates carrying mutations in FKS1 are echinocandin resistant and that routine in vitro testing of C. auris isolates for susceptibility to caspofungin by the broth microdilution method should be viewed cautiously or avoided.

scholarly journals Molecular Identification and Antifungal Susceptibility of Yeast Isolates Causing Fungemia Collected in a Population-Based Study in Spain in 2010 and 2011

2013 ◽  
Vol 58 (3) ◽  
pp. 1529-1537 ◽  
Author(s):  
Jesús Guinea ◽  
Óscar Zaragoza ◽  
Pilar Escribano ◽  
Estrella Martín-Mazuelos ◽  
Javier Pemán ◽  
...  
Keyword(s):  
Hot Spot ◽  
Antifungal Susceptibility ◽  
Point Mutations ◽  
Population Based ◽  
Candida Guilliermondii ◽  
Population Based Study ◽  
Content Type ◽  
Candida Lusitaniae ◽  
Species Specific

ABSTRACTWe report the molecular identifications and antifungal susceptibilities of the isolates causing fungemia collected in the CANDIPOP population-based study conducted in 29 Spanish hospitals. A total of 781 isolates (from 767 patients, 14 of them having mixed fungemia) were collected. The species found most frequently wereCandida albicans(44.6%),Candida parapsilosis(24.5%),Candida glabrata(13.2%),Candida tropicalis(7.6%),Candida krusei(1.9%),Candida guilliermondii(1.7%), andCandida lusitaniae(1.3%). OtherCandidaand non-Candidaspecies accounted for approximately 5% of the isolates. The presence of cryptic species was low. Compared to findings of previous studies conducted in Spain, the frequency ofC. glabratahas increased. Antifungal susceptibility testing was performed by using EUCAST and CLSI M27-A3 reference procedures; the two methods were comparable. The rate of fluconazole-susceptible isolates was 80%, which appears to be a decrease compared to findings of previous studies, explained mainly by the higher frequency ofC. glabrata. Using the species-specific breakpoints and epidemiological cutoff values, the rate of voriconazole and posaconazolein vitroresistance was low (<2%). In the case ofC. tropicalis, using the EUCAST procedure, the rate of azole resistance was around 20%. There was a correlation between the previous use of azoles and the presence of fluconazole-resistant isolates. Resistance to echinocandins was very rare (2%), and resistance to amphotericin B also was very uncommon. The sequencing of the hot spot (HS) regions fromFKS1orFKS2genes in echinocandin-resistant isolates revealed previously described point mutations. The decrease in the susceptibility to fluconazole in Spanish isolates should be closely monitored in future studies.

scholarly journals Molecular Analysis of Resistance and Detection of Non-Wild-Type Strains Using Etest Epidemiological Cutoff Values for Amphotericin B and Echinocandins for Bloodstream Candida Infections from a Tertiary Hospital in Qatar

2018 ◽  
Vol 62 (9) ◽  
Author(s):  
Saad J. Taj-Aldeen ◽  
Husam Salah ◽  
Winder B. Perez ◽  
Muna Almaslamani ◽  
Mary Motyl ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Hot Spot ◽  
Antifungal Susceptibility ◽  
Tertiary Hospital ◽  
Wild Type ◽  
Content Type ◽  
Cutoff Values ◽  
Echinocandin Resistance ◽  
Candida Infections ◽  
Type Strains

ABSTRACT A total of 301 Candida bloodstream isolates collected from 289 patients over 5 years at a tertiary hospital in Qatar were evaluated. Out of all Candida infections, 53% were diagnosed in patients admitted to the intensive care units. Steady increases in non-albicans Candida species were reported from 2009 to 2014 (30.2% for Candida albicans versus 69.8% for the other Candida species). Etest antifungal susceptibility testing was performed on all recovered clinical isolates to determine echinocandin (micafungin and anidulafungin) and amphotericin B susceptibilities and assess non-wild-type (non-WT) strains (strains for which MICs were above the epidemiological cutoff values). DNA sequence analysis was performed on all isolates to assess the presence of FKS mutations, which confer echinocandin resistance in Candida species. A total of 3.9% of isolates (12/301) among strains of C. albicans and C. orthopsilosis contained FKS hot spot mutations, including heterozygous mutations in FKS1. For C. tropicalis, the Etest appeared to overestimate strains non-WT for micafungin, anidulafungin, and amphotericin B, as 14%, 11%, and 35% of strains, respectively, had values above the epidemiological cutoff value. However, no FKS mutations were identified in this species. For all other species, micafungin best reported the echinocandin non-WT strains relative to the FKS genotype, as anidulafungin tended to overestimate non-wild-type strains. Besides C. tropicalis, few strains were classified as non-WT for amphotericin B.

scholarly journals Low Level of Antifungal Resistance in Iranian Isolates of Candida glabrata Recovered from Blood Samples in a Multicenter Study from 2015 to 2018 and Potential Prognostic Values of Genotyping and Sequencing of PDR1

2019 ◽  
Vol 63 (7) ◽  
Author(s):  
Amir Arastehfar ◽  
Farnaz Daneshnia ◽  
Kamiar Zomorodian ◽  
Mohammad Javad Najafzadeh ◽  
Sadegh Khodavaisy ◽  
...  
Keyword(s):  
Mortality Rate ◽  
Hot Spot ◽  
Large Subunit ◽  
Antifungal Susceptibility ◽  
Antifungal Drugs ◽  
Specific Gene ◽  
Missense Mutations ◽  
Content Type ◽  
Flight Mass Spectrometry ◽  
Crude Mortality Rate

ABSTRACT Establishing an effective empirical antifungal therapy requires that national surveillance studies be conducted. Herein, we report the clinical outcome of infections with and the microbiological features of Iranian isolates of Candida glabrata derived from patients suffering from candidemia. C. glabrata isolates were retrospectively collected from four major cities in Iran; identified by a 21-plex PCR, matrix-assisted laser desorption ionization–time of flight mass spectrometry, and large subunit of ribosomal DNA sequencing; and genotyped by amplified fragment length polymorphism (AFLP). Mutations in PDR1, ERG11, and hot spot 1 (HS1) of FKS1 and FKS2 were investigated, and antifungal susceptibility testing (AFST) was performed (by the CLSI M27-A3 and M27-S4 methods). Seventy isolates of C. glabrata were collected from 65 patients with a median age of 58 years. Fluconazole was the most widely used (29.23%) and least effective antifungal agent. The overall crude mortality rate was 35.4%. Only one strain was resistant to fluconazole, and 57.7% and 37.5% of the isolates were non-wild type (non-WT) for susceptibility to caspofungin and voriconazole, respectively. All isolates showed the WT phenotype for amphotericin B, posaconazole, and itraconazole. HS1 of FKS1 and FKS2 did not harbor any mutations, while numerous missense mutations were observed in PDR1 and ERG11. AFLP clustered our isolates into nine genotypes; among them, genotypes 1 and 2 were significantly associated with a higher mortality rate (P = 0.034 and P = 0.022, α < 0.05). Moreover, 83.3% of patients infected with strains harboring a single new mutation in PDR1, T745A, died despite treatment with fluconazole or caspofungin. Overall, Iranian isolates of C. glabrata were susceptible to the major antifungal drugs. Application of genotyping techniques and sequencing of a specific gene (PDR1) might have prognostic implications.

scholarly journals Trypanosome Alternative Oxidase Possesses both an N-Terminal and Internal Mitochondrial Targeting Signal

2014 ◽  
Vol 13 (4) ◽  
pp. 539-547 ◽  
Author(s):  
VaNae Hamilton ◽  
Ujjal K. Singha ◽  
Joseph T. Smith ◽  
Ebony Weems ◽  
Minu Chaudhuri
Keyword(s):  
Amino Acid ◽  
Alternative Oxidase ◽  
Heterologous Protein ◽  
Amino Acid Residues ◽  
Mitochondrial Targeting ◽  
Content Type ◽  
Targeting Signal ◽  
Internal Signal ◽  
Trypanosome Alternative Oxidase

ABSTRACTRecognition of mitochondrial targeting signals (MTS) by receptor translocases of outer and inner membranes of mitochondria is one of the prerequisites for import of nucleus-encoded proteins into this organelle. The MTS for a majority of trypanosomatid mitochondrial proteins have not been well defined. Here we analyzed the targeting signal for trypanosome alternative oxidase (TAO), which functions as the sole terminal oxidase in the infective form ofTrypanosoma brucei. Deleting the first 10 of 24 amino acids predicted to be the classical N-terminal MTS of TAO did not affect its import into mitochondriain vitro. Furthermore, ectopically expressed TAO was targeted to mitochondria in both forms of the parasite even after deletion of first 40 amino acid residues. However, deletion of more than 20 amino acid residues from the N terminus reduced the efficiency of import. These data suggest that besides an N-terminal MTS, TAO possesses an internal mitochondrial targeting signal. In addition, both the N-terminal MTS and the mature TAO protein were able to target a cytosolic protein, dihydrofolate reductase (DHFR), to aT. bruceimitochondrion. Further analysis identified a cryptic internal MTS of TAO, located within amino acid residues 115 to 146, which was fully capable of targeting DHFR to mitochondria. The internal signal was more efficient than the N-terminal MTS for import of this heterologous protein. Together, these results show that TAO possesses a cleavable N-terminal MTS as well as an internal MTS and that these signals act together for efficient import of TAO into mitochondria.

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 The Intracellular Cyclophilin PpiB Contributes to the Virulence ofStaphylococcus aureusIndependently of Its Peptidyl-Prolylcis/transIsomerase Activity

2018 ◽  
Vol 86 (11) ◽  
Author(s):  
Rebecca A. Keogh ◽  
Rachel L. Zapf ◽  
Richard E. Wiemels ◽  
Marcus A. Wittekind ◽  
Ronan K. Carroll
Keyword(s):  
Staphylococcus Aureus ◽  
Amino Acid ◽  
Single Amino Acid ◽  
Amino Acid Residues ◽  
Prolyl Isomerase ◽  
Ppiase Activity ◽  
Content Type ◽  
Allelic Exchange ◽  
Isomerase Activity

ABSTRACTTheStaphylococcus aureuscyclophilin PpiB is an intracellular peptidyl prolylcis/transisomerase (PPIase) that has previously been shown to contribute to secreted nuclease and hemolytic activity. In this study, we investigated the contribution of PpiB toS. aureusvirulence. Using a murine abscess model of infection, we demonstrated that appiBmutant is attenuated for virulence. We went on to investigate the mechanism through which PpiB protein contributes to virulence, in particular the contribution of PpiB PPIase activity. We determined the amino acid residues that are important for PpiB PPIase activity and showed that a single amino acid substitution (F64A) completely abrogates PPIase activity. Using purified PpiB F64A proteinin vitro, we showed that PPIase activity only partially contributes to Nuc refolding and that PpiB also possesses PPIase-independent activity. Using allelic exchange, we introduced the F64A substitution onto theS. aureuschromosome, generating a strain that produces enzymatically inactive PpiB. Analysis of the PpiB F64A strain revealed that PPIase activity is not required for hemolysis of human blood or virulence in a mouse. Together, these results demonstrate that PpiB contributes toS. aureusvirulence via a mechanism unrelated to prolyl isomerase activity.

scholarly journals Identification and Antifungal Susceptibility of Penicillium-Like Fungi from Clinical Samples in the United States

2016 ◽  
Vol 54 (8) ◽  
pp. 2155-2161 ◽  
Author(s):  
Marcela Guevara-Suarez ◽  
Deanna A. Sutton ◽  
José F. Cano-Lira ◽  
Dania García ◽  
Adela Martin-Vicente ◽  
...  
Keyword(s):  
United States ◽  
Antifungal Susceptibility ◽  
The United States ◽  
Growth Conditions ◽  
Antifungal Drugs ◽  
Health Science ◽  
Clinical Samples ◽  
Science Center ◽  
Content Type

Penicilliumspecies are some of the most common fungi observed worldwide and have an important economic impact as well as being occasional agents of human and animal mycoses. A total of 118 isolates thought to belong to the genusPenicilliumbased on morphological features were obtained from the Fungus Testing Laboratory at the University of Texas Health Science Center in San Antonio (United States). The isolates were studied phenotypically using standard growth conditions. Molecular identification was made using two genetic markers, the internal transcribed spacer (ITS) and a fragment of the β-tubulin gene. In order to assess phylogenetic relationships, maximum likelihood and Bayesian inference assessments were used. Antifungal susceptibility testing was performed according to CLSI document M38-A2 for nine antifungal drugs. The isolates were identified within three genera, i.e.,Penicillium,Talaromyces, andRasamsonia. The most frequent species in our study werePenicillium rubens,P. citrinum, andTalaromyces amestolkiae. The potentin vitroactivity of amphotericin B (AMB) and terbinafine (TRB) and of the echinocandins againstPenicilliumandTalaromycesspecies might offer a good therapeutic alternative for the treatment of infections caused by these fungi.

scholarly journals In Vitro Antifungal Susceptibility Profiles of 12 Antifungal Drugs against 55 Trichophyton schoenleinii Isolates from Tinea Capitis Favosa Patients in Iran, Turkey, and China

2016 ◽  
Vol 61 (2) ◽  
Author(s):  
Shuwen Deng ◽  
Saham Ansari ◽  
Macit Ilkit ◽  
Haleh Rafati ◽  
Mohammad T. Hedayati ◽  
...  
Keyword(s):  
Antifungal Agents ◽  
Antifungal Susceptibility ◽  
Antifungal Drugs ◽  
Content Type ◽  
Microdilution Method ◽  
Broth Microdilution Method ◽  
In Vitro Antifungal Susceptibility ◽  
Trichophyton Schoenleinii ◽  
Susceptibility Profiles

ABSTRACT Trichophyton schoenleinii is an anthropophilic dermatophyte mainly causing tinea favosa of the scalp in certain regions of the world, especially Africa and Asia. We investigated the in vitro susceptibilities of 55 T. schoenleinii isolates collected over the last 30 years from Iran, Turkey, and China to 12 antifungals using the CLSI broth microdilution method. Our results revealed that terbinafine and ketoconazole were the most potent antifungal agents among those tested, independently of the geographic regions where strains were isolated.

scholarly journals Rapid Development of Candida krusei Echinocandin Resistance during Caspofungin Therapy

2015 ◽  
Vol 59 (11) ◽  
pp. 6975-6982 ◽  
Author(s):  
A. Forastiero ◽  
V. Garcia-Gil ◽  
O. Rivero-Menendez ◽  
R. Garcia-Rubio ◽  
M. C. Monteiro ◽  
...  
Keyword(s):  
Hot Spot ◽  
Rapid Development ◽  
Lymphocytic Leukemia ◽  
Molecular Evidence ◽  
Intrinsic Resistance ◽  
Content Type ◽  
Rapid Acquisition ◽  
Clinical Resistance ◽  
Echinocandin Resistance

ABSTRACTIn invasive candidiasis, there has been an epidemiological shift fromCandida albicansto non-albicansspecies infections, including infections withC. glabrata,C. parapsilosis,C. tropicalis, andC. krusei. Although the prevalence ofC. kruseiremains low among yeast infections, its intrinsic resistance to fluconazole raises epidemiological and therapeutic concerns. Echinocandins havein vitroactivity against mostCandidaspp. and are the first-line agents in the treatment of candidemia. Although resistance to echinocandin drugs is still rare, individual cases ofC. kruseiresistance have been reported in recent years, especially with strains that have been under selective pressure. A total of 15C. kruseistrains, isolated from the blood, urine, and soft tissue of an acute lymphocytic leukemia patient, were analyzed. Strains developed echinocandin resistance during 10 days of caspofungin therapy. The molecular epidemiology of the isolates was investigated using two different typing methods: PCR-based amplification of the species-specific repetitive polymorphic CKRS-1 sequence and multilocus sequence typing. All isolates were genetically related, and the mechanism involved in decreased echinocandin susceptibility was characterized. Clinical resistance was associated with an increase in echinocandin MICsin vitroand was related to three different mutations in hot spot 1 of the target enzyme Fks1p. Molecular evidence of the rapid acquisition of resistance by different mutations inFKS1highlights the need to monitor the development of resistance inC. kruseiinfections treated with echinocandin drugs.

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