Rapid Identification of Pseudallescheria and Scedosporium Strains by Using Rolling Circle Amplification

Lackner, Michaela; Najafzadeh, Mohammad Javad; Sun, Jiufeng; Lu, Qiaoyun; Hoog, G. Sybren de

doi:10.1128/aem.05280-11

Cited by 43 publications

(29 citation statements)

References 38 publications

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“…For clinical practice, a simple and robust diagnostic method is required. RCA has been applied to the identification of diverse human-pathogenic fungi (21,30,31). According to previous studies, the method proved to be easily operated, rapid, costeffective, sensitive, and specific.…”

Section: Discussionmentioning

confidence: 99%

“…Subsequently, rolling circle amplification (RCA) is developed as a sensitive, rapid, and costeffective technique to identify the cutaneous Cyphellophora and Phialophora species. RCA is an isothermal amplification method which has been applied for molecular diagnosis of microbial human pathogens (21)(22)(23). We developed seven RCA padlock probes for the main species in the clade of cutaneous agents of Cyphellophora and Phialophora.…”

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confidence: 99%

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Identification and Typing of Isolates of Cyphellophora and Relatives by Use of Amplified Fragment Length Polymorphism and Rolling Circle Amplification

et al. 2013

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hThe species diversity and identification of black fungi belonging to Cyphellophora and Phialophora, which colonize and infect human skin and nails, were studied using amplified fragment length polymorphism (AFLP). A total of 76 Cyphellophora and Phialophora isolates were evaluated, and their delimitation was compared to earlier studies using multilocus sequencing. The results of the AFLP analysis and sequencing were in complete agreement with each other. Seven species-specific padlock probes for the most prevalent species were designed on the basis of the ribosomal DNA internal transcribed spacer region, and identification of the respective species could easily be achieved with the aid of rolling circle amplification.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Identification and Typing of Isolates of Cyphellophora and Relatives by Use of Amplified Fragment Length Polymorphism and Rolling Circle Amplification

et al. 2013

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“…Genomic DNA of each A. fumigatus isolate was extracted from 3-day-old cultures (24). PCRs were carried out in a 25-l volume, containing 10.5 l of water, 12.5 l of Taq Kapa 2G Robust Ready (Kapa Biosystems Inc., Wilmington, MA, USA), 0.5 l of each primer (10 M), and 50 ng of genomic DNA.…”

Section: Fungal Isolatesmentioning

confidence: 99%

In Vivo Synergy of Amphotericin B plus Posaconazole in Murine Aspergillosis

Martín‐Vicente

Capilla

Guarro

2016

Antimicrob Agents Chemother

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Aspergillus fumigatus is the main mold causing invasive fungal infection that shows high mortality rates. Therapeutic failure and the increase in drug resistance make it necessary to explore alternative treatments for this infection. We have evaluated the efficacy of amphotericin B at 0.8 mg/kg or 0.3 mg/kg of body weight combined with 40 mg/kg of posaconazole against three A. fumigatus isolates in a murine model of disseminated infection. The combination of the polyene and the azole led to a greater increase in survival and a significantly greater reduction in tissue burden than monotherapies.A spergillus fumigatus is the most common mold causing invasive fungal infection (IFI) in immunocompromised patients (1), especially in those with hematological malignancies, with high mortality rates (2-4). Voriconazole (VRC) is the first-line therapy for the treatment of aspergillosis, but in patients with infections that are refractory to this drug, therapy options include other azoles such as itraconazole or posaconazole (PSC), lipid formulations of amphotericin B (LAMB), or echinocandins (5). Because of the relatively limited efficacy of the current antifungal treatments, an exploration of alternative strategies against this difficult-to-treat infection is crucial. Combinations of antifungal agents are not common therapies but might be good alternatives for infections by resistant organisms or when the standard treatments fail (6-11). Synergistic interactions of two drugs with different targets on the fungal cell can be more effective than each drug working alone. In addition, combined therapies can allow lower doses to be administered, with lower toxicity, faster cure, and probably lower costs. Since the efficacies of different antifungal combinations have been demonstrated by several studies in patients with aspergillosis (6, 7, 12), we were interested in evaluating the in vivo efficacy of the combination of amphotericin B (AMB) plus PSC against A. fumigatus. This combination had already been tested in a murine model of invasive aspergillosis caused by Aspergillus flavus (13), although no improvement over the PSC monotherapy was observed. Another study demonstrated the efficacy of suboptimal doses of VRC plus anidulafungin in a murine model of A. fumigatus infection (14), suggesting that combined therapies might have an important role as alternative treatments against systemic aspergillosis, allowing a reduction of the doses administered. One of the isolates tested in the present study (FMR 10528) had already been used in previous studies but showed a poor in vivo response to VRC when administered at 25 mg/kg of body weight despite having a low MIC (15, 16). The goals of this study were (i) to evaluate the efficacy of the combination of AMB plus PSC against isolates of A. fumigatus in a murine model of disseminated aspergillosis, comparing the results with those of the corresponding monotherapies and VRC, (ii) to investigate the presence of CYP51A gene mutations that might explain the poor in vivo response of such a...

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“…Two studies used ITS-directed RCA assays as a means of identifying Scedosporium species [43,44]. Zhou et al [43] clearly differentiated S. prolificans from S. apiospermum/ P. boydii, whilst Lackner et al [44] used RCA to distinguish at least seven to eight species within the Scedosporium/ Pseudallescheria species complex.…”

Section: Rolling Circle Amplification (Rca)mentioning

confidence: 99%

The Genus Scedosporium and Pseudallescheria: Current Challenges in Laboratory Diagnosis

Ramsperger

Duan

Sorrell

et al. 2014

Curr Clin Micro Rpt

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Scedosporium and Pseudallescheria fungi are increasingly encountered fungal pathogens. Species identification is important as there are differences in epidemiology, virulence, and antifungal susceptibility that vary according to species or species groups. Histological and culture techniques are limited by low sensitivity and specificity but improved culture approaches employing selective media have improved the recovery of these fungi from clinical samples. Molecular-and proteomics-based methods are increasingly explored and used for species identification. These include multiplex polymerase chain reaction (PCR) and other PCR-based methods, oligonucleotide arrays, DNA sequencing, rolling circle amplification, and matrix-assisted laser desorption time-of-flight mass spectrometry. These techniques have been harnessed for the detection and identification of Scedosporium and Pseudallescheria species directly from clinical samples and from cultures. This review summarizes the methods currently used to enable an informed choice of detection and/or identification techniques in the clinical mycology laboratory.

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Rapid Identification of Pseudallescheria and Scedosporium Strains by Using Rolling Circle Amplification

Cited by 43 publications

References 38 publications

Identification and Typing of Isolates of Cyphellophora and Relatives by Use of Amplified Fragment Length Polymorphism and Rolling Circle Amplification

Identification and Typing of Isolates of Cyphellophora and Relatives by Use of Amplified Fragment Length Polymorphism and Rolling Circle Amplification

In Vivo Synergy of Amphotericin B plus Posaconazole in Murine Aspergillosis

The Genus Scedosporium and Pseudallescheria: Current Challenges in Laboratory Diagnosis

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