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

Abstract

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.

Molecular identification of Histoplasma capsulatum using rolling circle amplification

“…Padlock probe HcPL2 presented positive amplification only when the DNA target region of H. capsulatum was used, and no cross‐reaction with strains of species included as a negative controls was observed (Fig. d) corroborating previous results on the effectiveness of this region for identification of other pathogenic fungi . Our data showed that the RCA technique with padlock probe is applicable for rapid and correct identification of H. capsulatum var.…”

“…Nevertheless, padlock probe HcPL1 demonstrated cross‐linking with the above negative controls, showing insufficient specificity and appeared to be unsuitable for H. capsulatum detection. Similar problems with cross‐reactions with probes designed for specific detection are commonly reported . Padlock probe HcPL2 presented positive amplification only when the DNA target region of H. capsulatum was used, and no cross‐reaction with strains of species included as a negative controls was observed (Fig.…”

Molecular identification of unusual Mycetoma agents isolated from patients in Venezuela

“…Cyphellophora is phylogenetically close to Phialophora , and is unusual as causing mycetoma. It has been reported in human skin, and mycetoma in a horse with osteolysis in maxillary region . The patient from whom we isolated C. oxyspora was a young man.…”

“…Interestingly one isolate misidentified as E. jeanselmei was identified as C. oxyspora (case 2). Cyphellophora is a genus of black yeast‐like fungi characterised by having simple phialides with septate conidia . Cyphellophora is phylogenetically close to Phialophora , and is unusual as causing mycetoma.…”

Multiplex detection and genotyping of pathogenic bacteria on paper-based biosensor with a novel universal primer mediated asymmetric PCR

“…The method presented here not only can be used for multiplex detection, it also can be used for genotyping pathogenic bacteria. Compared with traditional discriminatory subtyping methods, such as pulsed-field gel electrophoresis (PFGE) (Chenal-Francisque et al, 2013), amplified fragment length polymorphism (AFLP) (Feng et al, 2013), and multilocus variable-number tandem repeat analysis (MLVA) (Saleh-Lakha et al, 2013), this method is simpler, more efficient and easier to carry out using inexpensive, biodegradable and renewable materials.…”