Abstract
Molecular methods have been established to make better sensitivity and specificity for differentiation of pathogenic and allergenic fungi. The purpose of this study was to obtain an appropriate molecular pattern for differentiation of Penicillium species understudy. In this study, 30 isolates of four Penicillium species including Penicillium citrinum,Penicillium notatum, Penicillium oxalicum and Penicillium frequentans were used. First, isolates were cultured. Then, for obtaining the fungal mats, fungi were grown into Sabouraud glucose broth. Freeze and Thaw, and glass beads were used for cell disruption, and DNA was extracted by phenol-chloroform method. RAPD-PCR technique was performed using three primers. The results showed that each primer produced a different reproduction pattern. A total of 2 to 17 bands were observed in all the isolates ofPenicillium, except P. citrinum C2 and P. frequentans F7. According to primer 1, molecular patterns of Penicillium species were approximately similar, but multiplied patterns were not completely similar. Having used primer 2, multiplied patterns in P. oxalicum and P. notatum were to some extent similar, but there were some observed differences between P. frequentans and two other species P. oxalicum and P. notatum.P. citrinum, P. oxalicum and P. notatum had approximately the same patterns using primer 3, but some differences were observed in P. frequentans with three other species.In conclusion, the interspecies differences were demonstrated with all the primers. Primers 1 and 2 were able, to some extent, to differentiate these species, while primers 1 and 3 separated P. frequentans from the others. Key words: Penicillium, molecular pattern, random amplified polymorphic DNA-polymerase chain reaction, molecular bands.
Highlights
Penicillium species classified in subgenus Penicillium are among the most common fungi which spoil food and contaminate indoor environments (Carlile et al, 2001)
Bands of ~510 bp (500 to 550 bp) and 700 bp were observed with frequencies of 16.6 and 26.6%, respectively, while band of ~510 bp was not demonstrated in P. frequentans (Table 2)
Molecular patterns of four Penicillium species were to some extent similar with the primer 1, while the reproduced pattern was not identical and some differences were observed between P. frequentans with the others
Summary
Penicillium species classified in subgenus Penicillium are among the most common fungi which spoil food and contaminate indoor environments (Carlile et al, 2001). These organisms are commonly considered as contaminants but may cause infections, in immunocompromised hosts. The wide spread distribution of Penicillium conidia in the air can cause allergic asthma in atopic individuals (Hoffman, 1984). Species identification relies on evaluation of macromorphological characters, microscopic observation of reproductive structures (De Hoog et al, 2000), recognizable secondary metabolite production (Smedsgaard and Frisvad, 1997) and isoenzyme profiling.
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