Molecular identification and characterization of filamentous fungi and yeasts isolated in a pharmaceutical industry environment

Abstract

Microbiological monitoring of pharmaceutical industry aseptic areas is an important procedure to assess the efficiency of contamination control measures in these areas. Once the permitted microbiological level has been exceeded, the microbial contaminant should be identified, and the actions to eliminate this agent should be adopted. The objective of this study was to identify the filamentous fungi and yeasts isolated from the pharmaceutical industry environment by the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) proteomic approach as well as to analyze fungal universal region polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) capacity in discriminating inter-and intraspecies differences in these isolates. The MALDI-TOF method was able to identify 100% of the samples to the genus level; however, only 68.42% of the samples were identified to the species level. Candida guilliermondii, Penicillium spp., Rhodosporidium toruloides, and Aspergillus species were the most prevalent microorganisms among the samples. The PCR-RFLP analysis of the fungal universal region revealed a heterogeneous profile in the analyzed samples. The isolates of the same species, collected from different sampling points, presented the same restriction profile. MALDI-TOF and PCR-RFLP methods of the fungal universal region were able to, respectively, identify and characterize the genetic diversity and differentiate interspecifically the yeast and fungi samples analyzed in this study. The genetic profiles established by the PCR-RFLP methods of this study can be used to create a database for in-house identification of yeasts and fungi in the laboratory, where this study was carried out.