Departamento de Higiene y Tecnología de los Alimentos, Universidad de León, 24071 León, Spain.
Food Microbiol. 2013 Dec;36(2):481-7. doi: 10.1016/j.fm.2013.07.009. Epub 2013 Aug 2.
The aim of this work was the genetic characterization at the strain level of 39 presumed Geotrichum candidum isolates isolated throughout the artisanal manufacturing and ripening of Armada cheese and tentatively identified at genus and/or species level by phenotypic characteristics. The molecular identification of the strains included among others the amplification and sequencing of the D1/D2 domains of the 26S rRNA gene. A restriction fragment length polymorphism (RFLP) analysis with the ITS1-5.8S-ITS2 PCR amplicons and a randomly amplified polymorphic DNA (RAPD) analysis with five different primers were carried out. The bands pattern profile obtained through RFLP by enzymatic restriction with HinfI was the same for all the strains studied, which confirmed the classification of the strains at species level. A RAPD-PCR analysis with three different primers was applied to assess the intraspecific diversity, in this way 16 band profiles were obtained for the 39 strains studied by the combined use of primers Ari1 and Omt1. This study contributes to know the occurrence and genotypic biodiversity of G. candidum in Armada cheese.
本工作的目的是对在 Armada 奶酪的手工制作和成熟过程中分离出的 39 株假定的近平滑念珠菌菌株进行种水平的遗传特征分析,这些菌株最初根据表型特征鉴定为属和/或种水平。菌株的分子鉴定包括扩增和测序 26S rRNA 基因的 D1/D2 结构域。进行了 ITS1-5.8S-ITS2PCR 扩增子的限制性片段长度多态性 (RFLP) 分析和 5 种不同引物的随机扩增多态性 DNA (RAPD) 分析。用 HindIII 酶切进行 RFLP 获得的带型图谱在所有研究的菌株中均相同,这证实了菌株在种水平上的分类。应用 RAPD-PCR 分析了三个不同的引物,以评估种内多样性,从而通过组合使用引物 Ari1 和 Omt1 获得了 39 株研究菌株的 16 个条带图谱。本研究有助于了解 Armada 奶酪中近平滑念珠菌的发生和基因型多样性。