Rocchi Steffi, Valot Benoit, Reboux Gabriel, Millon Laurence
UMR CNRS 6249 Chrono-environnement, Université Bourgogne Franche-Comté, Besançon, France; Service de parasitologie mycologie, CHRU Jean Minjoz, Besançon, France.
UMR CNRS 6249 Chrono-environnement, Université Bourgogne Franche-Comté, Besançon, France.
J Microbiol Methods. 2017 Aug;139:107-112. doi: 10.1016/j.mimet.2017.05.014. Epub 2017 May 27.
DNA metabarcoding is increasingly being used to characterize the microbiological composition of both the indoor and outdoor environments of dwellings. Our study aimed to evaluate metabarcoding and bioinformatic analysis resulting from calibrated samples and samples collected by an electrostatic dust collector (EDC) in dwellings with no moisture problems. Thus, the fungal communities of 14 dwellings (eastern France, Franche-Comté region) were analyzed by Illumina MiSeq technology after amplification of the ITS2 region. Using the standard samples of 11 species of yeasts and molds allowed us to validate the Operational taxonomic units (OTU) assignment. These calibrated samples also showed a low amplification bias, a low rate of sequencing errors and the semi-quantitative nature of the technique. Only one species from the calibrated samples (Lichtheimia corymbifera) was less amplified probably due to the presence of two mismatches in its3 primer. EDC analysis identified 3594OTU with 75% of reads corresponding to 30 genera. The main genera are those usually found by culture techniques (Penicillium, Aspergillus and Cladosporium), but findings also indicate others less commonly isolated in culture such as Epicoccum, the fourth detected genus in our study. The type of heating systems was correlated with fungal diversity. We found less diversity in the dwellings with wood heating and larger quantities of Epicoccum nigrum verified by qPCR. DNA metabarcoding analysis applied to EDC seems promising. However, we think that it must be used along with qPCR, to obtain a more global view of microbial ecology and relative quantification of species of interest within communities.
DNA宏条形码技术越来越多地被用于描述住宅室内和室外环境的微生物组成。我们的研究旨在评估来自校准样本以及通过静电除尘器（EDC）在无潮湿问题的住宅中收集的样本的宏条形码技术和生物信息学分析。因此，在扩增ITS2区域后，利用Illumina MiSeq技术对14所住宅（法国东部，弗朗什-孔泰地区）的真菌群落进行了分析。使用11种酵母和霉菌的标准样本使我们能够验证操作分类单元（OTU）的归属。这些校准样本还显示出低扩增偏差、低测序错误率以及该技术的半定量性质。校准样本中只有一个物种（伞枝犁头霉）扩增较少，可能是由于其3引物中存在两个错配。EDC分析确定了3594个OTU，其中75%的读数对应于30个属。主要的属是那些通常通过培养技术发现的属（青霉属、曲霉属和枝孢属），但研究结果也表明还有其他在培养中较少分离到的属，如在我们的研究中检测到的第四大属附球菌属。供暖系统的类型与真菌多样性相关。我们发现使用木材供暖的住宅中多样性较低，并且通过qPCR验证了有大量的黑附球菌。应用于EDC的DNA宏条形码分析似乎很有前景。然而，我们认为它必须与qPCR一起使用，以获得更全面的微生物生态学观点以及群落中感兴趣物种的相对定量。
