Hubad Barbara, Lapanje Aleš
Institute of Microbial Sciences and Technologies, Domžale, Slovenia.
PLoS One. 2013 Dec 20;8(12):e82186. doi: 10.1371/journal.pone.0082186. eCollection 2013.
Cultivation-based microbiological methods are a gold standard for monitoring of airborne micro-organisms to determine the occupational exposure levels or transmission paths of a particular infectious agent. Some highly contagious microorganisms are not easily culturable but it is becoming evident that cultivation and molecular methods are complementary and in these cases highly relevant. We report a simple and efficient method for sampling and analyzing airborne bacteria with an impactor-type high-flow-rate portable air sampler, currently used for monitoring culturable bacteria or fungi. A method is reported for extraction of nucleic acids from impacted cells without prior cultivation and using agarose as a sampling matrix. The DNA extraction efficiency was determined in spiked samples and, samples taken from a wastewater treatment plant and an alpine area. The abundance, diversity and quantity of total bacteria were analysed by a quantitative polymerase chain reaction, and by construction and analysis of clone libraries. The method does not interfere with downstream PCR analysis and can cover the gap between traditional culture and molecular techniques of bioaerosol monitoring.
基于培养的微生物学方法是监测空气传播微生物以确定特定传染源的职业暴露水平或传播途径的金标准。一些高传染性微生物不易培养，但越来越明显的是，培养方法和分子方法是互补的，在这些情况下具有高度相关性。我们报告了一种使用撞击式高流量便携式空气采样器对空气传播细菌进行采样和分析的简单有效方法，该采样器目前用于监测可培养细菌或真菌。本文报道了一种无需事先培养，以琼脂糖作为采样基质从撞击细胞中提取核酸的方法。在加标样品、取自污水处理厂和高山地区的样品中测定了DNA提取效率。通过定量聚合酶链反应以及构建和分析克隆文库，分析了总细菌的丰度、多样性和数量。该方法不会干扰下游的PCR分析，能够填补传统生物气溶胶监测培养技术和分子技术之间的空白。
