Efficiency of fluorescence in situ hybridization for bacterial cell identification in temporary river sediments with contrasting water content
Efficiency of fluorescence in situ hybridization for bacterial cell identification in temporary river sediments with contrasting water content
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DOI：
10.1016/j.syapm.2007.03.003
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发表时间：
2007-09-01
影响因子：
3.4
通讯作者：
Pernthaler, Jakob
中科院分区：
文献类型：
--
作者：
Fazi, Stefano;Amalfitano, Stefano;Pernthaler, Jakob
研究了两种杂交技术在不同沉积物含水率下分析底栖细菌群落组成的效率。微观世界是用来自欧洲四条临时河流的沉积物建立的。湿沉淀物被干燥，干沉淀物被人工再湿。荧光单标记探针(FISH)荧光原位杂交检测到的细菌细胞百分比从干沉积物到湿沉积物显著增加，与通过掺入H-3亮氨酸测量的群落活性呈正相关。FISH和催化报告沉积信号放大技术(CARD-FISH)能更好地检测干沉积物中活性较低的细胞。通过应用优化的细胞渗透方案，在干燥和潮湿条件下，CARD-FISH杂交细胞的百分比显示出可比值。这种方法与H-3亮氨酸参入率无关。此外，优化的方案允许显著更好地可视化研究样本中的革兰氏阳性放线杆菌。因此，CARD-FISH被提出为一种有效的技术，用于比较居住在沉积物中的细菌群落与不同水分含量的细菌群落，而不考虑目标细胞活动状态的差异。考虑到欧洲临时河流中洪水和干旱循环的频率越来越高，我们的方法可能有助于更好地了解此类系统中微生物群落的动态。(C)2007年爱思唯尔股份有限公司。版权所有。
We studied the efficiency of two hybridization techniques for the analysis of benthic bacterial community composition under varying sediment water content. Microcosms were set up with sediments from four European temporary rivers. Wet sediments were dried, and dry sediments were artificially rewetted. The percentage of bacterial cells detected by fluorescence in situ hybridization with fluorescently monolabeled probes (FISH) significantly increased from dry to wet sediments, showing a positive correlation with the community activity measured via incorporation of H-3 leucine. FISH and signal amplification by catalyzed reporter deposition (CARD-FISH) could significantly better detect cells with low activity in dried sediments. Through the application of an optimized cell permeabilization protocol, the percentage of hybridized cells by CARD-FISH showed comparable values in dry and wet conditions. This approach was unrelated to H-3 leucine incorporation rates. Moreover, the optimized protocol allowed a significantly better visualization of Gram-positive Actinobacteria in the studied samples. CARD-FISH is, therefore, proposed as an effective technique to compare bacterial communities residing in sediments with contrasting water content, irrespective of differences in the activity state of target cells. Considering the increasing frequencies of flood and drought cycles in European temporary rivers, our approach may help to better understand the dynamics of microbial communities in such systems. (c) 2007 Elsevier GmbH. All rights reserved.