Survival of bacterial DNA and culturable bacteria in archived soils from the Rothamsted Broadbalk experiment
Survival of bacterial DNA and culturable bacteria in archived soils from the Rothamsted Broadbalk experiment
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DOI：
10.1016/j.soilbio.2007.11.021
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发表时间：
2008-05-01
影响因子：
9.7
通讯作者：
Hirsch, Penny R.
中科院分区：
文献类型：
--
作者：
Clark, Ian M.;Hirsch, Penny R.
多年来，来自许多来源的干燥土壤样品被保存在世界各地的档案馆中，但很少有人研究它们对研究微生物种群的价值。自1843年以来，从洛桑斯特德的Broadbalk作物营养田间实验中收集的样本已被用来记录随着农业实践的发展而发生的土壤结构和组成的变化，还提供了从前到后工业时代环境变化的宝贵记录。在英国。到目前为止，这些土壤的微生物群落还没有被研究，部分原因是由于干燥土壤中细菌可培养性的下降。然而，基于PCR扩增直接从土壤中提取的DNA的现代分子方法不需要细菌细胞是活的或完整的，并且可以允许调查样品收集时存在的细菌的遗留物。在初步研究中，为了确定干燥的土壤是否可以提供细菌群落的历史记录，调查了Broadbalk土壤档案中可追溯到1868年的样品，并比较了用农家肥（FYM）或无机肥（NPK）处理的地块。正如预期的那样，空气干燥和研磨的过程大大降低了细菌的活力，而DNA产量下降较少，并可能通过干燥保存。一个较高比例的可培养细菌存活的归档过程中FYM土壤，可能受到保护的土壤有机质的增加。大多数存活的细菌是厚壁菌门，无论是在2003年还是在1914年收集的，但是使用PCR和16 S rRNA基因的DGGE从样品中提取的DNA中检测到广泛的属。DGGE图谱分析表明，两个小区保持不同的群体。从1914年收集的样品中，从DGGE凝胶上切下的条带的序列分析显示，DNA来自α-和β-变形菌以及厚壁菌门。在最近收集的样品中，使用氨氧化细菌特异性引物的PCR在两种处理中显示出相似的条带谱，然而来自NPK图的较老样品显示出更大的分歧。设计了假单胞菌属特异性引物，并用于实时定量PCR，以表明1868年收集的存档土壤中含有的假单胞菌DNA比新鲜土壤少10倍，约为10（5）个基因组g（-1）土壤。在碾磨之前，与FYM地块相比，从NPK最近收集的风干土壤中提取的假单胞菌DNA显著减少;否则，两个地块遵循相似的趋势。总体细菌丰度，多样性和生存在存档过程中不同的两种土壤，可能是由于粘土和土壤有机质含量的差异。然而，研究结果表明，风干土壤可以保护微生物DNA超过150年，并为未来的研究提供了宝贵的资源。(C)2007爱思唯尔有限公司保留所有权利。
Dried soil samples from many sources have been stored in archives world-wide over the years, but there has been little research on their value for studying microbial populations. Samples collected since 1843 from the Broadbalk field experiment on crop nutrition at Rothamsted have been used to document changes in the structure and composition of soils as agricultural practices evolve, also offering an invaluable record of environmental changes from the pre- to post-industrial era in the UK. To date, the microbial communities of these soils have not been studied, in part due to the well-documented drop in bacterial culturability in dried soils. However, modern molecular methods based on PCR amplification of DNA extracted directly from soil do not require bacterial cells to be viable or intact and may allow investigations into the legacy of bacteria that were present at the time of sample collection.In a preliminary study, to establish if dried soils can provide a historical record of bacterial communities, samples from the Broadbalk soil archive dating back to 1868 were investigated and plots treated with either farmyard manure (FYM) or inorganic fertilizer (NPK) were compared. As anticipated, the processes of air-drying and milling greatly reduced bacterial viability whilst DNA yields declined less and may be preserved by desiccation. A higher proportion of culturable bacteria survived the archiving process in the FYM soil, possibly protected by the increased soil organic matter. The majority of surviving bacteria were firmicutes, whether collected in 2003 or in 1914, but a wide range of genera was detected in DNA extracted from the samples using PCR and DGGE of 16S rRNA genes. Analysis of DGGE band profiles indicated that the two plots maintained divergent populations. Sequence analysis of bands excised from DGGE gels, from a sample collected in 1914, revealed DNA from alpha- and beta-proteobacteria as well as firmicutes. PCR using Primers specific for ammonia oxidizing bacteria showed similar band profiles across the two treatments in recently collected samples, however older samples from the NPK plot showed greater divergence. Primers specific for the genus Pseudomonas were designed and used in real-time quantitative PCR to indicate that archived soil collected in 1868 contained 10-fold less pseudomonad DNA than fresh soil, representing around 10(5) genomes g(-1) soil. Prior to milling, dramatically less pseudomonad DNA was extracted from recently collected air-dried soil from the NPK compared to the FYM plot; otherwise, the two plots followed similar trends. Overall bacterial abundance, diversity and survival during the archiving process differed in the two soils, possibly due to differences in clay and soil organic matter content. Nevertheless, the results demonstrate that air-dried soils can protect microbial DNA for more than 150 years and offer an invaluable resource for future research. (C) 2007 Elsevier Ltd. All rights reserved.