Johnson Philip J, Hargreaves Leeza L, Zobrist Chelsea N, Ericsson Aaron C
University of Missouri College of Veterinary Medicine, 900 E. Campus Dr., Columbia, MO 65211, USA.
University of Missouri Metagenomics Center (MUMC), 4011 Discovery Dr., Columbia, MO 65201, USA; University of Missouri College of Veterinary Medicine, 900 E. Campus Dr., Columbia, MO 65211, USA.
J Microbiol Methods. 2018 Mar;146:1-6. doi: 10.1016/j.mimet.2018.01.007. Epub 2018 Feb 3.
While recent advances in culture-independent sequencing approaches have revitalized the field of microbiology, rapid collection and preservation of microbial DNA in samples like feces is critical to avoid degradation of target DNA via nuclease activity and proliferation of aerotolerant microbes. Common laboratory practices to ameliorate such changes rely on prompt freezing of samples or dispersion in nuclease-inhibiting reagents. As many of the microbial enzymes associated with nuclease activity and bacterial proliferation are hydrolases, prompt desiccation of samples offers an attractive alternative to freezing and liquid reagents for field collection of samples in remote areas. Herein, we evaluated the utility of a portable desiccant chamber with a rechargeable cartridge, for preservation of equine fecal samples for downstream microbial profiling via 16S rRNA amplicon sequencing. Controls included matched samples promptly frozen at -80 °C or left at room temperature for an equivalent period of time. While samples held at room temperature showed a significant reduction in richness and proliferation of several facultative anaerobes, desiccated samples showed minimal change from promptly frozen samples, with the exception of increased abundance of Acinetobacter spp. in desiccated samples relative to frozen samples. The data support the utility of portable desiccant chambers for the preservation of microbial field samples intended for downstream sequencing approaches.
尽管免培养测序方法的最新进展重振了微生物学领域，但在粪便等样本中快速收集和保存微生物DNA对于避免目标DNA因核酸酶活性和气耐氧微生物增殖而降解至关重要。改善此类变化的常见实验室做法依赖于迅速冷冻样本或将其分散在核酸酶抑制试剂中。由于许多与核酸酶活性和细菌增殖相关的微生物酶都是水解酶，因此迅速干燥样本为偏远地区现场采集样本提供了一种有吸引力的替代冷冻和液体试剂的方法。在此，我们评估了一种带有可充电盒的便携式干燥剂室用于保存马粪便样本的效用，以便通过16S rRNA扩增子测序进行下游微生物分析。对照包括立即在-80°C冷冻或在室温下放置相同时间的匹配样本。虽然室温下保存的样本显示出丰富度显著降低，并且几种兼性厌氧菌增殖，但干燥样本与立即冷冻的样本相比变化最小，只是干燥样本中不动杆菌属的丰度相对于冷冻样本有所增加。数据支持便携式干燥剂室用于保存用于下游测序方法的微生物现场样本的效用。
