Comparison of stool versus rectal swab samples and storage conditions on bacterial community profiles.
Comparison of stool versus rectal swab samples and storage conditions on bacterial community profiles.
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DOI：
10.1186/s12866-017-0983-9
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发表时间：
2017-03-31
期刊：
影响因子：
4.2
通讯作者：
CDC Prevention Epicenters Program
中科院分区：
文献类型：
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作者：
Bassis CM;Moore NM;Lolans K;Seekatz AM;Weinstein RA;Young VB;Hayden MK;CDC Prevention Epicenters Program
从住院患者中采集用于肠道微生物群分析的样本可能具有挑战性。采集方法和储存条件是变异性的潜在来源。在这项研究中，我们比较了在不同条件下储存的粪便中的细菌微生物群，以及粪便和拭子样本，以评估由于样本储存条件和采集方法而导致的差异。使用细菌16 S rRNA基因序列分析，我们比较了在各种条件下储存和收集的粪便样本的微生物群谱。来自两家医院的三名不同患者的粪便样品（2份液体，1份形成）各自在以下条件下进行评价：立即在-80 ° C下冷冻，在-80 ° C下冷冻之前在4°C下储存1 - 2 -48小时，以及在-80 ° C下储存之前在-20 ° C下储存1-2个解冻循环。此外，从一家医院的8名住院患者中采集了8份粪便和30份直肠拭子样本。采用Yue和克莱顿相异指数（θYC距离）和分子方差分析（AMOVA）评估微生物群差异。无论储存条件如何，基于θYC距离（样本内θYC距离中位数：0.035，IQR：0.015-0.061），来自相同粪便样本的等分试样的细菌群落与来自不同粪便样本的等分试样（样本间θYC距离中位数：0.93，IQR：0.85-0.97）非常相似（Wilcoxon检验p值：<0.0001）。对于粪便和直肠拭子比较，来自不同患者的样品，无论样品收集方法如何，均显著不同（AMOVA p值：<0.001-0.029），相比之下，所有粪便和拭子样品之间无显著差异（AMOVA p值：0.976）。个体内拭子和粪便样本之间的θYC相异指数（中位数0.17，IQR：0.10-0.27）显著低于个体间（中位数0.93，IQR：0.85-0.97）（Wilcoxon检验p值：<0.0001），表明采样期间从同一个体采集的粪便和拭子样本之间的差异极小。 对于基于细菌16 S rRNA基因序列分析的胃肠道微生物群研究，在4 °C或-20 °C下临时储存粪便样本，而不是在-80 °C下立即储存，不会显著改变结果。此外，来自同一受试者的粪便和直肠拭子微生物群落高度相似，表明这些采样方法可互换使用，以评估远端胃肠道的群落结构。
Sample collection for gut microbiota analysis from in-patients can be challenging. Collection method and storage conditions are potential sources of variability. In this study, we compared the bacterial microbiota from stool stored under different conditions, as well as stool and swab samples, to assess differences due to sample storage conditions and collection method. Using bacterial 16S rRNA gene sequence analysis, we compared the microbiota profiles of stool samples stored and collected under various conditions. Stool samples (2 liquid, 1 formed) from three different patients at two hospitals were each evaluated under the following conditions: immediately frozen at -80°C, stored at 4°C for 12-48 hours before freezing at -80°C and stored at -20°C with 1-2 thaw cycles before storage at -80°C. Additionally, 8 stool and 30 rectal swab samples were collected from 8 in-patients at one hospital. Microbiota differences were assessed using the Yue and Clayton dissimilarity index (θYC distance) and analysis of molecular variance (AMOVA). Regardless of the storage conditions, the bacterial communities of aliquots from the same stool samples were very similar based on θYC distances (median intra-sample θYC distance: 0.035, IQR: 0.015-0.061) compared to aliquots from different stool samples (median inter-sample θYC distance: 0.93, IQR: 0.85-0.97) (Wilcoxon test p-value: <0.0001). For the stool and rectal swab comparison, samples from different patients, regardless of sample collection method, were significantly different (AMOVA p-values: <0.001-0.029) compared to no significant difference between all stool and swab samples (AMOVA p-value: 0.976). The θYC dissimilarity index between swab and stool samples was significantly lower within individuals (median 0.17, IQR: 0.10-0.27) than between individuals (median 0.93, IQR: 0.85-0.97) (Wilcoxon test p-value: <0.0001), indicating minimal differences between stool and swab samples collected from the same individual over the sampling period. For gastrointestinal microbiota studies based on bacterial 16S rRNA gene sequence analysis, interim stool sample storage at 4 °C or -20 °C, rather than immediate storage at -80 °C, does not significantly alter results. Additionally, stool and rectal swab microbiotas from the same subject were highly similar, indicating that these sampling methods could be used interchangeably to assess the community structure of the distal GI tract.