使用实时聚合酶链式反应(PCR)对高微生物丰度和低微生物丰度海绵中的细菌和古菌共生体进行定量分析。

Quantification of bacterial and archaeal symbionts in high and low microbial abundance sponges using real-time PCR.

作者信息

Bayer Kristina, Kamke Janine, Hentschel Ute

机构信息

Department of Botany II, Julius-von-Sachs Institute for Biological Sciences, University of Wuerzburg, Wuerzburg, Germany.

出版信息

FEMS Microbiol Ecol. 2014 Sep;89(3):679-90. doi: 10.1111/1574-6941.12369. Epub 2014 Jul 9.

Abstract

In spite of considerable insights into the microbial diversity of marine sponges, quantitative information on microbial abundances and community composition remains scarce. Here, we established qPCR assays for the specific quantification of four bacterial phyla of representative sponge symbionts as well as the kingdoms Eubacteria and Archaea. We could show that the 16S rRNA gene numbers of Archaea, Chloroflexi, and the candidate phylum Poribacteria were 4-6 orders of magnitude higher in high microbial abundance (HMA) than in low microbial abundance (LMA) sponges and that actinobacterial 16S rRNA gene numbers were 1-2 orders higher in HMA over LMA sponges, while those for Cyanobacteria were stable between HMA and LMA sponges. Fluorescence in situ hybridization of Aplysina aerophoba tissue sections confirmed the numerical dominance of Chloroflexi, which was followed by Poribacteria. Archaeal and actinobacterial cells were detected in much lower numbers. By use of fluorescence-activated cell sorting as a primer- and probe-independent approach, the dominance of Chloroflexi, Proteobacteria, and Poribacteria in A. aerophoba was confirmed. Our study provides new quantitative insights into the microbiology of sponges and contributes to a better understanding of the HMA/LMA dichotomy.

摘要

尽管对海洋海绵的微生物多样性有了相当深入的了解,但关于微生物丰度和群落组成的定量信息仍然匮乏。在此,我们建立了定量聚合酶链反应(qPCR)检测方法,用于特异性定量四种具有代表性的海绵共生细菌门以及真细菌和古细菌界。我们能够表明,在高微生物丰度(HMA)海绵中,古细菌、绿弯菌门和候选菌门“孔杆菌门”的16S rRNA基因数量比低微生物丰度(LMA)海绵中的高4 - 6个数量级,并且放线菌的16S rRNA基因数量在HMA海绵中比LMA海绵中的高1 - 2个数量级,而蓝细菌的基因数量在HMA和LMA海绵之间保持稳定。对嗜气艾氏海绵组织切片进行荧光原位杂交证实了绿弯菌门在数量上占主导地位,其次是孔杆菌门。检测到的古细菌和放线菌细胞数量要少得多。通过使用荧光激活细胞分选这种不依赖引物和探针的方法,证实了绿弯菌门、变形菌门和孔杆菌门在嗜气艾氏海绵中的主导地位。我们的研究为海绵微生物学提供了新的定量见解,并有助于更好地理解HMA/LMA二分法。

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