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

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.

Marine sponges as Chloroflexi hot-spots: Genomic insights and high resolution visualization of an abundant and diverse symbiotic clade

“…Members of Chloroflexi are members of HMA sponge microbiomes, with representatives of classes SAR202, Anaerolineae, and Caldilineae, being most abundant (32). Visualization of Chloroflexi by fluorescence in situ hybridization (FISH) revealed bright and abundant signals (33, 34). Because Chloroflexi likely play an important role in the HMA sponge holobiont, we aimed (i) to assess the relative abundances and distribution in diverse HMA sponge species by using the largest dataset currently available (EMP sponge microbiome), (ii) to provide an updated phylogenetic analysis, (iii) to characterize the functional gene repertoire with a particular focus on carbon degradation and symbiotic lifestyle, and, (iv) to visualize Chloroflexi in mesohyl tissues at ultrastructural resolution by FISH-CLEM methodology.…”

Pangenomic comparison of globally distributed Poribacteria associated with sponge hosts and marine particles

“…Phylogenetic trees constructed from partial 16S rRNA sequences suggest that sponge-associated Poribacteria may fall into four [16,23] or five [24] distinct subclasses, but no correlations have been observed between these classes and sponge host taxonomy or geographical location. Several studies analyzing sponge-associated microbial communities with specifically targeted 16S rRNA gene primers observed Poribacteria-related sequences at relative abundances of 20-30% [1,12,[25][26][27]. Other studies, using only broader "universal" primers, found much lower relative abundances in samples from the same sponge host species [19,28].…”

Taxonomical Structure of Black Soil Bacterial Community on the Level of Phyla

“…The methods used to analyze soil microflora include cultivation of microorganisms on differential media, realtime taxon-specific PCR (RT PCR), metagenomic sequencing of evolutionary conserved sequences, transcriptomic, proteomic and other methods [1,2]. The strengths of taxon-specific RT PCR include rapidity and relatively low cost of analysis, the simplicity of sample preparation, processing and interpretation of the results.…”