Captured metagenomics: large-scale targeting of genes based on ‘sequence capture’ reveals functional diversity in soils

Abstract

Microbial enzyme diversity is a key to understand many ecosystem processes. Whole metagenome sequencing (WMG) obtains information on functional genes, but it is costly and inefficient due to large amount of sequencing that is required. In this study, we have applied a captured metagenomics technique for functional genes in soil microorganisms, as an alternative to WMG. Large-scale targeting of functional genes, coding for enzymes related to organic matter degradation, was applied to two agricultural soil communities through captured metagenomics. Captured metagenomics uses custom-designed, hybridization-based oligonucleotide probes that enrich functional genes of interest in metagenomic libraries where only probe-bound DNA fragments are sequenced. The captured metagenomes were highly enriched with targeted genes while maintaining their target diversity and their taxonomic distribution correlated well with the traditional ribosomal sequencing. The captured metagenomes were highly enriched with genes related to organic matter degradation; at least five times more than similar, publicly available soil WMG projects. This target enrichment technique also preserves the functional representation of the soils, thereby facilitating comparative metagenomics projects. Here, we present the first study that applies the captured metagenomics approach in large scale, and this novel method allows deep investigations of central ecosystem processes by studying functional gene abundances.

Microbial energy and matter transformation in agricultural soils

“…Total OC (%) plays a strong role in shaping the relative abundance of GH families in situ, as shown by greater richness of GH families involved in cellulose, hemicellulose, lignin and pectin degradation in relatively C rich organic soil layers compared to a greater relative abundance of phenol and protein degradation in C poor mineral soil layers (Cardenas et al, 2015;de Menezes et al, 2015;Uroz et al, 2013). Comparative analysis of a relatively C rich grassland soil showed preferential enrichment of GH families involved in cellobiose and amine degradation while a relatively C poor wheat cropping soil enriched for GH families involved in chitin, b-N-acetylglucosamine and glycoside degradation (Manoharan et al, 2015). Similarly, a comparison of conventional cropping versus low input management of a corn-soybean-winter wheat rotational cropping soil showed that where SOC (%) was significantly lower under conventional management, there was a loss in relative abundances of functional genes involved in the degradation of starch, hemicellulose, cellulose, aromatic C compounds and an endochitinase (Xue et al, 2012).…”

“…Conversely, where TOC (%) does not change between management practices (conventional versus growth of N-fixing legumes during a fallow period) the activity of cellobiohydrolase and b-glucosidase does not change (Bossio et al, 2005). In these examples, GHs could be predominantly associated to the potentially copiotrophic phyla Proteobacteria, Bacteroides and Actinobacteria, reflecting their dominance in agricultural systems (Uroz et al, 2013;de Vries et al, 2015;Manoharan et al, 2015). Thus the composition of the microbial community, the relative abundance of functional genes and the activity of C degrading enzymes closely follow SOM quantity and, presumably, availability.…”

Genomics and metagenomics technologies to recover ribosomal DNA and single-copy genes from old fruit-body and ectomycorrhiza specimens

“…We believe that such low-coverage genomics analyses represent a feasible option to generate multi-gene phylogenomic data sets for tens to hundreds of specimens or mining for the presence and diversity of certain gene families such as carbohydrate active enzymes (CAZymes), antibiotics resistance genes and unique metabolic pathways, but not for routine identification. Targeted enrichment using biotin-linked DNA/RNA probes enables even greater throughput and direct focus on selected markers (Carpenter et al 2013;Moriarty Lemmon and Lemmon 2013;Manoharan et al 2015). The full metagenome data also enable to construct draft genomes of prokaryotes and viruses associated with the fruit-body 'mycosphere' and soil 'mycorrhizosphere' that shed light on putative functions and metabolic pathways of these co-occurring microorganisms.…”

Selection for novel metabolic capabilities inSalmonella enterica

“…; Manoharan, et al. ; Anantharaman et al. ; Locey and Lennon ), where the interconnectedness of different biochemical pathways (Bhattacharya et al.…”

“…Microorganisms exhibit diverse metabolic functions (Barberán et al 2014;Gilbert et al 2014;Manoharan, et al 2015;Anantharaman et al 2016;Locey and Lennon 2016), where the interconnectedness of different biochemical pathways (Bhattacharya et al 2003;Nielsen 2003;Fani and Fondi 2009) as well as secondary (promiscuous) activities of existing enzymes (Copley 2003;Nobeli et al 2009;Khersonsky and Tawfik 2010) further increases an organism's metabolic repertoire. The underlying genetic architecture generating this complexity in biochemical pathways and functions has been well appreciated (Tavazoie et al 1999;Jeong et al 2000;Ravasz et al 2002;Patil and Nielsen 2005;Deutscher et al 2006).…”