Metasecretome-selective phage display approach for mining the functional potential of a rumen microbial community

Ciric, Milica; Moon, Christina D.; Leahy, Sinead C.; Creevey, Christopher J.; Altermann, Eric; Attwood, Graeme T.; Rakonjac, Jasna; Gagić, Dragana

doi:10.1186/1471-2164-15-356

Cited by 15 publications

(14 citation statements)

References 79 publications

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“…Sarkosyl resistance is used as a basis for selection in metasecretome phage display. Single stranded DNA (ssDNA) purified from Sarkosyl R virions after the selection can be used to obtain the amplified metasecretome plasmid library for preliminary assessment of metasecretome diversity by next-generation sequencing [Taken from ( Ciric et al, 2014 ) with permission].…”

Section: Phage Displaymentioning

confidence: 99%

Exploring the Secretomes of Microbes and Microbial Communities Using Filamentous Phage Display

et al. 2016

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Microbial surface and secreted proteins (the secretome) contain a large number of proteins that interact with other microbes, host and/or environment. These proteins are exported by the coordinated activities of the protein secretion machinery present in the cell. A group of bacteriophage, called filamentous phage, have the ability to hijack bacterial protein secretion machinery in order to amplify and assemble via a secretion-like process. This ability has been harnessed in the use of filamentous phage of Escherichia coli in biotechnology applications, including screening large libraries of variants for binding to “bait” of interest, from tissues in vivo to pure proteins or even inorganic substrates. In this review we discuss the roles of secretome proteins in pathogenic and non-pathogenic bacteria and corresponding secretion pathways. We describe the basics of phage display technology and its variants applied to discovery of bacterial proteins that are implicated in colonization of host tissues and pathogenesis, as well as vaccine candidates through filamentous phage display library screening. Secretome selection aided by next-generation sequence analysis was successfully applied for selective display of the secretome at a microbial community scale, the latter revealing the richness of secretome functions of interest and surprising versatility in filamentous phage display of secretome proteins from large number of Gram-negative as well as Gram-positive bacteria and archaea.

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Section: Phage Displaymentioning

confidence: 99%

Exploring the Secretomes of Microbes and Microbial Communities Using Filamentous Phage Display

et al. 2016

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“…The first reports of metagenomic analysis of ruminal digesta demonstrated the power of the technology, focussing in functional terms on fibrolytic enzymes [ 12 – 14 ]. Since then, several further reports have appeared, applying various metagenomics methods again to fibrolytic enzymes [ 15 – 17 ], and to lipases [ 18 , 19 ], virulence and antibiotic resistance genes [ 20 ], polyphenol oxidase [ 21 ], CRISPR elements [ 22 ] and secretome proteins [ 23 ].…”

Section: Introductionmentioning

confidence: 99%

The rumen microbial metagenome associated with high methane production in cattle

et al. 2015

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BackgroundMethane represents 16 % of total anthropogenic greenhouse gas emissions. It has been estimated that ruminant livestock produce ca. 29 % of this methane. As individual animals produce consistently different quantities of methane, understanding the basis for these differences may lead to new opportunities for mitigating ruminal methane emissions. Metagenomics is a powerful new tool for understanding the composition and function of complex microbial communities. Here we have applied metagenomics to the rumen microbial community to identify differences in the microbiota and metagenome that lead to high- and low-methane-emitting cattle phenotypes.MethodsFour pairs of beef cattle were selected for extreme high and low methane emissions from 72 animals, matched for breed (Aberdeen-Angus or Limousin cross) and diet (high or medium concentrate). Community analysis was carried out by qPCR of 16S and 18S rRNA genes and by alignment of Illumina HiSeq reads to the GREENGENES database. Total genomic reads were aligned to the KEGG genes databasefor functional analysis.ResultsDeep sequencing produced on average 11.3 Gb per sample. 16S rRNA gene abundances indicated that archaea, predominantly Methanobrevibacter, were 2.5× more numerous (P = 0.026) in high emitters, whereas among bacteria Proteobacteria, predominantly Succinivibrionaceae, were 4-fold less abundant (2.7 vs. 11.2 %; P = 0.002). KEGG analysis revealed that archaeal genes leading directly or indirectly to methane production were 2.7-fold more abundant in high emitters. Genes less abundant in high emitters included acetate kinase, electron transport complex proteins RnfC and RnfD and glucose-6-phosphate isomerase. Sequence data were assembled de novo and over 1.5 million proteins were annotated on the subsequent metagenome scaffolds. Less than half of the predicted genes matched matched a domain within Pfam. Amongst 2774 identified proteins of the 20 KEGG orthologues that correlated with methane emissions, only 16 showed 100 % identity with a publicly available protein sequence.ConclusionsThe abundance of archaeal genes in ruminal digesta correlated strongly with differing methane emissions from individual animals, a finding useful for genetic screening purposes. Lower emissions were accompanied by higher Succinovibrionaceae abundance and changes in acetate and hydrogen production leading to less methanogenesis, as similarly postulated for Australian macropods. Large numbers of predicted protein sequences differed between high- and low-methane-emitting cattle. Ninety-nine percent were unknown, indicating a fertile area for future exploitation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2032-0) contains supplementary material, which is available to authorized users.

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“…Ciric et al . 2014 used a similar technology for the enrichment for DNA inserts that contained an endogenous signal peptide and analyzed this secretome enriched library by next generation sequencing, but this approach did not include any functional screening 52 53 . Here, we present an approach that allows to identify immunogenic proteins from ORF-enriched phage display libraries based on specific protein interactions including functional selection from proteins of a metaproteome independent of cultivation, expression level or antigen size.…”

Section: Discussionmentioning

confidence: 99%

Mining gut microbiome oligopeptides by functional metaproteome display

Zantow

Just

Lagkouvardos

et al. 2016

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Pathogen infections, autoimmune diseases, and chronic inflammatory disorders are associated with systemic antibody responses from the host immune system. Disease-specific antibodies can be important serum biomarkers, but the identification of antigens associated with specific immune reactions is challenging, in particular if complex communities of microorganisms are involved in the disease progression. Despite promising new diagnostic opportunities, the discovery of these serological markers becomes more difficult with increasing complexity of microbial communities. In the present work, we used a metagenomic M13 phage display approach to select immunogenic oligopeptides from the gut microbiome of transgenic mice suffering from chronic ileitis. We constructed three individual metaproteome phage display libraries with a library size of approximately 107 clones each. Using serum antibodies, we selected and validated three oligopeptides that induced specific antibody responses in the mouse model. This proof-of-concept study provides the first successful application of functional metaproteome display for the study of protein-protein interactions and the discovery of potential disease biomarkers.

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Metasecretome-selective phage display approach for mining the functional potential of a rumen microbial community

Cited by 15 publications

References 79 publications

Exploring the Secretomes of Microbes and Microbial Communities Using Filamentous Phage Display

Exploring the Secretomes of Microbes and Microbial Communities Using Filamentous Phage Display

The rumen microbial metagenome associated with high methane production in cattle

Mining gut microbiome oligopeptides by functional metaproteome display

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