Insights into the Human Virome Using CRISPR Spacers from Microbiomes

Abstract

Due to recent advances in next-generation sequencing over the past decade, our understanding of the human microbiome and its relationship to health and disease has increased dramatically. Yet, our insights into the human virome, and its interplay with important microbes that impact human health, is relatively limited. Prokaryotic and eukaryotic viruses are present throughout the human body, comprising a large and diverse population which influences several niches and impacts our health at various body sites. The presence of prokaryotic viruses like phages, has been documented at many different body sites, with the human gut being the richest ecological niche. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and associated proteins constitute the adaptive immune system of bacteria, which prevents attack by invasive nucleic acid. CRISPR-Cas systems function by uptake and integration of foreign genetic element sequences into the CRISPR array, which constitutes a genomic archive of iterative vaccination events. Consequently, CRISPR spacers can be investigated to reconstruct interplay between viruses and bacteria, and metagenomic sequencing data can be exploited to provide insights into host-phage interactions within a niche. Here, we show how the CRISPR spacer content of commensal and pathogenic bacteria can be used to determine the evidence of their phage exposure. This framework opens new opportunities for investigating host-virus dynamics in metagenomic data, and highlights the need to dedicate more efforts for virome sampling and sequencing.

Keywords: CRISPR spacers; CRISPR-Cas systems; microbiome; phages; virome.

Figure 1

Human microbiome and virome. Different environments showing commensal and pathogenic bacteria that carry Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas immune systems as defense mechanisms against invasive nucleic acids from phages and plasmids (left panel). Distribution of the human microbiome in the most representative body sites, including some of the species that contain CRISPR-Cas immune systems (right panel) (adapted from Lloyd-Price and co-workers [23,24]).

Figure 2

CRISPR-Cas systems: Mode of action. CRISPR-Cas are DNA-encoded, RNA-mediated nucleic acid targeting systems that operate via a three step process. The first step is acquisition, where Cas1 and Cas2 proteins copy and paste a short sequence of the invasive DNA which is added as a new spacer at the 5′ end of the CRISPR array, close to the leader region. During expression, the second step, the CRISPR array is transcribed to generate the mature CRISPR RNAs (crRNA), which are the repeat-spacer pairs, and the trans-activated CRISPR RNA (tracrRNA). Finally, the interference stage is mediated by the duplex crRNA:tracrRNA that drives the Cas9 protein towards the complementary sequence of the spacer in the invasive nucleic acid to bind and cleave DNA after PAM recognition.

Figure 3

Phage-bacteria dynamics. Interaction and evolution of phage and bacteria developing next generation defense mechanisms, and escape strategies. Bacteria have developed CRISPR-Cas immune systems, among others, that defend against predatory attack from phages whereas the phages also evolve (i) mutating their genome to escape CRISPR targeting; (ii) introducing mutations into CRISPR systems or (iii) developing anti-CRISPR proteins to inactivate the Cas proteins.