Report

Teaser, summary, work performed and final results

Periodic Reporting for period 2 - RuMicroPlas (The Plasmidome: a Driving Force of Rumen Microbial Evolution from Birth to Adulthood)

Teaser

The goal of my project is to understand the community structure of the microbiome, its drivers and the role(s) of mobile genetic elements (plasmids) within it.In recent years, the mammalian gut, including the ruminant gut, has emerged as a fundamentally important microbial...

Summary

The goal of my project is to understand the community structure of the microbiome, its drivers and the role(s) of mobile genetic elements (plasmids) within it.
In recent years, the mammalian gut, including the ruminant gut, has emerged as a fundamentally important microbial environment. The intricate relationships between mammalian hosts and their microbial communities have been shown to play a central role in the host\'s well-being The rumen environment is an anaerobic compartment in the ruminant digestive system that accommodates heterogeneous microbial communities. This complex microbiome is comprised of Protozoa, Archaea and Bacteria co-residing at a density greater than 1010 ml-1. The rumen, together with its microbial symbionts, is responsible for the ruminant\'s remarkable ability to convert indigestible plant mass into digestible food products. These microorganisms are entirely responsible for the degradation and fermentation of the plant material, consisting mainly of indigestible sugar polymers such as cellulose and hemicellulose, consequently enabling the conversion of plant fibers into chemical compounds that can be digested by the animal. In this sense, ruminants are completely dependent on their rumen microbiome for their existence. This cooperative relationship between the ruminant and its resident microbiome has evolved over millions of years and has implications for our everyday lives with respect to food sustainability, environment, renewable energy, and economics. Ruminants hold enormous significance for man, as they convert the energy stored in plant biomass polymers, which are indigestible for humans, to digestible food products. Humans domesticated these animals for this purpose in the Neolithic era and have been farming them ever since for the production and consumption of animal protein in the form of meat and milk. In today\'s extensive production regimes, ruminants consume 30% of the crops grown on earth and occupy another 30% of the earth\'s land mass. These animals also emit methane—a highly potent greenhouse gas—to the atmosphere and are considered to be responsible for a considerable portion of its emission because of anthropogenic activities. Hence, an understanding of this complex microbial ecosystem and the evolutionary rules that govern it is of major interest. The complexity of the rumen microbial environment and its key role in animal physiology raises intriguing questions regarding the genetics and mobility of its functions among its microbial members. Lateral gene transfer (LGT)—the process by which microbial species donate and receive genetic material—is a major determinant of genetic novelty and genome evolution in prokaryotes. Mobile genetic elements serve as DNA vehicles for the communal gene pool. Plasmids are self-replicating, extrachromosomal, mobile genetic elements that operate as “gene ferries”, transferring genes from one host to another. Plasmids have been recognized as key vectors of genetic exchange between microbial chromosomes. Their high abundance in microbial populations sampled from various habitats indicates that they have an important ecological role. Plasmids are composed of a conserved DNA backbone that includes replication and mobilization genes, which are important for plasmid maintenance within the host and transfer among hosts. They also carry a variable assortment of accessory genes, which often contribute to the phenotypic diversity of their host. Plasmids isolated from different ecological niches encode a versatile array of accessory functions, ranging from antibiotic resistance to nitrogen fixation. These plasmid-borne functions may confer an advantage to their host in its niche, making the burden of carrying the plasmid worthwhile. An understanding of plasmid biology and biodiversity is expected to greatly contribute to our understanding of microbial ecology and evolution in diverse environments.
An ecological and mechanistic understanding of the r

Work performed

The main results achieved so far showed several fundamental and global aspects connected to the functioning of the rumen ecosystem, host-microbiome interactions , its plasmid composition and key components such as individual microbes and metabolites.
1. Ecosystem functions and composition
We identified several fundamental aspects that are connected to the functioning of the rumen ecosystem (publications 8, 9, 10), providing support to findings from microbial communities worldwide. Within the rumen ecosystem, we explored the microbial interactions as a response to diet and feeding cycle (fibre degradation and other extracellular matrices).
Using dietary intervention experiments, we revealed that diet affects the most abundant taxa within the microbiome and that a specific group of methanogenic archaea of the order Methanomicrobiales is highly sensitive to changes. Using metabolomic analyses together with in vitro microbiology approaches and whole-genome sequencing of Methanomicrobium mobile, a key species within this group, we identified that redox potential changes with diet, is the main factor that causes these dietary-induced alternations in this taxon’s abundance. Our genomic analysis suggests that the redox potential effect stems from a reduced number of anti-reactive oxygen species proteins coded in this taxon\'s genome. Our study highlighted redox potential as a pivotal factor that could serve as a sculpturing force of community assembly within anaerobic gut microbial communities (publication 8). Moreover, we investigated the metabolic potential and taxonomic composition of the rumen methanogens that play a key role in sustaining host metabolism and function. We discovered that the methanogenesis process changes with age and that the early methanogenic community is characterized by a high activity of methylotrophic methanogenesis. These findings were highlighted by science journal as they suggest that environmental filtering acts on the archaeal communities and select for different methanogenic lineages during different growth stages, affecting the functionality of this ecosystem (publication 9).
We next studied the postprandial diurnal community oscillatory patterns of the rumen microbiome, in order to understand what affects the community composition during the feeding cycle. We showed that metabolites produced by the rumen microbiome serve to condition its environment and lead to dramatic diurnal changes in community composition and function. These changes in community composition were accompanied by changes in pH and methane partial pressure, suggesting a strong functional connection. Our experiments showed that the metabolites released by microbes are sufficient to reproduce changes in community function comparable to those observed in vivo. These findings highlighted microbiome niche modification as a deterministic process that drives diurnal community assembly via environmental filtering (publication 10)

2. Host-microbiome axis
In order to understand the associations between the rumen host and its essential microbiome, we aimed to link the rumen microbial components to the cow’s ability to extract energy from their feed, termed as feed efficiency (publications 1, 3, 4). We discovered that rumen microbiome components are tightly linked to the cows\' feed efficiency as well as methane emission. This work is seminal and of high importance in the field and has high relevance to our every life therefore is also highly cited (publication 1). Moreover, we further aimed to understand the role of rumen genetics on the microbiome composition. We found that genetics and physiology are correlated with microbiome structure and that host genetics may shape the microbiome landscape by enriching for phylogenetically related taxa (publication 3). We found that the gut microbiomes of hosts that were genetically selected are different ones that were not selected, although subjected to the same environmental conditions. More

Final results

We have developed two novel methodologies that enables us to detect plasmids in their natural environments – the Recycler - as well as a new FISH (Fluorescence In-Situ Hybridization) method, combining single molecule FISH (smFISH), used to target mRNA molecules with single fluorophores per probe, together with Catalyzed Reporter Deposition FISH (CARD FISH), which amplifies fluorescent signals. Moreover, we have developed a method to predict the microbial community temporal dynamics based on the community composition at previous time stamps (MTV-LMM).