Research
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Elucidation of the survival mechanism of bifidobacteria in the intestine (Fukiya)
One of the fundamental issues in intestinal bacteria research is how each intestinal bacterium survives among the hundreds to thousands of intestinal bacteria present in the individual gut microbiota. We are attempting to elucidate this issue for bifidobacteria, one of the representative intestinal bacteria that are beneficial to human health. To elucidate this issue, it is essential to select genes that are important for survival in the intestine from the approximately 2,000 genes that bifidobacteria possess and to clarify their functions.
Therefore, we have been working on the development of a genetic manipulation system for bifidobacteria, which has been difficult to manipulate, and have established genetic manipulation systems that allow for the construction of gene-deletion mutants and a comprehensive mutagenesis system using transposons. Additionally, we have developed a survivability assessment system in conventionally raised mice. Using these innovative techniques, we have identified key genes that are essential for bifidobacterial survival in the gut. We are now investigating the functions of these genes both in vitro and in vivo, using mice as a model organism. By uncovering the secrets of bifidobacterial survival, we hope to develop new strategies for maintaining a healthy gut microbiota.
Elucidation of the mechanism of secondary bile acid production by intestinal bacteria (Fukiya)
Bile acids are synthesized in the liver from the cholesterol and aid in the digestion and absorption of lipids in the small intestine in humans and animals. Bile acids act on bacterial cell membranes and exert bactericidal activity due to their detergent properties. When bile acids flow into the large intestine, they undergo transformation reactions by intestinal bacteria to produce secondary bile acids.
Our previous research has revealed that deoxycholic acid (DCA), a representative secondary bile acid in humans, exhibits high bactericidal activity among bile acid molecular species and is an important factor in regulating the gut microbiota structure. DCA increases with the intake of a high-fat diet and can cause colorectal cancer and liver cancer. In contrast, it also plays a role in preventing the onset of intestinal infectious diseases and inflammatory bowel diseases, making it an important molecule that exhibits pleiotropic functions in our body.
However, many unknowns exist about the mechanism of DCA production in the intestine. Although identification of the species of bacteria that produce DCA has proceeded, it is expected that in the actual intestine, various intestinal bacteria and DCA-producing bacteria interact with each other, suppressing and activating production, ultimately resulting in DCA production. We aimed to elucidate the whole mechanism of DCA production in the intestine by conducting the following basic research.
- Screening of novel DCA-producing bacteria and their characterization
- Population analysis of DCA-producing bacteria in human feces
- Identification of intestinal bacteria that interact with DCA-producing bacteria and elucidation of their interaction mechanism
Through the basic research, we hope to develop methods to control DCA production in the human intestine and contribute to maintaining health.
Laboratory Evolution of Antimicrobial Resistance in Bacteria (Maeda)
Widespread of antimicrobial resistance (AMR) is a growing concern for global public health because of the depletion of the pool of effective antimicrobial drugs. This evolutionary phenomenon involves pathogens adapting to counteract antibiotics, emphasizing the need for understanding the drug resistance evolution process and developing strategies to impede it. Since the emergence of AMR is based on evolutionary dynamics, quantitative analysis of antibiotic resistance evolution as well as genome analysis of clinical isolates has been extensively performed using representative pathogens. These researches revealed that AMR is complex, involving multiple mutations and significant intracellular changes. To unravel the complexity of the AMR mechanism, the quantitative analysis of phenotypic and genotypic changes faces limitations when using clinical isolates due to the absence of nearest ancestral strains and the presence of numerous neutral mutations. To overcome these limitations, we employ a method known as laboratory evolution, followed by whole-genome sequencing and phenotyping assay.
Our previous study using E. coli revealed that resistance strategies are not specific to each drug but rather involve a limited set of strategies. Moreover, resistance to one drug can influence resistance to unrelated drugs, and conversely, increased sensitivity to one drug may constrain resistance to others. Utilizing such evolutionary constraints in bacterial evolution, we seek novel methods to prevent bacterial drug resistance. Our research extends to various bacteria, including non-pathogenic mycobacteria, contributing to understanding and controlling drug resistance evolution.
Understanding the Principle Rules that determine the structure of microbial ecosystems (Maeda)
Various environments on Earth, such as soil, freshwater, seawater, and the human intestinal tract, harbor complex microbial communities with hundreds to thousands of microbial species coexisting. The advent of high-throughput sequencing has unveiled the community compositions of microbes within these microbial communities.
According to the competitive exclusion principle, also known as Gause’s law, multiple species occupying the same niche cannot stably coexist. Nevertheless, the coexistence of hundreds to thousands of microbes within microbial communities in the same environment remains a fascinating phenomenon. To comprehend the rules governing microbial community assembly, a detailed analysis of the microbial interactions is essential.
In pursuit of a quantitative theory that integrates selective and stochastic ecological processes into a predictive framework, we are intentionally constructing bacterial communities by manipulating and combining specific species under controlled environmental conditions. Through the analysis of microbial interactions and characteristics within the artificially assembled community, along with an exploration of their correlation with environmental factors, our objective is to unveil the foundational principles governing microbial community assembly.
To understand a quantitative theory that integrates selective and stochastic ecological processes into a predictive framework, we are artificially assembling bacterial communities by controlling and combining specific strains under controlled environmental conditions. By analyzing the interactions and characteristics of microbes within the artificially assembled community and investigating their correlation with environmental factors, we are trying to unveil the fundamental principles behind microbial community assembly. Through this foundational research, we aim to develop methodologies for controlling and engineering microbial ecosystems.
Analysis of Central Metabolism and its Application to Bioproduction (Maeda)
With a focus on sustainability and environmental conservation, the necessity to transition from petrochemical production to bioproduction using renewable biomass is increasingly evident. Dr. Atsushi Yokota, a former professor in our laboratory, previously found that inhibiting oxidative phosphorylation, a pivotal process in energy synthesis within Escherichia coli, enhances sugar metabolism. ATP synthase, a critical component for ATP synthesis, relies on proton motive force (PMF). In E. coli, respiratory chain enzymes exhibit multiple isozymes with varying PMF-generating capabilities, finely controlled to maintain a harmonious PMF reflective of external and cellular conditions. Additionally, the regulation of the redox state of NADH and NADPH, crucial for driving redox reactions in sugar metabolism and amino acid synthesis, is of paramount importance.
Nevertheless, the intricate regulatory mechanisms governing these energy and redox levels remain elusive. Currently, using E. coli and Corynebacterium glutamicum as model organisms, we have engineered single and multiple knockout strains targeting enzymes involved in ATP synthesis, as well as the redox state of NADH and NADPH. Through a comprehensive analysis of their effects on metabolism and resulting phenotypic changes, our objective is to optimize energy and redox states for the more efficient bioproduction of valuable chemicals.