Rodolphe Barrangou
Professor
Todd R. Klaenhammer Distinguished Professor
Partners Building II 2300
Bio
Area(s) of Expertise
Our laboratory focuses on the biology and genetics of CRISPR-Cas immune systems in bacteria. Using microbiology, molecular biology and genomics approaches, we investigate the use of CRISPR-Cas systems for three types of applications:
- The exploitation of CRISPR spacer hypervariability for genotyping and phylogenetic studies of beneficial and pathogenic bacteria
- Leveraging CRISPR-mediated interference for building up phage resistance in probiotic strains and starter cultures used in food manufacturing
- Harnessing of Cas9-mediated, re-programmable dsDNA cleavage for genome editing in bacteria
These activities provide insights into the genetic and molecular processes that drive CRISPR-mediated adaptive immunity in bacteria and generate novel tools for the manipulation of industrially relevant organisms for food and biotechnological applications.
Teaching Activities
- Introduction to Biopharmaceutical Sciences (BBS 201)
- MR Supervised Research (FS 693)
- Fermentation Microbiology (FS 725)
- DR Supervised Research (FS 893)
Publications
- Lactobacillus acidophilus Expressing Murine Rotavirus VP8 and Mucosal Adjuvants Induce Virus-Specific Immune Responses , VACCINES (2023)
- Bile salt hydrolases shape the bile acid landscape and restrict Clostridioides difficile growth in the murine gut , NATURE MICROBIOLOGY (2023)
- CRISPR Technology and its Many Applications with Select Examples Related to Animal Agriculture , JOURNAL OF ANIMAL SCIENCE (2023)
- CasPEDIA Database: a functional classification system for class 2 CRISPR-Cas enzymes , NUCLEIC ACIDS RESEARCH (2023)
- Genome-Wide Comparative Analysis of Lactiplantibacillus pentosus Isolates Autochthonous to Cucumber Fermentation Reveals Subclades of Divergent Ancestry , FOODS (2023)
- Impact of Pomegranate on Probiotic Growth, Viability, Transcriptome and Metabolism , MICROORGANISMS (2023)
- Mining microbial organisms to discover and characterize novel CRISPR-Cas systems , CURRENT OPINION IN BIOMEDICAL ENGINEERING (2023)
- Multiplex CRISPR editing of wood for sustainable fiber production , SCIENCE (2023)
- The Expanding Dissemination and Distribution Patterns of Diverse CRISPR Plasmids by Addgene , CRISPR JOURNAL (2023)
- Broad-spectrum CRISPR-Cas13a enables efficient phage genome editing , NATURE MICROBIOLOGY (2022)
Grants
The m-CAFEs SFA (Microbial Community Analysis and Functional Evaluation in Soils Science Focus Area) is a multi-institutional project that will advance our understanding of the mechanisms and interactions or rhizosphere microbiomes governing nutrient cycling. This will be achieved by developing powerful new in situ community manipulation capabilities to test model predictions and establish causal mechanisms. Our work builds upon a two-year pilot effort that has developed fabricated ecosystems (EcoFABs) enabling reproducible plant growth and colonization within these controlled environments designed for systems biology, imaging, and community manipulation technologies. We have used genetic manipulation approaches within the EcoFABs to understand the molecular mechanisms underpinning these processes and provide much needed insights into the ~50% of microbial genes of unknown function. In this three-year SFA period, we will rapidly develop novel CRISPR-Cas and phage-based rhizosphere microbiome editing technologies to investigate uncultivated microbial activities and interactions within controlled lab environments. We will use both defined microbial assemblies and native soil-derived enriched microbial communities to further develop and apply this powerful approach to dissecting functions of plant-rhizosphere microbiomes for eventual extension to fully-complex communities. The defined microbial assemblies will enable detailed characterization of constituent isolates whereas the enrichments of native microbial communities will enable investigation of community functions of uncultivated microbes. Given the vast number of unknown microbial interactions and gene functions within these communities, we will use powerful modeling approaches combined with our EcoFAB data to simulate interventions to prioritize targeted manipulations that provide new insights into activities and interactions of microbes including uncultivated species. Ultimately, the new experimental tools and detailed understanding organism interactions obtained by the m-CAFEs team will enable the predictable control of rhizosphere microbiomes, with important implications for sustainable energy production and environmental health.
This study proposes to continue to investigate Lactobacillus species for probiotic properties using microbiological and genetic approaches. The genome sequence information of two significant probiotic species, Lactobacillus acidophilus and Lactobacillus gasseri, will be the primary model organisms for this study. Other lactobacilli will also be investigated when appropriate. The impact of environmental signals encountered in foods and the gastrointestinal tract (food-dairy components, acid, bile, heat, cold, oxidative stresses, etc) will be evaluated using whole genome microarrays, for these model organisms and for other lactobacilli for which genome information is available. Discovery and confirmation of genetic traits that direct beneficial activities and outcomes of probiotic Lactobacillus species is expected to promote the use of effective cultures in a variety of functional food, and dairy food scenarios and, thereby, sustain future expansion of this category.
Diarrheal disease is the second leading cause of death in children under the age of 5 worldwide with rotavirus responsible for 40 percent of hospitalizations due to diarrheal illness1. It is estimated that rotavirus killed approximately 215,000 children in 2013. The World Health Organization recommends including a rotavirus vaccine in all global vaccination protocols and there are currently two vaccines licensed worldwide2. The global implementation is ongoing but in countries where data is available, vaccination has resulted in a 33% reduction in hospitalization due to rotavirus morbidities. Unfortunately, both vaccines have limited efficacy (50-60%) in developing countries and are associated with a low level risk of intussusception3. Next generation vaccines are under development and will need to address several key concerns and limitations inherent with the current modified-live vaccines. IgA has been shown to be important in protection against rotaviral infection in humans and in animal models4-9. An orally delivered vaccine that induces protective mucosal and systemic antibody responses would be ideal for use in the campaign against rotavirus. Induction of IgA through systemic immunization has proven to be difficult. The mucosal immune system is, in many respects, independent of the systemic immune system. For example, ninety percent of intestinal IgA is produced locally and induction of mucosal immunity is best achieved via mucosal infection or vaccination10-12. Commensal organisms of the intestinal tract have evolved to cope with the hostile environment presented by the gastrointestinal tract and some commensals, now considered probiotics, have been shown to enhance health via beneficial interaction with the mucosal immune system13,14. Appropriately selected commensal organisms could be powerful vectors for the delivery of therapeutics and vaccines. We have developed an orally-delivered mucosal vaccine platform that employs the commensal bacteria Lactobacillus acidophilus (LA). We have brought together several adjuvant and antigen-expression strategies in unique constructs that show great potential to induce mucosal and systemic antibody responses. This platform offers several important feasibility advantages as it employs a commensal designated as GRAS (generally regarded as safe) by the FDA, is inexpensive to produce, does not require cold-chain, and is needleless. Investigation of immunodominant epitopes from the rotavirus envelope have identified two regions, a 4 amino acid fragment of the VP8 trypsin cleavage fragment of VP4 and a 14 amino acid fragment corresponding to AA242-259 from VP6, that induce protective IgA15-17. In the proposed studies, we will utilize CRISPR-Cas to engineer a state-of-the-art vaccine construct and then determine the immunogenicity and efficacy of this novel vaccine in a Balb/c mouse model. We will explore correlates of protection and determine the influence of the interaction between vaccine, host, and intestinal microbiome on immunization and challenge outcomes.
The m-CAFEs SFA (Microbial Community Analysis and Functional Evaluation in Soils Science Focus Area) is a multi-institutional project that will advance our understanding of the mechanisms and interactions or rhizosphere microbiomes governing nutrient cycling. This will be achieved by developing powerful new in situ community manipulation capabilities to test model predictions and establish causal mechanisms. Our work builds upon a two-year pilot effort that has developed fabricated ecosystems (EcoFABs) enabling reproducible plant growth and colonization within these controlled environments designed for systems biology, imaging, and community manipulation technologies. We have used genetic manipulation approaches within the EcoFABs to understand the molecular mechanisms underpinning these processes and provide much needed insights into the ~50% of microbial genes of unknown function. In this three-year SFA period, we will rapidly develop novel CRISPR-Cas and phage-based rhizosphere microbiome editing technologies to investigate uncultivated microbial activities and interactions within controlled lab environments. We will use both defined microbial assemblies and native soil-derived enriched microbial communities to further develop and apply this powerful approach to dissecting functions of plant-rhizosphere microbiomes for eventual extension to fully-complex communities. The defined microbial assemblies will enable detailed characterization of constituent isolates whereas the enrichments of native microbial communities will enable investigation of community functions of uncultivated microbes. Given the vast number of unknown microbial interactions and gene functions within these communities, we will use powerful modeling approaches combined with our EcoFAB data to simulate interventions to prioritize targeted manipulations that provide new insights into activities and interactions of microbes including uncultivated species. Ultimately, the new experimental tools and detailed understanding organism interactions obtained by the m-CAFEs team will enable the predictable control of rhizosphere microbiomes, with important implications for sustainable energy production and environmental health.
CRISPR-Cas systems are revolutionary technologies that enable precise genome editing in a broad variety of organisms, encompassing humans, animals, plants and bacteria. Here, we focus on developing novel enzymes and proteins for use in altering the human genome to treat diseases via gene therapies.
Nitrogenous fertilizers are critical for sustaining small to large farms in the US. The Haber-Bosch process generates the majority of fixed nitrogen, but it comes at a high cost, both in terms of dollars and environmental impact. Requiring temperatures between 400-500oC and pressures of 150-250 bar, this process consumes 1-2% of global energy. Reliance on fossil fuels to power this process translates into unstable fertilizer prices and a significant release of greenhouse gases. Low-cost and carbon-neutral ammonia fertilizer production is therefore needed to improve the sustainability of our food production systems. Biological nitrogen (N2) fixation, as practiced in the farming of legumes, is attractive because of its low-energy demand, operation under ambient conditions, and point-of-use production; however, slow fixation rates and, in the case of non-legume crops, a lack of abundant N2 fixing symbiotic diazotrophs in the soil, limit the large-scale feasibility of this approach. Moreover, options to accelerate symbiotic N2 fixation rates to the industrial levels needed to compete with the Haber-Bosch process are lacking. As an alternative, we propose investigating a hybrid microbial electrochemical system to electrically enhance microbial N2 fixation rates. Bacteria in these systems consume organic matter (such as waste biomass) and generate electrical current when they respire (breathe) on anode electrodes. By exploiting their physiology, we hypothesize that we can electrically ????????????????boost??????????????? N2 fixation rates in these organisms. The overall objective of this proposal is to determine the influence of the electrical driving force on the rates, mechanisms, and pathways of microbial N2 fixation. The rationale is that with this knowledge, we can improve N2 fixation rates in these communities and optimize a scalable technological platform to produce fixed nitrogen from small-scale farms to industrial-scale applications.
CRISPR-Cas systems are revolutionary technologies that enable precise genome editing in a broad variety of organisms, encompassing humans, animals, plants and bacteria. Here, we focus on developing novel enzymes and proteins for use in altering the human genome to treat diseases via gene therapies.
The primary objective of the project is to screen for a natural LTA- variant in Lactobacillus acidophilus NCFM, as to find a natural equivalent of NCK2025. As published, the NCK 2025 strain is a variant of NCFM in which the LTA gene was deleted as to positively impact the host (human) immune system to generate a strain with valuable anti-inflammatory functional properties.
The soil microbiome is a complex network that plays a critical role in determining the fate of carbon (C) within soils. An overarching science challenge is to understand the molecular mechanisms controlling the forms and flow of C through the soil microbiome in response to environmental perturbations. Here, we focus on determining microbial functional roles at hot spots and hot moments during rhizosphere development and punctuated water events that account for a significant percentage of ecosystem carbon transformation and respiration. We will construct ecosystem fabrications (EcoFABs) to recapitulate microbial networks based on studies conducted at the UC Angelo Coastal Reserve, where extensive genomic, proteomic and metabolomic data have provided insights regarding the partitioning of organisms and metabolic capacity with soil depth and changing water regimes. Our EcoFABs are designed to reproduce key physical, chemical, and biological features of soils and can be integrated with systems biology and high throughput genetics approaches, enabling ???????????????bottom-up?????????????????? investigation of microbial networks. To discover causal microbial mechanisms associated with punctuated water events and carbon flow in microbial communities, we will pioneer break-through genetic technologies that rapidly classify proteins of unknown function and interrogate the ecological function of microbial communities. Critically, we will simultaneously develop CRISPR-Cas and RNAi community editing technologies, enabling top-down interrogation of microbe function within intact communities. These approaches will provide foundational systems for developing predictive understanding of the biochemical ecology and the role of C flow in soil microbiomes in response to environmental perturbations.
CRISPR-based genome editing is a disruptive technology enabling the rapid and flexible alteration of virtually any genome in any organism in a programmable manner. Select organisms of industrial interest are used as manufacturing workhorses for the production of industrial enzymes. Several Bacillus species in particular are used to manufacture diverse enzymes by Novozymes. Here, we will assess the occurrence and potential of endogenous CRISPR-Cas systems in select industrial Bacillus species and test the ability of endogenous or exogenous systems to trigger genome editing.