CRISPR/Cas9 technology has emerged as a revolutionary tool in the field of genetics and biotechnology, allowing for precise and targeted editing of the genome of various organisms, including bacteria. This technology has been widely used in a variety of applications, such as the development of new medicines, improvements in agriculture, and the production of biofuels. In this article, we will explore a multitude of examples of how CRISPR/Cas9 technology has been used to edit the genome of bacteria. This showcase demonstrates the versatility and potential of this technology, highlighting its impact in various fields. Whether it’s for creating bacteria with desirable metabolic activities, correcting genetic mutations, or developing new methods for biofuel production, CRISPR/Cas9 technology has proven to be an indispensable tool in modern biotechnology.

Research:

• Re-sensitisation of antibiotic-resistant bacteria:

With the help of CRISPR/Cas9 technology, researchers are able to target and modify bacterial genes responsible for resistance to one or more antibiotics. Jun-Seob Kim et. al. were successful in re-sensitising antibiotic-resistant E. coli that harboured extended-spectrum β-Lactamases. Ref1

• Research and Development:

One area where CRISPR/Cas9 is making a significant impact is the development of new research tools for studying bacterial biology and the molecular mechanisms underlying bacterial pathogenesis. By using CRISPR/Cas9 technology, scientists can introduce specific mutations into the genome of bacteria, allowing them to study the effects of these mutations on bacterial growth and behaviour. This can provide valuable insights into the molecular mechanisms underlying bacterial pathogenesis, and help researchers to identify new targets for the development of antibacterial therapies. Ref2

Food & Agriculture:

• Enhancement of plant resistance to pests and disease:

Agrobacterium is a type of bacteria used to deliver DNA into plants. It can also be used to deliver CRISPR/Cas9 into plant cells and target the modification of the plant’s genome. The bacteria serve as cost-effective and efficient vectors for delivering disease and pest resistance traits to crops. The ability to enhance plant resistance through use of CRISPR/Cas9 represents a significant advancement in the field of agriculture and has the potential to have a profound impact on food security and sustainability. Ref 3&4

• Development of new biopesticides:

By modifying the genes in bacteria that control pest resistance, CRISPR/Cas9 can be used to develop new biopesticides that are safe and effective alternatives to traditional chemical pesticides. (Pazmiño-Ibarra et al, 2019)

• Improving food preservation:

Through use of CRISPR/Cas9 to modify the genes in bacteria that control spoilage and spoilage resistance, preservation of food products can be improved.

Therapeutics & Diagnostics:

• Development of new diagnostic tools:

CRISPR/Cas9 can be used to develop new diagnostic tools that can detect the presence of specific bacterial and viral pathogens. Ref6

• Development of new vaccines & immunotherapies:

Immunotherapy approaches to treating human diseases have grown immensely over the past 10 years. One potential application of CRISPR/Cas9 in immunotherapy is to engineer bacteria to produce proteins or molecules that can activate the immune system to recognize and attack cancer cells. For example, researchers have used CRISPR/Cas9 to modify the genome of bacteria to produce proteins that activate T-cells, which play a critical role in fighting cancer.

Another approach is to use CRISPR/Cas9 to modify the genome of bacteria to produce molecules that can inhibit or suppress the immune system. This approach could be used to treat autoimmune diseases, where the immune system attacks healthy cells in the body. By modifying the genes in bacteria associated with the human immune system, it may be possible to develop new immunotherapies that can selectively target and regulate the immune response.

• Development of new cancer treatments:

CRISPR/Cas9 can be used to develop new cancer treatments, such as bacteria-based cancer vaccines. These vaccines use live bacteria to stimulate the immune system to recognize and attack cancer cells. By modifying the genes in these bacteria, researchers can increase their ability to trigger an immune response against cancer cells. (Abedi et al., 2022)

• Modification of the oral microbiome:

CRISPR/Cas9 can be used to prevent tooth decay, gum disease, and oral cancer. The oral microbiome is a complex ecosystem of microorganisms that inhabit the mouth, including bacteria, fungi, and viruses. These microorganisms play a crucial role in maintaining oral health, but imbalances in the microbiome can lead to a range of oral diseases, including tooth decay, gum disease, and oral cancer. CRISPR/Cas9 gene editing technology has the potential to modify the genes in oral bacteria to prevent and treat these diseases.

Tooth decay is caused by the production of acid by bacteria in the mouth that erodes the enamel on the teeth. By modifying the genes in these bacteria, researchers can reduce their ability to produce acid, thereby preventing tooth decay. For example, researchers have used CRISPR/Cas9 to modify the genome of the bacterium Streptococcus mutans, which is a major contributor to tooth decay, to prevent it from producing acid. (Gong T et al., 2018)

• Modification of the gut microbiome:

By modifying the genes in gut bacteria, CRISPR/Cas9 can be used to improve gut health and prevent diseases such as inflammatory bowel disease, obesity, and colon cancer.

• Development of new anti-inflammatory treatments:

By modifying the genes in bacteria that are associated with inflammation, CRISPR/Cas9 can be used to develop new anti-inflammatory treatments for conditions such as rheumatoid arthritis and psoriasis.

• Development of new probiotics:

By modifying the genes in bacteria that are associated with beneficial properties, CRISPR/Cas9 can be used to develop new probiotics with improved health benefits.

The Environment:

• Bioremediation:

By modifying the genes in bacteria that control the degradation of specific pollutants, CRISPR/Cas9 can be used to improve bioremediation technologies for cleaning up contaminated environments. Ref 7

• Production of biodegradable plastics:

By modifying the genes in bacteria that interact with certain plastic polymers, CRISPR/Cas9 can be used to improve the efficiency of plastic manufacturing and recycling and reduce environmental waste. Si Liue et al were able to synthesise the biodegradable polyester polyhydroxyalkanaotes in Pseudomonas putida using CRISPR/Cas9. Ref 8

Synthetic Biology:

• Biofuel production:

By modifying the genes that control metabolic pathways in bacteria, CRISPR/Cas9 can be used to increase the efficiency of biofuel production. Ref 9 & 10

• Fine chemical production:

By editing specific genes in bacteria, CRISPR/Cas9 can be used to improve the production of fine chemicals, such as enzymes, proteins and other products used in the pharmaceutical and food industries. Ref 11

Concluding Remarks:

CRISPR/Cas9 technology has proven to be a powerful tool in the field of genetics and biotechnology, particularly in the editing of the genome of bacteria. The examples explored in this article demonstrate the versatility and potential of this technology, highlighting its impact in various fields such as medicine, agriculture, and biotechnology. As research and development in CRISPR/Cas9 technology continues to advance, it is likely that even more innovative and impactful applications will emerge, further advancing our understanding of genetics and the potential to treat and cure diseases. The use of CRISPR/Cas9 technology in the editing of the genome of bacteria is a testament to the progress that can be achieved through collaboration and innovation in science and technology. ERS Genomics has partnered with several industrial partners to enable them to manipulate bacteria with CRISPR/Cas9 and commercialize their innovative applications.

 

References:

Abedi, M.H., Yao, M.S., Mittelstein, D.R. et al. Ultrasound-controllable engineered bacteria for cancer immunotherapy. Nat Commun 13, 1585 (2022). https://doi.org/10.1038/s41467-022-29065-2

Chowdhury, S., Castro, S., Coker, C. et al. Programmable bacteria induce durable tumor regression and systemic antitumor immunity. Nat Med 25, 1057–1063 (2019). https://doi.org/10.1038/s41591-019-0498-z

Gong T, Tang B, Zhou X, Zeng J, Lu M, Guo X, Peng X, Lei L, Gong B, Li Y. Genome editing in Streptococcus mutans through self-targeting CRISPR arrays. Mol Oral Microbiol. 2018 Dec;33(6):440-449. doi: 10.1111/omi.12247. Epub 2018 Oct 24. PMID: 30329221.

Hille Frank and Charpentier Emmanuelle, 2016 CRISPR-Cas: biology, mechanisms and relevancePhil. Trans. R. Soc. B3712015049620150496http://doi.org/10.1098/rstb.2015.0496

Kim JS, Cho DH, Park M, Chung WJ, Shin D, Ko KS, Kweon DH. CRISPR/Cas9-Mediated Re-Sensitization of Antibiotic-Resistant Escherichia coli Harboring Extended-Spectrum β-Lactamases. J Microbiol Biotechnol. 2016 Feb;26(2):394-401. doi: 10.4014/jmb.1508.08080. PMID: 26502735.

Mimee, M., Tucker, A. C., Voigt, C. A., & Lu, T. K. (2015). Programming a human commensal bacterium, Bacteroides thetaiotaomicron, to sense and respond to stimuli in the murine gut microbiota. Cell Systems, 1(1), 62-71. https://doi.org/10.1016/j.cels.2015.06.001

Nina Gouba, Yeri Esther Hien, Marie Laure Guissou, Maxime Descartes Mbogning Fonkou, Yves Traoré, Zekiba Tarnagda,

Digestive tract mycobiota and microbiota and the effects on the immune system,

Human Microbiome Journal, Volume 12, 2019,100056,ISSN 2452-2317, https://doi.org/10.1016/j.humic.2019.100056.

Pazmiño-Ibarra V, Mengual-Martí A, Targovnik AM, Herrero S. Improvement of baculovirus as protein expression vector and as biopesticide by CRISPR/Cas9 editing. Biotechnol Bioeng. 2019 Nov;116(11):2823-2833. doi: 10.1002/bit.27139. Epub 2019 Sep 2. PMID: 31403180.

Wenzhi Jiang, Huanbin Zhou, Honghao Bi, Michael Fromm, Bing Yang, Donald P. Weeks, Demonstration of CRISPR/Cas9/sgRNA-mediated targeted gene modification in Arabidopsis, tobacco, sorghum and rice, Nucleic Acids Research, Volume 41, Issue 20, 1 November 2013, Page e188, https://doi.org/10.1093/nar/gkt780