Team:MIT/Safety

Safety

Overview

  1. B. subtilis

  2. Sporulation

  3. Probiotic

B. subtilis

JMA222 is the strain of Bacillus subtilis that we are working with, generously provided by the Grossman Lab at MIT. This strain does not contain any integrative and conjugative elements (ICEs), a type of mobile genetic element that transfers between cells via conjugation in some strains of B. subtilis. We initially looked into using a current commercial probiotic strain of B. subtilis, but due to a lengthy approval process to acquire the strain, we decided to proceed using JMA222. The ultimate goal would be to test our system in a probiotic strain of B. subtilis which would be safe to consume and could therefore be made into a probiotic pill or incorporated into other foods such as natto.

Sporulation

B. subtilis is known to produce highly resistant spores that grow under a wide range of conditions, which is advantageous for surviving the acidic stomach environment, but also means released strains may survive outside of containment. In order to prevent this, we took care to abide by appropriate lab safety protocols, including treating E. coli and B. subtilis waste liquid culture with the proper disinfectant, autoclaving plates and solid culture, wearing proper PPE, and keeping surfaces and containers sterile. We also closely communicated with an experienced graduate student mentor in the Grossman Lab.

Probiotic

We intend our bacteria to be ingested as a probiotic and reside in the human gut microbiome. Our engineered bacteria and parts could spread to the environment through the gut, but this is unlikely as our organism potentially have reduced fitness due to constitutive expression of many large protein complexes. In the future, we may look into different kill switches and auxotrophy, potentially de-activating natural BCAA synthesis pathways with the recombinase switch.
Ingestion of a probiotic may also change the composition of the human gut microbiome, which could lead to altered immune and metabolic function. If we were to continue to develop our project, we would need to test for safety and efficacy in clinical trials and comply with USA FDA regulations to conduct these trials. Many biotech companies are working to bring similar probiotics to the market: Synlogic, a Cambridge based synthetic biology biotech company is currently conducting Phase II clinical trials for their phenylketonuria (PKU) probiotic SYNB1618 which is designed to break down Phe in the GI tract. We contacted Dr. Vockley from UPMC Children’s Hospital of Pittsburgh, who specializes in MSUD treatment, to learn more about the process of FDA approval; in order of approval, the age brackets tested are over 18, over 12, over 6, then finally over the age of 2. Thus, we first focused on creating a probiotic for adults, with the long-term aim of expanding its safety to teens and children. Additionally, recent studies (1) of the probiotic effects of probiotic strain Bacillus subtilis DE111 in children 2-6 years of age show that there are subtle but positive effects on the composition of the gut microbiome, demonstrating the safety of B. subtilis in our target demographic.

References

  1. https://www.wageningenacademic.com/doi/10.3920/BM2020.0022