IBD is very complex class of diseases and given that some bacterial species might worsen the condition for patients, it is of the utmost importance to take the necessary precautions to make our bacteria interact as little as possible with the disease. In the following paragraphs we will summarize the interview with Prof. Rogler, where we address these issues. The full transcript in German can be downloaded here:
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Our project is currently implemented in E. coli, however E. coli lipopolysaccharides are known to increase the immune reaction in IBD patients and are sometimes even connected to pathogenesis. One strain of E. coli, E. coli Nissle, is currently being used as a probiotic in IBD patients and does not increase inflammation. The problem with E. coli Nissle is that it cannot take up any plasmids and it is therefore not a feasible candidate host for our project.

Prof. Rogler therefore suggests using Lactobacillus or Lactococcus lactis as hosts. These genera are not known to increase immune response. The former are already being used as probiotics in IBD patiens and the latter have been used in clinical trials with only minor adverse effects.^[2]

On the topic of current methods to investigate microbiome composition, Prof. Rogler stated that tissue samples are taken during endoscopy. Through 16S RNA sequencing the distribution of bacterial populations can be asserted on the phylum level and in some cases even species level. However, such an analysis cannot discern between bacteria that are present and functional and those that are non-functional. A dysbiosis is found in all cases.

Our analysis with nitric oxide (NO) and N-acyl homoserine lactone (AHL) might be a useful tool to investigate simultaneously the state of inflammation and microbiome composition, yet it all depends on the sensitivity of the system. Currently, inflammation is measured through calprotectin levels in the feces of patients; one would have to compare the calprotectin analysis to the measurement of NO in order to assert its effectiveness.

Finally, in order to make the project clinically feasible, we would need to encapsulate our bacteria. There are certain properties in the pill that we need to look out for to make it safe for administration. The material needs to withstand, even partial, digestion by stomach acid and pancreas enzymes. The bacteria need to be prevented from migrating out of the pill and colonizing the gut. Although, the colonization itself is not a safety issue, as a clinical study of L. lactis showed^[2], it is certainly not a desirable trait.

Our project relies heavily on the sensing of bacterial signaling molecules and metabolites in order to draw conclusions on the factors that affect IBD. Prof. Lacroix was able to point us to interesting markers that our system could sense, in order to determine what role they play in IBD.

The first suggestion he made in order to improve our system was to introduce a health marker that could be replaced with the inflammation marker nitric oxide (NO). This will be useful once we want to compare healthy individuals with IBD patients, in order to establish the difference between the two states. His suggestion was to use butyrate as this candidate marker, as it is a metabolite whose concentration is reduced significantly during IBD.

The most interesting suggestion was that of sensing metabolic intermediates, such as lactate. These will give our project a significant advantage to fecal analysis, as these intermediates will not be present in the feces. Nevertheless, they will allow us to paint the profile of, for example, lactate metabolizing bacteria and determine their dysbiotic behavior in IBD.

In order to make our project feasible, we would need to make sure that our encapsulated bacteria stay close to the mucus, where NO is produced. This will probably be necessary due to NO being relatively short lived.

Prof. Lacroix was also kind enough to offer us access to his gut simulator, a machine imitating the mucus of the gut, on which we could test our system and to their IBD affected mice, in case we would arrive that far in the project.