Systemic antibody responses against human microbiota flagellins are overrepresented in chronic fatigue syndrome patients, 2022, Vogl et al

[Andy]

Abstract

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a debilitating disease with an unclear etiology and pathogenesis. Both an involvement of the immune system and gut microbiota dysbiosis have been implicated in its pathophysiology. However, potential interactions between adaptive immune responses and the microbiota in ME/CFS have been incompletely characterized.

Here, we profiled antibody responses of patients with severe ME/CFS and healthy controls against microbiota and viral antigens represented as a phage-displayed 244,000 variant library. Patients with severe ME/CFS exhibited distinct serum antibody epitope repertoires against flagellins of Lachnospiraceae bacteria. Training machine learning algorithms on this antibody-binding data demonstrated that immune responses against gut microbiota represent a unique layer of information beyond standard blood tests, providing improved molecular diagnostics for ME/CFS.

Together, our results point toward an involvement of the microbiota-immune axis in ME/CFS and lay the foundation for comparative studies with inflammatory bowel diseases and illnesses characterized by long-term fatigue symptoms, including post–COVID-19 syndrome.

Open access, https://www.science.org/doi/10.1126/sciadv.abq2422

 

[Andy]

The SMC not only deliberately calling it CFS/ME, rather than either CFS or ME/CFS, as used in the paper itself, but also allowing Alan Carson to give his 'expert' opinion, https://www.sciencemediacentre.org/...chnospiraceae-bacteria-in-people-with-cfs-me/

 

[Hutan]

Funding Information
Austrian Science Fund (FWF): Erwin Schrödinger fellowship J4256
Solve ME/CFS Initiative (SMCI): Ramsay Award

 

[Hutan]

severe ME/CFS cases uniformly assembled by the U.K. ME/CFS Biobank (UKMEB)

case selection is probably good then

Therefore, we leveraged a high-throughput antibody profiling technology, PhIP-Seq (46), to test for reactivity against 244,000 peptide antigens (47) in a target-agnostic way (50). This antigen library encompasses diverse bacterial and viral antigens originating from pathogenic, probiotic, and commensal bacteria,

That's a lot of peptides. Presumably there is the possibility that these little bits of protein antigens might join together to make a protein that people with ME/CFS are actually reacting to, but there may be no reaction to the protein fragments. It might be like a key that fits into a lock - if you take the key and smash it up into little pieces, none of the pieces are going to open the lock. I don't know if there are many examples of this technology finding useful things?

Comparing general metrics such as the number of overall antibody-bound peptides or the diversity of the Ig epitope repertoires did not show any significant differences between patients with severe ME/CFS and healthy controls (Fig. 1C). Mann-Whitney-Wilcoxon tests indicated that the difference in both number of significantly bound peptides per individual and Shannon α-diversity were not statistically significant (P > 0.3 and P > 0.9, respectively). In addition, no single peptides were bound at significantly different rates in patients with severe ME/CFS and healthy controls after false discovery rate correction (Fig. 1D, Fisher’s exact test for differences between the two groups). However, the majority of peptides, which were more frequently bound in patients with severe ME/CFS, originated from bacterial flagellins (Fig. 1D and table S1 for a full list).

So, that's sounding a bit disappointing. Clearly peptides are binding in both healthy controls and the people with ME/CFS. And no single peptides bound at significantly different rates in the two groups.

This figure shows the interesting finding:


ME/CFS patients were found to be a lot more likely to have antibody responses to peptides associated with bacterial flagella (the 'tails' that some bacteria have to help with movement). Also Figure 1E, showing that the ratio of antibody responses in ME/CFS and controls is about 3 to 1 for the flagella-related antigens, but pretty much 1 to 1 for the rest of the antigen groups.

 

[Hutan]

So, then they tried to narrow down the particular bacteria that might be eliciting the antibody response.

Next, we studied the phylogenetic origin of the antibody-bound flagellins in greater detail. The overrepresented flagellins were frequently originating from the order of Clostridiales and mostly the family of Lachnospiraceae (species such as Roseburia inulinivornas or Roseburia faecis) as well as some Eubacteriaceae (genus Eubacterium). In addition, some flagellins of Gammaproteobacteria (e.g., genera of Salmonella, Pseudomonas, and Escherichia) were more frequently bound by antibodies in patients with severe ME/CFS.

Some of the detail is given in Table 2a below. Note the ratios on the right (incidence in ME/CFS vs controls). The biggest ratio is for a Legionella flagella with 15% of ME/CFS patients responding and only 2.5% of healthy controls responding. I'm not too sure what to make of this, as the differences don't look too compelling. It's certainly not a situation of overwhelming levels of response in the ME/CFS people versus basically no response in controls.


They then went on to use machine learning to develop a sort of biomarker signature that supposedly would identify ME/CFS patients. Here's the 15 top performing peptides:

In some cases, even multiple peptides from the same protein appeared in the SHAP analysis (e.g., peptide nos.106628 and 209928; table S3), making it highly unlikely that these Ig responses against very specific bacterial taxa and protein groups would occur by chance.

The story, if there really is one here, is looking pretty complicated. And I wonder to what extent different diets might be influencing things.

They also looked at parameters in the blood, finding lower creatine kinase in the ME/CFS group (as previously found in UK-MEB samples), but not finding much else of note. We didn't get very excited by that finding when it was first reported, because of the risk that the levels are associated with activity - and there are big differences in activity levels between the severe ME/CFS patients and the controls.

They next tried making machine-learning generated algorithms to identify the ME/CFS individuals from the controls based on the blood test parameters, and found that it sorted the patients from controls better than the antibody models did. But, that might just be largely based on the creatine kinase, with the activity level confounding. And then, of course, they made biomarker models with the antibody results and the blood test results.

 

[Hutan]

Discussion section

So, this is interesting - an increased response to Lachnospiraceae flagellins was found in the ME/CFS group, and has been reported in Crohn's disease.

In this study, we have detected an overrepresentation of systemic anti-flagellin Ig responses in patients with severe ME/CFS by leveraging a target-agnostic (50) PhIP-Seq screening approach (47). A similar overrepresentation of serum Ig responses against Lachnospiraceae flagellins has been reported in Crohn’s disease (CD) (64, 65). CD is a chronic inflammatory disease of the intestinal tract associated with intestinal microbial dysbiosis and immune system dysregulation (66). Lachnospiraceae and other Clostridiales produce short-chain fatty acids that mediate a range of beneficial effects in the gut such as enhancing tolerance and the epithelial barrier function, anti-inflammatory effects, as well as activating regulatory T cells (64, 67). Hence, excessive, mistargeted immune reactions against these favorable commensals are conceptually in line with CD and irritable bowel syndrome, a common comorbidity reported by patients with severe ME/CFS (10). Notably, CD patients (compared to healthy individuals) have an increased risk to also develop ME/CFS (68).

They do discuss the possibility that the overlapping findings in the two diseases might be due to problems in the lab processes, but they don't think so. They also discuss the concern about creatine kinase just being a marker of activity, and the possibility that diet, or long periods of not eating might be contributing to the differences.

All up, the antibodies to some bacteria proteins is interesting, but with no clear differences between patients and controls (there's a lot of overlap), I'm less excited about this than some other recent findings.

 

[Jonathan Edwards]

Thanks for a very careful analysis, @Hutan.

I am quite interested in the idea that PWME might be making immune responses to endocytosed bacterial components in gut more than healthy people. This would not be autoimmunity, or need to be anything specific to an antigen.

The gut immune system is normally suppose to 'ignore' bacterial antigens an just chew them up or keep them out. If it got preoccupied with making antibodies that might have systemic signalling effects.

This would likely be due to a shift in T cell control - maybe cytotoxic CD57 or regulatory T.
The antibodies would be part of a generalised response under T cell permission. This would be reminiscent of the situation in Reiter's syndrome or Ankylosing Spondylitis. And of course Crohn's disease is in that group.

Like you, I am not very convinced this will pan out but I think the pattern of data might mean something interesting.

 

[FMMM1]

From Wikipedia:
"Endocytosis is a cellular process in which substances are brought into the cell. The material to be internalized is surrounded by an area of cell membrane, which then buds off inside the cell to form a vesicle containing the ingested material. Endocytosis includes pinocytosis (cell drinking) and phagocytosis (cell eating). It is a form of active transport."

I think Maureen Hanson did some work on vesicles - of course this is potentially an immune response to the vesicle contents(?) i.e. rather than abnormalities in vesicles (numbers of vesicles, sizes ---). So Hanson's work wouldn't necessarily have turned up anything.

Again (common response from me) I wonder if GWAS will turn up something e.g. flag up immune genes which increase/reduce risk --- highlights the potential benefits of GWAS.

 

[Dolphin]

New publication by Vogl. May use same data?
https://www.s4me.info/threads/syste...y-in-crohns-disease-cfs-2024-bourgonje.37042/