One thing about dealing with the microbiome is the need to constantly check and test your assumptions and revised from the results of tests. The series of A new specialized selection of suggestions links shattered several reasonable assumptions that appears to be false for most people, namely:

• A surplus of specific bacteria is not the cause (no statistical significance was found)
• A surplus of specific enzymes is not the cause (no statistical significance was found)
• Abundance or deficiency of certain compounds rarely has any statistical significance.

What keeps showing up as statistically significant are:

• An insufficient number of “good guy” bacteria — I do NOT mean traditional internet-lore good guy bacteria.
• An insufficient number of specific enzymes being produced.

In every study we have had 99% of the items being statistically significant at the 99% level being deficiency, i.e. with a condition, the mean of those with the condition is less than the mean of the reference group. The 1% that show up as too high are usually at relatively low z-score and is a surprising good match to the number expected from the False Detection Rate. In other words, they were false significant.

This means that some refactoring is planned. The first one is to give an option to get suggestions ONLY from the undergrowth. The second one, revised the probiotic suggestions from using the genomic data from KEGG: Kyoto Encyclopedia of Genes and Genomes to match the pattern. This has just been done.

A common pattern that I have seen in the enzyme data has been that the mean of the study group was about 30% of the mean of the reference group. So, the shotgun approach is to flag in a sample all of the enzymes that are less than 30% of the refence group (using lab specific data). From the flagged enzymes, we look at all of the available probiotics as well as potential probiotics species and see which ones produces the most of the missing enzymes. My expectation is that often the best of these probiotics may “kick ass”, in the case of a ME/CFS person “Just started it and wow does this one give me crazy dreams and headaches. I even tried it in the morning and still the same result.  Did you just power though or any pointers to deal with the herx” 

PLACEHOLDER — VIDEO COMING

This is a common symptom for our times. This is reported often in samples, and thus being examined if it reaches our threshold for inclusion as defined in A new specialized selection of suggestions links. It does.

Anxiety is likely a part of a feedback look — anxiety chemicals modify the microbiome resulting in more anxiety triggering chemicals being released.

Study Populations:

We have 3 symptom annotations that could be included

• Comorbid: High Anxiety [8.5 z-score x 45 samples]
• DePaul University Fatigue Questionnaire : Anxiety/tension [6.5 z-score x 63 samples]
• Condition: Generalized anxiety disorder [just 7 samples – so we exclude]

Taking the first two together we get 85 samples with an max z-score of 5.7

Symptom	Reference	Study
High Anxiety and Anxiety/tension	1115	85

• Bacteria Detected with z-score > 2.6: found 160 items, highest value was 5.7
• Enzymes Detected with z-score > 2.6: found 290 items, highest value was 5.7
• Compound Detected with z-score > 2.6: found No items

This hints that the enzymes being produced are likely more significant than the bacteria.

Interesting Significant Bacteria

All bacteria found significant (except 1) had too low levels. This is a common pattern found with these studies, it is not “bad bacteria bogie man bacteria” but an absence of “upstanding citizens bacteria”.

Bacteria	Reference Mean	Study	Z-Score
Actinobacillus pleuropneumoniae (species)	56	21	5.7
Haemophilus parainfluenzae (species)	1633	543	5.5
Haemophilus (genus)	1632	545	5.4
Actinobacillus porcinus (species)	183	68	5.2
Pasteurellaceae (family)	2001	760	5.1
Pasteurellales (order)	2001	760	5.1

This is echoed in some studies

• Early-Life Stress Modulates Gut Microbiota and Peripheral and Central Inflammation in a Sex-Dependent Manner [2021]

Interesting Enzymes

All enzymes found significant had too low levels.

Enzyme	Reference Mean	Study Mean	Z-Score
HslU—HslV peptidase; (3.4.25.2)	7177	3218	5.7
UDP-N-acetyl-alpha-D-glucosamine hydro-lyase (configuration-retaining; UDP-2-acetamido-2,6-dideoxy-alpha-D-xylo-hex-4-ulose-forming) (4.2.1.135)	1896	519	5.6
propanoate:CoA ligase (AMP-forming) (6.2.1.17)	1921	614	5.4
[RNA] 5′-hydroxy-ribonucleotide-3′-[RNA fragment]-lyase (cyclicizing; [RNA fragment]-3′- nucleoside-2′,3′-cyclophosphate-forming and hydrolysing) (4.6.1.19)	1541	534	5.3
(2S,3R)-3-hydroxybutane-1,2,3-tricarboxylate pyruvate-lyase (succinate-forming) (4.1.3.30)	1474	464	5.3
(S)-2-hydroxyglutarate:quinone oxidoreductase (1.1.5.13)	2241	710	5.3
propanoyl-CoA:oxaloacetate C-propanoyltransferase (thioester-hydrolysing, 1-carboxyethyl-forming) (2.3.3.5)	1519	500	5.3
CMP-N-acetyl-beta-neuraminate:beta-D-galactosyl-(1->4)-N-acetyl-beta-D-glucosaminyl-R (2->3)-N-acetyl-alpha-neuraminyltransferase (configuration-inverting) (2.4.99.6)	1630	549	5.3
4-hydroxybutanoate:NAD+ oxidoreductase (1.1.1.61)	2370	680	5.3
ATP:(Kdo)-lipid IVA 3-deoxy-alpha-D-manno-oct-2-ulopyranose 4-phosphotransferase (2.7.1.166)	1535	555	5.2
n/a (3.1.25.1)	1573	556	5.1
(S)-3-amino-2-methylpropanoate:2-oxoglutarate aminotransferase (2.6.1.22)	2424	876	5.1
5-aminopentanoate:2-oxoglutarate aminotransferase (2.6.1.48)	2431	883	5.1

Usually I leave the enzymes to nerds, but in this I decided to do a little exploration. I recall that Lactobacillus Casei has been shown to reduce anxiety in studies. So I looked up what enzymes are in it. In the above list of too low, I found the first one above, 3.4.25.2 in it. It opens up a possible model for picking probiotics a priori to address anxiety.

There was only one lactobacillus with more than one of these enzymes, Lactobacillus yamanashiensis – not available as a retail probiotic. There were three bifidobacterium which had more than one of these enzymes:

• Bifidobacterium dentium (3)
• Bifidobacterium thermophilum (3)
• Bifidobacterium subtile (2)

Unfortunately, none are available as a retail probiotic. HOWEVER, there is one commercial probiotic that appears to have EIGHT (8) of these enzymes: Escherichia coli, available as Mutaflor (E.Coli Nisse 1917) or Symbioflor-2. So if you have anxiety, the theoretically best probiotic for you to take is an E.Coli one!!

A surprise!

I went to look at the a priori suggestions and was shocked/delighted that the only probiotic on the suggestion list was Symbioflor-2. Even with the complexities of 160 bacteria to adjust, somehow our enzymes calculations and this totally study based approach came to agreement! I repeat, I was shocked —

This selecting probiotics based on enzyme deficiency has an interesting scent and I will likely follow up on future posts.

A small number of possible ones are reported in the literature, but the quality of the results are suspect.

Tax Rank	Tax Name	Shift	Citation Link
genus	Enterococcus (NCBI:1350 )	High	📚 PubMed
genus	Klebsiella (NCBI:570 )	High	📚 PubMed
genus	Prevotella (NCBI:838 )	High	📚 PubMed
genus	Salmonella (NCBI:590 )	High	📚 PubMed
genus	Staphylococcus (NCBI:1279 )	High	📚 PubMed
genus	Streptococcus (NCBI:1301 )	High	📚 PubMed
phylum	Firmicutes (NCBI:1239 )	Low	📚 PubMed
species	Acinetobacter baumannii (NCBI:470 )	High	📚 PubMed
species	Bifidobacterium longum (NCBI:216816 )	Low	📚 PubMed
species	Enterococcus faecalis (NCBI:1351 )	High	📚 PubMed
species	Enterococcus faecalis (NCBI:1351 )	Low
species	Enterococcus faecium (NCBI:1352 )	High	📚 PubMed
species	Escherichia coli (NCBI:562 )	High	📚 PubMed
species	Escherichia coli (NCBI:562 )	High
species	Klebsiella pneumoniae (NCBI:573 )	High	📚 PubMed
species	Methanobrevibacter smithii (NCBI:2173 )	High	📚 PubMed
species	Methanobrevibacter smithii (NCBI:2173 )	High
species	Pseudomonas aeruginosa (NCBI:287 )	High	📚 PubMed

The usual method of testing is from breath tests. From KEGG.JP we can get a list of bacteria that produces the compounds detected in the breath — the number is huge.

This page/video was suggested reading/viewing by a reader.

Question: How do you search on Microbiome Prescription for H2 etc?

On My Own Sample

To me, the more information about the microbiome that you have, the better it is to identify issues and build a treatment course. Below, the numbers will speak for themselves.

Second Class Tests

I deem these as 2nd class for several reasons: the number of bacteria reported is low (compare to others), the lab method requires the bacteria to be culturable (hence many bacteria will never be reported), they do no provide a suitable CSV file for upload to Microbiome Prescription (allowing 2nd opinions for treatment), they mechanism of measurement is not compatible to Microbiome Prescription (and most recent microbiome studies on the US National Library of Medicine)

Lab Name	BacteriaReported
Bioscreen (cfu/gm)	17
Biovis Microbiome Plus (cfu/g)	40
DayTwo	76
Diagnostic Solution GI-Map (cfu/gm)	38
GanzImmun Diagnostic A6 (cfu/gm)	76
GanzImmun Diagnostics AG Befundbericht	25
Genova Gi Effects (cfu/g)	28
Genova Parasitology (cfu/g)	7
GI EcologiX (Invivo)	55
GI360 Stool (UK)	67
Gut Zoomer (vibrant-wellness)	152
InVitaLab (cfu/gm)	23
Kyber Kompakt (cfu/g)	11
Medivere: Darm Mikrobiom Stuhltest (16s limited)	16
Medivere: Darn Magen Diagnostik (16s Limited)	16
Medivere: Gesundsheitscheck Darm (16s Limited)	17
Metagenomics Stool (De Meirleir) (16s Limited)	53
Smart Gut (ubiome 16s – Limited Taxonomy)	23
Verisana (cfu/ml) aka (kbe/ml)	11
Viome (No objective measures)	29

First Class Tests

These are first class because: they allow an easy upload to Microbiome Prescription, use an appropriate measurement process, report based on bacteria DNA/RNA and not cultured count. The key words are 16s and shotgun analysis and not cultured.

There can be considerable cost differences between these labs (links to my first choices are below).

Lab/Processor	Low	Typical	High	Uploads
AmericanGut	73	156	213	18
BiomeSight	11	570	1305	1283
BiomeSightRdp	279	656	862	11
Nirvana / CosmosId	6	430	705	33
es-xenogene	1461	2880	5225	4
Medivere	530	721	934	7
Microba	53	122	153	16
SequentiaBiotech	166	313	460	36
Ombre Labs	185	667	2238	1066
uBiome (Out of Business)	6	249	589	813

Most Useful Lab?

Microbiome Prescription does some Fuzzy Logic Artificial Intelligence (FLAI) a.k.a. Dr. Artificial Intelligence. The accuracy is a function of the data available to him, that is the number of uploads from a specific lab. The newer lab BiomeSight is the winner here. This preference is made stronger because they will ship worldwide while #2, OmbreLabs (according to feedback) will only ship to the US. Microbiome Prescription mission is to support people worldwide .. so the handwriting on the statistics board is clear.

We have a growing number of special studies using BiomeSight data.

1. 🗺️ Allergic Rhinitis (Hay Fever) ( 165 candidate bacteria) with highest z-score of 9.5
2. 🗺️ Autism ( 192 candidate bacteria) with highest z-score of 8.3
3. 🗺️ Bloating ( 126 candidate bacteria) with highest z-score of 5.4
4. 🗺️ Brain Fog ( 93 candidate bacteria) with highest z-score of 5.2
5. 🗺️ Chronic Fatigue Syndrome (CFS/ME) ( 170 candidate bacteria) with highest z-score of 6.6
6. 🗺️ Cold Extremities ( 145 candidate bacteria) with highest z-score of 12.7
7. 🗺️ COVID19 (Long Hauler) ( 218 candidate bacteria) with highest z-score of 12.6
8. 🗺️ Easily irritated ( 248 candidate bacteria) with highest z-score of 9.9
9. 🗺️ High Anxiety ( 170 candidate bacteria) with highest z-score of 5.9
10. 🗺️ Histamine or Mast Cell issues ( 138 candidate bacteria) with highest z-score of 8.4
11. 🗺️ Inflammatory bowel disease ( 245 candidate bacteria) with highest z-score of 12.5
12. 🗺️ irritable bowel syndrome ( 150 candidate bacteria) with highest z-score of 6.7
13. 🗺️ ME/CFS with IBS ( 193 candidate bacteria) with highest z-score of 8.4
14. 🗺️ ME/CFS without IBS ( 251 candidate bacteria) with highest z-score of 7.3
15. 🗺️ Post-exertional malaise ( 175 candidate bacteria) with highest z-score of 6
16. 🗺️ Tinnitus (ringing in ear) ( 133 candidate bacteria) with highest z-score of 6.7
17. 🗺️ Unrefreshed sleep ( 165 candidate bacteria) with highest z-score of 6.9

Social Media Questions

For other tests not covered above, see 16s Providers

On Sun Genomics

On Gut Zoomer

“Show me the beef!” This appears to be pure marketing hype — or more specifically, a substance for microbiome analysis that is high in pathogens!

Gut Zoomer [Est. 2016] – 170 -200 species (per their advertising) must be ordered thru a physician

While Xerogene typically reports TEN TIMES as many!!! It is DEFINITELY NOT THE MOST COMPLETE, it is infact at the bottom of their first class competitors but the top of second class competitors.

What about Biohm? 

Based on their official View a sample report. they report on only 44 bacteria items.

We have IBS annotated in three different ways:

• Autonomic Manifestations: irritable bowel syndrome
• Official Diagnosis: Irritable Bowel Syndrome
• Condition: ME/CFS with IBS

I ran each possible combination of the above and Official Diagnosis: Irritable Bowel Syndrome gave the strongest results and this will be reported here. When other things are combined, it is common for associations to become diffused/weaker. This is part of  A new specialized selection of suggestions links.

Study Populations:

Symptom	Reference	Study
Official Diagnosis: Irritable Bowel Syndrome	1122	56

• Bacteria Detected with z-score > 2.6: found 148 items, highest value was 6.7
• Enzymes Detected with z-score > 2.6: found 218 items, highest value was 6.1
• Compound Detected with z-score > 2.6: found No items

Interesting Significant Bacteria

The results are very striking — low bifidobacterium across the board at the top! Almost all of the bacteria associated are low.

Bacteria	Reference Mean	Study	Z-Score
Bifidobacterium gallicum (species)	3741	587	6.7
Bifidobacterium kashiwanohense PV20-2 (strain)	325	62	6.6
Bifidobacterium catenulatum subsp. kashiwanohense (subspecies)	315	62	6.6
Thermosediminibacterales (order)	49	16	6.3
Lactiplantibacillus pentosus (species)	123	28	5
Escherichia (genus)	5983	1397	4.9
Bifidobacterium cuniculi (species)	81	30	4.7
Bifidobacterium angulatum (species)	184	32	4.7

Interesting Enzymes

As is often seen with various symptoms/conditions, the associations are due to insufficiency and not surplus.

Enzyme	Reference Mean	Study Mean	Z-Score
substrate,NADPH—hemoprotein reductase:oxygen oxidoreductase (RH-hydroxylating or -epoxidizing) (1.14.14.1)	51	19	6.1
NADPH:hemoprotein oxidoreductase (1.6.2.4)	51	19	6.1
propanoyl-CoA:oxaloacetate C-propanoyltransferase (thioester-hydrolysing, 1-carboxyethyl-forming) (2.3.3.5)	1480	381	6.1
(2S)-3-(4-hydroxyphenyl)-2-isocyanopropanoate,2-oxoglutarate:oxygen oxidoreductase (decarboxylating) (1.14.20.10)	131	26	6
(2S)-3-(4-hydroxyphenyl)-2-isocyanopropanoate,2-oxoglutarate:oxygen oxidoreductase (1.14.20.9)	131	26	6
(2S,3R)-3-hydroxybutane-1,2,3-tricarboxylate pyruvate-lyase (succinate-forming) (4.1.3.30)	1432	375	5.9
ATP:amicoumacin A 2-phosphotransferase (2.7.1.230)	41	18	5.7
(E)-4-(trimethylammonio)but-2-enoyl-CoA:L-carnitine CoA-transferase (2.8.3.21)	1354	388	5.7
gamma-butyrobetainyl-CoA:electron-transfer flavoprotein 2,3-oxidoreductase (1.3.8.13)	1365	390	5.7
L-carnitine:CoA ligase (AMP-forming) (6.2.1.48)	1339	443	5.4
hydrogen-sulfide:ferredoxin oxidoreductase (1.8.7.1)	9912	4683	5.3
NADPH:acceptor oxidoreductase (1.6.99.1)	5684	2284	5.3
(2S,3S)-2-hydroxybutane-1,2,3-tricarboxylate hydro-lyase [(Z)-but-2-ene-1,2,3-tricarboxylate-forming] (4.2.1.79)	1284	407	5.2
S-methyl-5′-thioadenosine:phosphate S-methyl-5-thio-alpha-D-ribosyl-transferase (2.4.2.28)	3638	1668	5.1
acyl-CoA,ferrocytochrome b5:oxygen oxidoreductase (6,7 cis-dehydrogenating) (1.14.19.3)	1062	284	5
butanoyl-CoA:acetoacetate CoA-transferase (2.8.3.9)	3892	2229	5
L-carnitinyl-CoA hydro-lyase [(E)-4-(trimethylammonio)but-2-enoyl-CoA-forming] (4.2.1.149)	1422	362	5

Bottom Line

Looking at the bacteria, the probiotic solution would be:

• Bifidobacterium (a wide variety)
• E. Coli Probiotic (Mutaflor or Symbioflor-2)

Lactobacillus are HOSTILE to E.Coli, so Lactobacillus probiotics should likely be avoided.

Looking at some of the recommendations, we see Human milk oligosaccharides (prebiotic, Holigos, Stachyose) which are known to greatly encourage bifidobacterium. We also see saccharomyces boulardii which is also known to increase Bifidobacterium [2020].

As always, using your own 16s samples would produce the best suggestions.