High-throughput 16S rRNA gene sequencing reveals alterations of intestinal microbiota in myalgic encephalomyelitis/chronic fatigue syndrome patients

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Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09
study design
Citation
PMID PubMed identifier for scientific articles.
DOI Digital object identifier for electronic documents.
URI Uniform resource identifier for web resources.
Authors
Frémont M, Coomans D, Massart S, De Meirleir K
Journal
Anaerobe
Year
2013
Human intestinal microbiota plays an important role in the maintenance of host health by providing energy, nutrients, and immunological protection. Intestinal dysfunction is a frequent complaint in myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) patients, and previous reports suggest that dysbiosis, i.e. the overgrowth of abnormal populations of bacteria in the gut, is linked to the pathogenesis of the disease. We used high-throughput 16S rRNA gene sequencing to investigate the presence of specific alterations in the gut microbiota of ME/CFS patients from Belgium and Norway. 43 ME/CFS patients and 36 healthy controls were included in the study. Bacterial DNA was extracted from stool samples, PCR amplification was performed on 16S rRNA gene regions, and PCR amplicons were sequenced using Roche FLX 454 sequencer. The composition of the gut microbiota was found to differ between Belgian controls and Norwegian controls: Norwegians showed higher percentages of specific Firmicutes populations (Roseburia, Holdemania) and lower proportions of most Bacteroidetes genera. A highly significant separation could be achieved between Norwegian controls and Norwegian patients: patients presented increased proportions of Lactonifactor and Alistipes, as well as a decrease in several Firmicutes populations. In Belgian subjects the patient/control separation was less pronounced, however some abnormalities observed in Norwegian patients were also found in Belgian patients. These results show that intestinal microbiota is altered in ME/CFS. High-throughput sequencing is a useful tool to diagnose dysbiosis in patients and could help designing treatments based on gut microbiota modulation (antibiotics, pre and probiotics supplementation).

Experiment 1


Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: WikiWorks

Revision editor(s): WikiWorks

Subjects

Location of subjects
Belgium
Host species Species from which microbiome was sampled (if applicable)
Homo sapiens
Body site Anatomical site where microbial samples were extracted from according to the Uber Anatomy Ontology
Feces Cow dung,Cow pat,Droppings,Dung,Excrement,Excreta,Faeces,Fecal material,Fecal matter,Fewmet,Frass,Guano,Matières fécales@fr,Merde@fr,Ordure,Partie de la merde@fr,Piece of shit,Porción de mierda@es,Portion of dung,Portion of excrement,Portion of faeces,Portion of fecal material,Portion of fecal matter,Portion of feces,Portion of guano,Portion of scat,Portionem cacas,Scat,Spoor,Spraint,Stool,Teil der fäkalien@de,Feces
Condition The experimental condition / phenotype studied according to the Experimental Factor Ontology
chronic fatigue syndrome CFS,chronic fatigue syndrome,myalgic encephalitis,myalgic encephalomeyelitis/chronic fatigue syndrome,Myalgic Encephalomyelitis,myalgic encephalomyelitis,Postviral fatigue syndrome,systemic exertion intolerance disease
Group 0 name Corresponds to the control (unexposed) group for case-control studies
Belgian controls
Group 1 name Corresponds to the case (exposed) group for case-control studies
Norweigan controls
Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group
Patients were diagnosed for CFS according to the clinical criteria of Fukuda et al
Group 0 sample size Number of subjects in the control (unexposed) group
19
Group 1 sample size Number of subjects in the case (exposed) group
17
Antibiotics exclusion Number of days without antibiotics usage (if applicable) and other antibiotics-related criteria used to exclude participants (if any)
4 weeks

Lab analysis

Sequencing type
16S
16S variable region One or more hypervariable region(s) of the bacterial 16S gene
V5-V6
Sequencing platform Manufacturer and experimental platform used for quantifying microbial abundance
Roche454

Statistical Analysis

Statistical test
Mann-Whitney (Wilcoxon)
Significance threshold p-value or FDR threshold used for differential abundance testing (if any)
0.05
MHT correction Have statistical tests be corrected for multiple hypothesis testing (MHT)?
No


Alpha Diversity

Shannon Estimator of species richness and species evenness: more weight on species richness
unchanged

Signature 1

Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: Shaimaa Elsafoury

Revision editor(s): WikiWorks

Source: table 1

Description: Differentiately abundant mictobiota between Norwegian controls and Belgian controls.

Abundance in Group 1: increased abundance in Norweigan controls

NCBI Links
Roseburia
Holdemania

Revision editor(s): WikiWorks

Signature 2

Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: Shaimaa Elsafoury

Revision editor(s): WikiWorks

Source: table 1

Description: Differentiately abundant mictobiota between Norwegian controls and Belgian controls

Abundance in Group 1: decreased abundance in Norweigan controls

NCBI Links
Bacteroides
Alistipes
Barnesiella
Prevotella
Parabacteroides

Revision editor(s): WikiWorks

Experiment 2


Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: WikiWorks

Revision editor(s): WikiWorks

Subjects

Location of subjects
Belgium
Host species Species from which microbiome was sampled (if applicable)
Homo sapiens
Body site Anatomical site where microbial samples were extracted from according to the Uber Anatomy Ontology
Feces Cow dung,Cow pat,Droppings,Dung,Excrement,Excreta,Faeces,Fecal material,Fecal matter,Fewmet,Frass,Guano,Matières fécales@fr,Merde@fr,Ordure,Partie de la merde@fr,Piece of shit,Porción de mierda@es,Portion of dung,Portion of excrement,Portion of faeces,Portion of fecal material,Portion of fecal matter,Portion of feces,Portion of guano,Portion of scat,Portionem cacas,Scat,Spoor,Spraint,Stool,Teil der fäkalien@de,Feces
Condition The experimental condition / phenotype studied according to the Experimental Factor Ontology
chronic fatigue syndrome CFS,chronic fatigue syndrome,myalgic encephalitis,myalgic encephalomeyelitis/chronic fatigue syndrome,Myalgic Encephalomyelitis,myalgic encephalomyelitis,Postviral fatigue syndrome,systemic exertion intolerance disease
Group 0 name Corresponds to the control (unexposed) group for case-control studies
Norweigan controls
Group 1 name Corresponds to the case (exposed) group for case-control studies
Norweigan CFS/ME patients
Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group
Patients were diagnosed for CFS according to the clinical criteria of Fukuda et al
Group 0 sample size Number of subjects in the control (unexposed) group
17
Group 1 sample size Number of subjects in the case (exposed) group
25
Antibiotics exclusion Number of days without antibiotics usage (if applicable) and other antibiotics-related criteria used to exclude participants (if any)
4 weeks

Lab analysis

Sequencing type
16S
Sequencing platform Manufacturer and experimental platform used for quantifying microbial abundance
Roche454

Statistical Analysis

Statistical test
Mann-Whitney (Wilcoxon)
Significance threshold p-value or FDR threshold used for differential abundance testing (if any)
0.05
MHT correction Have statistical tests be corrected for multiple hypothesis testing (MHT)?
No


Alpha Diversity

Shannon Estimator of species richness and species evenness: more weight on species richness
unchanged

Signature 1

Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: Shaimaa Elsafoury

Revision editor(s): WikiWorks

Source: table 2

Description: Differentiately abundant mictobiota between Norwegian CFS patients and Norwegian controls

Abundance in Group 1: increased abundance in Norweigan CFS/ME patients

NCBI Links
Alistipes
Lactonifactor

Revision editor(s): WikiWorks

Signature 2

Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: Shaimaa Elsafoury

Revision editor(s): WikiWorks

Source: table 2

Description: Differentiately abundant mictobiota between Norwegian CFS patients and Norwegian controls

Abundance in Group 1: decreased abundance in Norweigan CFS/ME patients

NCBI Links
Roseburia
Holdemania
Dialister
Syntrophococcus

Revision editor(s): WikiWorks

Experiment 3


Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: WikiWorks

Revision editor(s): WikiWorks

Subjects

Location of subjects
Belgium
Host species Species from which microbiome was sampled (if applicable)
Homo sapiens
Body site Anatomical site where microbial samples were extracted from according to the Uber Anatomy Ontology
Feces Cow dung,Cow pat,Droppings,Dung,Excrement,Excreta,Faeces,Fecal material,Fecal matter,Fewmet,Frass,Guano,Matières fécales@fr,Merde@fr,Ordure,Partie de la merde@fr,Piece of shit,Porción de mierda@es,Portion of dung,Portion of excrement,Portion of faeces,Portion of fecal material,Portion of fecal matter,Portion of feces,Portion of guano,Portion of scat,Portionem cacas,Scat,Spoor,Spraint,Stool,Teil der fäkalien@de,Feces
Condition The experimental condition / phenotype studied according to the Experimental Factor Ontology
chronic fatigue syndrome CFS,chronic fatigue syndrome,myalgic encephalitis,myalgic encephalomeyelitis/chronic fatigue syndrome,Myalgic Encephalomyelitis,myalgic encephalomyelitis,Postviral fatigue syndrome,systemic exertion intolerance disease
Group 0 name Corresponds to the control (unexposed) group for case-control studies
Belgian controls
Group 1 name Corresponds to the case (exposed) group for case-control studies
Belgian CFS/ME patients
Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group
Patients were diagnosed for CFS according to the clinical criteria of Fukuda et al
Group 0 sample size Number of subjects in the control (unexposed) group
19
Group 1 sample size Number of subjects in the case (exposed) group
18
Antibiotics exclusion Number of days without antibiotics usage (if applicable) and other antibiotics-related criteria used to exclude participants (if any)
4 weeks

Lab analysis

Sequencing type
16S
Sequencing platform Manufacturer and experimental platform used for quantifying microbial abundance
Roche454

Statistical Analysis

Statistical test
Mann-Whitney (Wilcoxon)
Significance threshold p-value or FDR threshold used for differential abundance testing (if any)
0.05
MHT correction Have statistical tests be corrected for multiple hypothesis testing (MHT)?
No


Alpha Diversity

Shannon Estimator of species richness and species evenness: more weight on species richness
unchanged

Signature 1

Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: Shaimaa Elsafoury

Revision editor(s): WikiWorks

Source: table 3

Description: Differentiately abundant mictobiota between Belgian CFS patients and Belgian controls

Abundance in Group 1: increased abundance in Belgian CFS/ME patients

NCBI Links
Lactonifactor

Revision editor(s): WikiWorks

Signature 2

Reviewed Marked as Reviewed by Shaimaa Elsafoury on 2021/02/09

Curated date: 2021/01/10

Curator: Shaimaa Elsafoury

Revision editor(s): WikiWorks

Source: table 3

Description: Differentiately abundant mictobiota between Belgian CFS patients and Belgian controls

Abundance in Group 1: decreased abundance in Belgian CFS/ME patients

NCBI Links
Asaccharobacter

Revision editor(s): WikiWorks