Probiotic supplementation mitigates sex-dependent nociceptive changes and gut dysbiosis induced by prenatal opioid exposure

Study information

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

study design

laboratory experiment

Citation

PMID PubMed identifier for scientific articles.

39950489

DOI Digital object identifier for electronic documents.

https://doi.org/10.1080/19490976.2025.2464942

URI

https://pubmed.ncbi.nlm.nih.gov/39950489/

Authors

Singh S, Abu Y, Antoine D, Gomez D, Tao J, Truitt B, Roy S

Journal

Gut microbes

Year

2025

Keywords:

Prenatal opioid exposure, dysbiosis, gut microbiome, nociception, sex-based differences

The gut microbiome has emerged as a promising target for modulating adverse effects of opioid exposure due to its significant role in health and disease. Opioid use disorder (OUD) has become increasingly prevalent, specifically in women of reproductive age, contributing to an increased incidence of offspring exposed to opioids in utero. Recent studies have shown that prenatal opioid exposure (POE) is associated with notable changes to the maternal gut microbiome, with subsequent implications for the offspring's microbiome and other adverse outcomes. However, the role of the gut microbiome in mediating sex-based differences in pain sensitivity has not yet been investigated. In this study, both male and female C57BL/6 offspring were used to determine sex-based differences in nociception and gut microbial composition as a result of POE. Our data reveals significant sex-based differences in offspring prenatally exposed to opioids. The gut microbiome of opioid-exposed females showed an enrichment of commensal bacteria including Lactobacillus compared to opioid-exposed males. Additionally, POE females demonstrated decreased nociceptive sensitivity, while males demonstrated increased nociceptive sensitivity. RNA sequencing of the prefrontal cortex showed sex-based differences in several canonical pathways, including an increase in the opioid signaling pathway of opioid-exposed females, which was not observed in males. Microbiome modification via maternal probiotic supplementation attenuated sex-based differences throughout the early stages of life. Together, our study provides further insight on sex-based differences arising from POE and highlights the pivotal role of the gut microbiome as a modifiable target for mitigating its negative effects.

Experiment 1

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/03/19

Curator: Dorcas-od

Revision editor(s): Dorcas-od, Miss Lulu

Subjects

Location of subjects

United States of America

Host species Species from which microbiome was sampled. Contact us to have more species added.

Mus musculus

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,feces

Condition The experimental condition / phenotype studied according to the Experimental Factor Ontology

Treatment outcome measurement Treatment outcome measurement,treatment outcome measurement

Group 0 name Corresponds to the control (unexposed) group for case-control studies

Female offspring prenatally exposed to saline group (CSAL_F)

Group 1 name Corresponds to the case (exposed) group for case-control studies

Female offspring prenatally exposed to opioids group (MSAL_F)

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group

Offspring born to dams that were exposed to opioids (hydromorphone pre-gestationally followed by methadone during gestation), representing the prenatal opioid exposure condition

Group 0 sample size Number of subjects in the control (unexposed) group

10

Group 1 sample size Number of subjects in the case (exposed) group

10

Lab analysis

Sequencing type

16S

16S variable region One or more hypervariable region(s) of the bacterial 16S gene

V4

Sequencing platform Manufacturer and experimental platform used for quantifying microbial abundance

Illumina

Statistical Analysis

Data transformation Data transformation applied to microbial abundance measurements prior to differential abundance testing (if any).

relative abundances

Statistical test

LEfSe

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

LDA Score above Threshold for the linear discriminant analysis (LDA) score for studies using the popular LEfSe tool

2

Matched on Factors on which subjects have been matched on in a case-control study

age

Alpha Diversity

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

increased

Signature 1

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 2C

Description: Microbiome analysis of female offspring (CSAL_F vs. MSAL_F) (n = 10 per group in all experiments).

Abundance in Group 1: increased abundance in Female offspring prenatally exposed to opioids group (MSAL_F)

NCBI	Quality Control	Links
Eubacterium xylanophilum
Lachnospiraceae bacterium NK4A136
Lactobacillus
Monoglobus
Romboutsia
Ruminococcus
UGC_005UGC_005

Revision editor(s): Miss Lulu

Signature 2

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 2C

Description: Microbiome analysis of female offspring (CSAL_F vs. MSAL_F) (n = 10 per group in all experiments).

Abundance in Group 1: decreased abundance in Female offspring prenatally exposed to opioids group (MSAL_F)

NCBI	Quality Control	Links
Bacteroides
Enterorhabdus
Eubacterium ventriosum
Ligilactobacillus
Lachnospiraceae UGC-001Lachnospiraceae UGC-001
Prevotellaceae UGC-001Prevotellaceae UGC-001
Clostridium
Rikenellaceae RC9 gut groupRikenellaceae RC9 gut group

Revision editor(s): Miss Lulu

Experiment 2

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Differences from previous experiment shown

Subjects

Group 0 name Corresponds to the control (unexposed) group for case-control studies

Male offspring prenatally exposed to saline group (CSAL_M)

Group 1 name Corresponds to the case (exposed) group for case-control studies

Male offspring prenatally exposed to opioids group (MSAL_M)

Lab analysis

Statistical Analysis

Alpha Diversity

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

increased

Signature 1

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 3C

Description: Microbiome analysis of male offspring (CSAL_M vs. MSAL_M) (n = 10 per group in all experiments).

Abundance in Group 1: increased abundance in Male offspring prenatally exposed to opioids group (MSAL_M)

NCBI	Quality Control	Links
Alloprevotella
Muribaculum
Odoribacter
Parvibacter
Tyzzerella
Lachnospiraceae bacterium NK4A136
UGC-009UGC-009
UGC-005UGC-005
Prevotellaceae-Ga6A1Prevotellaceae-Ga6A1

Revision editor(s): Miss Lulu

Signature 2

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 3C

Description: Microbiome analysis of male offspring (CSAL_M vs. MSAL_M) (n = 10 per group in all experiments).

Abundance in Group 1: decreased abundance in Male offspring prenatally exposed to opioids group (MSAL_M)

NCBI	Quality Control	Links
Akkermansia
Eubacterium ventriosum
Staphylococcus

Revision editor(s): Miss Lulu

Experiment 3

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu, Dorcas-od

Differences from previous experiment shown

Subjects

Group 0 name Corresponds to the control (unexposed) group for case-control studies

Control female offspring without prenatal exposure to probiotics(CSAL_F)

Group 1 name Corresponds to the case (exposed) group for case-control studies

Control female offspring with prenatal exposure to probiotics(CPRO_F)

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group

Female offspring from control group exposed prenatally to probiotics

Lab analysis

Statistical Analysis

Alpha Diversity

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

unchanged

Signature 1

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu, Tosin

Source: Figure 6C

Description: Microbiome analysis of control female offspring with or without prenatal exposure to probiotics (CPRO_F vs CSAL_F) (n = 10 per group in all experiments).

Abundance in Group 1: increased abundance in Control female offspring with prenatal exposure to probiotics(CPRO_F)

NCBI	Quality Control	Links
Anaeroplasma
Eubacterium xylanophilum
Lachnoclostridium
Lactobacillus
Ligilactobacillus
Parvibacter
Romboutsia
Clostridium sp. MD294

Revision editor(s): Miss Lulu, Tosin

Signature 2

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 6C

Description: Microbiome analysis of control female offspring with or without prenatal exposure to probiotics (CPRO_F vs CSAL_F) (n = 10 per group in all experiments).

Abundance in Group 1: decreased abundance in Control female offspring with prenatal exposure to probiotics(CPRO_F)

NCBI	Quality Control	Links
Akkermansia
Bacteroides
Eubacterium ventriosum
Faecalibaculum
Odoribacter
Turicibacter
Rikenellaceae RC9 gut groupRikenellaceae RC9 gut group

Revision editor(s): Miss Lulu

Experiment 4

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu, Dorcas-od, Folakunmi

Differences from previous experiment shown

Subjects

Group 0 name Corresponds to the control (unexposed) group for case-control studies

POE female offspring without prenatal exposure to probiotics(MSAL_F)

Group 1 name Corresponds to the case (exposed) group for case-control studies

POE female offspring with prenatal exposure to probiotics(MPRO_F)

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group

Prenatal Opioid exposure female offspring with prenatal exposure to probiotics

Lab analysis

Statistical Analysis

Alpha Diversity

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

unchanged

Signature 1

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 7C

Description: Microbiome analysis of POE female offspring with or without prenatal exposure to probiotics (MPRO_F vs. MSAL_F) (n = 10 per group in all experiments).

Abundance in Group 1: increased abundance in POE female offspring with prenatal exposure to probiotics(MPRO_F)

NCBI	Quality Control	Links
Desulfovibrio
Rikenellaceae RC9 gut groupRikenellaceae RC9 gut group

Revision editor(s): Miss Lulu

Signature 2

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Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu, Svetlana up

Source: Figure 7C

Description: Microbiome analysis of POE female offspring with or without prenatal exposure to probiotics (MPRO_F vs. MSAL_F) (n = 10 per group in all experiments).

Abundance in Group 1: decreased abundance in POE female offspring with prenatal exposure to probiotics(MPRO_F)

NCBI	Quality Control	Links
Monoglobus
[Eubacterium] brachy
A2A2

Revision editor(s): Miss Lulu, Svetlana up

Experiment 5

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu, Dorcas-od

Differences from previous experiment shown

Subjects

Group 0 name Corresponds to the control (unexposed) group for case-control studies

Control Male offspring without prenatal exposure to probiotics(CSAL_M)

Group 1 name Corresponds to the case (exposed) group for case-control studies

Control Male offspring with prenatal exposure to probiotics(CPRO_M)

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group

Male offspring from control group exposed prenatally to probiotics

Lab analysis

Statistical Analysis

Significance threshold p-value or FDR threshold used for differential abundance testing (if any)

0.005

Alpha Diversity

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

decreased

Signature 1

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Svetlana up, Folakunmi, Tosin, Miss Lulu

Source: Figure 8C

Description: Microbiome analysis of control male offspring with or without prenatal exposure to probiotics (CPRO_M vs. CSAL_M) (n = 10 per group in all experiments).

Abundance in Group 1: increased abundance in Control Male offspring with prenatal exposure to probiotics(CPRO_M)

NCBI	Quality Control	Links
Anaerotruncus
Colidextribacter
Intestinimonas
Lachnospiraceae bacterium NK4A136
Ligilactobacillus
Oscillibacter
Lachnospiraceae_UGC_006Lachnospiraceae_UGC_006
Prevotellaceae UGC-001Prevotellaceae UGC-001
Clostridium sp. MD294

Revision editor(s): Svetlana up, Folakunmi, Tosin, Miss Lulu

Signature 2

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 8C

Description: Microbiome analysis of control male offspring with or without prenatal exposure to probiotics (CPRO_M vs. CSAL_M) (n = 10 per group in all experiments).

Abundance in Group 1: decreased abundance in Control Male offspring with prenatal exposure to probiotics(CPRO_M)

NCBI	Quality Control	Links
Monoglobus
Eubacterium ventriosum
Bifidobacterium
Faecalibaculum
Lactobacillus

Revision editor(s): Miss Lulu

Experiment 6

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu, Dorcas-od

Differences from previous experiment shown

Subjects

Group 0 name Corresponds to the control (unexposed) group for case-control studies

POE Male offspring without prenatal exposure to probiotics(MSAL_M)

Group 1 name Corresponds to the case (exposed) group for case-control studies

POE Male offspring with prenatal exposure to probiotics(MPRO_M)

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group

Prenatal Opioid exposure Male offspring with prenatal exposure to probiotics

Lab analysis

Statistical Analysis

Alpha Diversity

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

increased

Signature 1

EditHistoryDelete

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu, Tosin

Source: Figure 9C

Description: Microbiome analysis of POE male offspring with or without prenatal exposure to probiotics (MPRO_M vs. MSAL_M) (n = 10 per group in all experiments).

Abundance in Group 1: increased abundance in POE Male offspring with prenatal exposure to probiotics(MPRO_M)

NCBI	Quality Control	Links
Akkermansia
Alistipes
Bacteria incertae sedis
Desulfovibrio
Intestinimonas
Ligilactobacillus
Roseburia
A2A2
Rikenellaceae RC9 gut groupRikenellaceae RC9 gut group
Prevotellaceae UGC-001Prevotellaceae UGC-001
Clostridium sp. MD294

Revision editor(s): Miss Lulu, Tosin

Signature 2

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Reviewed Marked as Reviewed by Folakunmi on 2025-4-10

Curated date: 2025/04/09

Curator: Miss Lulu

Revision editor(s): Miss Lulu

Source: Figure 9C

Description: Microbiome analysis of POE male offspring with or without prenatal exposure to probiotics (MPRO_M vs. MSAL_M) (n = 10 per group in all experiments).

Abundance in Group 1: decreased abundance in POE Male offspring with prenatal exposure to probiotics(MPRO_M)

NCBI	Quality Control	Links
Prevotella_Ga6A1Prevotella_Ga6A1
Alloprevotella
Turicibacter
Lachnospiraceae_UGC_006Lachnospiraceae_UGC_006

Revision editor(s): Miss Lulu