Information Flow and Genetics

TWiM #190 Exosomes in Your Nose and in Your Gut

Podcast and Annotation Information
  • Annotation by Leonardo Baumgartner, Martin Leyhe, Triston Walsh, Mel Melendrez-Vallard, and Nancy Boury.
  • Podcast audio by TWiM: Listen to TWiM #190 Podcast
  • Podcast transcript by Sarah Morgan: Access TWiM #190 Transcript
  • Papers Discussed:
    • Nocera AL, Mueller SK, Stephan JR, Hing L, Seifert P, Han X, Lin DT, Amiji MM, Libermann T, Bleier BS. 2019. Exosome swarms eliminate airway pathogens and provide passive epithelial immunoprotection through nitric oxide. J Allergy Clin Immunol. 143(4):1525-1535.e1. doi: 10.1016/j.jaci.2018.08.046.
    • Teng Y, Ren Y, Sayed M, Hu X, Lei C, Kumar A, Hutchins E, Mu J, Deng Z, Luo C, Sundaram K, Sriwastva MK, Zhang L, Hsieh M, Reiman R, Haribabu B, Yan J, Jala VR, Miller DM, Van Keuren-Jensen K, Merchant ML, McClain CJ, Park JW, Egilmez NK, Zhang HG. 2018. Plant-Derived Exosomal MicroRNAs Shape the Gut Microbiota. Cell Host Microbe. 24(5):637-652.e8. doi: 10.1016/j.chom.2018.10.001.

1. Paper Abstracts

1.1. Snippet paper; discussion starts at 1:42 minutes

The Most Interesting Things (according to students)

Small vacuoles called exosomes are released by epithelial cells in the nose in response to detection of a pathogen; these help fight pathogens and can alert other cells to the presence of danger, like a human ‘smoke detector’.

The abstract cannot be copied due to licensing restrictions. Please see licensing information and links to the article at the journal’s web page and/or PubMed in Section 8.1.

1.2. Main paper; discussion starts at 22:56 minutes

The Most Interesting Things (according to students)

The vegetables we eat can influence our gut microbiome composition, due to exosomes from the plants.

The abstract cannot be copied due to licensing restrictions. Please see licensing information and links to the article at the journal’s web page and/or PubMed in Section 8.1.

2. Vision and Change Core Concepts and 2024 ASM Fundamental Statements

Snippet Main
Vision and Change Topics
  • Structure and Function (V&C_SF)
  • Information Flow and Genetics (V&C_IFG)
  • Impact of Microorganisms (V&C_IM)
  • Microbial Ecology (V&C_ME)
  • Structure and Function (V&C_SF)
  • Metabolic Pathways (V&C_MP)
ASM Fundamental Statements
  • Fundamental Statement 6 (ASM_6): The distinct structures and processes in microbes can be targets for interspecies competition, antimicrobial treatments, and host immunity.
  • Fundamental Statement 17 (ASM_17): The regulation of gene expression is influenced by external and internal molecular cues and/or signals.
  • Fundamental Statement 13 (ASM_13):  Intrinsic factors, such as genotype, metabolism, and cell structures, impact the survival and growth of microbes.
  • Fundamental Statement 14 (ASM_14):  Extrinsic factors, such as abiotic and biotic interactions in the environment, can impact survival and growth of microbes.
  • Fundamental Statement 18 (ASM_18): The regulation of gene expression is influenced by external and internal molecular cues and signals.
  • Fundamental Statement 22 (ASM_22): Most microbes interact with hosts in beneficial or neutral ways, with a minority having a detrimental impact on their host.
  • Fundamental Statement 27 (ASM_27): The extent of microbial diversity is largely unknown, and exploration of this diversity is critical to understanding microbes and their role in the biosphere.

3.  Potential Learning Objectives for the Podcast

The student will be able to: Paper1 Order2
  • Identify and order the sequence of events leading to activation of an immune response in the nose.
  • Define exosome.
 S L
  • Predict and discuss whether nasal decongestant drugs would help, hamper, or not affect exosome production.
 S H
  • Explain how different plant exosomes end up in different organs when eaten.
  • Link bacterial family to food source uptake by exosomes.
 M L
  • Predict ways exosomes can be used to promote a better intestinal health to an average consumer.
 M H

1 Papers: Snippet (S) or Main (M)

2 Learning Objectives: Lower Order or Higher Order (H)

4. Techniques Described (with Time Stamps)

Here is a link to a bio-dictionary that has many, but not all definitions if you need a definition: Explore Biology Bio-Dictionary

4.1. Snippet Paper

  • Fluorescence Microscopy (18:30–18:55): This is a type of microscopy that is used to follow or locate a fluorescent molecules.  Here, labeled DNA allowed the researchers to visualize the transfer of genetic material in exosomes.

4.2. Main Paper

  • Mouse Experiments (31:11–31:45; 32:42–33:21; 34:31–35:02): Mice were fed ginger exosomes for a week and then fecal samples were taken for microbiome analysis (qPCR). In the colitis experiments with mice, they fed the mice dextran sulfate sodium (DSS) which compromises the barrier of the gut, fed the ginger exosomal RNA, then analyzed the gut microbiome, cytokine numbers, and DSS outcomes.  The researchers compared these values in treated (ginger exosomal RNA) and untreated (negative control) groups.
  • 16S rRNA Sequencing (32:47–32:53; 38:10–38:44; 39:10–39:50): This is a method for identifying genetic sequences of ribosomal rRNA (small subunit) that was used to analyze the mice’s intestinal fecal microbiome after being fed ginger exosomes and to identify what kinds of bacteria took up the ginger exosomes.
  • Human Experiments (34:41; 36:30–37:15): Humans were fed ginger extract (or not) in a double-blind controlled experiment.  Researchers then gut microbiomes were analyzed using qPCR on fecal samples.
  • Fluorescent Microscopy and Flow Cytometry (37:48–38:18) These methods are used to follow, locate, or identify cell features using a fluorescent molecular probe. Here these methods were used to confirm the exosomes were being taken up by bacteria.
  • Culture Experiments (41:38-43:30): The researchers cultured Lactobacillus rhamnosus (LGG) and showed its growth is promoted by ginger exosomes and reduced by grapefruit exosomes. In supernatant experiments, a supernatant of LGG treated with ginger was found to inhibit Listeria, E. coli, B. fragilis but not itself.
  • mRNA Sequencing/Gene Profiling (43:36–44:40): This is a sequencing technique used to examine all mRNAs produced in a cell (transcriptomics).  It was used to compare mRNA from mice that were fed ginger exosomes to mRNA from mice that were not.
  • Metabolomics (51:40–52:21): Similar to other -omics techniques, this one examines all metabolites produced in a cell.  The researchers used it to identify how ginger exosomal RNA leads to induction of IL-22.

5. Connections to General Microbiology Processes/Concepts (with Time Stamps)

5.1. Snippet Paper

  • Cell Structures (5:15): LPS, Toll like receptor IV (TLR4, the toll-like receptor that recognizes LPS).
  • Host Responses (5:50–6:15; 23:40–24:30): TLRs detect signatures indicating bacterial presence.
  • Pathogen Associated Molecular Patterns (PAMPs) (6:15): These are pathogen molecules that are recognized by the immune system (e.g. LPS, flagellin, dsRNA).
  • Secretion, Innate Immunity (7:25–8:00): Exosomes bud off nasal epithelial cells and are stimulated by bound TLR4.
  • Exosomes (7:25–24:24 ): Small fluid filled sacs that contain tetraspanin; released from the cell in the lining of the nasal passage. Embedded proteins have transmembrane 4 superfamily type canonical structure.
  • Cell-Cell Signaling (8:00–8:30, 11:15–12:15): Exosomes cycle to naïve epithelial cells to offer their protective cargo.
  • Innate Immune Response (9:55–10:20): Antimicrobial peptide (AMP), nitric oxide, production upregulated upon stimulation and are packaged in exosomes
  • Bactericidal (10:04 ): Kills bacteria–in this context, the antimicrobial peptides, colistin, and nitric oxide.
  • Gene Expression (13:44–13:54; 14:40–15:00): This was mentioned but not discussed at length, but they were looking at genes that were ‘off’ versus ‘on’. A gene that is ‘on’ is making protein. An example they discussed was the nitric oxide synthetase gene.
  • Antibiotics (14:55–16:33): Molecules that prevent the growth or kill bacteria.  In this paper, they used colistin, a surface acting antibiotic that targets gram negative organisms.
  • Evolution of Immunity (22:50–23:10): The innate immune system evolved early; adaptive, antigen-specific responses evolved later.

5.2. Main Paper

  • Microbiome (25:55–26:37; 33:00–34:00; 37:04–37:14; 41:25–42:30):The gut microbiome is composed of “good” and “bad” bacteria, and its composition is partly shaped by diet (e.g. Lactobacilli are associated with healthy foods, and Clostridia are associated with “danger”),  Waste products of bacteria that were treated with exosomes can inhibit growth of other bacteria.
  • Colitis (27:00 –  27:42; 46:32–51:30): Colitis is an inflammation of the gut,  Mice that were fed ginger exosomes were protected against DSS induced colitis.
  • Human Response to Environmental Contamination (27:02–27:45): Humans have a cellular pathway called aero-hydrocarbon receptor pathway (AHR) that is activated when we are exposed to halogenated or polycyclated aromatic hydrocarbons (PAHs). IL-22 is induced by AHR which suppresses inflammation.
  • Cytokines (27:25–27:45; 49:29–54:45): IL-22 is a beneficial cytokine that suppresses inflammation that is discussed at length. Other cytokines and chemokines typically involved in colitis include TNFalpha and IL1 beta.
  • microRNA (29:48–31:18; 43:39–43:48): Typically 21 nucleotides long, microRNAs are the main component within the ginger derived exosome like nanoparticles (ELNs) found when they sequenced them. microRNA regulates gene expression by targeting mRNA and degrading it.
  • Exosomes: (29:50–32:40; 39:50–40:03; 42:44–44:39): Exosomes from different plants (ginger, in this study is the focus) end up in different organs when ingested, likely due to their lipid composition. Probiotic/ prebiotic research is focusing on exosomes as vehicles to deliver microRNAs to the targeted gut bacteria  The varying lipid composition is responsible for targeting specific bacteria. Plant exosomes deliver microRNA that can alter gene expression in targeted bacteria.
  • SOS Response (44:35–46:20): LexA maintains the SOS response inactive until it is cleaved by RecA. Plant exosome microRNA can target LexA, reducing its expression.
  • Bacterial Structures (54:50 – 57:05) LGG (Lactobacillus rhamnosus) don’t migrate in the mice that were fed the exosomes. Bacterial motility was impaired due to the down-regulation of a pilus specific protein called SPAC.

6. Podcast Questions

  1. Define exosome.
    1. Small fluid filled sacs that are released from the cell in the lining of the nasal passage.
    2. Small fluid filled sacs that contain tetraspanin and have a lipid bilayer that has a transmembrane 4 superfamily type canonical structure. They are released from the cell in the lining of the nasal passage.
    3. Small fluid filled sacs that contain connexins and a lipid bilayer that has a transmembrane 4 superfamily type canonical structure. They are released from the cell in the lining of the nasal passage.
    4. Small fluid filled sacs that contain tetraspanin and a lipid bilayer that has a transmembrane 4 superfamily type canonical structure. They are released from the cell in the lining of the trachea and bronchi.
  2. Order the events in exosome release and immune activation.
    1. Innate immune system activated
    2. Nasal cells detect LPS, a PAMP
    3. Microbes are killed
    4. TLR4 activated
    5. Exosomes released
    6. Adaptive immune system activated
    1. b, d, a, e, c
    2. b, e, a, d, c
    3. f, e, d, a, b
    4. b, a, d, c, e
    5. e, a, d, b, c

3. A patient is experiencing a respiratory infection caused by a bacterial infection. The doctor prescribes a nasal decongestant to alleviate symptoms of the infection. What effect will these nasal decongestants have on exosome action in nasal passages?

    1. decongestants will help exosome action.
    2. decongestants will help exosome action.
    3. decongestants will have no effect on exosome action.

4. Given findings within this podcast; that Lactobacillus growth is promoted by ginger exosomes, Yun Teng’s daughter Lucy conducted a science fair experiment to determine the survivability of Lactobacillus cultures in acidic media (mimicking the stomach acid environment). Based on the results of her study and her thoughts afterwards, what advice might a medical professional give to a patient who had their gut microbiome wiped out by antibiotics?

    1. Eat yogurt containing Lactobacillus cultures on an empty stomach.
    2. Eat yogurt containing Lactobacillus cultures with food.
    3. Eat yogurt containing Lactobacillus cultures with milk.
    4. Take a probiotic containing Clostridia and Ruminococcus bacteria on an empty stomach.

5. Unidentified bacterial exosomes are provided to you, and you are asked to determine what is the most likely bacterial family they are from. You decide to feed them garlic, grapefruit, and turmeric. Results: you saw uptake of garlic and grapefruit but not turmeric. What is your determination of the bacterial family for the unidentified exosomes? Select all that apply.

    1. Lactobacilliaceae
    2. Clostridiaceae
    3. Ruminococcaceae
    4. Bacteroidales

7. Figure Reading Exercises

The following are two figure reading exercises, one from the snippet (Figure 1) and one main paper (Figure 1).

7.1. First Figure Reading Exerise

7.1.1. Learning Objectives

Students will be able to:

  • Determine correlation from a graph.
  • Estimate size in an electron micrograph image
  • Analyze data to determine the best workflow stopping point considering time, money, purity, and recovery.
  • Defend an experimental result with evidence.
Experimental Background (Nocera et al., Figure 1)

Inhalation is a common route for pathogens to enter the body.  The nasal cavity is therefore a prime location for immune defense.  Exosomes are small vesicles containing a variety of immune defense proteins that Nocera et al. (2019) are interested in identifying and characterized from this prime immune environment.   First, they show data in which they defend their choice of using a single marker for identifying exosomes.  Using Enzyme Linked Immunosorbent Assay (ELISA), they compared levels of CD81 to CD63, two common exosome proteins (panel A).   Next, they isolated primary nasal tissue and mucus in order to purify nasal exosomes,  which they term this Nasal Derived Mucosal Exosomes (NDME).  The researchers quantified levels of common exosome or known contaminant components at various points in their preparation.  They tested whole mucus, mucus subjected to ultracentrifugation (UCF), and mucus subjected to one of three different density gradient centrifugation levels (OptiPrep) for calnexin (panel B) and CD81 (panel C).  An important confirmation to make is that the vesicles purified and identified as nasal exosomes are near the size expected for exosomes.  The researchers do this using electron microscopy and gold labeling for CD81 (panel D).

7.1.2. Questions

  1. What is the correlation between CD63 and CD81 and what does this indicate?
    1. It is a positive correlation; when CD63 is present, CD81 is also present showing CD81 is a good exosome marker.
    2. It is a negative correlation; when CD63 is present, CD81 is absent showing CD63 is a good exosome marker.
    3. There is no correlation; when CD63 is present, CD81 is also present, so neither can be used alone as markers.
    4. There is weak positive correlation; when CD81 is present, CD63 is sometimes present, so it is a useful marker.
  2. Contaminant (calnexin; panel B) and exosome marker (CD81; panel C) are shown for different steps of the exosome preparation protocol.  What is the best prepration taking into consideration the need to use money and time wisely without sacrificing purity and yield?
    1. Whole mucus prep
    2. Ultracentrifugation (UCF)
    3. OptiPrep 1.096 g/mL
    4. OptiPrep 1.11 g/mL
    5. OptiPrep 1.15 g/mL
  3. An important confirmation to make is that the vesicles purified and identified as nasal exosomes have features consistent with known exosomes. The researchers do this using electron microscopy and gold labeling for CD81 (panel D).  Here, let’s estimate the size of the vesicle in the NEG image. The black bar (ruler) is 100 nm long.
    1. 10 nm
    2. 50 nm
    3. 100 nm
    4. 500 nm
  4. What features in the electron micrographs support that Nocera et al. (2019) successfully isolated NDME?
    1. The size and shape are consistent with known exosome size and shape.
    2. The high level of CD63 and CD81 inside the exosome structure.
    3. The presence of CD81, as detected by gold labeling, inside the exosome.
    4. The fact that CD63 is absent and CD81 is present inside the exosome.

7.2. Second Figure Reading Exercise

7.2.1. Learning Objectives

Students will be able to:

  • Identify microbial abundances in biodiversity stacked bar charts.
  • Interpret qPCR results as it relates to bacterial abundance.
  • Differentiate bacterial families distinct and/or enriched in microbiome compositions
  • Defend or refute an experimental result with evidence.
Experimental Background (Teng et al., Figure 1 ABD)

The gut microbiome plays a significant role in health, particularly in nutrition and immune function.  Diet affects the gut microbiome regardless of age, but humans have vastly different diets across the globe.  To investigate the role of specific diet elements on microbiomes of adults, Teng et al. (2018) populated the gut of mice with microbiome flora of human volunteers between the ages of 25-46 and fed a group of each either saline (PBS) as a control  or ginger-embedded plant exosomes-like nanoparticles (GELNs).  Bacterial DNA from feces was analyzed using 16S rDNA sequencing to determine microbiome composition (panel A) or specific bacterial abundance (panel B left), quantitative PCR (qPCR; panel B right). To determine whether the mouse model was an accurate representation in human then the examined the microbiome compositions of humans fed GELN or Saline (panel D).

7.2.2. Questions

  1. Which bacterial family is highly enriched in mice fed ginger exosome like nanoparticles (GELN) compared to the mice fed PBS (panel A)?
    1. Clostridiaceae
    2. Ruminococacceae
    3. Clostridiales
    4. Lactobacilliaceae
    5. S24-7
  2. The authors saw a marked decrease in Clostridiaceae in the GELN-fed mice compared to PBS-fed mice, what specific result(s) support(s) this the best?
    1. Figure 1A abundance bar charts
    2. Figure 1B 16S rDNA sequencing
    3. Figure 1B qPCR verification
    4. All of the above
  3. What does the upward direction of the bar (from the 0 point) indicate for the qPCR results (panel B right)?
    1. A positive fold change indicating higher abundance in GELN compared to PBS-fed mice.
    2. A negative fold change indicating higher abundance in PBS compared to GELN-fed mice.
    3. No fold change indicating the GELN and PBS-fed mice have equivalently low values.
    4. No fold change indicating the GELN and PBS-fed mice have equivalently high values.
  4. Experiments in panels A-C focus on a mouse model. The researchers next tested their hypothesis in humans by feeding humans GELN or Saline (panel D). Do these data indicate that the mouse model was accurate for humans? What is your evidence?
    1. Lactobacilliaceae decreased, Clostridiaceae increased, Bacteroidaceae decreased, and Ruminococcaceae increased.
    2. Lactobacilliaceae increased, Clostridiaceae decreased, Bacteroidaceae increased, and Ruminococcaceae decreased.
    3. Lactobacilliaceae increased, Clostridiaceae decreased, Bacteroidaceae decreased, and Ruminococcaceae increased.
    4. Lactobacilliaceae, Clostridiaceae, Bacteroidaceae, and Ruminococcaceae all increased.
    5. Lactobacilliaceae, Clostridiaceae, Bacteroidaceae, and Ruminococcaceae all decreased.

8. Paper Information and Licensing

8.1. Snippet paper

  • Nocera AL, Mueller SK, Stephan JR, Hing L, Seifert P, Han X, Lin DT, Amiji MM, Libermann T, Bleier BS. 2019. Exosome swarms eliminate airway pathogens and provide passive epithelial immunoprotection through nitric oxide. J Allergy Clin Immunol. 143(4):1525-1535.e1. doi: 10.1016/j.jaci.2018.08.046.
  • This article is not licensed for Creative Commons use; thus, the abstract and figures cannot be copied here. Please see the article at the journal’s web page: https://www.jacionline.org/article/S0091-6749(18)31351-4/fulltext

8.2. Main paper

  • Teng Y, Ren Y, Sayed M, Hu X, Lei C, Kumar A, Hutchins E, Mu J, Deng Z, Luo C, Sundaram K, Sriwastva MK, Zhang L, Hsieh M, Reiman R, Haribabu B, Yan J, Jala VR, Miller DM, Van Keuren-Jensen K, Merchant ML, McClain CJ, Park JW, Egilmez NK, Zhang HG. 2018. Plant-Derived Exosomal MicroRNAs Shape the Gut Microbiota. Cell Host Microbe. 24(5):637-652.e8. doi: 10.1016/j.chom.2018.10.001.
  • This article is not licensed for Creative Commons use; thus, the abstract and figures cannot be copied here. Please see the article at the journal’s web page: https://www.cell.com/cell-host-microbe/fulltext/S1931-3128(18)30523-7?

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Podcast Annotation and Resources in Microbiology Copyright © 2025 by Rebecca Seipelt-Thiemann; Nancy Boury; Gwendowlyn S. Knapp; Amaya Garcia Costas; and Patrick Armstrong is licensed under a Creative Commons Attribution 4.0 International License, except where otherwise noted.

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