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Revealing microbial recognition by specific antibodies.

Simón-Soro Á, D'Auria G, Collado MC, Džunková M, Culshaw S, Mira A - BMC Microbiol. (2015)

Bottom Line: We show the power of the technique in human faecal, saliva, oral biofilm and breast milk samples, labeled with fluorescent anti-IgG or anti-IgA antibodies.Using Fluorescence-Activated Cell Sorting (FACS), bacterial cells were separated depending on whether they are coated with IgA or IgG antibodies.Each bacterial population was PCR-amplified and pyrosequenced, characterizing the microorganisms which evade the immune system and those which were recognized by each immunoglobulin.

View Article: PubMed Central - PubMed

Affiliation: Department of Health and Genomics, FISABIO Foundation, Center for Advanced Research in Public Health, Avda. Cataluña 21, 46020, Valencia, Spain. simon.aurea@gmail.com.

ABSTRACT

Background: Recognition of microorganisms by antibodies is a vital component of the human immune response. However, there is currently very limited understanding of immune recognition of 50 % of the human microbiome which is made up of as yet un-culturable bacteria. We have combined the use of flow cytometry and pyrosequencing to describe the microbial composition of human samples, and its interaction with the immune system.

Results: We show the power of the technique in human faecal, saliva, oral biofilm and breast milk samples, labeled with fluorescent anti-IgG or anti-IgA antibodies. Using Fluorescence-Activated Cell Sorting (FACS), bacterial cells were separated depending on whether they are coated with IgA or IgG antibodies. Each bacterial population was PCR-amplified and pyrosequenced, characterizing the microorganisms which evade the immune system and those which were recognized by each immunoglobulin.

Conclusions: The application of the technique to healthy and diseased individuals may unravel the contribution of the immune response to microbial infections and polymicrobial diseases.

No MeSH data available.


Related in: MedlinePlus

Diversity of Ig-coated and uncoated bacteria in human saliva. Saliva samples collected 24 h after toothbrushing (n = 16) were stained with fluorescent markers for bacterial DNA, IgA and IgG, and sorted in three groups: IgA-coated bacteria, IgG-coated bacteria and uncoated, non-opsonized bacteria. a Bacterial composition at the genus level for total saliva samples, as well as for the IgA- and IgG-coated fractions. The bacterial composition appeared to be different between the different fractions. b Rarefaction curves relating pyrosequencing effort to the estimated number of species (OTUs at 97 % sequence identity). The non-opsonized fractions display a lower diversity and different taxonomic composition to opsonized populations
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Fig5: Diversity of Ig-coated and uncoated bacteria in human saliva. Saliva samples collected 24 h after toothbrushing (n = 16) were stained with fluorescent markers for bacterial DNA, IgA and IgG, and sorted in three groups: IgA-coated bacteria, IgG-coated bacteria and uncoated, non-opsonized bacteria. a Bacterial composition at the genus level for total saliva samples, as well as for the IgA- and IgG-coated fractions. The bacterial composition appeared to be different between the different fractions. b Rarefaction curves relating pyrosequencing effort to the estimated number of species (OTUs at 97 % sequence identity). The non-opsonized fractions display a lower diversity and different taxonomic composition to opsonized populations

Mentions: The sum of the proportion of IgA- and IgG coated bacteria for the same samples was in most cases higher than 100 %, indicating that a large proportion of cells were coated by both immunoglobulins, as suggested by other studies [16, 17]. However, the bacterial composition in the IgA- and IgG-coated fractions was different (Fig. 5). As shown by a Principal Coordinates Analysis in saliva samples, the composition of the IgA- and IgG-opsonized fractions did not cluster together. In addition, the opsonized and non-opsonized populations occupied different positions in the PCoA space, indicating that the taxonomic groups coated by antibodies are different from those that are ignored or undetected by them. For instance, the genera Delftia or Propionibacterium were found at low frequencies in the saliva samples, but were highly opsonized by both IgA and IgG (Fig. 5a). Streptococcus, on the other hand, was very frequent in the same saliva samples, but moderately opsonized. Differential opsinization was found for Haemophilus, which was found to be coated by IgG but not IgA (Fig. 5a). Data for saliva samples from individual CA021 show that the most common genera in the IgA-opsonized fraction were Propionibacterium, Streptococcus, Arthrobacter, Veillonella and Atopostipes, whereas the latter three were absent in the non-opsonized fraction (Fig. 4). In the future, the sequencing of IgA-, IgG- and IgM-coated microbes in larger numbers of samples should confirm whether there is Ig-specific opsonization.Figure 5


Revealing microbial recognition by specific antibodies.

Simón-Soro Á, D'Auria G, Collado MC, Džunková M, Culshaw S, Mira A - BMC Microbiol. (2015)

Diversity of Ig-coated and uncoated bacteria in human saliva. Saliva samples collected 24 h after toothbrushing (n = 16) were stained with fluorescent markers for bacterial DNA, IgA and IgG, and sorted in three groups: IgA-coated bacteria, IgG-coated bacteria and uncoated, non-opsonized bacteria. a Bacterial composition at the genus level for total saliva samples, as well as for the IgA- and IgG-coated fractions. The bacterial composition appeared to be different between the different fractions. b Rarefaction curves relating pyrosequencing effort to the estimated number of species (OTUs at 97 % sequence identity). The non-opsonized fractions display a lower diversity and different taxonomic composition to opsonized populations
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4489363&req=5

Fig5: Diversity of Ig-coated and uncoated bacteria in human saliva. Saliva samples collected 24 h after toothbrushing (n = 16) were stained with fluorescent markers for bacterial DNA, IgA and IgG, and sorted in three groups: IgA-coated bacteria, IgG-coated bacteria and uncoated, non-opsonized bacteria. a Bacterial composition at the genus level for total saliva samples, as well as for the IgA- and IgG-coated fractions. The bacterial composition appeared to be different between the different fractions. b Rarefaction curves relating pyrosequencing effort to the estimated number of species (OTUs at 97 % sequence identity). The non-opsonized fractions display a lower diversity and different taxonomic composition to opsonized populations
Mentions: The sum of the proportion of IgA- and IgG coated bacteria for the same samples was in most cases higher than 100 %, indicating that a large proportion of cells were coated by both immunoglobulins, as suggested by other studies [16, 17]. However, the bacterial composition in the IgA- and IgG-coated fractions was different (Fig. 5). As shown by a Principal Coordinates Analysis in saliva samples, the composition of the IgA- and IgG-opsonized fractions did not cluster together. In addition, the opsonized and non-opsonized populations occupied different positions in the PCoA space, indicating that the taxonomic groups coated by antibodies are different from those that are ignored or undetected by them. For instance, the genera Delftia or Propionibacterium were found at low frequencies in the saliva samples, but were highly opsonized by both IgA and IgG (Fig. 5a). Streptococcus, on the other hand, was very frequent in the same saliva samples, but moderately opsonized. Differential opsinization was found for Haemophilus, which was found to be coated by IgG but not IgA (Fig. 5a). Data for saliva samples from individual CA021 show that the most common genera in the IgA-opsonized fraction were Propionibacterium, Streptococcus, Arthrobacter, Veillonella and Atopostipes, whereas the latter three were absent in the non-opsonized fraction (Fig. 4). In the future, the sequencing of IgA-, IgG- and IgM-coated microbes in larger numbers of samples should confirm whether there is Ig-specific opsonization.Figure 5

Bottom Line: We show the power of the technique in human faecal, saliva, oral biofilm and breast milk samples, labeled with fluorescent anti-IgG or anti-IgA antibodies.Using Fluorescence-Activated Cell Sorting (FACS), bacterial cells were separated depending on whether they are coated with IgA or IgG antibodies.Each bacterial population was PCR-amplified and pyrosequenced, characterizing the microorganisms which evade the immune system and those which were recognized by each immunoglobulin.

View Article: PubMed Central - PubMed

Affiliation: Department of Health and Genomics, FISABIO Foundation, Center for Advanced Research in Public Health, Avda. Cataluña 21, 46020, Valencia, Spain. simon.aurea@gmail.com.

ABSTRACT

Background: Recognition of microorganisms by antibodies is a vital component of the human immune response. However, there is currently very limited understanding of immune recognition of 50 % of the human microbiome which is made up of as yet un-culturable bacteria. We have combined the use of flow cytometry and pyrosequencing to describe the microbial composition of human samples, and its interaction with the immune system.

Results: We show the power of the technique in human faecal, saliva, oral biofilm and breast milk samples, labeled with fluorescent anti-IgG or anti-IgA antibodies. Using Fluorescence-Activated Cell Sorting (FACS), bacterial cells were separated depending on whether they are coated with IgA or IgG antibodies. Each bacterial population was PCR-amplified and pyrosequenced, characterizing the microorganisms which evade the immune system and those which were recognized by each immunoglobulin.

Conclusions: The application of the technique to healthy and diseased individuals may unravel the contribution of the immune response to microbial infections and polymicrobial diseases.

No MeSH data available.


Related in: MedlinePlus