Transcript Microbial diversity and virulence probing of five different body sites

Microbial diversity and virulence probing of five different body sites
Anu Rebbapragada, Pub. Health Ontario Central Lab. Canada
Wei-Jen Lin, Cal State Polytech. Univ. Pomona, USA
Thomas Keller, Oregon Health &Science Univ., USA
Anna Edlund, JCVI, USA
Questions:
-What are the differences in taxonomic & functional
diversity between and within body sites?
-Is there a difference in the diversity of virulence genes
between body sites?
Sample selection in IMG/HMP
- Downloaded 10 metagenomes
- Selected 2 metagenomes per body site
Same person analyzed for
skin and stool sample
Table for General Statistics
•Compare Genomes
•Genome Statistics
•Breakdown by selected
genomes, general statistics
Bar Graph for % GC
•Created in Excel
Radial Tree by Class
Radial Tree
•Compare Genomes
•Phylogenetic
Distribution
•Radial Tree of 5
selected
metagenomes
•Customize by
Phyla, Class,
Domain etc
categories
Bacterial diversity at class level
14
5
Skin
Pie charts and tables
created by clicking
node on Radial Tree
2
11
4
Nose
Diversity of Archaea by Phyla and Class levels
Pie charts and tables created by clicking
node on Radial Tree. Radial tree customized
to display at either Phyla or Class level.
Bacterial diversity behind your ear
Left ear
Selected only 2 metagenomes (ear samples) to generate radial tree.
Pie charts and tables created by clicking node on Radial Tree.
Radial tree customized to display at either Phyla or Class level.
Right ear
Functional diversity
across the metagenomes
Compare Genomes
Genome Clustering, Correlation Matrix
Select metagenomes of interest
Clustering method: Correlation Matrix
Function: COG
COG functions
Display: r2 values by pairwise
comparison across all metagenomes
PCA 2-D COG groups
Compare Genomes
Genome Clustering, Correlation Matrix
Stool
Select metagenomes of interest
Clustering method: PCA Principle
component analysis
Tongue
Function: COG (clustering by COG
pathways)
Skin
Nose
Display: 2D plot, mouse over redX to
identify the metagenome identity
Vagina
Created figure in powerpoint to show
clustering of COGs across
metagenomes
•Can also show Hierarchical clustering
by Function (COG, Pfam etc) or
Taxonomy (Genus, Family etc) or
Function Categories (COG, KEGG,
Pfam)
PCA COG pathways
Nose
Compare Genomes
Genome Clustering,
Correlation Matrix
Select metagenomes
of interest
Clustering method:
PCA Principle
component analysis
Tongue
Function Category:
COG pathway
Skin
Stool
Now the Nose samples don’t cluster together
suggesting differences in pathways
Vagina
Display: 2D plot,
mouse over redX to
identify the
metagenome identity
Created figure in
powerpoint to show
clustering of COGs
across metagenomes
Hierarchical Clustering across Metagenomes
by COG
Compare Genomes
Genome Clustering,
Correlation Matrix
Select metagenomes
of interest
Clustering method:
Hierarchical
Function: COG
Display: Phylogram
showing distribution of
metagenomes
Compare COG function
between Metagenomes
Compare genomes
•function comparison tool
•abundance profile
•function comparison
•select query genome
•Export table
Secretion Pathways
Select Metagenomes of
interest
Find Functions
Search All pathways, Select
KEGG pathways only
Type “secretion” as
Keyword
Click on Pathway OID for
bacterial secretion
View Map
Gene Products in
Type III Secretion pathway
Select Metagenomes of interest
Find Functions, Function Search
Keyword: “Secretion”, Gene Product name, Click GO
Text Filter: “Type III”, click Apply
Select MetaGenomes
View Profile
Output: Table of Function ID (gene product name) and abundance (gene
counts) in each metagenome
Gene Products in
Type Vb secretion (hemolysin) pathway
Select Metagenomes of interest
Find Functions, Function Search
Keyword: “Secretion”, Gene Product name, Click GO
Text Filter: “ShlB”, click Apply
Select MetaGenomes
View Profile
Output: Table of Function ID (gene product name) and abundance (gene
counts) in each metagenome
Gene Counts Across Metagenomes for Type VI Secretion Pathway
Distribution of antibiotic resistance genes by
human body sites
Nose
Ears
Stool
Tongue Vagina
Beta-lactamase
Cephalosporin resistance
Vancomycin resistance
Aminoglycoside resistance
Tetracycline resistance
Select Metagenomes of interest
Find Functions, Function Search
Keyword: individual searches for each antibiotic resistance category under
Gene Product name, Click GO
Output: Table of Function ID (gene product name) and abundance (gene
counts) in each metagenome
Exported Table, Tally of totals in spreadsheet and created PieCharts in Excel
Erythromycin resistance
Chloramphenicol resistance
Quinolone resistance
Antibiotic resistance in human body sites
Normalized to counts/10,000 genes
45
40
Beta-lactamase
35
Cephalosporin resistance
30
Vancomycin resistance
25
Aminoglycoside resistance
20
Tetracycline resistance
15
Erythromycin resistance
10
Chloramphenicol resistance
5
Quinolone resistance
0
Nose 1
Nose 2
Ear Left
Ear Right
Stool 1
Stool 2
Tongue 1
Tongue 2
Vagina 1
Vagina 2
US Antibiotic Usage Data
Google search :)
www.cddep.org/ResistanceMap
Study Limitations
&
Future Analytical Directions