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, USAThomas Keller, Oregon Health &Science Univ., USAAnna 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

Table for General Statistics•Compare Genomes•Genome Statistics•Breakdown by selected genomes, general statistics

Bar Graph for % GC•Created in Excel

Same person analyzed for skin and stool sample

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 2 411

5

Skin

Nose

Pie charts and tables created by clicking node on Radial Tree

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 Right 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.

Functional diversity across the metagenomes

COG functions

Compare Genomes

Genome Clustering, Correlation Matrix

Select metagenomes of interest

Clustering method: Correlation Matrix

Function: COG

Display: r2 values by pairwise comparison across all metagenomes

PCA 2-D COG groups

Stool

Tongue

Skin

Nose Vagina

Compare Genomes

Genome Clustering, Correlation Matrix

Select metagenomes of interest

Clustering method: PCA Principle component analysis

Function: COG (clustering by COG pathways)

Display: 2D plot, mouse over redX to identify the metagenome identity

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

Stool

Tongue

Skin

Nose

Vagina

Compare Genomes

Genome Clustering, Correlation Matrix

Select metagenomes of interest

Clustering method: PCA Principle component analysis

Function Category: COG pathway

Display: 2D plot, mouse over redX to identify the metagenome identity

Created figure in powerpoint to show clustering of COGs across metagenomesNow the Nose samples don’t cluster together suggesting

differences in pathways

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 PathwaysSelect Metagenomes of interestFind FunctionsSearch All pathways, Select KEGG pathways onlyType “secretion” as KeywordClick on Pathway OID for bacterial secretionView Map

Gene Products in Type III Secretion pathway

Select Metagenomes of interestFind Functions, Function Search

Keyword: “Secretion”, Gene Product name, Click GO

Text Filter: “Type III”, click ApplySelect 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 interestFind Functions, Function Search

Keyword: “Secretion”, Gene Product name, Click GO

Text Filter: “ShlB”, click ApplySelect 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

Select Metagenomes of interestFind 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

Antibiotic resistance in human body sitesNormalized to counts/10,000 genes

US Antibiotic Usage Data

Google search :)www.cddep.org/ResistanceMap

Study Limitations &

Future Analytical Directions