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• The microbial world is a complex place - How to make sense of “diversity”?
• The molecular view of the natural microbial world.
• Sampling the indoor environment – aerosols.
Kim Ross
Kim Ross
Making Sense of Sequences: Molecular Phylogeny
1. Align sequences so that “homologous” residues are juxtaposed.
2. Count the number of differences between pairs of sequences; this is some measure of “evolutionary distance” that separates the organisms.
3. Calculate the “tree”, the relatedness map, that most accurately represents all the pairwise differences.
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“Metagenomics”
Sample DNA rDNA PCR library clone sequence
“ “ “ Next-gen multiplex
Expansion of the Bacterial Tree
Strain A
Strain C
E.g. Gene contents of different “strains” of “Escherichia coli”
Strain B
Indoor Microbiology Project (sponsored by A.P. Sloan Fndn.)
Laura Baumgartner • NYC Subway system aerosols (7 stations, 3 connected sites, multiple seasons.
Piret Koll • School aerosols (3 in Denver–Boulder area) • Homeless shelter aerosols (2 in Denver–Boulder area) • Health setting aerosols (2 hospitals, 1 clinic in CO) • Hotel HVAC (1, Atlanta) • Office building aerosols (2, Atlanta) • University setting aerosols (3 in TX, several in CO)
Kim Ross • Municipal water distribution, source-to-POU (many
samples in Boulder and Centennial systems, some overlap with a concurrent 454 source water survey.
Kristen Peterson • Public swimming facility aerosols and biofilms (5 in CO,
2 in TX)
Sampling aerosols: NON-TRIVIAL!
Evolution of the Sceptor Industries Omni 3000
What are we finding generally?
• The microbiology of the human environment phylogene:cally is rela:vely simple, mainly only ~5 of ~100 known phyla. Few “candidate phyla” in ~100K sequences.
• Sequences are seldom iden:cal to a previously determined sequence, but many “bin” with cultured species at the “genus” level (95%) or even “species” level (97%).
• Every site sampled differs in the detail, but generali:es emerge for each kind of environment.
• Indoor aerosol microbiology is significantly/mainly human-‐shed microbiology.
• Water microbiology is idiosyncra:c with source and system, and in some seSngs is of poten:al health concern.
How similar are our samples to the Human Microbiome?
Where in the home? -
Any place moist
Who’s in your showerhead?
Mycobacterium avium Positive -cause of MAC pulmonary infections -have seen in 35/54 Showerheads analyzed
VSM5Q1u showerhead swab
N=85
Mycobacterium gordonae
Mycobacterium otherMethylobacterium
sp.
Delftia acidovorans
Escherichia coli
Sphingomonas yanoikuyae
Staphylococcus epidermidis
Staphylococcus capitis
Moraxella osloensisDietzia maris
Streptococcus oralis
Mycobacterium avium
Blastomonas natatoria
BSD2Q1ushowerhead swab
N=81
Mycobacterium gordonae
7%
Flavobacterium2%
Methylococcaceae2%
Novel Group2%
Chlamydiae1%
Sphingomonas16%
Uncultured Beta Proteo13%
Bacteroidetes11%
Bacillus7%
Methylocystis34%
M. avium Negative
Examples of Showerhead Biofilm Swab Results
Water/Showerhead Swab Comparison
VSM5Q1u showerhead swab
N=85
Mycobacterium gordonae
Mycobacterium otherMethylobacterium sp.
Delftia acidovorans
Escherichia coli
Sphingomonas yanoikuyae
Staphylococcus epidermidis
Staphylococcus capitis
Moraxella osloensisDietzia maris
Streptococcus oralis
Mycobacterium avium
Blastomonas natatoria
VPM5W1u, water from pipe pre-showerhead
N=86
Methylobacter psychrophilus
Blastomonas natatoria
Mycobacterium avium
Novosphingobium lentum
Escherichia coli
Methylobacterium sp.
Mycobacterium gordonae
VSM5W1u, water collected post-showerhead
N=90
Methylobacterium sp.
Methylobacter psychrophilus
Novosphingobium sp.
Sphingomonas sp.Rhodobacter blastica
Bdellovibrio bacteriovorus
Blastomonas natatoriaLegionella waltersii
Mycobacterium gordonae
Dir. of Water Flow
