Bacterial Assemblage Differences SAME14 between Urban ... · crobial communities and urbanization....
Transcript of Bacterial Assemblage Differences SAME14 between Urban ... · crobial communities and urbanization....
Ryan J. Newton Sandra L. McLellan
Bacterial Assemblage Differences between
Urban-impacted Waterways & Oligotrophic Lake Michigan
BACKGROUND PRIMARY FINDINGS
CENTRAL QUESTION
THE RESULTS
Linwood
2 Mile 5 Mile
Bradford
McKinley
Kinnickinnic River
Menomonee River
Milwaukee River
N
Junction
3.5 Mile1 Mile
Doctor’sIn
Doctor’sMid
Doctor’sOut
Harbor wallsSampling site
How does urbanization interface with & impact aquatic microbial communities? Lake Michigan
Urban Estuary and Lake Michigan Communities are Distinct
Stormwater - A Unique Urban Bacterial Assemblage
to contact, email - [email protected]
ACKNOWLEDGEMENTS & REFERENCES
Within-Genus Habitat Specialization
OLIGOTYPING EXPLAINED1
www.uwm.edu/freshwater
Urbanization alters watershed ecosystem functioning through the movement, magnitude, and content of surface water runo�. However, there is still much to be learnt about the relationship between aquatic mi-crobial communities and urbanization. For example: What are the microbial biodiversity patterns associ-ated with urban areas? and Do urbanized waterways promote the maintenance of organisms that impact human health or well-being? With urban areas rapidly expanding and cities looking to become more sustainable, it will be important to integrate aquatic microbial ecology into the development of urban ecological theory and practice.
Here we aim to further our understanding of the bacterial connectivity between urban discharge and surface waters and the in�uence urbanized waterways have on community assemblages. We examine the prevalence of bacteria from urban stormwater in receiving water bodies and identify community assem-blage di�erences between a heavily urbanized estuary and a low urban-impacted oligotrophic lake.
We used massively-parallel sequencing (454 and illumina) of bacterial 16S rRNA genes (v6 region) ampli�ed from whole-community DNA extractions to characterize and compare the surface water bac-terial assemblages from stormwater, Milwaukee’s estuary, and nearshore Lake Michigan.
Stormwater area
Stormwater area 5,000 meters
Top left: Milwaukee estuary channel, looking toward Lake Michigan. Bottom left: Estuary discharge into Lake Michi-gan. Right: Map of sampling locations.
SAME14
acI-A
acI-B
Sediminibacterium
LD12
Limnohabitans
Polynucleobacter 1 10 100 1000 10000
110
100
1000
1000
0
Count in Lake Michigan Samples (abundance + 1)
Coun
t in
Urb
an E
stua
ry S
ampl
es
(abu
ndan
ce +
1)
-0.4 -0.2 0.0 0.2 0.4
-0.3
-0.2
-0.1
0.0
0.1
0.2
0.3
MDS1
MD
S2
Lake Michigan
Urban Estuary
Stormwater
NMDS plot of the bacterial taxonomic composition in samples from Lake Michigan (blue), the Milwaukee es-tuary (green), & Milwaukee stormwater outfalls (gold). Taxonomic a�liations for sequence reads were assigned with GAST5. Stress=0.03, dim. = 5
Minimum entropy decomposition2 was used to cluster se-quences into oligotypes/OTUs. LEfSe3 was then used to identify biomarkers (i.e. OTUs) that di�erentiated the urban stormwater environment from the other aquatic envi-ronments4. �e distribution of those biomarker OTUs are shown below in a boxplot (min, median, max, 1st, & 3rd quartiles indicated) & barplot (mean & std. dev. depicted).
20%
15%
10%
5%
0%
Between Enivronments
*
Urban EstuaryPreferred
Lake Michigan Preferred
4 2 2 4 6 8 10 121
**
*
*
*
*
*
AbundanceRatio
acI-AacI-BacI-C
acI-TH1acI-STL
AquilunaArcicella
AlgoriphagusFlavobacterium
FluviicolaSediminibacterium
LD12Sphingopyxis
HydrogenophagaLimnohabitans
PolynucleobacterRhodobacterLuteolibacter
Sample EnvironmentUrban EstuaryLake Michigan
Mann-Whitney U
68 ± 4037 ± 14
379*
Inverse Simpson Tail6
584 ± 463145 ± 85
370*
Mean & standard deviation are reported for diversity measures. Sequence data was grouped by closed-reference clustering. * indicates p<0.01
Right: Magnitude of a “habitat preference” for common freshwater genera/lineages. Habitat preference is deter-mined by the ratio of the mean relative abundance of each genus/lineage in the urban estuary samples vs. the Lake Michigan samples. Samples are normalized to max abundance for each sample transect. An asterisk indicates a signi�cant association with habitat (Mann-Whitney U test, p<0.01).
Alpha Diversity Estimates Habitat Preference
Urban Estuary Lake Michigan GeneralistSpecialist - Urban EstuarySpecialist - Lake MIRare
Left: Bar plot of oligotype composition within common freshwater genera/clades. 6 of 18 examined genera/clades are illustrated. Not depicted: acI-C, acI-TH1, acSTL, Aquiluna, Arcicella, Algoriphagus, Flavobacterium, Fluviicola, Sphin-gopyxis, Hydrogenophaga, Rhodobacter, & Luteolibacter. Right: Oligotype habitat preference classi�cation results derived from the CLAM7 statistical approach, which implements a multinomial model for group classi�cation.
Important Points:• 80 of 351 oligotypes classi�ed as habitat specialists 51 for urban estuary and 29 for Lake Michigan
• Numerous urban estuary specialists from: Flavobacterium, Hydrogenophaga, Limnohabitans, & Rhodobacter
• Numerous Lake Michigan specialists from: acIA, acIB, acIC, acTH1, & Fluviicola
• 7 of 18 genera/lineages harbored oligotypes with ≥96% sequence identity, but opposite habitat specialization
4. For more info see Fisher et al., (2015) Elementa 3:000064
5. Huse et al., (2008) PLoS Genet. 4.6. Li et al., (2012) PLoS ONE 7(6):e32118.
1. Modi�ed from: Eren et al., (2013), Method. Ecol. Evol. 4:1111-2. Eren et al., (2015) ISME J. 9:968- 3. Segata et al., (2011) Genome Biol. 12
We would like to thank the crew of the R/V Neeskay and everyone in the McLellan lab who has contributed to sampling e�orts. We also thank Mitch Sogin’s group at the Marine Biological Laboratory for DNA sequening and data processing assistance and Jen Fisher for her work on the stormwater data.
DatasetGAACCTCTAAGAAAGGTTGAACCTCTAAGAAAGGTTGAACCACTAAGAAACGTTGAACCACTAAGAAACGTTGAACCTCTAAGAAAGGTTGAACCTCTAAGAAAGGTTGAACCACTAAGAAACGTTGAACCCCGGAGAAAGGTT
Sample 1
Sample 2
Entropy Analysis
GAACCTCTAAGAAAGGTT
1
Ent
ropy
OligotypingSample 1
0
Sample 2
OligotypesT A C
If further entropy
decomposition is needed
or
Stop
Repeat
• Urban stormwater systems contain a diverse & variable bacterial assemblage, but is an environment with its own identi�able bacterial community signature.
• Stormwater contributes signi�cantly to the organisms present in Milwaukee’s urban estuary bacterial assemblage (massive immigration - up to 20% of bacterial 16S rRNA genes).
• Milwaukee’s urban estuary community was distinct from that of Lake Michigan: 1. Estuary harbors a more diverse & variable community 2. Shifted numerical dominance among cosmopolitan freshwater lineages 3. Within-genus composition change - habitat specialization
Lake MI Estuary
% o
f bac
teria
l com
mun
ity
15%
10%
5%
0%Dry Rain
Estuary - Weather Scenarios
CSO
Dry = no rain in a 48 hr. period.CSO = sewage over�ow
-0.1
0.0
0.1
0.2
0.3
NM
DS
2
-0.2 -0.1 0.0 0.1 0.2MDS1
0.3
Lake Michigan
Oligotrophic lakes
Eutrophic lake
Urban Estuary
NMDS plot of oligo-type composition, for the 18 freshwater taxa mentioned above. Samples include three additional Wisconsin lakes (squares): Lake Mendota, Trout Lake, & Sparkling Lake.
SCHOOL OF FRESHWATER SCIENCES
What about other lakes?
• Further exploration - do urban areas select for genetic compositions or functional traits that di�er from un-impacted waters? or promote the maintenance of microbes that impact human health?
7. Chazdon et al., (2011) Ecology 92:1332-
Refs:
to download - scan