ACTIVITY 2:

SIZE

AND

SCALE

MATTER!

Original drawings by John Tenniel

http://www.ca.gov/

Terrestrial environment: Soil Crumb

Photo and Diagram from Brock Biology of Microorganisms, Madigan and Martinko

Aquatic Environment:Marine “snow”

‘Hot spots’ of bacterial concentration and activity, contributing to global cycling of carbon and nutrients

Azam and Long, Nature, 2001

Sampling soil microbes in a relatively static soil community

100 cm200 cm

200 cm

A2 A3

A4 A5 A6

A7 A8 A9

A1

mer A

mer A

mer ACtl

Ctl

Ctl

A1-1 A1-2

A1-3 A1-4WT

A1

Reference Position}Populus deltoides type

wt = Wild Type

mer A = presence of mer A gene

Ctl = No Plants

Approximate soil sampling location

Isopleth - A

Plot Key (Enlarged to show detail)

wt

wt

wt

Plot ID

Water Direction

Water Direction

North Street

Ba

rnum

Co

urt

59.70 ft

187.46 ft

80.60 ft

20.30 ft

81.19 ft

17.91 ft

43.58 ft

53.13 ft

115.22 ft

Kohanza B

rook

Location of Proposed Phytormediation Cells

Intermediate Mercury Isopltth (29-50 ppm)

Low Mercury Isopltth (1-20 ppm)

High Mercury Isopltth (96-320 ppm)

(Isopleth areas are approximate)

Examples of microbial communities requiring different sampling considerations:

1.Small, isolated (non-interacting), moderate density, highly diverse (Drop-soda lakes, hypersaline mats)

2.Small, isolated but interacting, high density, moderately diverse (Gut, Ley)

3.Small, isolated but interacting, low density, moderately diverse (Finlay and Fenchel)

4.Large, continuous, highly diverse????? What do you suggest?

Drop-size soda lakes (Qvit-Raz , Genetics, 2008)

This is an amzing study showing actual differences in complex communities that go through successional stages in tiny drops that form from dew and exudates of the Tamarax tree which releases salty compounds.

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Ley et al., 2006

GUT MICROBIAL COMMUNITY

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BIOGEOGRAPHY:

IS EVERYTHING EVERYWHERE???

Fenchel and Finlay’s work, as reported in Science, 2005

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PNAS, 2007

ACTIVITY #2:

Let’s “sample” some diagrammatic representations of microbial communities!

STATISTICAL METHODS AND PRIMER, ENVIRONMENTAL APPLICATIONS:

http://epa.gov/bioindicators/statprimer/

BRAINSTORM: HOW DOES ONE MAKE DECISIONS ABOUT SAMPLING?

1. Size of population, community or system, area of interest

2. Scope of study?

3. Budget?

4. Variability (standard deviation, error)? depends on heterogeneity, abundance, distribution, both spatial and temporal, method, etc.

5. Technological ability (Can one directly observe organisms or cells? Can one target individuals or groups with specificity? How precise are the units of measurement?)

6. Experimental approach

7. SCALE! (many issues)

EXAMINE YOUR BACTERIAL “COMMUNITY “ and answer the following questions:

1. Would it be more appropriate to sample a large number of small plots, or fewer large plots? Considering the scale, how many large plots would be feasible to measure? Would sampling a larger number of smaller plots be less or more costly or time consuming?

2.Would using a line transect approach be appropriate? What would be the advantages or disadvantages to using this method, compared to plot sampling?

3.Describe a microbial ecosystem or community that you feel most closely matches the hypothetical “community” that your diagram represents.

D

A

C

B

v

vv

C D

Site C Small plot Large plot count: count: Transects:

Species 1 2 3 4 5 6 7 8 9 1 1 2 3A 0 0 0 3 0 0 1 0 0 0 0 0 0B 0 0 0 5 0 0 0 0 0 0 0 0 0C 0 0 0 1 0 0 0 0 0 0 0 0 0D 0 0 0 1 0 0 0 0 0 0 0 0 0E 8 0 0 0F 18 0 0 0G 2 0 0 0H 1 0 0 0

Site D Small plot Large plot count: count: Transects:

Species 1 2 3 4 5 6 7 8 9 1 1 2 3A 8 0 0 0 0 0 1 0 0 6 1 1 0B 3 2 1 0 0 0 1 0 2 5 2 1 0C 2 2 1 1 1 0 2 0 0 4 0 0 0D 1 3 0 0 0 0 1 0 0 1 0 1 0E 1 4 0 0 0 0 0 1 0 1 0 0 0F 1 16 0 1 0 0 0 0 5 12 1 0 0G 2 1 6 0 0 0 0 1 0 4 3 0 1H 0 1 4 0 2 0 4 0 0 1 2 0 2I 0 0 5 0 0 2 0 0 0 1 5 1 1J 0 0 0 0 0 1 0 0 0 0 0 1 0K 0 0 0 0 0 2 0 0 0 7 0 2 0L 0 0 5 0 14 0 0 0 0 8 1 0 2M 0 0 3 11 10 0 0 0 1 3 0 0 3

SOME SAMPLE DATA

Findlay 1982:Both approaches yield similar estimates of abundance, but larger scale sampling tends to give poor representationof distribution/patchiness

WHAT ABOUT MICROBIAL COMMUNITIES?

Ranjard et al. 2003 tested effect of SIZE of soil sample used in molecular community analysis.

For bacteria, SIZE OF SAMPLE did not affect ability to distinguish unique communities from one another. For fungi, samples <1g may not be adequate.

HOWEVER, in larger samples, technique of DNA extraction is biased toward dominant organisms and may not accurately assess diversity.

FOR BACTERIA: Many subsamples are often taken, and it is the selection of the sampling location that will give best indicator of larger scale abundance

and diversity

Desmarais 2002

When is use of transect appropriate?

•One example: when one hypothesizes a gradient relationship and wishes to test it using regression analysis, see below!

•In this study, researchers looked for change in numbers of fecal indicator organisms across various transects in relation to water’s edge.

•Keep in mind, most aquatic depth sampling is, in effect – transect sampling

FINAL THOUGHTS:•The question being asked drives the experimental design.

•The practical limitations of sampling limit the type of question that can be asked – and answered!

•Pilot sampling is always essential to assess the nature of the system!!

Students will better understand scientific process when they appreciate the considerations that go into every

scientist’s sampling and measurement plan!