Enhanced Biodegradation at a 

Fractured Rock Site

USEPA-USGS Fractured Rock Workshop

EPA Region 2

14 January 2014

Tom Imbrigiotta and Claire Tiedeman, USGS
Bioaugmentation Basics

 Concept

 Inject bacteria and food

 Bacteria populations increase, and

thus degradation rates increase

 Advantages

 Chlorinated solvents degraded in situ –

substantial concentration reductions can be 

achieved

 Possible reduced need for pump & treat – lower 

energy and treatment costs 

 Challenges

 Difficult to distribute amendments over large

volumes of the subsurface

 Uncertainty in this distribution, due to transport 

path uncertainty in fractured rock

 Sustainability of enhanced biodegradation –

electron donor (food) may become limiting

 Possible biofouling

Enhanced Biodegradation at a Fractured Rock Site 2

TCE  cisDCE  VC  Ethene

+ Cl- + Cl- + ClReductive Dechlorination
Inject Pump

Electron 

Donor &

Microbes

Bioaugmentation Experiment in

Highly Contaminated Mudstones

Enhanced Biodegradation at a Fractured Rock Site 3
 Questions related to hydrogeology

 Expected transport paths?

 Pumping rate at extraction well?

 Where to monitor?

 Volume of amendments to inject?

 Characterization activities

 Detailed stratigraphic framework

 Single & cross-hole hydraulic testing

 Cross-hole tracer testing

 Flow and transport modeling

 Push-pull tracer testing

Characterization for 

Bioaugmentation Design

Inject Pump

Electron 

Donor &

Microbes

Enhanced Biodegradation at a Fractured Rock Site 4
Cross-Hole Tracer Testing

Enhanced Biodegradation at a Fractured Rock Site 5

Inject 3700 mg/L Bromide Pump

Conclusions:

• Huge dilution at pumped well: 

only small amount of pumped 

water is coming from the 

region between 36BR & 15BR.

• > 80% of mass still in aquifer 

after 5 months

?
Tracer Testing: Bromide in Aquifer

7 Months after Injection

Conclusions:

• Most of mass is in 

downdip region 

low-K rocks/fractures 

strongly retain tracer.

Enhanced Biodegradation at a Fractured Rock Site 6
 No need to change 

pumping rate at 15BR

 Must inject large enough 

volume so amendments 

are pushed to 73BR

 Subsequent push-pull 

tracer test established 

needed volume

Tracer Testing Informed 

Bioaugmentation Design

Inject Pump

Enhanced Biodegradation at a Fractured Rock Site 7
Bioaugmentation Implementation

Enhanced Biodegradation at a Fractured Rock Site 8

Injection 

bladders

EOS

Water-quality 

monitoring
 Can geochemical & microbiological changes 

be observed downgradient?

 How much an effect will it have and how long 

will the effects last?

 Do the bacteria and electron donor move? 

 How far away are lateral effects felt? 

Questions About Effectiveness of 

Bioaugmentation in Fractured Rock 

Enhanced Biodegradation at a Fractured Rock Site 9
Can organic geochemical changes be 

observed in downgradient wells?

Enhanced Biodegradation at a Fractured Rock Site

- Significant cisDCE 

increases seen in these 

same wells

- Significant TCE decreases 

seen in wells 18 m and 30 m 

down the flow path

31,000

340

19,000

900

11,000

4,900

5,000

3,600

Red: median TCE conc (μg/l) pre-bioaug

Blue: TCE conc 4.8 years post-bioaug

TCE Reductions

- Significant TCE decreases 

seen in wells 18 m and 30 m 

down the flow path

3,600

59,000

6,400

22,000

2,400

6,000

5,000

3,400

Red: median cisDCE conc (μg/l) pre-bioaug

Blue: cisDCE conc 4.8 years post-bioaug

cisDCE Production

10
Can organic geochemical changes be 

observed in downgradient wells?

Enhanced Biodegradation at a Fractured Rock Site

- Significant TCE decreases 

seen in wells 18 m and 30 m 

down the flow path

2,700

5,100

4,900

130,000

1,600

6,000

1,400

1,400

Red: median Fe+2 conc (μg/l) pre-bioaug

Blue: Fe+2 conc 4.8 years post-bioaug

Ferrous Iron Production

- Significant TCE decreases 

seen in wells 18 m and 30 m 

down the flow path

21

2

17

<1

28

25

55

47

Red: median SO4 conc (mg/l) pre-bioaug

Blue: SO4 conc 4.8 years post-bioaug

Sulfate Reduction

- Fe+2 increases & SO4

decreases seen in wells 18 

m and 30 m down the flow 

path

11
How much of an effect will the 

bioaugmentation have?

Enhanced Biodegradation at a Fractured Rock Site 12

- Cl production/ 

CVOC degradation 

peaked ~7 months 

after injection.

- Bioaugmentation

has increased Cl

production/CVOC 

degradation by 

factor of 4-5 times 

natural attenuation.
How much of an effect will the 

bioaugmentation have?

Enhanced Biodegradation at a Fractured Rock Site 13

- Cl production 

in first downgradient well is 

combination of 

upgradient Cl

and bioaug Cl.

-Cl production/

CVOC degradation continuing 

at high level.

- Implies 

continuing TCE 

source and 

favorable redox 

conditions.
How long will the effects of 

bioaugmentation last?

Enhanced Biodegradation at a Fractured Rock Site 14

- TCE degraded 

& cisDCE 

produced 

quickly

- VC & ethene 

produced after 

lag period

- cisDCE = VC 

since ~1 yr 

post-injection

- Continuing 

source of TCE
How long will the effects of 

bioaugmentation last?

Enhanced Biodegradation at a Fractured Rock Site 15

- Increased DOC 

(EOS surrogate) 

not yet back to 

pre-injection 

levels

- Lactic acid used 

up after ~2.5 

years
Do the bacteria move?

Enhanced Biodegradation at a Fractured Rock Site

- Significant TCE decreases seen in wells 18 m and 30 

m down the flow path

Dhc

104

108

Red: median bacteria copies/L pre-bioaug

Blue: max bacteria conc 3.8 years post-bioaug

Geo

106

1010

Dhc

105

107

Geo

107

109

Dhc

106

107

Geo

106

108

Dhc

105

105

Geo

105

105

Dehalococcoides and Geobacter Increases

- Dhc & Geo increases seen 

in both 18 m and 

30 m downgradient wells16
Does the electron donor move?

Enhanced Biodegradation at a Fractured Rock Site 17

- Significant TCE decreases seen in wells 18 m and 30 

m down the flow path

5.9

73.7

Red: median DOC conc (mg/L) pre-bioaug

Blue: max DOC conc 3.8 years post-bioaug

20.0

44.5

0.9

1.6

1.2

2.2

DOC Concentrations
15BR

71BR

73BR

36BR

80BR

Lateral effects

Enhanced Biodegradation at a Fractured Rock Site 18

10 m
What is the lateral extent of the effect 

of bioaugmentation?

Enhanced Biodegradation at a Fractured Rock Site 19

- Packers separate 

GW with very 

different WQ 

effectively

- Areal effect of 

bioaugmentation is 

<10 m laterally

- Bacteria from 

bioaugmentation 

have not reached 

any zone in 80BR

- TCE:cisDCE >1 in 

80BR

TCE 820

cisDCE 2700

Dhc 105

TCE 120

cisDCE 22000

Dhc 106

TCE 5700

cisDCE 9600

Dhc 104

TCE 2000

cisDCE 1200

Dhc <101

TCE 4300

cisDCE 1900

Dhc 102

TCE 2300

cisDCE 1300

Dhc <101

80BR 73BR

A

B

C

D1

D2

E

10 m
 Fractured rock flow paths must be identified 

using aquifer tests and tracer tests.

 Geochemical effects of bioaugmentation

were observed up to 30 m downgradient

from injection well along a flow path. 

 Effects noted included increased reductive 

dehalogenation, more reducing conditions, 

breakdown of electron donor, and presence 

of increased bacterial concentrations.

Conclusions

Enhanced Biodegradation at a Fractured Rock Site 20
 Electron donor and bacteria can be 

transported a limited distance along flow 

path by injection and pumping.

 Bioaugmentation increased Cl production/ 

CVOC degradation by factor of 4-5x over 

natural attenuation.

 Bioaugmented biodegradation was still 

significant in injection well and still strong 18 

m downgradient after almost 5 years.

Conclusions

Enhanced Biodegradation at a Fractured Rock Site 21
 Bioaugmented biodegradation of TCE 

frequently stalls at cisDCE after relatively 

short period of time (6-12 months).

 Lateral spread of bioaugmentation effects 

depends on heterogeneity of flow and 

transport paths in the fractured rock.

Conclusions

Enhanced Biodegradation at a Fractured Rock Site 22