Transport of Bacterial Endospores in Silica Sand

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Transport of Bacterial Endospores in Silica Sand

• Sibylle Tesar, Fulbright Scholar
• Dr. Barbara Williams, Faculty
• Dr. Robin Nimmer, Res. Supp. Sci.
• Angelina Cernick, Undergraduate
• Kristina Beaulieau, NSF REU

Department of Biological and Agricultural
Engineering University of Idaho
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Outline

• Research Goals transport mechanisms /
  endospores
• Background transport mechanisms / endospores
• Research Questions preliminary
• Methods sporulation / saturated column tests /
  breakthrough curves / depth distribution data
• Preliminary Results
• Preliminary Conclusions
• Future Work B. cereus, other microbes

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Research Goals Spore Transport in Porous Media

• Mechanistic Goal Contribute to the lively
  debate of attachment versus straining
• Microbe-specific goal Bacterial endospore
• Practical Applications
• Drinking water protection - groundwater
• Shallow recharge
• Septic drainfield setbacks
• Surface water filtration
• Riverbank or riverbed filtration

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Outline

• Research Goals transport mechanisms /
  endospores
• Background transport mechanisms / endospores
• Research Questions preliminary
• Methods sporulation / saturated column tests /
  breakthrough curves / depth distribution data
• Preliminary Results
• Preliminary Conclusions
• Future Work B. cereus, other microbes

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Terminology Mechanisms for Retention

• Attachment adhesion sorption
• Function of collision, collector efficiency,
  sticking efficiency
• Mechanical filtration complete retention of
  particles that are larger than all of the soil
  pores (formation of filter cake)
• Straining physical trapping in geometric
  corners
• Particles can be smaller than smallest pore
  openings
• Requires grain-grain contact
• Only occurs in some fraction of soil pore space,
  transport occurs elsewhere

Bradford et al, WRR, 2006
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Strained versus Mechanically Filtered
dp/d50 ? .005
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Background

• Clean-bed Filtration Theory
• Depends on mechanism of attachment / detachment
• Deviation from Clean Bed Filtration Theory
• Unfavorable attachment condition neg-neg
• Fine sand and large colloids (dp/d50 ? .005)

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Explanations for Deviation from CFT

• Attachment w/ porous media charge variability
  Johnson and Elimelech, 1995
• Attachment w/ heterogeneity in surface charge
  characteristics of colloids Li et al, 2004
• Attachment w/ deposition of colloids in a
  secondary energy minimum Tufenkji et al. 2003,
  Redman et al., 2004
• All of the above Tufenkji and Elimelech, 2005
• Attachment w/ straining Foppen et al, 2005,
  Bradford et al, 2006a, b

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Theory (cont.)
Aqueous Phase Colloid Mass Balance Equation-
Bradford et al., 2003

• Where
• ?w volumetric water content -
• t time T
• C colloid concentration in the aqueous phase
  N L-3
• JT total colloid flux N L-2 T-1
• EattSW colloid attachment mass transfer
  between solid/water phases N L-3 T-1
• EstrSW colloid straining mass transfer between
  solid/water phases N L-3 T-1

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Research Goals Endospore Transport

• Endospore-forming bacteria have two viable modes
• Vegetative cell (growing)
• Endospore (dormant) formed as survival
  mechanism
• Endospores have the potential to be more mobile
  than their vegetative cell counterparts
• smaller size
• potentially less adhesion

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Bacterial Endospores

• Formed as a survival mechanism
• Cryptobiotic no sign of life - dormant mode

http//www.textbookofbacteriology.net/
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Differences between endospores and vegetative
cells in Bacillus species
http//www.textbookofbacteriology.net
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Differences between endospores and vegetative
cells in Bacillus species
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In terms of physical passage through the pore
space
the spore has a shorter aspect ratio than the
vegetative cell.

• B. cereus spore properties
• Food poisoning pathogen
• Length 1-2 mm, Width 0.5-0.75 mm
• Hydrophobic
• Isoelectric point pH 3

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Outline

• Research Goals transport mechanisms /
  endospores
• Background transport mechanisms / endospores
• Research Questions preliminary
• Methods sporulation / saturated column tests /
  breakthrough curves / depth distribution data
• Preliminary Results
• Preliminary Conclusions
• Future Work B. cereus, other microbes

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Preliminary Research Questions

• Do spores obey CFT, exhibiting more retention in
  higher ionic strength solution or does spore
  transport deviate from CFT theory as do other
  negatively charged particles (unfavorable
  attachment)?
• Future Do vegetative cells and endospores have
  a different charge?
• Future Do vegetative cells exhibit more
  attachment than endospores?

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Outline

• Research Goals transport mechanisms /
  endospores
• Background transport mechanisms / endospores
• Research Questions preliminary
• Methods sporulation / saturated column tests /
  breakthrough curves / depth distribution data
• Preliminary Results
• Preliminary Conclusions
• Future Work B. cereus, other microbes

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Materials Sand Properties

• Saturated conductivity Ksat 1.8x10-4 m/sec
• Dry bulk density
• rb 1.65 g/cm3
• Porosity
• n 0.34
• dp/d50 ? .0017

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Method Constant Head, Sand Column
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Breakthrough (C/Co) of B. cereus spores as a
function of ionic strength
Artificial groundwater
DDI
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Column Dissection
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Column Dissection

• Drain column to field capacity, in the flow
  direction.
• Dissect into seven 2 cm increments
• STR 1 Gently place sand, allowing bridging and
  loose packing, in a funnel that has been plugged
  with Scotchbritetm pad
• STR 2 Wash off the strained bacteria by pouring
  the solution (the solution used in that
  particular experiment) over the sand into a
  graduated cylinder
• ATT To remove the attached bacteria, place a
  known amount of 2 Tweentm 80 solution into a
  beaker containing the sand. Stir then sonicate.
• Used optical density (OD) measurements in
  addition to plate counting to enumerate.

(Tween and sonication proven not to affect
germination efficiency)
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Depth Distribution Data
Strained spores in AGW run
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Depth Distribution Data
Attached spores in AGW run
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Depth Distribution Data
Strained and attached fractions combined
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Outline

• Research Goals transport mechanisms /
  endospores
• Background transport mechanisms / endospores
• Research Questions preliminary
• Methods sporulation / saturated column tests /
  breakthrough curves / depth distribution data
• Preliminary Results
• Preliminary Conclusions
• Future Work B. cereus, other microbes

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Preliminary Conclusions

• Breakthrough curve data are consistent with CFT
  higher ionic strength, more retention
• Depth distribution data show deviation from CFT
  not exponential with depth

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Outline

• Research Goals transport mechanisms /
  endospores
• Background transport mechanisms / endospores
• Research Questions preliminary
• Methods sporulation / saturated column tests /
  breakthrough curves / depth distribution data
• Preliminary Results
• Preliminary Conclusions
• Future Work B. cereus, other microbes

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Future Work

• Compare attachment/straining of spores versus
  vegetative cells
• Column experiments
• Micromodels and photographs
• Wet AFM
• Compare zeta potential
• pH and more ionic strength effects
• Different endospore bacteria, such as S.
  pasteurii, for biomineralization

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Acknowledgements

• Dr. Ron Crawford, Director, Environmental
  Biotechnology Institute, UI
• Nick Benardini, PhD Candidate, MMBB
• Elizabeth Scherling, MS, BAE
• Dr. Markus Tuller, PSES
• David Christian, Research Support Sci.

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Funding Acknowledgements

• Fulbright Scholars Program
• USDA Hatch
• UI URO Seed Grant Program
• NSF REU program

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References

• Bradford, S.A., J. imunek, M. Bettahar, M.
  vanGenuchten, and S.R. Yates. 2003. Modeling
  colloid attachment, straining, and exclusion in
  saturated porous media. Environmental Science
  and Technology 37 2242-2250.
• Bradford, S.A., J. imunek, M. Bettahar, M.Th.
  vanGenuchten, and S.R. Yates. 2006a.
  Significance of straining in colloid deposition
  evidence and implications. Water Resources
  Research, 42doi10.1029/2005WR004791.
• Bradford, S.A., J. imunek, and S.L. Walker.
  2006b. Transport and straining of E. coli
  0157H7 in saturated porous media. Water
  Resources Research (in review).
• Li, X., TD. Scheibe, and W.P. Johnson. 2004.
  Apparent decreases in colloid deposition rate
  coefficient with distance of transport under
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• Redman, J.A., S.L. Walker, and M. Elimelech.
  2004. Bacterial adhesion and transport in porous
  media Role of the secondary energy minimum,
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• Tufenkji, N., J.A. Redman, and M. Elimelech.
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• Tufenkji, N., Elimelech, M. 2005. Breakdown of
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  energy minimum and surface charge
  heterogeneities. Langmuir 21 841-852.