Microbiological Diagnostics

Cutting edge microbiological tools from:
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Site Characterization

Determine the presence of microbes and desired biological processes

Monitor distribution of contaminant degrading populations

Demonstrate the feasibility of monitored natural attenuation (MNA)

Site Management

Evaluate remedial action alternatives with quantitative data

Monitor the effectiveness of an implemented bioremediation strategy

Optimize current corrective actions

In-Situ Microcosm Investigation

Passive groundwater sampling tools for collection of microbes over time for the purpose of analysis and understanding biodegradation potential

The Bio-Trap Sampler in essence becomes an in situ microcosms for routine groundwater sampling, assessment of monitored natural attenuation, and quantitative evaluation of enhanced bioremediation alternatives.

Samples collected by Bio-Trap can be analysed using a wide variety of methods including all the Molecular Biological Tools (CENSUS®, PLFA, DGGE, SIP) and more!

The key to the Bio-Trap® approach is a unique sampling matrix, Bio-Sep® beads - an engineered composite of Nomex® and powdered activated carbon (PAC). When a Bio-Trap® sampler is deployed in a monitoring well, the Bio-Sep® beads absorb contaminants and nutrients present in the aquifer essentially becoming an in situ microcosm with an incredibly large surface area which is readily colonized by subsurface microorganisms. Once recovered from a monitoring well (30-60 days after deployment), DNA, RNA, or phospholipid fatty acids (PLFA) can be extracted from the beads for CENSUS® or PLFA assays to evaluate the microbial community

Standard Bio-Trap simply provide a large surface area for the microbes to colonize and form biofilms. The technique is essentially a replacement for collecting conventional groundwater samples, recognising that most microbes prefer to be attached to a surface rather than free floating. The results generated using this approach have been shown to minimize the variability associated with traditional sampling methods and to reflect the spatial and time dependent changes in the aquifer microbial community structure which is not always evident from groundwater analysis.

Baited Bio-Trap® Samplers are impregnated with various amendments or compounds to answer site-specific questions and screen remedial alternatives. For example, they can be baited with specific contaminants of concern, various electron donors such as HRC®, EOS®, or lactate, or with inorganic amendments like nitrate or sulphate.

Bio-Trap® In-Situ Microcosm Studies provide microbial, chemical, and geochemical evidence to cost-effectively evaluate biodegradation and screen remedial alternatives. This approach is an effective solution to major limitations found with the two traditional approaches:

Duplication of in situ conditions in the laboratory is difficult and the results often do not correlate to the field.
Pilot studies are performed in the field but are often prohibitively expensive as an investigative tool

Bio-Trap® In Situ Microcosms can be tailored to investigate a wide variety of remediation approaches but often consist of three units each corresponding to one of the most common bioremediation options:

Applications

MNA at chlorinated solvent- and petroleum hydrocarbon- impacted sites
Evaluate the effectiveness of electron acceptor addition (e.g. oxygen, sulfate)
Assess the feasibility of electron donor addition to stimulate reductive dechlorination
Assess the need and effectiveness of bioaugmentation
Can be used in combination with Stable Isotope Probing (SIP) or Compound Specific Isotope Analysis (CSIA)

	Control MNA Unit: The Control Unit contains no additional electron donor or amendments and represents MNA or existing site conditions.
	Biostimulation BioStim Unit: The BioStim Unit contains a specified electron donor (sodium lactate,EOS®, HRC®, molasses, etc), electron acceptor (e.g. oxygen, nitrate, sulfate), or other amendment.
	Bioaugmentation BioAug Unit: The BioAug Unit is pre-inoculated with a commercial culture (e.g. Dehalococcoides) and amended with an electron donor.

Molecular Biological Tools (MBT)

Superior genetic and chemical diagnostic tools are used to aid in the understanding and management of microbiological processes in remediation of groundwater and other water systems.

CENSUS is a nucleic acid-based approach to quantify specific microorganisms, groups of microorganisms, or functional genes involved in bioremediation or other biological processes. CENSUS targets include bacteria and functional genes responsible for biodegradation of chlorinated solvents and petroleum products among others. For example, at a site impacted by chlorinated solvents like PCE or TCE, CENSUS quantification of Dehalococcoides enables project managers to:

Evaluate the feasibility of monitored natural attenuation (MNA)
Evaluate the efficacy of enhanced bioremediation approaches
Assess the need for bioaugmentation

Phospholipid Fatty Acid (PLFA) is a broad-based biochemical approach that provides direct information on the entire microbial community in three key areas:

Viable (living) biomass concentrations
Community composition or population “Fingerprint”
Insight into the metabolic status or “health” of the microbial community

In addition, PLFA analysis can be coupled to an innovative technique called Stable Isotope Probing (see below) to show conclusive evidence of biodegradation.

All cells have membranes which consist mainly of phospholipid fatty acids. PLFA biomarkers break down quickly when a cell dies, so intact PLFA extracted from an environmental sample (groundwater, soil, sediment or Bio-Trap®) is only from living (viable) organisms and is expressed as cells per unit of sample. The chemical composition of the PLFA biomarkers differs depending on the type of organism and therefore can be used to generate a “fingerprint” of the microbial community composition.

Stable Isotope Probing (SIP) is an innovative method to conclusively determine if biodegradation is occurring.

SIP incorporates a 13C labelled contaminant (acting as a tracer) into microbial biomass and dissolved inorganic carbon. The label is detected in the end products of biodegradation - new biomass and CO2 or dissolved inorganic carbon. With the SIP method, a Bio-Trap sampler is baited with a specially synthesized form of the contaminant containing 13C labelled carbon. Since 13C is normally so rare, the labelled compound can be readily differentiated from the contaminants present at the site.

SIP studies can be performed for any compound that microbes use as a carbon source. Some of the more common include Benzene, Chlorobenzen, Toluene, Xylenes, Napthalene, MTBE and TBA.

Denaturing Gradient Gel Electrophoresis (DGGE) is a DNA-based technique which generates a genetic profile or “fingerprint” of dominant members of the microbial community.

Individual DNA sequences or “bands” from this profile can be sequenced to identify the dominant members of the population. The technique has been used to investigate microbial responses in a wide variety of applications, including:

Bioremediation assessment
Wastewater treatment
Drinking water treatment
Biofilm formation
Microbial induced corrosion
Identification of microbial contaminants in commercial/industrial products

For bioremediation assessment, DGGE analysis is commonly used for evaluating “shifts” in microbial community composition over time or following a treatment. For example, DGGE can be used to determine the differences in the dominant bacterial groups in contaminated versus non-contaminated groundwater monitoring wells to evaluate which groups are enriched in impacted zones. Likewise, DGGE can help determine which bacterial groups are stimulated following a corrective action such as addition of a growth substrate or nutrient.