University of Colorado
Environmental Engineering

Research Projects

Biological Load in Bioaerosols

The organic fraction of airborne particulate matter is heterogenous in chemistry and origin. We estimate that up to 25% of this fraction originates from biological sources. In order to assess the biological contribution, we measure biopolymers (carbohydrates, phospholipids, proteins, and DNA), and construct 16S clone libraries to understand the source of biological matter. As biological particles have a pronounced health effect, we also assess the genotoxicity of particulate matter to bacterial and eukaryal cells.


Airborne particulate matter will be analyzed for its total biological load—carbohydrate6, protein5,
phospholipid and DNA content—as well as its total carbon content (both inorganic and organic).
Particulate matter will be eluted from filters into sterile, pyrogen-free water. Carbohydrates are measured
using a sensitive colorimetric assay of phenol conjugation to monomeric sugars following sulfuric acid
digestion using glucose as a standard. Protein associations with atmospheric aerosols are determined
using selected quantitation of Nano-Orange dye from a sensitive fluorescence assay, which uses bovine
serum albumin as a standard (Invitrogen). Phospholipid associations with atmospheric aerosols are
determined using a simple colorimetric assay of airborne particulate matter extracted into chloroformmethanol
mixtures. The lipids present in the solvent extracts are quantitatively subject to a persulfate
digestion, which is calibrated against a ß-glycerol phosphate standard.


The gulf project compares particulate matter collected from Grand Isle, LA and Sea Rim, TX in order to investigate how the biological fraction has been affected by the Deepwater Horizon oil spill. The University of Colorado published an article describing our work.

 

The Manitou project characterizes the biological fraction of particulate matter from a semi-pristine high-alpine outdoor environment, with the goal of understanding the baseline for outdoor bioaerosols.

 

In summer 2012, we collected aerosol samples downwind of a forest fire on the Colorado Front Range. These samples will be analyzed for bioaerosol load and toxicity.

Personnel: Alina Handorean, Jane Turner, Bharath Prithiviraj, Odessa Gomez

Past Personnel: Mari Rodriguez

 

A Comprehensive Toxicological Suite for the Analysis of Airborne Particulate Matter


Epidemiological studies have provided compelling statistical evidence for associations between airborne particulate matter (PM) exposure and negative health effects. Additionally, numerous toxicological studies have demonstrated that beyond certain thresholds, airborne PM has the potential to introduce DNA damage, cytotoxicity and oxidative stress. Most recent toxicological research on indoor and outdoor PM focuses primarily on a single biomarker, generalizing cellular damage pathways to a solitary mechanism. There is a need for the development of broader screening approaches which are capable of quickly and comprehensively describing the conglomerate of biological activity that PM may carry. In response to this need, a suite of cytometric toxicological assays, employing human cell lines (lung epithelial and phagocytic monocytes) has been adapted to assess the major toxicological modes associated with airborne PM. The assay panel addresses the following classes of fundamental cellular responses previously linked with PM exposure: necrotic and apoptotic cytotoxicity, genotoxicity and oxidative stress. These assays are relatively rapid and can deliver an overview of individual toxicity mechanisms, as well as allow for the analysis of synergistic effects. By developing and testing this analytic suite, we hope to move toward providing a template for future mechanistic-based PM regulations.

 

Personnel: Jane Turner, Diedra Gustafson

Past Personnel: Kevin McCabe

 

Microbially-Induced Concrete Corrosion: Characterization and solutions

The microbially-induced corrosion of concrete in wastewater infrastructure costs wastewater utilities billions of dollars in replacement and rehabilitation costs and negatively impacts public relations. We aim to better understand the microbial players in this community and to design a biological-based approach to inhibiting their growth.

 

In one phase of this study, we collected corroded concrete biofilm samples from ten cities acrosso the United States. We will use V4V5 16S amplicon sequencing on the Illumina MiSeq platform and will collect environmental data such as pH, moisture content, headspace gasses, and sulfur content to assess how the corrosion community differs across environmental gradients.

 

In the second phase of this work, a metal-based antimicrobial coating will be designed and tested in lab and in field manholes. We exposed treated and untreated concrete coupons in a working sanitary manhole for time periods of 1 month to 12 months. These samples were analyzed for microbial community composition, surface chemistry, and extent of corrosion. Additionally, we will perform laboratory inhibition studies of the product using sulfide-oxidizing bacterial cultures.

Personnel: Alison Ling, Alejandro Caicedo-Ramirez

Past Personnel: Anna Campbell

 

Airborne Pathogen Transmission: Survival and inactivation

Bordatella pertussis
In collaboration with the Food and Drug Administration Laboratory of Respiratory Pathogens we are developing novel methods to study the response of B. pertussis, the causative agent of whooping cough, to aerosol environmental stress. We have demonstrated a gene level response to aerosolization, or being coughed out of the host, to enhance survival in the air. The pathway controlling virulence gene expression in B. pertussis is regulated by environmental cues. A set of genes of unknown function, regulated oppositely the virulence genes, appears to convey resistance to aerosolization stress. Utilizing mutant strains affecting the gene regulation of these putative survival genes and a completely novel method for the collection of an "aerosolized RNA profile" we have identified gene level response to aerosolization. When B. pertussis is in a host, the virulence genes are on and these environmental survival genes are off. When then aerosolized, or coughed into the air, the bacteria sense the environmental change, and turn off the virulence genes, in favor of a set of genes that enhance survival in the air. When these microbes encounter a new host, they again sense a change in environment and alter gene expression, away from the environmental survival factors, and back to the virulence factors.

 

Past Personnel: Kevin McCabe, Jane Turner,Jordan Peccia, Sarahann Dow, Lars Angenent

 

Electro-Magnetic Fields: Effect on cell growth

Our work on electromagnetic fields (EMF) arose from our studies of electrically enhanced filter systems and their ability, not only to capture bioaerosols, but also inactivate microbes on the filter surface. This intensively collaborative endeavor, in conjunction with Frank Barnes in Electrical Engineering at CU Boulder, seeks to identify the molecular mechanism/s by which EMF exert effects on biological systems. We have demonstrated changes in growth kinetics of cancer cell lines and, more recently, bacteria in response to shielding or reducing the Earth's magnetic field. These micro-Tesla level changes are on the order of what one might observe between indoor versus outdoor settings, or perhaps more importantly to the biological sciences community, between cell culture incubators. While large epidemiological studies of the effects of EMF on human health have been carried out for decades, these efforts are hampered by a lack of understanding of the most fundamental effects of EMF at the subatomic to molecular level.

 

Past Personnel: EJ Sheehan, Kevin McCabe

 

Benzotriazoles: Metal uptake and toxicity

Benzotriazoles are a anti-corrosion agent used in aircraft deicing fluid and other industrial applications. These compounds are resistant to environmental degradation and inhibitory to bacterial growth.

 

Benzotriazoles can form strong complexes with transition metals and enable the removal of toxic metals from industrial waste streams.

 

Past Personnel: Muna Abu-Dalo, Jeff Cornell, Cyndee Gruden, Ivette O'Brien