We use different molecular tools to collect, test and enhance wildtype and/or engineered bacterial strains for biodegradation of xenobiotics in soil and water. Our principal focus concerns organophosate (OP) pesticide remediation, with an added interest in the breakdown of several other high impact xenobiotics common to the Houston-metropolitan area including herbicides, oil products and byproducts as well as polychlorinated organic waste. In addition, we use our growing bacterial collection to actively engage our students in undergraduate research experiences to prepare them for the 21st century STEM workforce. To this end, research in our laboratory is currently divided into six major projects.
(1) Location and distribution of organophosphate degradation activity
As part of an ongoing project in organophosphate (OP) pesticide bioremediation, our students screen soil and water samples to isolate bacterial strains which biodegrade organophosphate (OP) compounds. The location of all collected isolates is pinned to Google maps and uploaded to an Environmental Sampling database. This database allows students to map and analyze the distribution of OP-degrading activity across the Houston-metropolitan area as well as any other regions samples have been taken. In the near future we will be collaborating with other institutions to expand our collection both nationally and globally. A representative list of identified OP degraders we have collected to date is shown below.
Xenobiotic-degrading microorganisms collected to date:
Pseudomonas aeruginosa, Pseudomonas putida, Pseudomonas stuterzi, Stenotrophomonas maltophilia, Exiguobacterium indicum, Acinetobacter calcoaceticus, Acinetobacter haemolyticus, Achromobacter xylosoxidans, Achromobacter denitrificans, and Ochrobactrum anthropi as well as unknown species of Citrobacter, Arthrobacter, Rhizobium, Agrobacterium, and Delftia.
(2) Molecular characterization of OP-degrading microorganisms
Positive isolates are tested against different OP substrates including parathion, paraoxon, methyl parathion, chlorpyrifos, and malathion. Degradation kinetics data is then compared under different environmental and nutritional parameters to determine how field remediation of a particular OP may be enhanced. In addition to OPs, we have also begun preliminary testing on biodegradation of common industrial solvents, toluene and benzene, the herbicides glyphosate and atrazine, petroleum hydrocarbons, 1, 4 dioxane, and toxic cogeners of dibenzo-p-dioxin and dibenzofuran to identify putative broad-spectrum degraders among the strains previously collected by our students.
(3) Development of new databases to compile and maintain research data of collected microorganisms
We are in the process of building three new databases to host the research data we have compiled on our bacterial isolates. A genomic database will store 16S rDNA sequences for identification purposes as well as sequences of key xenobiotic degradation genes. A target molecule database will list the toxic compound(s) that a particular bacterium degrades, while the degradation kinetics database will store information on how fast an individual bacterium can break down its target molecule(s). Once these databases have been crosslinked to our Environmental Sampling database, all pertinent data on a bacterium may be accessed by simply clicking on one of the pins representing its sample collection site.
(4) Using compiled research data to enhance student STEM skillsets
Research data is made available to students for a variety of STEM-building activities. These activities serve as a learning tool to enhance skillsets in bioinformatics, quantitative literacy, hypothesis testing, data interpretation, critical thinking, and problem solving. Degradation analyses familiarize students with the statistical significance of quantitative scientific data and employ statistical methods from simple T-tests and ANOVA to linear regression analysis to predict outcomes of an experiment based on past trials, interpret their results, and confirm or refute a hypothesis. Sequence and geographic information system data is used to conduct simple phylogenetic and phylogeographic analyses to illustrate evolutionary relationships between bacterial strains and/or their xenobiotic degradation genes. Finally, proteomic and structural data is compared to determine sites critical for enzyme function or to identify mutagenic hotspots for protein engineering studies.
(5) Virulence attenuation of Stenotrophomonas maltophilia isolates
Several bacterial isolates part of our collection are noted for their potential pathogenicity in addition to their capacity for xenobiotic detoxification. We are currently investigating how to reduce the potential virulence of our Stentrophomonas maltophilia isolates, which harbor a conserved metallo-B-lactamase enzyme with the capacity to degrade organophosphate substrates. The attenuated microorganisms that exhibit insignificant changes in growth and degradation dynamics will be used for further bioremediation applications.
(6) Investigation of northeast Harris County cancer clusters and their possible relationship with the San Jacinto dioxin waste pits
Potential cancer clusters localized throughout Harris County may be linked to the presence of an EPA Superfund site alongside the San Jacinto River near I-10 that holds two dioxin waste pits. A combination of informatics data obtained online from EPA and Texas government reports is currently being used to compare dioxin concentration levels with elevated cancer statistics in the area on both a spatial and temporal basis. Recent efforts are now underway to expand this project to include sample collection from along the San Jacinto River and other impacted sites such as the Houston Ship Channel and nearby tributaries to map dioxin degradation activity in a similar manner. Furthermore, by incorporating genomic data consisting of 16S rDNA and conserved xenobiotic degradation genes extracted from the environmental isolates obtained at these sites, we will seek to determine how genetic and environmental factors influence the phylogeography of different bacteria along the San Jacinto River and Houston Ship Channel as well as the evolution of their degradation genes.
Iyer, R., Iken, B., Tamez, T. (2011). Isolation, Molecular and Biochemical Identification of Paraoxon-Metabolizing Pseudomonas Species. Journal of Biodegradation and Bioremediation, Vol. 2(5), e132.
Iyer, R., Stepanov, V., Iken, B. (2013) Isolation and Molecular Characterization of Novel Pseudomonas putida Strain Capable of Degrading Organophosphate and Aromatic Compounds. Journal of Advances in Biological Chemistry. (Vol 3), 564-578
Iyer, R., Wales, M. (2013) "Identification of Water-borne bacterial isolates for Potential Remediation of Organophosphate Contamination", Journal of Advances in Biological Chemistry, Vol. 3, 146-152.
Iyer, R., Smith, K., Kudrle, B., and Leon, A. (2015) Detection and Location of OP-degrading Activity: A model to Integrate Education and Research. New Biotechnology. Vol. 32(4), 403-411.
Iyer, R., Iken, B., and Leon, A. (2015). Characterization and Comparison of Putative Stenotrophomonas maltophilia Methyl Parathion Hydrolases. Accepted for publication. Bioremediation Journal.