Date of Award

Spring 5-12-2017

Document Type


Degree Name

Master of Science (MS)



First Advisor

Dr. Anuradha Ghosh

Second Advisor

Dr. Dixie Smith

Third Advisor

Dr. Ram Gupta


Bacterial Diversity at an Abandoned Coal Mine in

Southeast Kansas

An Abstract of the Thesis by

Rachel Bechtold

Acid mine drainage (AMD) is found in areas of abandoned coal mines in southeast Kansas as a result of mine waste rocks and tailings create problems for the local environment. Soil bacterial population acts as a reliable indicator of ecosystem health in these human-perturbed areas. The goals of the present study were to assess the bacterial diversity of an AMD site over a two-year period and to isolate acid-tolerant bacterial species for bioremediation purpose.

In fall (2015) and summer (2016), soil samples were aseptically collected from five locations representing diverse topography at an AMD site in southeast Kansas. Soil texture was evaluated and samples were chemically digested for physicochemical analysis using inductively-coupled plasma optical emission spectroscopy. Concentration of bacterial isolates was determined by counting CFUs after dilution plating on tryptic soy agar. Up to thirty morphologically different colonies from each sampling were characterized using physiological and biochemical tests and were further identified at the species level using 16S rRNA gene sequencing. In addition, acidophilic bacterial strains were screened using selective differential media.

Preliminary data showed that soil pH ranged from 2.5-6.8 and contained varied concentrations of arsenic, manganese, and iron. Total bacterial concentration was 102-108 CFU/g of soil over two samplings. Biochemical tests revealed a diverse metabolic potential of the bacterial population. Bacterial isolates for both fall and summer samples were grown in citric acid buffer with varying pH of 3, 4, 5, and 6 and several were found to be acidophilic. PCR amplification of 16S rRNA gene and purification were completed before sending samples off for sequencing to Kansas State University; a phylogenetic analysis was completed for both sampling times and subsequently illustrated with neighbor-joining phylogenetic trees constructed using MEGA 7 software. Baseline measurements of bacterial diversity as well as soil chemistry in AMD sites in this region are novel and the findings may have potential use in bioremediation of contaminated AMD sites.



To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.