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Construction and Characterization of Cyanobacterial Bioreporters to Assess Nutrient (P, Fe) Availability in Marine Environments

by Boyanapalli, Ramakrishna Bharadwaj

Abstract (Summary)
Nutrient deficiency especially phosphorus (P) and iron (Fe), are well documented in world oceans, particularly associated with oligotrophic oceanic gyres and “high nutrient, low chlorophyll” (HNLC) regions. As a corresponding approach to identify bioavailable Fe in marine systems, my research has focused on the development, characterization and implementation of a whole-cell cyanobacterial Fe bioreporter to be used to assess the bioavailability of Fe in marine environments. A Fe responsive whole-cell marine bioreporter was developed by integrating a construct of the isiAB promoter fused to bacterial luciferase luxAB into the marine cyanobacterium Synechococcus PCC 7002. Dose-response characterization of the bioreporter was made in trace-metal buffered synthetic seawater medium containing Fe concentrations in the range of pFe [-log (Fe^3+ free ferric)] 19.4 – 22.4. The luminescent response from the Fe bioreporter was best described by a sigmoidal dose-response curve. A comprehensive growth and physiological characterization of the bioreporter was conducted prior to implementing it to assess Fe bioavailability from various marine samples. The bioreporters response was optimum when it was incubated at 25 °C under an irradiance of 45 µmol photon m-2 s-1. Varying the amount of bioreporter inoculum or salinity of the test medium did not affect cellular luminescence over the 12 h assay period. Testing the ability of the bioreporter to acquire Fe^3+ bound to ligands, the bioreporter was able to acquire bound Fe from rhodotorulic acid but, was unable to acquire Fe from ligands such as desferrioxamine B and N, N’-diethylenediamine-N,N’-diacetic acid. We assessed Fe availability in environmental samples from the Baltic Sea using the cyanobacterial bioreporter and demonstrated that the bioavailable Fe was one order of magnitude lower than the chemically-determined dissolved Fe and that samples from depth showed higher available Fe than at the surface. The Fe bioreporter was also implemented for assessing available Fe in samples collected during the mesoscale Fe fertilization experiment SERIES conducted in the HNLC eastern subarctic Pacific Ocean, and in samples collected from oligotrophic waters of the central north Pacific gyre during the ROMP study. The luminescent response from these open ocean samples was consistently higher than the corresponding calibration standard. As a result, the cyanobacterial Fe bioreporter can only be considered a qualitative tool with which to assess Fe availability in open ocean environments. Despite this constraint, we observed genuine differences in Fe availability from samples collected during each study.
Bibliographical Information:

Advisor:

School:Bowling Green State University

School Location:USA - Ohio

Source Type:Master's Thesis

Keywords:bioreporter cyanobacteria synechococcus pcc 7002 iron bioavailability chelators oxidative stress isiab luxab luciferase

ISBN:

Date of Publication:01/01/2006

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