ORIGINAL PAPER Biological and chemical analyses of a laboratory-scale biofilter for oxygenate bioremediation in simulated groundwater J. Purswani • G. A. Silva-Castro • I. M. Guisado • J. Gonza ´lez-Lo ´pez • C. Pozo Received: 11 July 2012 / Revised: 29 April 2013 / Accepted: 26 May 2013 / Published online: 26 June 2013 Ó Islamic Azad University (IAU) 2013 Abstract Microbial remediation of methyl tert-butyl ether-contaminated aquifers has been widely studied since their cost/efficiency ratios are lower than other remediating techniques. Based on previous studies, simultaneous assessment of two inocula (Acinetobacter calcoaceticus M10 and a co-culture between strain M10 and Rhodococ- cus ruber E10) was performed for methyl tert-butyl ether bioremediation in a designed pilot-scale biofilter. A non- inoculated biofilter was included for comparison. Tempo- rary bioremediation of methyl tert-butyl ether (up to 80 % removal) from the biofilter inoculated with the consortium was observed within 44 days. In addition, the taxonomic profile (temporal temperature gradient gel electrophoresis from RNA extracts) from the latter biofilter contained more active strains than in the other two systems. The presence of strains M10 and E10 in temporal temperature gradient gel electrophoresis analysis from RNA extracts and high presence of strain E10 in temporal temperature gradient gel electrophoresis analysis of DNA extracts, along with alkB amplification of both strains in the biofilter, suggest that the co-culture inoculum was responsible for the methyl tert- butyl ether removal. Keywords Methyl tert-butyl ether Á Rhodococcus ruber Á Acinetobacter calcoaceticus Á Temporal temperature gradient gel electrophoresis Introduction Groundwater is the main drinking and irrigating source of water in most countries around the world. In several southern European countries such as Spain, Portugal, and Italy, the existing groundwater abstraction is equal to or greater than the groundwater availability in the country. In addition, desertification is on the increase too (UNEP 2004); thus, accidental spills or unnoticed leakages of unleaded gasoline will quite often contaminate essential aquifers and endanger this natural resource. Fuel oxygen- ates, such as methyl tert-butyl ether (MTBE), ethyl tert- butyl ether (ETBE), and tert-amyl methyl ether (TAME), have been used to replace tetraethyl lead, consequently reducing harmful emissions in motorized vehicles; how- ever, they are also the second most found contaminant detected in aquifers in the USA (de Lacy Costello et al. 2005). The European Environmental Agency’s acknowl- edgment of the problems arising from fuel oxygenate contamination has been given less importance than the American counterparts, even though oxygenate spills have been detected throughout Europe (Germany, Austria, Bel- gium, Switzerland, Spain, Italy) in different environmental water bodies (Rosell et al. 2007). The presence of tert-butyl alcohol is also monitored, since this MTBE metabolite is more toxic than the initial contaminant (Schmidt et al. 2004). Remediation technologies to palliate MTBE-contami- nated groundwater include adsorption via activated carbon and other synthetic sorbents (Aivalioti et al. 2010), air Electronic supplementary material The online version of this article (doi:10.1007/s13762-013-0348-3) contains supplementary material, which is available to authorized users. J. Purswani Á G. A. Silva-Castro Á I. M. Guisado Á C. Pozo Environmental Microbiology Group, Institute of Water Research, University of Granada, Granada, Spain J. Purswani Á J. Gonza ´lez-Lo ´pez Á C. Pozo (&) Department of Microbiology, University of Granada, C/Ramo ´n y Cajal no. 4, 18071 Granada, Spain e-mail: clpozo@ugr.es 123 Int. J. Environ. Sci. Technol. (2014) 11:1517–1526 DOI 10.1007/s13762-013-0348-3