Comparison of methods for monitoring bacterial transport in the subsurface

The purpose of this study was to compare in a laboratory experiment, a suit e of methods developed to track viable bacteria during field transport expe riments. The criteria for development and selection of these methods includ ed: (1) the ability to track bacteria within the environment from which the y were isolated; (2) the lack of any effect upon the viability or the trans port characteristics of the strain; (3) low detection limits;, (4) a quanti fication range that covered several orders of magnitude; and (5) an analyti cal cost and turnover time commensurate with the analysis of several thousa nds of samples in a few months. The approaches developed included: enumerat ion of bacteria labeled with a vital fluorescent stain (CFDA/SE) using micr oplate spectrofluorometry, flow cytometry, and ferrographic (immunomagnetic ) capture; enumeration of highly C-13-enriched bacteria using combustion-IR MS; and quantitative PCR. These methods were compared to direct microscopic enumeration and plate counts during a bacterial transport experiment perfo rmed in an intact sediment core and designed to simulate the field experime nt. Four of the seven methods had equivalent recoveries for the breakthroug h of a pulse of bacteria eluting from a 50-cm long sediment core, and all o f the methods detected the arrival of cells in the effluent prior to the co nservative tracer. Combustion IRMS and ferrographic enumeration had the low est quantification limits (similar to2 to 20 cells/ml), whereas microplate spectrofluorometry had the highest quantification limit (similar to 10(5) c ells/ml). These methods have the potential for numerous applications beyond tracking bacteria injected into the subsurface, (C) 2001 Elsevier Science B.V. All rights reserved.