However, the effect of RNAlater on IMS separation efficiency has not been explored previously. This study tested and developed a method that can be used to study the transcriptome of one species in mixed-species communities, including suspended and biofilm communities. Escherichia coli was selected as the target species in this study and Stenotrophomonas maltophilia Selleckchem BGB324 as a background species, because we are interested in the interactions between these two species when E. coli forms biofilms in drinking water distribution systems. E. coli is an important indicator of fecal contamination and is detected in some water distribution systems

[17]. S. maltophilia is a ubiquitous species in water systems. For example, the abundance of Stenotrophomonas spp. was 2-6% in a pilot drinking water distribution system [18]. Isolation of both E. coli and S. maltophilia from water filtration and distribution systems [19] suggests that they share the same niches in engineered systems and that interactions between them take place in such systems. The efficiency of IMS to separate E. coli from various

suspended mixtures and biofilms consisting of E. coli and S. maltophilia was evaluated in this study. The recovery and purity of separated E. coli cells were reported. Changes in the transcription check details profiles of E. coli cells due to sample processing and cell separation were quantified by cDNA microarray analysis and quantitative PCR (qPCR) to evaluate the effectiveness of the developed method. We also discussed that the method could be applied Luminespib in vitro to study other species of interest in mixed community systems and was not limited to the example species used in this study as long as a specific antibody for the target species is available. Results and Discussion Recovery rate of E. RAS p21 protein activator 1 coli The recovery rate of E. coli by immuno-magnetic separation (IMS) from a series of suspended cultures was determined first. A general antibody

of E. coli (polyclonal anti-E. coli antibody (ViroStat, Portland, ME)) was used in this study. Using this antibody, the recovery rate of E. coli was 74.4-98.2% when separated from suspended cultures with a density up to 1.9 × 108 CFU/ml (Figure 1). However, the recovery rate dropped to 59.8% for samples with ten-fold higher cells (1.9 × 109 CFU/ml), which may have exceeded the capacity of separation columns used in IMS (Figure 1). Therefore, E. coli cell densities in samples were adjusted to less than 2 × 108 CFU/ml for subsequent IMS. Figure 1 Recovery rates of E. coli cells after immuno-magnetic separation. Recovery rates of E. coli cells after one-step IMS from suspensions of E. coli with densities adjusted from approximately 104 to 109 CFU/ml. Error bars indicate standard deviations of triplicate plate counts. Determining the recovery rate of target species is important when IMS is used to separate target species for subsequent cDNA microarray analysis.