Portable rapid detection of pathogenic bacteria such as Bacillus is highly desirable for safety in food manufacture and under the current heightened risk of biological terrorism. Surface-enhanced Raman scattering (SERS) is becoming the preferred analytical technique for bacterial detection, due to its speed of analysis and high sensitivity. However in seeking methods offering the lowest limits of detection, the current research has tended toward highly confocal, microscopy-based analysis, which requires somewhat bulky instrumentation and precisely synthesized SERS substrates. By contrast, in this study we have improved SERS for bacterial analyses using silver colloidal substrates, which are easily and cheaply synthesized in bulk, and which we shall demonstrate permit analysis using portable instrumentation. All analyses were conducted in triplicate to assess the reproducibility of this approach, which was excellent. We demonstrate that SERS is able to detect and quantify rapidly the dipicolinate (DPA) biomarker for Bacillus spores at 5 ppb (29.9 nM) levels which are significantly lower than those previously reported for SERS and well below the infective dose of 10(4)B. anthracis cells for inhalation anthrax. Finally we show the potential of multivariate data analysis to improve detection levels in complex DPA extracts from viable spores.