The development of techniques that could be useful in fields other than biological warfare agents countermeasures such as medical diagnostics, industrial microbiology, and environmental applications have become a very important subject of research. Raman spectroscopy can be used in near field or at long distances from the sample to obtain fingerprinting information of chemical composition of microorganisms. In this research, biochemical components of the cell wall and endospores of Bacillus thuringiensis (Bt) were identified by surface-enhanced Raman scattering (SERS) spectroscopy using silver (Ag) nanoparticles (NPs) reduced by hydroxylamine and borohydride capped with sodium citrate. Activation of “hot spots”, aggregation and surface charge modification of the NPs, was studied and optimized to obtain signal enhancements from Bt by SERS. Slight aggregation of the NPs as well as surface charge modification to a more acidic ambient was induced using small-size borohydride-reduced NPs in the form of metallic suspensions aimed at increasing the Ag NP-Bt interactions. Hydroxylamine-reduced NPs required slight aggregation and no pH modifications in order to obtain high spectral quality results in bringing out SERS signatures of Bt. 1. Introduction Bioterrorism’s high potential for destruction has become a subject of great concern. Fast, efficient, and inexpensive detection techniques for microorganisms have become a very important subject in areas of national defense and homeland security. Techniques such as conventional Raman spectroscopy (RS) and surface-enhanced Raman scattering (SERS) require a minimum amount of sample for a fast detection [1] so that they can be implemented in near field or at long distances from the sample by obtaining fingerprinting information of the chemical composition of microorganisms. The time required for identification of pathogens is a determinant factor of infection related sickness. The development of these techniques could be useful in fields other than biological warfare agents countermeasures such as medical diagnostics [2], industrial microbiology [3], and environmental applications [4]. Bacillus thuringiensis (Bt), a Gram-positive bacterium, is widely renowned for its toxicity on insect larvae and is commercially used as insecticide. Bt, as a bacillus species, has a bacterial life cycle in which it grows as vegetative cell forming endospores as defense mechanism. The bacterial cell wall contains a peptidoglycan layer responsible for strengthening the wall. This rigid layer in a Gram-positive species cell wall is
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