Specific detection and enumeration of Salmonella enterica in food using conventional culture-based methods (CCBM) are time consuming and labor intensive. This study was conducted to develop a rapid S. enterica detection and enumeration method by combining fluorescence in situ hybridization (FISH) with micro-colony formation culture (FISHFC). Specificity tests of the SAL343 probe for S. enterica detection revealed that SAL343-associated fluorescent micro-colonies were observed specifically for S. enterica, but not for any other organisms. This finding suggests that SAL343 is highly specific for detecting S. enterica using FISHFC. For validation, FISHFC with SAL343 was compared to CCBM, with multiple selective agar, using spiked food samples; no significant differences in enumeration were found between FISHFC and CCBM (p > 0.05). The FISHFC method allowed enumeration of S. enterica within 10 h while CCBM allowed enumeration within 5 days. Therefore, the FISHFC method has potential application for more rapid and specific enumeration of S. enterica in food samples compared to other available methods.Keywords: Salmonella enterica, fluorescence in situ hybridization, FISHFC, rapid detection *To whom correspondence should be addressed. E-mail: yamasaki@fish.hokudai.ac.jp IntroductionSalmonella are Gram-negative, facultatively anaerobic, rod-shaped bacteria that exist ubiquitously in the environment and are commonly found in water, animal intestine, reptilian skin and food. Salmonella also causes salmonellosis in humans worldwide due to the consumption of contaminated food such as fresh fruits and vegetables (Matthews, 2006). The genus Salmonella currently consists of two species: S. enterica and S. bongori. S. enterica is comprised of six subspecies: enterica, salamae, arizonae, diarizonae, houtenae and indica, which are further subdivided into more than 2,500 serotypes (Agbaja et al., 2011;Su and Chiu, 2007). Most of the Salmonella responsible for disease in humans and animals are members of S. enterica. S. enterica serotype Enteritidis and Typhimurium are two of the main sources of human food poisoning. Although S. bongori has been reported to infect humans, it is generally associated with cold-blooded animals (Giammanco et al., 2002). Therefore, S. enterica are clinically important pathogens of humans.Routine methods for enumerating Salmonella are traditional microbiological methods based on plating using selective agar and biochemical identification. However, the biochemical characteristics of some isolates may differ from the common phenotypic properties of typical strains. Moreover, it is recommended that multiple selective agars be used for Salmonella detection in food samples (Nye et al., 2002). Thus, the conventional methods for detecting Salmonella are time-consuming and labor-intensive, taking at least several days to obtain the final results (Kim et al., 2007). To overcome technical problems associated with Salmonella detection, a number of molecular-based methods, such as PCR, quantitative PCR (qPCR), enzy...