An Improved Medium for the Detection of Salmonella and Shigella Species

0196-4399/00 (see frontmatter) © 2010 Elsevier Clinical Microbiology Newsletter 32:5,2010 The following describes our experience in developing an improved medium for the detection of Salmonella and Shigella species in clinical specimens. Initially, we attempted to build on past improvements of enteric plating agars containing the surfactant Niaproof 4, formerly known as Tergitol 4. However, despite their broad application for detection of Salmonella species in food safety and veterinary medical diagnostics (1-4), Niaproof 4-supplemented (NS) enteric plating agars are limited for use on human clinical specimens because they do not adequately support the growth of Shigella species. Our goal was to develop an NS-type agar that would be sensitive for detecting Shigella as well as Salmonella species and thus be appropriate for the diagnosis of two major human bacterial enteropathogens. We sought to retain the NS agars’ advantages in detecting Salmonella species, which include the ability to prevent overgrowth of members of the Enterobacteriaceae and, usually, complete inhibition of Proteus spp. The latter characteristic increases the medium’s sensitivity for Salmonella detection compared with traditional enteric agars, such as Hektoen enteric (HE), salmonella-shigella (SS), and xylose-lysinedesoxycholate (XLD) agars (1-3). We believed this advantage would apply to detection of Salmonella in human specimens plated directly or after enrichment. However, NS agars required modification to match, or even exceed, the sensitivity of the traditional media (HE, SS, and XLD) for detecting Shigella species and also to match their ability to suppress the growth of enteric contaminants. After numerous trials, we developed a new NS formulation, termed N4 agar (Table 1), that had the requisite sensitivity for isolating Shigella spp. The key factor in stimulating the growth and allowing easy detection of Shigella spp. on N4 Agar was the unusually high concentration (16 g/L) of yeast extract, which is not found in other media (Fig. 1A and 2A, B, and C). Other differences from most agars were the slightly lowered concentration (13.5 g/L) of granulated agar and the omission of NaCl, which further improved the sensitivity of N4 for detecting Shigella spp. The growth of Salmonella colonies remained excellent (Fig. 1B). The presence of “false Salmonellasuspect” (FSS), H2S-positive colonies is a serious problem with many enteric agars (5,6).Advantageously, the inclusion of cellobiose at a 10-g/L concentration in the N4 agar formulation controlled the growth of black FSS Citrobacter youngae colonies (Fig. 3A), whereas HE and SS agars did not (Fig. 3B and C). C. youngae and other Citrobacter spp. are commonly found in human fecal specimens (7,8), and their presence can result in time-consuming laboratory investigation of H2S-positive FSS colonies, a problem that was less common with N4 agar than with traditional agars. We have previously reported that a 10-g/L concentration of lactose controls the production of FSS Citrobacter freundii colonies (9). The same appears to be true for C. youngae with N4 agar’s 10 g/L concentration of cellobiose (Fig. 3A). We believe that, when used by a particular bacterial species, higher concentrations of