|Cox, Nelson - Nac|
|Cason Jr, John|
Submitted to: Meeting Abstract
Publication Type: Proceedings
Publication Acceptance Date: 10/5/2011
Publication Date: 10/12/2011
Citation: Cox Jr, N.A., Cason Jr, J.A., Cray, P.J. 2011. Influence of culture methods on recovery of Salmonella serotypes. Proceedings of the 46th National Meeting on Poultry Health and Processing. October 11-13, 2011, Ocean City, Maryland. p.119-122.
Technical Abstract: Current laboratory methods were developed to obtain the greatest number of Salmonella-positive samples at an acceptable cost and different types of samples can contain different microbial competitors, nutrients, and antimicrobial inhibitors. Each of these result in turn affect a different response regarding Salmonella growth and recovery. Further, the degree of stress or sub lethal injury that the bacteria have undergone are often underappreciated or more often ignored when selecting cultural methodology for Salmonella. Influence on serotype further compounds the problem and is discussed below. Quite interestingly, even for a relatively well-understood sample type such as a chilled poultry carcass, there is no internationally recognized standard methodology for Salmonella detection, either for sample size or culture. Use of different cultural media is not simply a matter of laboratory choice as methods can be decided by regional, national, or international organizations. It is well known that many microbiologists/laboratories use a different culture method which further complicates comparison of recovery rates. Classical cultural techniques for Salmonella detection generally include a nonselective preenrichment (which may be used depending on whether cells are stressed or are present in low numbers), a selective enrichment, isolation on selective agar media, biochemical screening with triple sugar and lysine iron agars, and serological confirmation with poly-O and poly-H antisera. Many different media formulations have been developed for the preenrichment, enrichment, and selective isolation steps in the detection sequence. A survey of diagnostic veterinary laboratories in the United States revealed that 17 different selective enrichment media were being used (Waltman & Mallinson 1995).There were also differences in incubation temperatures, whether samples were incubated for 24 h or 48 h or both, or whether samples were subjected to delayed secondary enrichment conditions. Selective enrichment cultures were inoculated onto 14 different plating media, with most labs using two or more different types of media to increase the likelihood of recovering Salmonella. Finally, the survey revealed considerable variation in numbers of colonies selected for further testing by the different laboratories, with many selecting and identifying only one colony. Types of liquid and solid media have also changed over time. Lactose broth, Gram negative broth, Rappaport (Vassiliadis) broth, selenite cystine broth, and tetrathionate broth were used years ago. While all of these broths are still used to some degree significant composition modifications have occurred and selenite cystine broth is use the least. Plating media for Salmonella isolation included brilliant green, bismuth sulfite, Hektoen enteric, Salmonella-Shigella, and XLD agars. Some of these are rarely used today while others have been modified. Brilliant green with sulfapyridine added is now called BGS. XLD has had many modifications, including addition of sodium thiosulfate and Tergitol 4 to make XLT4 agar, one of the more popular agars used today. Food microbiology laboratories usually use two or more plating media to reduce the occurrence of false-negative results (Cox & Berrang 2000). Rappaport’s enrichment medium was modified to become the widely used Rappaport-Vassiliadis (RV) broth (Vassiliadis 1983) and was then further modified with the development of modified semisolid RV (MSRV) for selection of motile Salmonella (De Smedt et al. 1986). The main reason for development and adoption of different media has always been to isolate the maximum number of colonies of Salmonella serotypes other than Typhi. The widespread use of MSRV in Europe during the past 20 years may have played a major role in the reported differences in isolation rates and prevalence of different serotypes