Inoculation Preparation Affects Survival of Salmonella enterica on Whole Black Peppercorns and Cumin Seeds Stored at Low Water Activity

Abstract

Salmonellosis has been increasingly associated with contaminated spices. Identifying inoculation and stabilization methods for Salmonella on whole spices is important for development of validated inactivation processes. The objective of this study was to examine the effects of inoculation preparation on the recoverability of Salmonella enterica from dried whole peppercorns and cumin seeds. Whole black peppercorns and cumin seeds were inoculated with S. enterica using one dry transfer method and various wet inoculation methods: immersion of spice seeds in tryptic soy broth (TSB) plus Salmonella for 24 h (likely leading to inclusion of Salmonella in native microbiota biofilms formed around the seeds), application of cells grown in TSB, and/or application of cells scraped from tryptic soy agar (TSA). Postinoculation seeds were dried to a water activity of 0.3 within 24 h and held for 28 days. Seeds were sampled after drying (time 0) and periodically during the 28 days of storage. Salmonella cells were enumerated by serial dilution and plated onto xylose lysine Tergitol (XLT4) agar and TSA. Recovery of Salmonella was high after 28 days of storage but was dependent on inoculation method, with 4.05 to 6.22 and 3.75 to 8.38 log CFU/g recovered from peppercorns and cumin seeds, respectively, on XLT4 agar. The changes in surviving Salmonella (log CFU per gram) from initial inoculation levels after 28 days were significantly smaller for the biofilm inclusion method (+0.142pepper, +0.186cumin) than for the other inoculation methods (−0.425pepper, −2.029cumin for cells grown on TSA; −0.641pepper, −0.718cumin for dry transfer; −1.998pepper for cells grown in TSB). In most cases, trends for reductions of total aerobic bacteria were similar to those of Salmonella. The inoculation method influenced the recoverability of Salmonella from whole peppercorns and cumin seeds after drying. The most stable inoculum strategies were dry transfer, 24-h incubation of Salmonella and spices in TSB (i.e., potential inclusion of Salmonella within native microbiota biofilms), and inoculation of Salmonella cells grown on TSA subsequent to drying. However, with the dry transfer method it was difficult to obtain the large amount of inoculum needed for inactivation studies.