Characterization of a Portable, Non-instrumented Incubator for Enrichment of Escherichia coli 0157:H7 and Salmonella Serovar Typhimurium and Detection by Loop Mediated Isothermal Amplification (LAMP)

摘要

We developed a handheld, non-instrumented incubator INI-incubatorJ to enable direct enrichment of pathogenic bacteria in the field. The incubator contains a custom cartridge, enclosed in a thermos, filled with a phase-change material that can store latent energy at the desired temperature (37°C) after addition of approximately 300 mL of boiling water. Temperature profiles recorded in four IMI-incubators over 12 hours (n = 3) illustrated that the devices maintained temperatures above 35°C for over nine hours. To quantify bacterial growth rates in Nl-incubators, cultures of Salmonella Typhimurium (ATCC 14028}, Escherichia coli 0157:H7 (C7927), Escherichia co//K12, and generic E. coli (environmental isolate) were grown in buffered peptone water and subjected to a temperature profile representative of those observed in IMI-incubators. The empirical growth characteristics were used to predict the minimum enrichment periods required to achieve reliable detection (> 95% positive) of a given quantity ofinoculum by loop mediated isothermal amplification (LAMP). Predictions were validated byinoculation of 5 g of spinach with 2.33 ± 0.58 CFU of E. coli 0157:H7 and of 5 g ground chicken inoculated with 4.5 + 0.5 CFU Salmonella Typhimurium, which could be reliably detected by LAMP after 6 and 8 hours (h), respectively (n = 3 each), of enrichment in IMI-incubators.