Dry and heat stress affects H2S production of Salmonella on selective plating media

Authors: RICHARDSON, KURT - Anitox Corp; Cox Jr, Nelson; Cosby, Douglas; Berrang, Mark; HOLCOMBE, NICHOLE - Anitox Corp; WELLER, CHERYL - Anitox Corp

Submitted to: Journal of Environmental Science and Health
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/29/2018
Publication Date: 1/29/2019
Citation: Richardson, K.E., Cox Jr, N.A., Cosby, D.E., Berrang, M.E., Holcombe, N.L., Weller, C.E. 2019. Dry and heat stress affects H2S production of Salmonella on selective plating media. Journal of Environmental Science and Health. 54:313-316.

Interpretive Summary: Preenrichment media used to analyze feeds for Salmonella can become very acidic during incubation. Salmonella in feed that have been stressed by heat and dryness exhibit different tolerances to acidity when compared to non-stressed cultures. For example, in the case of S. Typhimurium and S. Enteritidis desiccation and heat reduces their ability to tolerate acidity and they are rarely found in feed. In contrast, S. Senftenberg has an increased acid tolerance when exposed to heat and dry stress and is one of the more commonly isolated from feed. These results suggest that the acidic conditions that occur in feed analysis can influence which serotypes are detected.

Technical Abstract: The pH of Salmonella pre-enrichment media can become acidic (pH 4.0 to 5.0) when feeds and ingredients are incubated for 24 h. Salmonella in poultry feed that have been stressed by heat and desiccation exhibit different tolerances to the lower pHs than non-stressed cultures. Acidic conditions affect the biochemical pathways and injure or kill Salmonella. In this study, eight serotypes, four commonly found in feed (S. Montevideo (SM), S. Senftenberg (SS), S. Tennessee (STn) and S. Schwarzengrund (SSc)) and four commonly found in poultry processing plants (S. Typhimurium (STy), S. Enteritidis (SE), S. Infantis (SI), and S. Heidelberg (SH)), were grown in sterile meat and bone meal for 48 h at 35°C, centrifuged, and the sediment subjected to desiccation at 37°C for 36-48 h under partial vacuum to expose them to stress. Isolates were subsequently exposed to acidic pH from 4.0 to 7.0 in 0.5 pH increments (3 replicates/pH increment) in citrate buffer. At 6 and 24 h, serial dilutions were plated in duplicate on xylose lysine tergitol 4 (XLT4) agar. Four serotypes (SE, SI, SM and SSc) showed an impaired ability to decarboxylate lysine on XLT4. At a pH of 6.5, 56.6 - 57.1% of the colonies of unstressed cultures of SSc and SI were H2S negative after 6 h of incubation. This percentage of H2S negative colonies significantly (P<0.05) increased for SSc at 24 h. Stressed SE and SI exhibited the greatest overall change in the ability of the isolate to decarboxylate lysine on XLT4. At a pH of 6, 97.5% of the SE colonies were H2S positive at 6 h, however, at 24 h, 87.5% of the colonies were H2S negative. In the case of SI, 90.2% of the colonies were H2S negative at 6 h. Results at 24 h were not significantly different (P<0.05). At a pH of 6, stressed cultures of SM and SSc produced an approximate 50:50 ratio of H2S positive: H2S negative colonies at both 6 and 24 h. When the isolates ability to decarboxylate lysine was further examined using the API20 biochemical test strip, with the exception of SI, cultures were still able to decarboxylate lysine. This suggests that XLT4 agar contains a biochemical stressor(s) which affects the rate of decarboxylation by these Salmonella instead of the isolate inability to decarboxylate lysine. These results suggest that acidic conditions may influence the detection and confirmation of Salmonella in feed.