Effects of Above-Optimum Growth Temperature and Cell Morphology on Thermotolerance of Listeria monocytogenes Cells Suspended in Bovine Milk

Abstract

The thermotolerances of two different cell forms of Listeria monocytogenes (serotype 4b) grown at 37 and 42.8°C in commercially pasteurized and laboratory-tyndallized whole milk (WM) were investigated. Test strains, after growth at 37 or 42.8°C, were suspended in WM at concentrations of approximately 1.5 × 108 to 3.0 × 108 cells/ml and were then heated at 56, 60, and 63°C for various exposure times. Survival was determined by enumeration on tryptone-soya-yeast extract agar andListeria selective agar, and D values (decimal reduction times) and Z values (numbers of degrees Celsius required to cause a 10-fold change in the D value) were calculated. Higher average recovery and higher D values (i.e., seen as a 2.5- to 3-fold increase in thermotolerance) were obtained when cells were grown at 42.8°C prior to heat treatment. A relationship was observed between thermotolerance and cell morphology of L. monocytogenes. AtypicalListeria cell types (consisting predominantly of long cell chains measuring up to 60 μm in length) associated with rough (R) culture variants were shown to be 1.2-fold more thermotolerant than the typical dispersed cell form associated with normal smooth (S) cultures (P ≤ 0.001). The thermal death-time (TDT) curves of R-cell forms contained a tail section in addition to the shoulder section characteristic of TDT curves of normal single to paired cells (i.e., S form). The factors shown to influence the thermoresistance of suspended Listeria cells (P ≤ 0.001) were as follows: growth and heating temperatures, type of plating medium, recovery method, and cell morphology. Regression analysis of nonlinear data can underestimate survival of L. monocytogenes; the end point recovery method was shown to be a better method for determining thermotolerance because it takes both shoulders and tails into consideration. Despite their enhanced heat resistance, atypical R-cell forms of L. monocytogenes were unable to survive the low-temperature, long-time pasteurization process when freely suspended and heated in WM.

Carbon : nitrogen : phosphorus ratios influence biofilm formation byEnterobacter cloacaeandCitrobacter freundii

“…Higher concentrations of nutrients in the bulk fluid may have favoured cells remaining in the planktonic/free-swimming form as opposed to attaching to the GAC surfaces, thus accounting for this latter result. Furthermore, previous studies have suggested that environmental stresses may result in altered cell shapes and surface roughness (Rowan and Anderson 1998;Rowan et al 2000;To et al 2002) and that high C : N ratios may indicate inadequate N supplies (Lin and Lay 2004). Thus, the altered cell shapes observed in scanning electron micrographs of attached populations at higher C : N ratios may be due to stressed cell populations.…”

Cellular morphology of rough forms of Listeria monocytogenes isolated from clinical and food samples

“…The CAMP test was performed using standard procedures by streaking out bacteria perpendicular to Staphylococcus aureus on 5% sheep blood agar plates and observing zones of augmented haemolysis, as described elsewhere (Bubert et al 1997). Characteristic blue-green sheen from colonies by obliquely transmitted light and tumbling motility of Listeria cells were determined as described elsewhere (Rowan and Anderson 1998). The commercial biochemical API Listeria test (bioMerieux, Marcy l'Etoile, France) was used according to the manufactures instructions.…”

“…L. monocytogenes RIII, another rough mutant derived from a smooth strain of serovar 1/2a, was obtained from J. Potel (Institute for Medical Microbiology, Medical Academy, Hannover, Germany). The rough variants SURI and SURII were derived from parent S1 and S2 strains, respectively, under conditions of heat stress (Rowan and Anderson 1998). Stored bacteria were kept at À70 C in phosphate-buffered saline (PBS) with 20% glycerol (v/v) until used.…”

Pulsed electric field inactivation of diarrhoeagenic Bacillus cereus through irreversible electroporation

“…The BCSA enumerated data is also observed to be consistently lower than that of the TSYEA data for treatments of 1000 pulses and more. This difference in recovery is possibly due to the added stress of dyes and antibiotics in the selective plating medium (Rowan and Anderson 1998;Rowan 1999).…”