Temperature adaptation and plasmid transformation in Listeria monocytogenes ATCC 7644

Abstract

As a recently recognized food-borne pathogen, L. monocytogenes has caused great concerns in food industry due to its ability to survive and grow in many refrigerated foods. The first part of this thesis research was to examine the ability of L. monocytogenes to adapt to changing environmental temperature, and the second part was to develop a suitable plasmid transformation system using electroporation.

It was found that L. monocytogenes ATCC 7644 had a temperature profile of growth with a break point of 15$\sp\circ$C. Energetically, it was considerably more difficult for L. monocytogenes to grow at temperatures below 15$\sp\circ$C than above. Both superoxide dismutase and catalase, however, showed greater specific activities when cultures were adapted at 5$\sp\circ$C than at 37$\sp\circ$C at all assay temperatures. This may suggest the L. monocytogenes grown at lower temperatures might be more likely to survive phagocytosis and be a more able pathogen.

In the second part of this research, it was discovered that L. monocytogenes ATCC 7644 had a natural cryptic plasmid of 20 to 21 md, designated as pMW 01. AFter electroporating with plasmid pAM$\beta$1, three Em$\sp{\rm r}$ Listeria derivatives--T-1, T-2 and T-3, were categorized and characterized. In comparison to the parental strain, these three Em$\sp{\rm r}$ derivatives were morphologically, biochemically and genetically different. Due to the high frequency of appearance, they were not likely to be the results of spontaneous or electroporation induced mutations. All three Em$\sp{\rm r}$ derivatives lost their native pMW 01 plasmid, both T-1 and T-2 contain two new plasmids--pMW 11 (5.4 md), pMW 12 (16 md) in T-1, and pMW 21 (2.8 md), pMW 22 (15 md) in T-2. More interestingly, homology between pAM$\beta$1 or pMW 01 and the four new species of plasmids could not be demonstrated by the Southern blot hybridization experiments. More work was suggested for optimizing the electroporation process and for looking further into the genetics of Listeria.