Acid stress induced persistence of Listeria monocytogenes and the differential expression of stress response genes in revived persister cell survivors

Transcription

1 Acid stress induced persistence of Listeria monocytogenes and the differential expression of stress response genes in revived persister cell survivors Thulani Sibanda, Elna M. Buys Department of Consumer and Food Sciences, University of Pretoria, Private Bag X20, Hatfield 0028, South Africa South African Society of Dairy Technology (SASDT) 51 st Symposium Kievits Kroon, Country Estate, Kameeldrift-East, Pretoria

2 INTRODUCTION Listeria monocytogenes: The cause of listeriosis DoH and NICD statistics as of 20 April laboratory-confirmed cases Nearly 200 deaths 2

3 Food processing plants are major reservoirs of outbreak L. monocytogenes Persistence in processing plants. Isolation of the same sequence type from one plant over months and years Recurrent contamination of finished products 3

4 Listeria persistence has been explained in terms of resistance to sanitizing agents and ability to form biofims. Experimental data has not yielded any conclusive evidence to suggest that persistent strains are more resistant. Parallels exist between persistent infections that survive antibiotic therapy and persistent survival of stress in the environment. Persister cells survive antibiotic therapy without resistance.

5 Therefore, in this study, we sought to: Test the hypothesis that persister cell formation as a stress response mechanism, aids L. monocytogenes persistence. Examine the expression profiles of persitent L. monocytogenes survivors when presented with growth permitting conditions. 5

6 EXPERIMENTAL AND RESULTS L. monocytogenes Strain 159/10 Lethal lactic acid stress Survivors were tested for susceptibility L o g 1 0 C F U /m l 6 4 NS (p > 0.05) H r L a c tic A c id C o n c e n tra tio n (m M ) 6

7 L o g 1 0 C F U /m l Surviving cells were as susceptible as the parental generation NO RESISTANCE 4 3 NS (p > 0.05) W T P 1 P 2 P 3 P 4 P 5 P e rs is te r Is o la te s 7

8 Cell Counts Normalized cell counts Cell Counts Persistent survivors of L. monocytogenes are in a metabolically inactive dormant state fitting the definition of persister cells. B Control cells Treated cells Unstained cells CFDA Fluorescence 8

9 Persister cell formation is accompanied by formation of extracellular vesicles and filamentation Control cells Persister cells A B C 9

10 R e la tiv e E x p re s s io n F o ld C h a n g e (L o g 2 ) When persister cells are revived under cold growth, their relative expression levels of stress related genes are not different from their parental generation. A reduced expression of PTS genes responsible for sugar uptake A 4 W IL D -T Y P E P E R S IS T E N T s ig B g a d D 2 o p p A lm o lm o * 10

11 The reduced expression of PTS genes means a poorer ability to utilize nutrients and slow growth. Thus impacting on the potential detectability of such cells by growth dependent methods. 11

12 Conclusions This study concluded that ; Although persistence of L. monocytogenes is often attributable to resistance and biofilm formation our findings indicate that a dormant persister cell state makes the organism to be highly tolerant to stress. When dormant persister cells resume growth, their expression profiles of most stress response genes indicates a lack of heritable genetic differences. However persister cells possess a reduced expression of genes for uptake of sugars indicating a potential for slow growth. 12

13 Acknowledgements University of Pretoria. DST-NRF Centre of Excellence in Food Security. South African Society of Dairy Technology (SASDT) 13