Table 1

Efficiency of intracellular replication and cell-to-cell spread of L. monocytogenes wild-type strain EGD and isogenic mutants with in-frame deletions in various virulence genes

Strain (mutation in gene)Replication efficiency*Spreading efficiency
None (wt)1.01.0
hly 0.6 (± 0.15)0.01 (± 0.01)
actA 0.9 (± 0.2)0.0
plcA 0.9 (± 0.1)0.7 (± 0.2)
plcB 0.7 (± 0.2)0.5 (± 0.2)
plcA + plcB0.4 (± 0.1)0.3 (± 0.15)
plcA + hly + mpl + actA + plcB0.5 (± 0.1)0.0
inlA + inlB1.0 (± 0.05)1.0 (± 0.1)
inlC 0.8 (± 0.1)0.7 (± 0.2)
prfA 0.05 (± 0.01)0.0
hpt 0.1 (± 0.02)0.2 (± 0.1)
  • * Replication efficiency was determined 8 h postinjection of the corresponding strain. At this time, cell spreading into neighboring host cells was still at a low level, and the fluorescent bacteria within the primarily injected host cell could be easily seen and counted. The given values represent an average from 10 successfully microinjected Caco-2 cells in which efficient bacterial replication is seen (about 50% of all successfully microinjected cells). In such host cells microinjected with the wild-type strain (wt), >32 bacteria per microinjected cell was observed (giving a total of at least 320 bacteria per 10 cells). This value (32 or more bacteria per host cell within 8 h) was defined as replication efficiency 1.0, and the efficiency of replication of the mutant strains (all bacteria in 10 successfully injected cells, e.g., in prfA mutant a total of 16 bacteria) was calculated in relation to this value based on the counted intracellular bacteria (each experiment was performed four times; the numbers in parentheses mark the maximal deviation from the average value). 

  • Spreading efficiency was determined 24 h postinjection by counting all fluorescent bacteria within a focus surrounding the primary injected host cell. The given spreading efficiencies represent again average values based on bacterial counts from 20 microinjected Caco-2 cells with the surrounding spreading foci (if existing) related to the number of bacteria within the spreading foci of the wild-type strain (∼500 bacteria per focus), which was again set to 1.0. 

  • Statistically significant differences relative to the wild-type values, with P < 0.01 as determined by Student's t test.