Listeria monocytogenes serotype 4b strains replicate in monocytes/macrophages more than the other serotypes

Abstract

We analyzed the pathogenicity of various serotypes of Listeria monocytogenes using a Balb/c mouse intravenous injection model. The survival rates of mice inoculated with strains NS1/2b (serotype 1/2b), NS3b (serotype 3b) and NS 4b (serotype 4b) were 60, 63.6 and 63.6%, respectively. Although the survival rates were similar, the bacterial growth in the liver of NS3b-infected mice was 144.5-fold higher than that in the liver of NS4b-infected mice. Histopathological analyses suggest that the NS4b strain replicated more in monocytes/macrophages, whereas the NS3b strain replicated more in hepatocytes. These results raise a possibility that the serotype 4b strains replicated more in monocytes/macrophages compared to the other serotype strains. To assess this, we isolated CD11b-positive cells from mouse livers infected with EGDe (serotype 1/2a), NS1/2b, NS3b, NS4b and the serotype 4b strains 51414 and F17 and counted the number of live bacteria in these cells. CD11b-positive cells from the NS4b-, 51414- and F17-infected mice possessed 24.4- to 42.7-fold higher numbers of live bacteria than those from mice infected with EGDe and NS3b strains. These results suggest that serotype 4b strains replicated more in monocytes/macrophages than the other serotypes, and this may be involved in the pathogenicity of serotype 4b strains, particularly in the dissemination of L. monocytogenes through the host body.

Keywords: Listeria monocytogenes; monocyte/macrophage; pathogenicity; serotype.

Fig. 1.

Survival curve of Balb/c mice infected with 7 serotypes of L. monocytogenes. Eight to 11 mice were inoculated intravenously with each strain. Survival of mice was observed until 7 dpi.

Fig. 2.

Bacterial growth of 7 serotypes of L. monocytogenes. (a) Bacterial growth in the liver. (b) Bacterial growth in the spleen. Mice were inoculated intravenously with each strain. At 1, 3 and 6 dpi, the livers and spleens were harvested from 3 or 4 mice for each strain, and live bacteria were counted. The detection limit was 10² CFU/g tissue. The bacterial counts in the livers of EGDe-, NS3b- and NS4b-infected mice at 3 dpi were enclosed by dashed circles for emphasis. White dots, 1 dpi; light gray dots, 3 dpi; dark gray dots, 6 dpi. Bars indicate the means of three or four samples.

Fig. 3.

Histopathology of L. monocytogenes-infected Balb/c mice. (a) Abscess in the liver of NS1/2b-infected mouse. (b) Liver of mock-infected mouse. (c) Perivascular infiltration of macrophages in the liver of NS4b-infected mouse. Arrowheads surround the area with macrophage infiltration. (d) Magnified image of the area surrounded by arrowheads in (c). (e) Migration of macrophages in the splenic sinus of NS4b-infected mouse. (f) Spleen of mock-infected mouse. (g, h) Bacterial clumps in the dead mice, observed in the liver of NS4b-infected mouse (g) and in the kidney of NS3b-infected mouse (h). Arrowheads show the bacterial clumps.

Fig. 4.

Immunostaining for L. monocytogenes antigens. (a–g) Distribution of the L. monocytogenes antigens in the livers of EGDe-(a), NS3b-(b), mock-(c), NS-1/2b-(d, e) and NS4b-infected (f, g) mice. (g) Magnified image of the enclosed area of (f). (h) HE staining of the serial sections of (g). (i–k) Double immunofluorescence staining of L. monocytogenes antigens and Iba1 in the liver of NS4b-infected mouse. Green, L. monocytogenes; Red, Iba1; Blue, Nuclei. (l–n) Distribution of L. monocytogenes antigens in the spleens of NS3b-(l), NS4b-(m) and mock-infected (n) mice.

Fig. 5.

Bacterial counts in CD11b-positive cells isolated from L. monocytogenes-infected mouse livers. Three or four mice were inoculated intravenously with each strain. At 3 dpi, CD11b-positive cells were isolated from the liver, and live bacteria were counted. *P<0.05 vs EGDe (Dunnett’s test).