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. 2023 Jan 12;8(1):58.
doi: 10.3390/tropicalmed8010058.

In Vitro and In Vivo Virulence Study of Listeria monocytogenes Isolated from the Andalusian Outbreak in 2019

Andrea Vila Domínguez  1   2 , Marta Carretero Ledesma  1   2 , Carmen Infante Domínguez  1   2 , José Miguel Cisneros  1   2   3 , Jose A Lepe  1   2   3 , Younes Smani  1   3   4
Affiliations

In Vitro and In Vivo Virulence Study of Listeria monocytogenes Isolated from the Andalusian Outbreak in 2019

Andrea Vila Domínguez et al. Trop Med Infect Dis. .

Abstract

In 2019, the biggest listeriosis outbreak by Listeria monocytogenes (Lm) in the South of Spain was reported, resulting in the death of three patients from 207 confirmed cases. One strain, belonging to clonal complex 388 (Lm CC388), has been isolated. We aimed to determine the Lm CC388 virulence in comparison with other highly virulent clones such as Lm CC1 and Lm CC4, in vitro and in vivo. Four L. monocytogenes strains (Lm CC388, Lm CC1, Lm CC4 and ATCC 19115) were used. Attachment to human lung epithelial cells (A549 cells) by these strains was characterized by adherence and invasion assays. Their cytotoxicities to A549 cells were evaluated by determining the cells viability. Their hemolysis activity was determined also. A murine intravenous infection model using these was performed to determine the concentration of bacteria in tissues and blood. Lm CC388 interaction with A549 cells is non-significantly higher than that of ATCC 19115 and Lm CC1, and lower than that of Lm CC4. Lm CC388 cytotoxicity is higher than that of ATCC 19115 and Lm CC1, and lower than that of Lm CC4. Moreover, Lm CC388 hemolysis activity is lower than that of the Lm CC4 strain, and higher than that of Lm CC1. Finally, in the murine intravenous infection model by Lm CC388, higher bacterial loads in tissues and at similar levels of Lm CC4 were observed. Although a lower rate of mortality of patients during the listeriosis outbreak in Spain in 2019 has been reported, the Lm CC388 strain has shown a greater or similar pathogenicity level in vitro and in an animal model, like Lm CC1 and Lm CC4.

Keywords: Listeria; animal model; outbreak; pathogenicity; virulence.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
L. monocytogenes adherence and invasion in host cells. (A) Analysis of bacterial adherence in A549 cells incubated with L. monocytogenes Lm CC88, Lm CC1, Lm CC4 and ATCC 19115 strains. (B) Analysis of bacterial invasion in A549 cell incubated with L. monocytogenes Lm CC88, Lm CC1, Lm CC4 and ATCC 19115 strains. Data are the means of 3 repetitive assays. *: p < 0.05 versus Lm CC4 strain.
Figure 2
Figure 2
Cytotoxic activity of L. monocytogenes. Cell viability of A549 cells with L. monocytogenes Lm CC388, Lm CC1, Lm CC4 and ATCC 19115 strains for 24 h. Data are the means of 3 repetitive assays. *: p < 0.05 versus control, #: p < 0.05 versus ATCC 19115.
Figure 3
Figure 3
Hemolytic activity of L. monocytogenes. Hemolysis of murine erythrocytes after 4 h of incubation with L. monocytogenes Lm CC388, Lm CC1, Lm CC4 and ATCC 19115 strains. Data are the means of 3 different assays. *: p < 0.05 versus control, #: p < 0.05 versus ATCC 19115 or Lm CC1.
Figure 4
Figure 4
Bacterial burden in intravenous murine model by L. monocytogenes. Bacterial concentrations in spleen, lungs, liver, kidneys and in the murine model of peritoneal sepsis by L. monocytogenes Lm CC388, Lm CC1, Lm CC4 and ATCC 19115 strains at 5 log10 CFU/mL. Data are represented as mean ± SEM. *: p < 0.05 versus ATCC 19115, #: p < 0.05 versus Lm CC1.
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