Foci of Listeria monocytogenes persist in the bone marrow.
ABSTRACT Murine listeriosis is one of the most comprehensive and well-studied models of infection, and Listeria monocytogenes has provided seminal information regarding bacterial pathogenesis. However, many aspects of the mouse model remain poorly understood, including carrier states and chronic colonization which represent important features of the spectrum of host-pathogen interaction. Bone marrow has recently been shown to harbor L. monocytogenes, which spreads from this location to the central nervous system. Bone could, therefore, be an important chronic reservoir, but this infection is difficult to study because it involves only a few bacteria and the extent of infection cannot be assessed until after the animal is sacrificed. We employed in vivo bioluminescence imaging to localize L. monocytogenes bone infections over time in live mice, revealing that the bacteria grow in discrete foci. These lesions can persist in many locations in the legs of mice and are not accompanied by a histological indication such as granuloma or a neutrophil infiltratate. We demonstrate that highly attenuated hly mutants, which have defective intracellular replication, are capable of prolonged focal infection of the bone marrow for periods of up to several weeks. These results support the recently proposed hypothesis that the bone marrow is a unique niche for L. monocytogenes.
SourceAvailable from: Gitte Maegaard Knudsen[Show abstract] [Hide abstract]
ABSTRACT: Listeria monocytogenes can cause the serious infection, listeriosis, which despite antibiotic treatment has a high mortality. Understanding the response of L. monocytogenes to antibiotic exposure is therefore important to ensure treatment success. Some bacteria survive antibiotic treatment by formation of persisters, which are a dormant antibiotic-tolerant subpopulation. The purpose of this study was to determine whether L. monocytogenes can form persisters and how bacterial physiology affects the number of persisters in the population. A stationary phase culture of L. monocytogenes was adjusted to 10(8) CFU ml(-1) and 10(3)-10(4) CFU ml(-1) survived 72h treatment with 100 μg ml(-1) norfloxacin indicating a persister subpopulation. This survival was not caused by antibiotic-resistance as regrown persisters were as sensitive to norfloxacin as the parental strain. Higher number of persisters (10(5)-10(6)) was surviving when older stationary phase or surface-associated cells were treated with 100 μg ml(-1) norfloxacin. The number of persisters was similar when a ΔsigB mutant and the wild-type were treated with norfloxacin, but the killing rate was higher in the ΔsigB mutant. Dormant norfloxacin-persisters could be activated by the addition fermentable carbohydrates, and subsequently killed by gentamicin, however, a stable surviving subpopulation of 10(3) CFU ml(-1) remained. Nitrofurantoin that has a growth independent mode-of-action was effective against both growing and dormant cells suggesting that eradication of persisters is possible. Our study adds L. monocytogenes to the list of bacterial species capable of surviving bactericidal antibiotics in a dormant stage and this persister phenomenon should be borne in mind when developing treatment regimens.Applied and Environmental Microbiology 09/2013; DOI:10.1128/AEM.02184-13 · 3.95 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: Listeria monocytogenes (Lm) is a human intracellular pathogen widely used to uncover the mechanisms evolved by pathogens to establish infection. However, its capacity to perturb the host cell cycle was never reported. We show that Lm infection affects the host cell cycle progression, increasing its overall duration but allowing consecutive rounds of division. A complete Lm infectious cycle induces a S-phase delay accompanied by a slower rate of DNA synthesis and increased levels of host DNA strand breaks. Additionally, DNA damage/replication checkpoint responses are triggered in an Lm dose-dependent manner through the phosphorylation of DNA-PK, H2AX, and CDC25A and independently from ATM/ATR. While host DNA damage induced exogenously favors Lm dissemination, the override of checkpoint pathways limits infection. We propose that host DNA replication disturbed by Lm infection culminates in DNA strand breaks, triggering DNA damage/replication responses, and ensuring a cell cycle delay that favors Lm propagation.Cell cycle (Georgetown, Tex.) 01/2014; 13(6). DOI:10.4161/cc.27780 · 5.01 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: In vivo bioluminescence imaging (BLI) is a powerful method for the analysis of host-pathogen interactions in small animal models. The commercially available bioluminescent Listeria monocytogenes strain Xen32 is commonly used to analyse immune functions in knockout mice and pathomechanisms of listeriosis. To analyse and image listerial dissemination after oral infection we have generated a murinised Xen32 strain (Xen32-mur) which expresses a previously described mouse-adapted internalin A. This strain was used alongside the Xen32 wild type strain and the bioluminescent L. monocytogenes strains EGDe-lux and murinised EGDe-mur-lux to characterise bacterial dissemination in orally inoculated BALB/cJ mice. After four days of infection, Xen32 and Xen32-mur infected mice displayed consistently higher rates of bioluminescence compared to EGDe-lux and EGDe-mur-lux infected animals. However, surprisingly both Xen32 strains showed attenuated virulence in orally infected BALB/c mice that correlated with lower bacterial burden in internal organs at day 5 post infection, smaller losses in body weights and increased survival compared to EGDe-lux or EGDe-mur-lux inoculated animals. The Xen32 strain was made bioluminescent by integration of a lux-kan transposon cassette into the listerial flaA locus. We show here that this integration results in Xen32 in a flaA frameshift mutation which makes this strain flagella deficient. The bioluminescent L. monocytogenes strain Xen32 is deficient in flagella expression and highly attenuated in orally infected BALB/c mice. As this listerial strain has been used in many BLI studies of murine listeriosis, it is important that the scientific community is aware of its reduced virulence in vivo.Gut Pathogens 07/2013; 5(1):19. DOI:10.1186/1757-4749-5-19 · 2.07 Impact Factor