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Characterization of Bacillus anthracis persistence in vivo.

Jenkins SA, Xu Y - PLoS ONE (2013)

Bottom Line: Immunofluorescence staining of lung sections showed that spores associated with the alveolar and airway epithelium.We also showed that the anthrax toxins did not play a role in persistence.Together, the results suggest that B. anthracis spores have special properties that promote their persistence in the lung, and that there may be multiple mechanisms contributing to spore persistence.

View Article: PubMed Central - PubMed

Affiliation: Center for Infectious and Inflammatory Diseases, Institute of Biosciences and Technology, Texas A&M Health Science Center, Houston, Texas, USA.

ABSTRACT
Pulmonary exposure to Bacillus anthracis spores initiates inhalational anthrax, a life-threatening infection. It is known that dormant spores can be recovered from the lungs of infected animals months after the initial spore exposure. Consequently, a 60-day course antibiotic treatment is recommended for exposed individuals. However, there has been little information regarding details or mechanisms of spore persistence in vivo. In this study, we investigated spore persistence in a mouse model. The results indicated that weeks after intranasal inoculation with B. anthracis spores, substantial amounts of spores could be recovered from the mouse lung. Moreover, spores of B. anthracis were significantly better at persisting in the lung than spores of a non-pathogenic Bacillus subtilis strain. The majority of B. anthracis spores in the lung were tightly associated with the lung tissue, as they could not be readily removed by lavage. Immunofluorescence staining of lung sections showed that spores associated with the alveolar and airway epithelium. Confocal analysis indicated that some of the spores were inside epithelial cells. This was further confirmed by differential immunofluorescence staining of lung cells harvested from the infected lungs, suggesting that association with lung epithelial cells may provide an advantage to spore persistence in the lung. There was no or very mild inflammation in the infected lungs. Furthermore, spores were present in the lung tissue as single spores rather than in clusters. We also showed that the anthrax toxins did not play a role in persistence. Together, the results suggest that B. anthracis spores have special properties that promote their persistence in the lung, and that there may be multiple mechanisms contributing to spore persistence.

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Bacterial and spore burden in the lungs of mice at 2, 4, and 8 weeks post-intranasal inoculation.Mice were inoculated i.n. with ∼1.1×108 or ∼1.3×107 spores/mouse. Lungs were harvested at 2, 4, and 8 weeks, homogenized, and dilution plated with or without heat treatment. The results were combined from at least two independent experiments. Closed circles represent total viable bacteria and open circles heat-resistant dormant spores. *, p<0.05; **, p<0.01; compared to respective total bacteria and spore titers at 2 weeks.
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pone-0066177-g001: Bacterial and spore burden in the lungs of mice at 2, 4, and 8 weeks post-intranasal inoculation.Mice were inoculated i.n. with ∼1.1×108 or ∼1.3×107 spores/mouse. Lungs were harvested at 2, 4, and 8 weeks, homogenized, and dilution plated with or without heat treatment. The results were combined from at least two independent experiments. Closed circles represent total viable bacteria and open circles heat-resistant dormant spores. *, p<0.05; **, p<0.01; compared to respective total bacteria and spore titers at 2 weeks.

Mentions: BALB/c mice were challenged intranasally (i.n.) with sub-lethal doses of spores of B. anthracis Sterne strain 7702 (pXO1+, pXO2−). BALB/c mice are generally more resistant to the Sterne strain [18] and therefore provided us a model to assay for persistence of spores where mice can survive weeks post initial infection. The presence of vegetative bacilli and spores in different organs was evaluated over a period of 8 weeks. The results indicated that substantial amounts of bacteria were recovered from the lungs of mice at 2, 4 and 8 weeks post-inoculation (Fig. 1 and Table 1). The majority of the bacteria recovered were heat-resistant dormant spores; however heat-sensitive vegetative bacilli were also detected at all three time points. Immunofluorescence staining of lung sections with antibodies specific for vegetative bacilli also showed positive staining (Supplemental figure S1), indicating that spore germination occurred in the lung although at a relatively low frequency. We observed decreases in the total bacteria and spore titers in the lung over the experimental period, suggesting a continuous host clearance process. The decreases were significantly sharper during the earlier weeks and were relatively moderate from 4 to 8 weeks (Fig. 1 and Table 1).


Characterization of Bacillus anthracis persistence in vivo.

Jenkins SA, Xu Y - PLoS ONE (2013)

Bacterial and spore burden in the lungs of mice at 2, 4, and 8 weeks post-intranasal inoculation.Mice were inoculated i.n. with ∼1.1×108 or ∼1.3×107 spores/mouse. Lungs were harvested at 2, 4, and 8 weeks, homogenized, and dilution plated with or without heat treatment. The results were combined from at least two independent experiments. Closed circles represent total viable bacteria and open circles heat-resistant dormant spores. *, p<0.05; **, p<0.01; compared to respective total bacteria and spore titers at 2 weeks.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3672131&req=5

pone-0066177-g001: Bacterial and spore burden in the lungs of mice at 2, 4, and 8 weeks post-intranasal inoculation.Mice were inoculated i.n. with ∼1.1×108 or ∼1.3×107 spores/mouse. Lungs were harvested at 2, 4, and 8 weeks, homogenized, and dilution plated with or without heat treatment. The results were combined from at least two independent experiments. Closed circles represent total viable bacteria and open circles heat-resistant dormant spores. *, p<0.05; **, p<0.01; compared to respective total bacteria and spore titers at 2 weeks.
Mentions: BALB/c mice were challenged intranasally (i.n.) with sub-lethal doses of spores of B. anthracis Sterne strain 7702 (pXO1+, pXO2−). BALB/c mice are generally more resistant to the Sterne strain [18] and therefore provided us a model to assay for persistence of spores where mice can survive weeks post initial infection. The presence of vegetative bacilli and spores in different organs was evaluated over a period of 8 weeks. The results indicated that substantial amounts of bacteria were recovered from the lungs of mice at 2, 4 and 8 weeks post-inoculation (Fig. 1 and Table 1). The majority of the bacteria recovered were heat-resistant dormant spores; however heat-sensitive vegetative bacilli were also detected at all three time points. Immunofluorescence staining of lung sections with antibodies specific for vegetative bacilli also showed positive staining (Supplemental figure S1), indicating that spore germination occurred in the lung although at a relatively low frequency. We observed decreases in the total bacteria and spore titers in the lung over the experimental period, suggesting a continuous host clearance process. The decreases were significantly sharper during the earlier weeks and were relatively moderate from 4 to 8 weeks (Fig. 1 and Table 1).

Bottom Line: Immunofluorescence staining of lung sections showed that spores associated with the alveolar and airway epithelium.We also showed that the anthrax toxins did not play a role in persistence.Together, the results suggest that B. anthracis spores have special properties that promote their persistence in the lung, and that there may be multiple mechanisms contributing to spore persistence.

View Article: PubMed Central - PubMed

Affiliation: Center for Infectious and Inflammatory Diseases, Institute of Biosciences and Technology, Texas A&M Health Science Center, Houston, Texas, USA.

ABSTRACT
Pulmonary exposure to Bacillus anthracis spores initiates inhalational anthrax, a life-threatening infection. It is known that dormant spores can be recovered from the lungs of infected animals months after the initial spore exposure. Consequently, a 60-day course antibiotic treatment is recommended for exposed individuals. However, there has been little information regarding details or mechanisms of spore persistence in vivo. In this study, we investigated spore persistence in a mouse model. The results indicated that weeks after intranasal inoculation with B. anthracis spores, substantial amounts of spores could be recovered from the mouse lung. Moreover, spores of B. anthracis were significantly better at persisting in the lung than spores of a non-pathogenic Bacillus subtilis strain. The majority of B. anthracis spores in the lung were tightly associated with the lung tissue, as they could not be readily removed by lavage. Immunofluorescence staining of lung sections showed that spores associated with the alveolar and airway epithelium. Confocal analysis indicated that some of the spores were inside epithelial cells. This was further confirmed by differential immunofluorescence staining of lung cells harvested from the infected lungs, suggesting that association with lung epithelial cells may provide an advantage to spore persistence in the lung. There was no or very mild inflammation in the infected lungs. Furthermore, spores were present in the lung tissue as single spores rather than in clusters. We also showed that the anthrax toxins did not play a role in persistence. Together, the results suggest that B. anthracis spores have special properties that promote their persistence in the lung, and that there may be multiple mechanisms contributing to spore persistence.

Show MeSH
Related in: MedlinePlus