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Host cell-derived lactate functions as an effector molecule in Neisseria meningitidis microcolony dispersal

View Article: PubMed Central - PubMed

ABSTRACT

The development of meningococcal disease, caused by the human pathogen Neisseria meningitidis, is preceded by the colonization of the epithelial layer in the nasopharynx. After initial adhesion to host cells meningococci form aggregates, through pilus-pilus interactions, termed microcolonies from which the bacteria later detach. Dispersal from microcolonies enables access to new colonization sites and facilitates the crossing of the cell barrier; however, this process is poorly understood. In this study, we used live-cell imaging to investigate the process of N. meningitidis microcolony dispersal. We show that direct contact with host cells is not required for microcolony dispersal, instead accumulation of a host-derived effector molecule induces microcolony dispersal. By using a host-cell free approach, we demonstrated that lactate, secreted from host cells, initiate rapid dispersal of microcolonies. Interestingly, metabolic utilization of lactate by the bacteria was not required for induction of dispersal, suggesting that lactate plays a role as a signaling molecule. Furthermore, Neisseria gonorrhoeae microcolony dispersal could also be induced by lactate. These findings reveal a role of host-secreted lactate in microcolony dispersal and virulence of pathogenic Neisseria.

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Lactate- and CM-induced microcolony dispersal of N. meningitidis occurs independent of oxygen depletion.Samples were taken 10, 20 and 40 min after addition of DMEM (control), CM or L-lactate (10 mM) to FAM20 microcolonies. The concentration of ATP (A) and the NAD+/NADH (B) ratio were measured. (C) Microcolony stability in both DMEM and GC liquid during oxygen depletion. At 1.5 h the oxygen level was set to 0% by a 95% N2/5% CO2 flow. At 5 h, the oxygen level was reversed to 95% air/5% CO2. The dotted lines in the panel represent the 1.5 h and 5 h time points. (D) Addition of DMEM (control), L-lactate (10 mM) and pyruvate (10 mM) to preformed microcolonies. Data represents the mean ± SD of three independent experiments. A black horizontal line in panel D represents the 3 h time point.
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ppat.1006251.g008: Lactate- and CM-induced microcolony dispersal of N. meningitidis occurs independent of oxygen depletion.Samples were taken 10, 20 and 40 min after addition of DMEM (control), CM or L-lactate (10 mM) to FAM20 microcolonies. The concentration of ATP (A) and the NAD+/NADH (B) ratio were measured. (C) Microcolony stability in both DMEM and GC liquid during oxygen depletion. At 1.5 h the oxygen level was set to 0% by a 95% N2/5% CO2 flow. At 5 h, the oxygen level was reversed to 95% air/5% CO2. The dotted lines in the panel represent the 1.5 h and 5 h time points. (D) Addition of DMEM (control), L-lactate (10 mM) and pyruvate (10 mM) to preformed microcolonies. Data represents the mean ± SD of three independent experiments. A black horizontal line in panel D represents the 3 h time point.

Mentions: One factor that controls microcolony dispersal in N. gonorrhoeae is the oxygen concentration. Oxygen depletion leads to dispersal through pili retraction, which is mediated by depletion of the proton motive force (PMF) [15]. Since it has been shown that lactate stimulates growth and oxygen consumption in the presence of glucose [32], one hypothesis was that this increased oxygen consumption leads to oxygen depletion and dispersal. Changes in PMF affect the ATP level and the NAD+/NADH ratio. To determine if depletion of the PMF caused dispersal in response to lactate addition, we measured the concentration of ATP and the ratio of NAD+/NADH after induction of microcolony dispersal with either lactate or CM. Our results showed that the ATP concentration and the NAD+/NADH ratio remained similar or increased slightly (Fig 8A and 8B). The addition of carbonyl cyanide m-chlorophenyl hydrazone (CCCP, 25 μM) caused a decrease in ATP concentration (S7 Fig). This suggests that a depletion of the PMF caused by oxygen depletion was not the cause of dispersal. In addition, we were unable to induce dispersal by decreasing the environmental oxygen concentration of bacteria grown in DMEM (Fig 8C), in contrast to bacteria grown in GC as shown by Dewenter et al. [15]. However, one of the reasons for the observed differences in GC and DMEM for the role of oxygen depletion in microcolony dispersal may be due to differences in the rate at which oxygen depletion takes places in both the medium. Further investigation is required to conclusively determine the role of oxygen depletion in the lactate-induced accelerated microcolony dispersal.


Host cell-derived lactate functions as an effector molecule in Neisseria meningitidis microcolony dispersal
Lactate- and CM-induced microcolony dispersal of N. meningitidis occurs independent of oxygen depletion.Samples were taken 10, 20 and 40 min after addition of DMEM (control), CM or L-lactate (10 mM) to FAM20 microcolonies. The concentration of ATP (A) and the NAD+/NADH (B) ratio were measured. (C) Microcolony stability in both DMEM and GC liquid during oxygen depletion. At 1.5 h the oxygen level was set to 0% by a 95% N2/5% CO2 flow. At 5 h, the oxygen level was reversed to 95% air/5% CO2. The dotted lines in the panel represent the 1.5 h and 5 h time points. (D) Addition of DMEM (control), L-lactate (10 mM) and pyruvate (10 mM) to preformed microcolonies. Data represents the mean ± SD of three independent experiments. A black horizontal line in panel D represents the 3 h time point.
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ppat.1006251.g008: Lactate- and CM-induced microcolony dispersal of N. meningitidis occurs independent of oxygen depletion.Samples were taken 10, 20 and 40 min after addition of DMEM (control), CM or L-lactate (10 mM) to FAM20 microcolonies. The concentration of ATP (A) and the NAD+/NADH (B) ratio were measured. (C) Microcolony stability in both DMEM and GC liquid during oxygen depletion. At 1.5 h the oxygen level was set to 0% by a 95% N2/5% CO2 flow. At 5 h, the oxygen level was reversed to 95% air/5% CO2. The dotted lines in the panel represent the 1.5 h and 5 h time points. (D) Addition of DMEM (control), L-lactate (10 mM) and pyruvate (10 mM) to preformed microcolonies. Data represents the mean ± SD of three independent experiments. A black horizontal line in panel D represents the 3 h time point.
Mentions: One factor that controls microcolony dispersal in N. gonorrhoeae is the oxygen concentration. Oxygen depletion leads to dispersal through pili retraction, which is mediated by depletion of the proton motive force (PMF) [15]. Since it has been shown that lactate stimulates growth and oxygen consumption in the presence of glucose [32], one hypothesis was that this increased oxygen consumption leads to oxygen depletion and dispersal. Changes in PMF affect the ATP level and the NAD+/NADH ratio. To determine if depletion of the PMF caused dispersal in response to lactate addition, we measured the concentration of ATP and the ratio of NAD+/NADH after induction of microcolony dispersal with either lactate or CM. Our results showed that the ATP concentration and the NAD+/NADH ratio remained similar or increased slightly (Fig 8A and 8B). The addition of carbonyl cyanide m-chlorophenyl hydrazone (CCCP, 25 μM) caused a decrease in ATP concentration (S7 Fig). This suggests that a depletion of the PMF caused by oxygen depletion was not the cause of dispersal. In addition, we were unable to induce dispersal by decreasing the environmental oxygen concentration of bacteria grown in DMEM (Fig 8C), in contrast to bacteria grown in GC as shown by Dewenter et al. [15]. However, one of the reasons for the observed differences in GC and DMEM for the role of oxygen depletion in microcolony dispersal may be due to differences in the rate at which oxygen depletion takes places in both the medium. Further investigation is required to conclusively determine the role of oxygen depletion in the lactate-induced accelerated microcolony dispersal.

View Article: PubMed Central - PubMed

ABSTRACT

The development of meningococcal disease, caused by the human pathogen Neisseria meningitidis, is preceded by the colonization of the epithelial layer in the nasopharynx. After initial adhesion to host cells meningococci form aggregates, through pilus-pilus interactions, termed microcolonies from which the bacteria later detach. Dispersal from microcolonies enables access to new colonization sites and facilitates the crossing of the cell barrier; however, this process is poorly understood. In this study, we used live-cell imaging to investigate the process of N. meningitidis microcolony dispersal. We show that direct contact with host cells is not required for microcolony dispersal, instead accumulation of a host-derived effector molecule induces microcolony dispersal. By using a host-cell free approach, we demonstrated that lactate, secreted from host cells, initiate rapid dispersal of microcolonies. Interestingly, metabolic utilization of lactate by the bacteria was not required for induction of dispersal, suggesting that lactate plays a role as a signaling molecule. Furthermore, Neisseria gonorrhoeae microcolony dispersal could also be induced by lactate. These findings reveal a role of host-secreted lactate in microcolony dispersal and virulence of pathogenic Neisseria.

No MeSH data available.


Related in: MedlinePlus