Fig 1.
Survival and bacterial clearance of neonatal mice to MRSA.
(A) C57Bl/6 neonate (5 d) and adult mice (6 wks) were intranasally infected with increasing doses of USA300 (6 mice/group) and survival was determined over a 6 d period. (B) C57Bl/6 pups (5 d) and adult mice (6 wks) were intranasally infected with 1x107 CFU’s of MRSA and the level of bacteria in the lung was measured on 1, 2 3 and 6 dpi. NSA = neonatal / MRSA infection, ASA = adult / MRSA infection.
Fig 2.
Immune cell recruitment into the lungs of neonatal and adult mice infected with MRSA.
Neonatal (5 d) and adult mice (6 wks) were infected with 1x107 CFU’s of MRSA and BAL performed at 1 and 3 dpi. The cells recovered from the BALF were counted by differential staining. NSA = neonatal / MRSA infection, ASA = adult / MRSA infection.
Fig 3.
In vivo phagocytosis of MRSA by neonatal and adult mice.
Neonate and adult mice were infected with 2x107 CFU’s of TAMRA-MRSA or unlabeled MRSA. The lungs were removed 30 min pi and intracellular expression of TAMRA was measured by flow cytometry. The level of background fluorescence measured using unlabeled MRSA was 6.7%. NSA = neonatal / MRSA infection, ASA = adult / MRSA infection.
Fig 4.
Proinflammatory cytokine and anti-microbial peptide expression in neonatal and adult mice.
Neonate and adult mice were infected with 1x107 CFU’s of MRSA and cytokine gene expression measured at 6 and 24 hr pi (A-F). Expression of mRNA for cytokines and anti-microbial peptides was determined by qRT-PCR on RNA isolated from individual lung lobes (n = 5–6 mice per group) and relative expression is fold induction over WT unexposed mice of the same age. The level of IL6 (G)and IFNγ (H) in the lung homogenates was measured by ELISA. The data represent mean ± SEM of duplicate samples and significance was determined using one-way ANOVA with Tukey post-hoc test. NSA = neonatal / MRSA infection, NS = Neonatal / sham infection, ASA = adult / MRSA infection, AS = Adult sham infection.
Fig 5.
Neonatal mice mounted limited Th1 and Th17 response to MRSA infection.
Neonate and adult mice were infected with 1.24 x107 CFU’s of MRSA or vehicle. Lung single cells were isolated at 6 dpi and Th profile was measured by flow cytometry. (A) Th1/Th2 cell percentage of CD4+ T cells. (B) Representative flow plots of Th1/Th2 percentage of CD4+ T cells. (C) Th1/Th17 cell subsets. (D) Representative flow plots of Th1/Th17 subsets. NSA = neonatal / MRSA infection, NS = Neonatal / sham infection, ASA = adult / MRSA infection, AS = Adult sham infection.
Fig 6.
Limited recruitment and maturation of mDCs to the lungs in response to MRSA infection.
Neonate and adult mice were infected with 1.24 x107 CFU’s of MRSA or vehicle. Lung single cells were isolated at 6 dpi and the amount of mDCs and their maturation status were measured by flow cytometry. (A) The frequency of mDCs in the lungs as a percentage of total lung cells. (B) Representative flow plots of mDCs. (C) Maturation status of mDCs in the lung. (D) Histogram of CD86 on mDCs. FMO = fluorescence minus one and is used to identify and gate cells in the complex fluorescent dye panel. NSA = neonatal / MRSA infection, NS = Neonatal / sham infection, ASA = adult / MRSA infection, AS = Adult sham infection.