Short term Candida albicans colonization reduces Pseudomonas aeruginosa-related lung injury and bacterial burden in a murine model

Abstract

Introduction: Pseudomonas aeruginosa is a frequent cause of ventilator-acquired pneumonia (VAP). Candida tracheobronchial colonization is associated with higher rates of VAP related to P. aeruginosa. This study was designed to investigate whether prior short term Candida albicans airway colonization modulates the pathogenicity of P. aeruginosa in a murine model of pneumonia and to evaluate the effect of fungicidal drug caspofungin.

Methods: BALB/c mice received a single or a combined intratracheal administration of C. albicans (1 × 10(5) CFU/mouse) and P. aeruginosa (1 × 10(7) CFU/mouse) at time 0 (T0) upon C. albicans colonization, and Day 2. To evaluate the effect of antifungal therapy, mice received caspofungin intraperitoneally daily, either from T0 or from Day 1 post-colonization. After sacrifice at Day 4, lungs were analyzed for histological scoring, measurement of endothelial injury, and quantification of live P. aeruginosa and C. albicans. Blood samples were cultured for dissemination.

Results: A significant decrease in lung endothelial permeability, the amount of P. aeruginosa, and bronchiole inflammation was observed in case of prior C. albicans colonization. Mortality rate and bacterial dissemination were unchanged by prior C. albicans colonization. Caspofungin treatment from T0 (not from Day 1) increased their levels of endothelial permeability and lung P. aeruginosa load similarly to mice receiving P. aeruginosa alone.

Conclusions: P. aeruginosa-induced lung injury is reduced when preceded by short term C. albicans airway colonization. Antifungal drug caspofungin reverses that effect when used from T0 and not from Day 1.

Figure 1

C. albicans tracheobronchial colonization in mice and dose-dependent pathophysiological effects. A. C. albicans clearance from lungs. C. albicans loads in lungs two days after the intratracheal instillation of 1 × 10⁵ and 1 × 10⁶ CFU/mouse. CFU were counted on YPD plates. The data are means ± standard error (SE) (indicated by error bars). n = 5 mice per group. B. Effect of C. albicans on alveolar-capillary barrier permeability. Evaluation of endothelial permeability (EP) of the alveolar-capillary barrier to ¹²⁵I-labeled bovine serum albumin two days after the intratracheal instillation of 1 × 10⁵ and 1 × 10⁶ CFU/mouse of C. albicans. The data are means ± SE (indicated by error bars). n = 5 mice per group.

Figure 2

Effect of previous C. albicans tracheobronchial colonization on P. aeruginosa-related lung injury. A. BALB/c mice survival. Effect of C. albicans (Ca), P. aeruginosa (Pa), and P. aeruginosa after C. albicans (CaPa) on mouse survival during four days after intratracheal instillation of a dose of 1 × 10⁵ CFU/mouse of C. albicans at T0 and of 1 × 10⁷ CFU/mouse for P. aeruginosa administrated at Day 2 post-colonization. n = 8 mice per group. B. Effect of C. albicans and P. aeruginosa on alveolar-capillary barrier permeability. Evaluation of endothelial permeability (EP) of the alveolar-capillary barrier to ¹²⁵I-labeled bovine serum albumin four days after the intratracheal instillation of a saline solution (Ctr) and in Ca, Pa, CaPa groups. The data are means ± SE (indicated by error bars). n = 8 mice per group.

Figure 3

P. aeruginosa CFU count in lungs. Live P. aeruginosa count in lung homogenates (CFU/ml) at Day 4 in Ctr, Ca, Pa, CaPa groups and caspofungin-treated group at the dose of 1 mg/kg the first day and 0.8 mg/kg the following days until Day 4, from T0 (CaPaCasp0) or from Day 1 (CaPaCasp1). The data are means ± SE (indicated by error bars). n = 4 mice per group.

Figure 4

Lung histopathology after sequential infection with C. albicans and P. aeruginosa in mice. A. Histological score of lung sections from BALB/c mice on Day 4. Peribranchiol and perivascular lung inflammation in mice was measured by two independent blinded examiners. Data are expressed as mean ±SE for each group. P < 0.05 for CaPa vs Pa mice. B. Immunofluorescence and periodic acid Schiff staining for C. albicans localization in lungs of BALB/c mice on Day 4. (a) Representative section of lung from mice challenged with both C. albicans and P. aeruginosa stained with fluorescent galenthus nivalis lectin (GNL) specific for terminal α-D-mannosyl, preferentially α-1,3 residues of C. albicans. The scale bars represent 10 μm. (b) Lung section from mouse receiving C. albicans and P. aeruginosae stained with PAS (periodic acid Schiff). The scale bars represent 5 μm.

Figure 5

Effect of caspofungin on alveolar-capillary barrier permeability. Evaluation of endothelial permeability (EP) of the alveolar-capillary barrier to ¹²⁵I-labeled bovine serum albumin at Day 4 in Ctr, Ca, Pa, CaPa groups and caspofungin-treated group at the dose of 1 mg/kg the first day and 0.8 mg/kg the following days until Day 4, from T0 (CaPaCasp0) or from Day 1 (CaPaCasp1). The data are means ± SE (indicated by error bars). n = 8 mice per group.