INFLUENZA-INFECTED CELLS INCREASE LEVOFLOXACIN TOLERANCE OF STREPTOCOCCUS PNEUMONIAE

HERNANDEZ MORFA, MIRELYS; NICOLAS REINOSO VIZCAINO; NADIA B. OLIVERO; CORTES, PAULO; JAIME, ANDREA; ECHENIQUE J

Mendoza

Congreso; Reuniòn Anual de la Sociedad Argentina de nvestigacion en Bioquimica y Biol Molecular; 2022

SAIB

Streptococcus pneumoniae is a major bacterial pathogen that usually colonizes the upper respiratory tract and causes pneumonia, bacteremia, and meningitis in humans. Pneumococcal infections are generally treated with beta-lactams and fluoroquinolones (FQs), but FQ resistance has been reported in this pathogen. In our lab, we have reported that host cell oxidative stress induces FQ tolerance, an unusual ability to tolerate the antibiotic effects. We have also demonstrated that intracellular survival of S. pneumoniae increases in host cells that are coinfected with the influenza A virus (IAV). This synergistic mechanism depends on the SirRH two-component system, which controls the expression of certain stress genes needed for bacterial survival in host cells. The purpose of this work was to elucidate the role of SirRH in the mechanism of FQ tolerance and the impact of influenza A infection. To determine the contribution of SirR-regulated genes in the formation of FQ-tolerant pneumococci, we mutated genes encoding for enzymes involved in the oxidative stress response, such as psaB (encodes for a subunit of a manganese ABC transporter), nrdH (encodes for a glutaredoxin-like protein), and sirR. The three mutants showed a decreased FQ-tolerance induced by hydrogen peroxide in bacterial cultures, demonstrating that psaB, nrdH, and sirR play an important role in the mechanism of FQ-tolerance induced by oxidative stress. To analyze the effect of IAV infection on FQ tolerance induced by host cell oxidative stress, A549 pneumocytes were coinfected with the IAV and S. pneumoniae. We found a significant increase in FQ tolerance compared to A549 cells infected only with pneumococci. We determined that the cytoplasmic ROS levels were increased in IAV-infected cells, indicating that the IAV-induced respiratory burst is probably involved in this mechanism. When A549 cells were previously infected with IAV for 24 h, and then treated with NAC (a known ROS inhibitor) for 1 h before bacterial infection, we found a decrease in FQ tolerance of S. pneumoniae. To analyze the putative contribution of autophagy to the IAV-induced FQ tolerance in host cells, we coinfected MEF and MEF-atg5-KO (deficient in autophagy) cells with IAV and S. pneumoniae. We observed a similar FQ tolerance level to that observed in A549 cells, however, in MEF-atg5-KO we observed a decreased FQ tolerance related to MEF wt. These results suggest that oxidative stress genes are involved in the FQ tolerance mechanism in S. pneumoniae. Importantly, FQ tolerance is developed in coinfected cells in a ROS-dependent manner, and it occurs in autophagy-proficient cells only. We propose that our findings about FQ tolerance in S. pneumoniae are clinically relevant, due to this mechanism may cause complications in the antimicrobial treatment of pneumococcal infections.