The lipoprotein NlpD in Cronobacter sakazakii responds to acid stress and regulates macrophage resistance and virulence by maintaining membrane integrity: Running Title: Identification and characterization of a novel factor involved in acid tolerance and virulence in Cronobacter sakazakii
Xuemeng Ji,
Ping Lu,
Juan Xue,
Ning Zhao,
Yan Zhang,
Lu Dong,
Xuejiao Zhang,
Ping Li,
Yaozhong Hu,
Jin Wang,
Bowei Zhang,
Jingmin Liu,
Huan lv,
Shuo Wang
Affiliations
Xuemeng Ji
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Ping Lu
Institute of Radiation Medicine, Chinese Academy of Medical Science and Peking Union Medical Collage
Juan Xue
Institute of Infection and Immunity, Taihe Hospital, Hubei University of Medicine
Ning Zhao
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Yan Zhang
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Lu Dong
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Xuejiao Zhang
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Ping Li
Key Laboratory of Food Nutrition and Safety, Ministry of Education, Tianjin University of Science and Technology
Yaozhong Hu
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Jin Wang
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Bowei Zhang
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Jingmin Liu
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Huan lv
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Shuo Wang
Tianjin Key Laboratory of Food Science and Health, School of Medicine, Nankai University
Cronobacter sakazakii, an emerging opportunistic pathogen, is implicated in severe foodborne outbreak infections in premature and full-term infants. Generally, acid tolerance is vital for the pathogenesis of foodborne pathogens; however, its role in C. sakazakii virulence remains largely unknown. To screen out acid-tolerance determinants from transposon mutants, anovel counterselection method using gentamicin and acid was developed. Using the counterselection method and growth assay, we screened several acid-sensitive mutants and found that nlpD encodes an acid-resistance factor in C. sakazakii. Compared to the wild-type strain, the nlpD mutant exhibited attenuated virulence in a rat model. Using macrophage THP-1 cells and a pH probe, we verified that nlpD enables bacteria to resist macrophages by resisting acidification. Finally, we confirmed that nlpD maintains C. sakazakii membrane integrity in acid using propidium iodide permeabilization assays via flow cytometry. Our results confirm that nlpD is a novel virulence factor that permits C. sakazakii to survive under acid stress conditions. Considering that NlpD is a conserved lipoprotein located in the bacterial outer membrane, NlpD could be used as a target for drug development.
