Bordetella pertussis is a Gram-negative coccobacillus that causes whooping cough or pertussis, a respiratory disease that has recently experienced a resurgence. Upon entering the respiratory tract, B. pertussis colonizes the airway epithelium and attaches to ciliated cells. Here, we used primary human nasal epithelial cells (hNECs) cultured at the air-liquid interface and investigated their interaction with B. pertussis B1917, focusing on the role of the type III secretion system effector protein BteA. In this model, which resembles the epithelial cells of nasal epithelium in vivo, B. pertussis B1917 localized predominantly in the overlying mucus and scarcely colonized the cell cilia. The colonization led to a gradual decline in epithelial barrier function, as shown by measurements of transepithelial electrical resistance (TEER) and staining of the tight junction protein zonula occludens 1. The decrease in TEER occurred independently of the cytotoxic effector protein BteA. Transcriptomic and proteomic analyses of hNECs showed only moderate changes following infection, primarily characterized by increased mucus production, including upregulation of mucin MUC5AC. No profound response to BteA was detected. Furthermore, the infection did not induce production of inflammatory cytokines, suggesting that B. pertussis B1917 evades recognition by hNECs in this model system. These results suggest that the mucus may serve as a niche that allows B. pertussis B1917 to minimize epithelial recognition and damage. The lack of a robust immune response further indicates that additional components of the nasal mucosa, such as innate immune cells, are likely required to initiate an effective host defense.IMPORTANCEThe nasal epithelium is the initial site where Bordetella pertussis comes into contact with the host during respiratory tract infection. In this study, human nasal epithelial cells cultured at the air-liquid interface were established as an in vitro model to investigate the early stages of B. pertussis infection. We showed that the clinical isolate B. pertussis B1917 resides in the mucus during the early stages of colonization without disrupting the epithelial barrier function. Infection results in moderate transcriptomic and proteomic changes, characterized by increased mucus production and minimal inflammatory signaling. These results suggest that B. pertussis B1917 may evade early host recognition by residing in mucus and avoiding direct interaction with epithelial cells. They also highlight the importance of other components of the mucosal immune system, such as resident immune cells, for the initiation of an effective defense.

• Klíčová slova
• Bordetella pertussis, BteA effector, air-liquid interface culture, airway epithelium, human nasal epithelial cell, type III secretion system,
• MeSH
• bakteriální proteiny metabolismus   genetika   MeSH
• Bordetella pertussis * patogenita   genetika   fyziologie   MeSH
• epitelové buňky * mikrobiologie   imunologie   metabolismus   MeSH
• faktory virulence rodu Bordetella metabolismus   MeSH
• hlen mikrobiologie   metabolismus   MeSH
• kultivované buňky MeSH
• lidé MeSH
• mucin 5AC metabolismus   genetika   MeSH
• nosní sliznice * mikrobiologie   cytologie   imunologie   MeSH
• pertuse * mikrobiologie   imunologie   MeSH
• proteomika MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH
• faktory virulence rodu Bordetella MeSH
• mucin 5AC MeSH

The pertussis agent Bordetella pertussis produces a number of virulence factors, of which the filamentous hemagglutinin (FhaB) plays a role in B. pertussis adhesion to epithelial and phagocytic cells. Moreover, FhaB was recently found to play a crucial role in nasal cavity infection and B. pertussis transmission to new hosts. The 367 kDa FhaB protein translocates through an FhaC pore to the outer bacterial surface and is eventually processed to a ~220 kDa N-terminal FHA fragment by the SphB1 protease. A fraction of the mature FHA then remains associated with bacterial cell surface, while most of FHA is shed into the bacterial environment. Previously reported indirect evidence suggested that FHA, or its precursor FhaB, may bind the β2 integrin CD11b/CD18 of human macrophages. Therefore, we assessed FHA binding to various cells producing or lacking the integrin and show that purified mature FHA does not bind CD11b/CD18. Further results then revealed that the adhesion of B. pertussis to cells does not involve an interaction between the bacterial surface-associated FhaB and/or mature FHA and the β2 integrin CD11b/CD18. In contrast, FHA binding was strongly inhibited at micromolar concentrations of heparin, corroborating that the cell binding of FHA is ruled by the interaction of its heparin-binding domain with sulfated glycosaminoglycans on the cell surface.

• Klíčová slova
• Bordetella pertussis, CD11b/CD18, adenylate cyclase toxin, filamentous hemagglutinin, heparin, integrin,
• MeSH
• antigeny CD18 MeSH
• bakteriální adheze MeSH
• bakteriální adheziny metabolismus   MeSH
• Bordetella pertussis * metabolismus   MeSH
• faktory virulence rodu Bordetella MeSH
• glykosaminoglykany MeSH
• hemaglutininy metabolismus   MeSH
• heparin MeSH
• integriny MeSH
• lidé MeSH
• makrofágový antigen 1 MeSH
• pertuse * MeSH
• proteasy MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antigeny CD18 MeSH
• bakteriální adheziny MeSH
• faktory virulence rodu Bordetella MeSH
• glykosaminoglykany MeSH
• hemaglutininy MeSH
• heparin MeSH
• integriny MeSH
• makrofágový antigen 1 MeSH
• proteasy MeSH

The whooping cough agent, Bordetella pertussis, secretes an adenylate cyclase toxin-hemolysin (CyaA, ACT, or AC-Hly) that catalyzes the conversion of intracellular ATP to cAMP and through its signaling annihilates the bactericidal activities of host sentinel phagocytes. In parallel, CyaA permeabilizes host cells by the formation of cation-selective membrane pores that account for the hemolytic activity of CyaA. The pore-forming activity contributes to the overall cytotoxic effect of CyaA in vitro, and it has previously been proposed to synergize with the cAMP-elevating activity in conferring full virulence on B. pertussis in the mouse model of pneumonic infection. CyaA primarily targets myeloid phagocytes through binding of their complement receptor 3 (CR3, integrin αMβ2, or CD11b/CD18). However, with a reduced efficacy, the toxin can promiscuously penetrate and permeabilize the cell membrane of a variety of non-myeloid cells that lack CR3 on the cell surface, including airway epithelial cells or erythrocytes, and detectably intoxicates them by cAMP. Here, we used CyaA variants with strongly and selectively enhanced or reduced pore-forming activity that, at the same time, exhibited a full capacity to elevate cAMP concentrations in both CR3-expressing and CR3-non-expressing target cells. Using B. pertussis mutants secreting such CyaA variants, we show that a selective enhancement of the cell-permeabilizing activity of CyaA does not increase the overall virulence and lethality of pneumonic B. pertussis infection of mice any further. In turn, a reduction of the cell-permeabilizing activity of CyaA did not reduce B. pertussis virulence any importantly. These results suggest that the phagocyte-paralyzing cAMP-elevating capacity of CyaA prevails over the cell-permeabilizing activity of CyaA that appears to play an auxiliary role in the biological activity of the CyaA toxin in the course of B. pertussis infections in vivo.

• Klíčová slova
• Bordetella pertussis, RTX toxin, adenylate cyclase toxin, cAMP intoxication, lung colonization, lung inflammation, pore-forming activity, virulence,
• MeSH
• adenylátcyklasový toxin metabolismus   MeSH
• AMP cyklický metabolismus   MeSH
• Bordetella pertussis patogenita   fyziologie   MeSH
• fagocyty metabolismus   mikrobiologie   MeSH
• interakce hostitele a patogenu MeSH
• lidé MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• ovce MeSH
• permeabilita buněčné membrány MeSH
• pertuse metabolismus   mikrobiologie   patologie   MeSH
• virulence MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• adenylátcyklasový toxin MeSH
• AMP cyklický MeSH