scholarly journals Neutralizing Antibodies to Adenylate Cyclase Toxin Promote Phagocytosis of Bordetella pertussis by Human Neutrophils

2000 ◽  
Vol 68 (12) ◽  
pp. 7152-7155 ◽  
Author(s):  
Christine L. Weingart ◽  
Paula S. Mobberley-Schuman ◽  
Erik L. Hewlett ◽  
Mary C. Gray ◽  
Alison A. Weiss
Keyword(s):  
Monoclonal Antibodies ◽  
Adenylate Cyclase ◽  
Bordetella Pertussis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Immune Serum ◽  
Human Neutrophils ◽  
Wild Type ◽  
Human Immune ◽  
Adenylate Cyclase Toxin

ABSTRACT A previous study showed that opsonization with human immune serum could either promote or antagonize phagocytosis of Bordetella pertussis by human neutrophils depending on whether the bacteria expressed adenylate cyclase toxin. Opsonization of the wild-type strain inhibited phagocytosis relative to unopsonized controls. In contrast, mutants lacking adenylate cyclase toxin were efficiently phagocytosed when opsonized with human immune serum. In this study, we examined opsonization in the presence or absence of monoclonal antibodies to adenylate cyclase toxin. Addition of neutralizing monoclonal antibodies to adenylate cyclase toxin converted a serum that previously inhibited both attachment and phagocytosis of the wild-type strain to one that increased both attachment and phagocytosis compared to the no-serum control. Monoclonal antibodies that recognize the adenylate cyclase toxin but fail to neutralize activity were without effect. These results suggest that adenylate cyclase toxin inhibits both Fc receptor-mediated attachment and phagocytosis of B. pertussis by neutrophils.

scholarly journals Bordetella pertussis Virulence Factors Affect Phagocytosis by Human Neutrophils

2000 ◽  
Vol 68 (3) ◽  
pp. 1735-1739 ◽  
Author(s):  
Christine L. Weingart ◽  
Alison A. Weiss
Keyword(s):  
Adenylate Cyclase ◽  
Virulence Factors ◽  
Pertussis Toxin ◽  
Bordetella Pertussis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Human Neutrophils ◽  
Wild Type ◽  
Dermonecrotic Toxin ◽  
Adenylate Cyclase Toxin

ABSTRACT The interaction between human neutrophils and wild-typeBordetella pertussis or mutants expressing altered lipopolysaccharide or lacking virulence factors—pertussis toxin, adenylate cyclase toxin, dermonecrotic toxin, filamentous hemagglutinin (FHA), pertactin, or BrkA—was examined. In the absence of antibodies, the wild-type strain and the mutants, with the exception of mutants lacking FHA, attached efficiently to neutrophils. The addition of opsonizing antibodies caused a significant reduction (approximately 50%) in attachment of the wild-type strain and most of the mutants expressing FHA, suggesting that bacterium-mediated attachment is more efficient than Fc-mediated attachment. Phagocytosis was also examined. In the absence of antibodies, about 12% of the wild-type bacteria were phagocytosed. Opsonization caused a statistically significant reduction in phagocytosis (to 3%), possibly a consequence of reduced attachment. Phagocytosis of most of the mutants was similar to that of the wild type, with the exception of the mutants lacking adenylate cyclase toxin. About 70% of the adenylate cyclase toxin mutants were phagocytosed, but only in the presence of opsonizing antibody, suggesting that Fc receptor-mediated signaling may be needed for phagocytosis. These studies indicate that FHA mediates attachment ofB. pertussis to neutrophils, but adenylate cyclase toxin blocks phagocytosis.

scholarly journals Pertussis Toxin and Adenylate Cyclase Toxin Provide a One-Two Punch for Establishment of Bordetella pertussis Infection of the Respiratory Tract

2005 ◽  
Vol 73 (5) ◽  
pp. 2698-2703 ◽  
Author(s):  
Nicholas H. Carbonetti ◽  
Galina V. Artamonova ◽  
Charlotte Andreasen ◽  
Nicholas Bushar
Keyword(s):  
Adenylate Cyclase ◽  
Respiratory Tract ◽  
Pertussis Toxin ◽  
Bordetella Pertussis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Infection Model ◽  
Wild Type ◽  
Adenylate Cyclase Toxin ◽  
Tract Infections

ABSTRACT Previously we found that pertussis toxin (PT), an exotoxin virulence factor produced by Bordetella pertussis, plays an important early role in colonization of the respiratory tract by this pathogen, using a mouse intranasal infection model. In this study, we examined the early role played by another exotoxin produced by this pathogen, adenylate cyclase toxin (ACT). By comparing a wild-type strain to a mutant strain (ΔCYA) with an in-frame deletion of the cyaA gene encoding ACT, we found that the lack of ACT confers a significant peak (day 7) colonization defect (1 to 2 log10). In mixed-infection experiments, the ΔCYA strain was significantly outcompeted by the wild-type strain, and intranasal administration of purified ACT did not increase colonization by ΔCYA. These data suggest that ACT benefits the bacterial cells that produce it and, unlike PT, does not act as a soluble factor benefiting the entire infecting bacterial population. Comparison of lower respiratory tract infections over the first 4 days after inoculation revealed that the colonization defect of the PT deletion strain was apparent earlier than that of ΔCYA, suggesting that PT plays an earlier role than ACT in the establishment of B. pertussis infection. Examination of cells in the bronchoalveolar lavage fluid of infected mice revealed that, unlike PT, ACT does not appear to inhibit neutrophil influx to the respiratory tract early after infection but may combat neutrophil activity once influx has occurred.

scholarly journals The Adenylate Cyclase Toxin of Bordetella pertussis Binds to Target Cells via the αMβ2 Integrin (Cd11b/Cd18)

2001 ◽  
Vol 193 (9) ◽  
pp. 1035-1044 ◽  
Author(s):  
Pierre Guermonprez ◽  
Nadia Khelef ◽  
Eric Blouin ◽  
Philippe Rieu ◽  
Paola Ricciardi-Castagnoli ◽  
...  
Keyword(s):  
Cell Death ◽  
Monoclonal Antibodies ◽  
Adenylate Cyclase ◽  
Cell Lines ◽  
Bordetella Pertussis ◽  
Chinese Hamster Ovary Cells ◽  
Human Neutrophils ◽  
Target Cells ◽  
Intracellular Camp ◽  
Adenylate Cyclase Toxin

The adenylate cyclase toxin (CyaA) of Bordetella pertussis is a major virulence factor required for the early phases of lung colonization. It can invade eukaryotic cells where, upon activation by endogenous calmodulin, it catalyzes the formation of unregulated cAMP levels. CyaA intoxication leads to evident toxic effects on macrophages and neutrophils. Here, we demonstrate that CyaA uses the αMβ2 integrin (CD11b/CD18) as a cell receptor. Indeed, the saturable binding of CyaA to the surface of various hematopoietic cell lines correlated with the presence of the αMβ2 integrin on these cells. Moreover, binding of CyaA to various murine cell lines and human neutrophils was specifically blocked by anti-CD11b monoclonal antibodies. The increase of intracellular cAMP level and cell death triggered by CyaA intoxication was also specifically blocked by anti-CD11b monoclonal antibodies. In addition, CyaA bound efficiently and triggered intracellular cAMP increase and cell death in Chinese hamster ovary cells transfected with αMβ2 (CD11b/CD18) but not in cells transfected with the vector alone or with the αXβ2 (CD11c/CD18) integrin. Thus, the cellular distribution of CD11b, mostly on neutrophils, macrophages, and dendritic and natural killer cells, supports a role for CyaA in disrupting the early, innate antibacterial immune response.

scholarly journals Probing the Function of Bordetella bronchiseptica Adenylate Cyclase Toxin by Manipulating Host Immunity

1999 ◽  
Vol 67 (3) ◽  
pp. 1493-1500 ◽  
Author(s):  
Eric T. Harvill ◽  
Peggy A. Cotter ◽  
Ming Huam Yuk ◽  
Jeff F. Miller
Keyword(s):  
Adenylate Cyclase ◽  
Type Strain ◽  
Low Dose ◽  
Wild Type Strain ◽  
Bordetella Bronchiseptica ◽  
Polymorphonuclear Neutrophil ◽  
Wild Type ◽  
Phagocytic Cells ◽  
Adenylate Cyclase Toxin ◽  
Rag 1

ABSTRACT We have examined the role of adenylate cyclase-hemolysin (CyaA) by constructing an in-frame deletion in the Bordetella bronchiseptica cyaA structural gene and comparing wild-type andcyaA deletion strains in natural host infection models. Both the wild-type strain RB50 and its adenylate cyclase toxin deletion (ΔcyaA) derivative efficiently establish persistent infections in rabbits, rats, and mice following low-dose inoculation. In contrast, an inoculation protocol that seeds the lower respiratory tract revealed significant differences in bacterial numbers and in polymorphonuclear neutrophil recruitment in the lungs from days 5 to 12 postinoculation. We next explored the effects of disarming specific aspects of the immune system on the relative phenotypes of wild-type and ΔcyaA bacteria. SCID, SCID-beige, or RAG-1−/− mice succumbed to lethal systemic infection following high- or low-dose intranasal inoculation with the wild-type strain but not the ΔcyaA mutant. Mice rendered neutropenic by treatment with cyclophosphamide or by knockout mutation in the granulocyte colony-stimulating factor locus were highly susceptible to lethal infection by either wild-type or ΔcyaA strains. These results reveal the significant role played by neutrophils early in B. bronchiseptica infection and by acquired immunity at later time points and suggest that phagocytic cells are a primary in vivo target of theBordetella adenylate cyclase toxin.

scholarly journals Fluorescent Labels Influence Phagocytosis ofBordetella pertussis by Human Neutrophils

1999 ◽  
Vol 67 (8) ◽  
pp. 4264-4267 ◽  
Author(s):  
Christine L. Weingart ◽  
Gina Broitman-Maduro ◽  
Gary Dean ◽  
Simon Newman ◽  
Mark Peppler ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Virulence Factor ◽  
Fluorescent Protein ◽  
Fluorescein Isothiocyanate ◽  
Immune Serum ◽  
Human Neutrophils ◽  
Ofbordetella Pertussis ◽  
Adenylate Cyclase Toxin ◽  
Green Fluorescent

ABSTRACT To explore the role of neutrophil phagocytosis in host defense against Bordetella pertussis, bacteria were labeled extrinsically with fluorescein isothiocyanate (FITC) or genetically with green fluorescent protein (GFP) and incubated with adherent human neutrophils in the presence or absence of heat-inactivated human immune serum. In the absence of antibodies, FITC-labeled bacteria were located primarily on the surface of the neutrophils with few bacteria ingested. However, after opsonization, about seven times more bacteria were located intracellularly, indicating that antibodies promoted phagocytosis. In contrast, bacteria labeled intrinsically with GFP were not efficiently phagocytosed even in the presence of opsonizing antibodies, suggesting that FITC interfered with a bacterial defense. Because FITC covalently modifies proteins and could affect their function, we tested the effect of FITC on adenylate cyclase toxin activity, an important extracellular virulence factor. FITC-labeled bacteria had fivefold-less adenylate cyclase toxin activity than did unlabeled wild-type bacteria or GFP-expressing bacteria, suggesting that FITC compromised adenylate cyclase toxin activity. These data demonstrated that at least one extracellular virulence factor was affected by FITC labeling and that GFP is a more appropriate label forB. pertussis.

scholarly journals Pertussis Toxin Plays an Early Role in Respiratory Tract Colonization by Bordetella pertussis

2003 ◽  
Vol 71 (11) ◽  
pp. 6358-6366 ◽  
Author(s):  
Nicholas H. Carbonetti ◽  
Galina V. Artamonova ◽  
R. Michael Mays ◽  
Zoe E. V. Worthington
Keyword(s):  
Respiratory Tract ◽  
Pertussis Toxin ◽  
Bordetella Pertussis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Saline Control ◽  
Control Analysis ◽  
Wild Type ◽  
Respiratory Tract Colonization ◽  
Infection Experiments

ABSTRACT In this study, we sought to determine whether pertussis toxin (PT), an exotoxin virulence factor produced exclusively by Bordetella pertussis, is important for colonization of the respiratory tract by this pathogen by using a mouse intranasal infection model. By comparing a wild-type Tohama I strain to a mutant strain with an in-frame deletion of the ptx genes encoding PT (ΔPT), we found that the lack of PT confers a significant peak (day 7) colonization defect (1 to 2 log10 units) over a range of bacterial inoculum doses and that this defect was apparent within 1 to 2 days postinoculation. In mixed-strain infection experiments, the ΔPT strain showed no competitive disadvantage versus the wild-type strain and colonized at higher levels than in the single-strain infection experiments. To test the hypothesis that soluble PT produced by the wild-type strain in mixed infections enhanced respiratory tract colonization by ΔPT, we coadministered purified PT with the ΔPT inoculum and found that colonization was increased to wild-type levels. This effect was not observed when PT was coadministered via a systemic route. Intranasal administration of purified PT up to 14 days prior to inoculation with ΔPT significantly increased bacterial colonization, but PT administration 1 day after bacterial inoculation did not enhance colonization versus a phosphate-buffered saline control. Analysis of bronchoalveolar lavage fluid samples from mice infected with either wild-type or ΔPT strains at early times after infection revealed that neutrophil influx to the lungs 48 h postinfection was significantly greater in response to ΔPT infection, implicating neutrophil chemotaxis as a possible target of PT activity promoting B. pertussis colonization of the respiratory tract.

scholarly journals Characterization of Human Bactericidal Antibodies to Bordetella pertussis

1999 ◽  
Vol 67 (3) ◽  
pp. 1424-1431 ◽  
Author(s):  
Alison A. Weiss ◽  
Paula S. Mobberley ◽  
Rachel C. Fernandez ◽  
ChrisAnna M. Mink
Keyword(s):  
Immune Response ◽  
Bactericidal Activity ◽  
Bordetella Pertussis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Resistance Mechanism ◽  
Wild Type ◽  
Occupationally Exposed ◽  
Bactericidal Activities

ABSTRACT The Bordetella pertussis BrkA protein protects against the bactericidal activity of complement and antibody; however, some individuals mount an immune response that overcomes this bacterial defense. To further characterize this process, the bactericidal activities of sera from 13 adults with different modes of exposure toB. pertussis (infected as adults, occupational exposure, immunized with an acellular vaccine, or no identified exposure) against a wild-type strain and a BrkA complement-sensitive mutant were evaluated. All of the sera killed the BrkA mutant, suggesting past exposure to B. pertussis or cross-reactive organisms. Several samples had no or minimal activity against the wild type. All of the sera collected from the infected and occupationally exposed individuals but not all of the sera from vaccinated individuals had bactericidal activity against the wild-type strain, suggesting that some types of exposure can induce an immune response that can overcome the BrkA resistance mechanism. Adsorbing serum with the wild-type strain removed the bactericidal antibodies; however, adsorbing the serum with a lipopolysaccharide (LPS) mutant or an avirulent (bvg mutant) strain did not always result in loss of bactericidal activity, suggesting that antibodies to either LPS orbvg-regulated proteins could be bactericidal. All the samples, including those that lacked bactericidal activity, contained antibodies that recognized the LPS of B. pertussis. Bactericidal activity correlated best with the presence of the immunoglobulin G3 (IgG3) antibodies to LPS, the IgG subtype that is most effective at fixing complement.

Impact of sarA Mutation on Immune System Evasion and Stress Response in Staphylococcus aureus

2020 ◽  
Vol 29 (3) ◽  
pp. 105-112
Author(s):  
Yomna A. Hagag ◽  
Abdelaziz Elgaml ◽  
Ramadan Hassan ◽  
Hany I. Kenawy
Keyword(s):  
Staphylococcus Aureus ◽  
Immune System ◽  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Human Neutrophils ◽  
Bacterial Survival ◽  
Wild Type ◽  
The Impact

Background: Staphylococcus aureus is a major human pathogen responsible for a large number of infections. In S. aureus, SarA is an important global locus responsible for the regulation of virulence factors, as well as biofilm formation. Objectives: The aim of this work is to clarify the impact of SarA on biofilm formation, immune system evasion, as well as the survival of S. aureus under stress conditions. Methodology: A comparative study between S. aureus wild type strain, sarA mutant and complemented strains was established addressing the biofilm formation, opsonization, phagocytosis, as well as ability of the bacterium to survive in stressful environments including acidic pH, hyperosmotic and oxidative stress. The in vitro experiments were confirmed by challenging of mice via intraperitoneal injection with the wild type strain, sarA mutant and complemented strains. Results: Mutation of sarA diminished significantly biofilm formation. Moreover, this mutation resulted in a slight decrease in the deposition of the most important opsonin in complement-mediated immunity, named C3 on S. aureus cells. However, this mutation was associated with a significant enhancement of bacterial phagocytosis and killing by human neutrophils. Furthermore, this mutation altered bacterial survival in stressful conditions. It is also noteworthy that sarA mutation resulted in a significant higher survival rates during the challenging of mice. Conclusion: SarA plays a role as a key regulator of biofilm formation, which in turn has a great impact on immune system evasion through affecting opsonization and phagocytosis. In addition, SarA improves the ability of S. aureus to survive in stressful conditions.

scholarly journals Heme Transport Contributes to In Vivo Fitness of Bordetella pertussis during Primary Infection in Mice

2006 ◽  
Vol 74 (3) ◽  
pp. 1741-1744 ◽  
Author(s):  
Timothy J. Brickman ◽  
Carin K. Vanderpool ◽  
Sandra K. Armstrong
Keyword(s):  
Bordetella Pertussis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Relative Fitness ◽  
Transport Systems ◽  
Receptor Protein ◽  
Wild Type ◽  
Iron Starvation ◽  
Virulence Attenuation

ABSTRACT Bordetella pertussis, the causative agent of whooping cough or pertussis, is an obligate human pathogen with multiple high-affinity iron transport systems. Maximal expression of the dedicated heme utilization functions encoded by the hurIR bhuRSTUV genes requires an iron starvation signal to relieve Fur repression at the hurIR promoter-operator and an inducing signal supplied by heme for HurI-mediated transcriptional activation at the bhuRSTUV promoter. The BhuR outer membrane receptor protein is required for heme uptake and for heme sensing for induction of bhuRSTUV transcription. It was hypothesized that heme utilization contributed to the success of B. pertussis as a pathogen. In this study, virulence attenuation resulting from inactivation of the B. pertussis heme system was assessed using mixed infection competition experiments in a mouse model. As a measure of in vivo fitness, the ability of a B. pertussis heme utilization mutant to colonize and persist was determined relative to that of an isogenic coinfecting wild-type strain. Relative fitness of the mutant strain declined significantly after 7 days postinfection and continued to decline throughout the remainder of the 28-day infection time course. In parallel infections using inocula supplemented with an inducing 2 μM concentration of hemin chloride, hemin coadministration augmented the competitive advantage of the wild-type strain over the mutant. The results confirm that heme utilization contributes to the pathogenesis of B. pertussis in the mouse infection model and indicate that heme utilization may be most important for adaptation to host conditions existing during the later stages of infection.

scholarly journals Effect of Melanin and Carotenoids ofExophiala (Wangiella) dermatitidis on Phagocytosis, Oxidative Burst, and Killing by Human Neutrophils

1999 ◽  
Vol 67 (1) ◽  
pp. 94-101 ◽  
Author(s):  
Norbert Schnitzler ◽  
Heidrun Peltroche-Llacsahuanga ◽  
Nicole Bestier ◽  
Josef Zündorf ◽  
Rudolf Lütticken ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Oxidative Burst ◽  
Fungal Pathogen ◽  
Type Strain ◽  
Wild Type Strain ◽  
Redox Properties ◽  
Colony Count ◽  
Human Neutrophils ◽  
Wild Type ◽  
Neutrophil Oxidative Burst

ABSTRACT The black yeast Exophiala (Wangiella)dermatitidis is an increasingly recognized pathogen and a leading cause of severe pheohyphomycosis. Melanin is thought to contribute to the virulence of E. dermatitidis. Whereas the synthesis and the redox properties of melanin have been studied intensively, the influence of melanin and carotenoids on the phagocytosis, the oxidative burst, and the killing of E. dermatitidis by human neutrophils has not been studied. To study their effects on these phenomena, we applied a combination of flow cytometry and a colony-count-dependent method. Using E. dermatitidis wild-type strain 8565 and several melanin-deficient mutants that have been described previously, we demonstrate that melanin prevents this pathogen from being killed in the phagolysosome of the neutrophils. Melanin did not influence the phagocytosis or the oxidative burst of the neutrophils involved. The carotenoids torulene and torularhodine were not found to contribute to the prevention of killing. The ability of E. dermatitidis to block the effects of the neutrophil oxidative burst may critically impair the potential of the host to sufficiently eliminate this fungal pathogen and thus may play an important role in the pathogenesis of phaeohyphomycosis.

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