TNF-α production induced by a human-type PO-CpG ODN2006 was also

TNF-α production induced by a human-type PO-CpG ODN2006 was also increased by co-incubation

with DNase I-treated GpC ODN2006 or DNase I-treated ODN1720 in the cells (Supporting Information Fig. 2). To evaluate the involvement of TLR9 in the DNase I-treated DNA-mediated increase in cytokine production, similar experiments were carried out using splenic macrophages and the production of TNF-α (Fig. 1C) and IL-6 (Fig. 1D) was examined. The addition of LPS, a positive control, induced significant TNF-α production in splenic macrophages from both WT and TLR9 knockout (KO) mice, indicating the ability of these cells to produce cytokines. In the cells from WT mice, DNase I-treated DNA significantly increased the ODN1668-induced production of TNF-α and IL-6. find more However, no such increase was observed in splenic macrophages from TLR9 KO mice. Next, we evaluated the effect of DNase I-treated DNA on the TNF-α production induced by ligands other than ODN1668. The following ligands were selected and used: pCMV-Luc, a double-stranded circular DNA containing many CpG motifs; ODN2216, a CpG ODN with phosphorothioate (PS) bonds at the both ends; PS-1668, a PS-type CpG ODN having the same sequence as ODN1668; non-CpG lipoplex, a complex consisting of pCpG-ΔLuc and cationic liposomes, which was reported to be a ligand for cytosolic DNA

receptors 18, 19; polyI:C, a double-stranded RNA and a ligand for TLR3; LPS, a ligand for TLR4; and imiquimod, a ligand for TLR7 20, 21. Based on preliminary experiments, the concentration of each ligand was set at low levels to avoid saturation of TNF-α production in cells. Each ligand induced Saracatinib supplier significant TNF-α production in RAW264.7 cells at varying levels (Fig. 2, hatched bars). DNase I-treated ODN1720 significantly increased pCMV-Luc-induced TNF-α production, but it hardly affected TNF-α production induced by other ligands (Fig. 2, black bars). Dipeptidyl peptidase Again, ODN1720 showed no significant effects on the TNF-α production induced by any of these ligands (Fig. 2, gray bars). These results indicate that the DNase-I-treated DNA-mediated increase in cytokine production is specific to two TLR9 ligands, ODN1668

and pCMV-Luc. Additionally, we examined the effects of DNase I-treated DNA on TNF-α production induced by another 26-mer ODN containing three potent CpG motifs, 5′-TCGACGTTTTGACGTTTTGACGTTTT-3′. The addition of DNase I-treated ODN1720 also increased the TNF-α production induced by this CpG ODN (data not shown). Taken together, these results suggest that the effect of DNase I-treated ODN1720 on cytokine production is independent of the sequence and length of CpG DNA, and not restricted to single-stranded DNA. To examine which components of DNase I-treated DNA were responsible for the increase in the CpG motif-dependent TNF-α production, RAW264.7 cells were incubated with ODN1668 in the presence of nucleotides or nucleosides (Fig. 3A).

In contrast, circulating IgA levels in control wool lambs remaine

In contrast, circulating IgA levels in control wool lambs remained low and stable following initial larval exposure, and breed differences in circulating IgA were thus larger in control lambs. Greater responses in circulating IgA in response to transient exposure to parasite larvae in control hair lambs suggests a more robust response to this ‘natural’ vaccination protocol. IgA concentrations in hair sheep were somewhat higher than those reported for wool sheep in previous studies. Larval antigen-specific IgA production has been reported to peak between 1 and 2 weeks p.i. (42,44). However, total IgA in serum in our infected lambs continued to increase through 3 weeks

p.i. in both breeds. Previous studies have not observed higher serum IgA concentrations in resistant breeds including the Gulf EPZ-6438 mouse coast native (40), Santa Ines (17) and Barbados Blackbelly (34) compared with susceptible wool breeds, which may indicate that St. Croix hair sheep have a novel resistance mechanism that is absent in other resistant breeds. Globule leucocytes are described as partially

de-granulated intraepithelial mast cells (45) and have been suggested to be responsible for larvae damage and expulsion within the first few days of infection. Unlike eosinophils (46,47), mast cells bind IgE (41), leading to similar co-dependency to that suggested for eosinophils BMN 673 cell line and IgA. Breed differences in abomasal globule leucocytes were not significant in this study, but levels tended to be greater in hair sheep at 27 days p.i. This result is less striking than the 15- to 40-fold increase in globule leucocytes of younger infected hair compared with wool lambs reported by Gamble and Zajac (18). However, breed differences in concentration Protein kinase N1 of globule leucocytes have been reported to be minimal by 1 year of age (3) and hence age differences probably contribute to apparent inconsistencies among studies. Associations of increased globule leucocytes with lower FEC, lower worm numbers and decreased female worm length are present in young animals (11,15,48), suggesting a role for these cells in resistance and are consistent with our favourable association of globule

leucocyte numbers with IgE in the lymph nodes and PCV at 21 days p.i. Measurement of sheep IgE was first reported by Shaw et al. (49) and Kooyman et al. (8) and several studies have shown that sheep infected with GIN have increased total and worm-antigen-specific IgE (8,12,13,50,51). Mean serum IgE levels in our lambs exceeded 60 ng/mL through 16 days p.i. in infected lambs of both hair and wool types. IgE levels were similarly elevated through the first 16 days following exposure and subsequent de-worming in control hair lambs, but were only transiently elevated in control wool lambs over the same period. These patterns suggest a similar change in circulating IgE following infection in the two breeds with a potentially more robust vaccination response in hair lambs, similar to that observed for circulating IgA.

The role of STAT3

for normal signalling of the IL-6 recep

The role of STAT3

for normal signalling of the IL-6 receptor has important consequences for normal host defence. Together with other cytokines such as IL-1β and IL-23, the IL-6/STAT3 pathway is crucial for the normal development of CD4+–T helper type 17 (Th17) cells [6,7]. Because IL-17 has an important role in the activation of neutrophil-dependent immunity [8], defective Th17 generation as a result of STAT3 mutation may play an important role in the pathogenesis of HIES. In a recent paper, Milner et al. have demonstrated that T lymphocytes from patients with HIES are unable to differentiate into Th17 after mitogenic stimulation [9]. These data were supported by two reports that also showed defective generation of Th17 when anti-CD3/anti-CD28/IL-2

or cytokine cocktails were used [10,11]. These studies reported the defective generation of Th17 using mitogenic cocktails in patients with established Venetoclax manufacturer Selleck MK-1775 mutations in the SH2 and DNA-binding domains of STAT3. In contrast, patients with atopic dermatitis and high IgE, but without skin and respiratory infections and without STAT3 mutations, had normal Th17 responses [9,12]. In the present paper, we aimed to extend these initial findings by investigating the generation of Th17 cells and IL-17 production by relevant microbial stimuli for HIES. In addition, we assessed Th17 profiles in three distinct groups of patients: ‘classical’ HIES patients with STAT3 mutations in the SH2/DNA-binding domains, ‘classical’ HIES without STAT3 mutations and a family with ‘variant’ HIES that we described as having a milder clinical phenotype [13], with deletion of a triplet in the linker domain. The differences in the degree of IL-17 production defects after stimulation with Staphylococcus aureus or Candida albicans determined the severity of the clinical phenotype. Eight patients with a clinical diagnosis of HIES at the out-patient clinic for infectious diseases and immunodeficiencies of the Department of General Internal Medicine Ribose-5-phosphate isomerase of Radboud University Nijmegen Medical Centre were enrolled into the study. Three of these patients were family members. After

informed consent, blood was collected from eight healthy, non-smoking volunteers who were free of infectious or inflammatory disease and the enrolled HIES patients by venipuncture into 10 ml ethylenediamine tetraacetic acid (EDTA) syringes (Monoject; BD Vacutainer, Plymouth, UK). STAT3 mutation analysis was kindly performed in the Laboratory of Human Molecular Biology and Genetics, Catholic University of the Sacred Heart, Milan, Italy (head Professor Roberto Colombo). C. albicans American Type Culture Collection (ATCC) MYA-3573 (UC820), a strain well described elsewhere [14], was used. C. albicans was grown overnight in Sabouraud broth at 37°C, cells were harvested by centrifugation, washed twice and resuspended in culture medium (RPMI-1640 Dutch modification; ICN Biomedicals, Aurora, OH, USA) [15]. C.

Antibiotics can clear most infections and have a benefit for indi

Antibiotics can clear most infections and have a benefit for individual patients, but because of the large number of infected people and the increasing resistance to antibiotics, a more realistic

approach is the development of a vaccine. Granted that some experts doubt the possibility of making a protective H. pylori vaccine because the natural infection persists despite the host developing a strong immune response (Blanchard & Czinn, 2000). Yet, the fact that a postinfection immune response is not able to clear an infection does not necessarily negate the possibility that preinfection immunity may prevent the acquisition of a new infection. In fact, experimental animal selleck chemical data suggest that oral administration of Helicobacter-specific antibodies may be effective to prevent as well as to treat Helicobacter infection (Czinn et al., 1993; Casswall et al., 2002; Gorell & Robins-Browne, 2009). For 20 years, a number of researchers have been working toward the development of a vaccine to prevent H. pylori infection (Czinn & Nedrud, 1991). Of the various candidate antigens, the most promising is the B subunit of the urease protein (urease B), a 65-kDa protein encoded in a 1.7-kbp gene. The protein, which is exposed on the LBH589 research buy surface of the cell membrane, frequently

elicits an immune response (Futagami et al., 1998), and its activity (likely by counteracting the gastric acidity) is crucial for the survival of this bacterium, as shown by the fact that urease-deficient H. pylori mutants fail to colonize the gastric mucosa (Eaton et al., 1991). Ferrero et al. (1994) reported that Flavopiridol (Alvocidib) immunization with urease B resulted in 25–60% protection against Helicobacter felis (the Helicobacter species that naturally infects mice) challenge, as compared with no protection with urease A. Subsequent work has shown that mice immunized with whole-cell lysate or urease B purified protein (either natural or recombinant) results in protection against infection following challenge with either H. pylori SS1 (an H. pylori strain adapted

to colonize mice) (Kleanthous et al., 1998) or H. felis (Chen et al., 1992; Michetti et al., 1994). Despite these progresses, a vaccine for H. pylori remains elusive. Immunization of mice results in a reduction but rarely an elimination of Helicobacter organisms in the stomach (Sutton et al., 2000) and the few attempts to immunize human volunteers have not resulted in adequate immunogenicity (Kreiss et al., 1996; Michetti et al., 1999; Kotloff et al., 2001). Therefore, even though urease B remains an attractive candidate, its immunogenicity has to be improved. To achieve this goal, researchers have experimented with various strong adjuvants (such as Freund’s, cholera toxin or Escherichia coli labile toxin), but due to their toxicity, they have no human application.

Specific central memory CD4+ T cells, defined by CCR7 expression,

Specific central memory CD4+ T cells, defined by CCR7 expression, were virtually undetectable 2 months after vaccination. A change to central

memory phenotype may occur at a later post-vaccination time and this will be explored in future studies. In MVA85A-vaccinated subjects from the UK, Ag85A-specific T-cell proliferation peaked 6 months post-vaccination 32. Interestingly, in mice MVA-induced CD8+ T cells mostly convert to a central memory phenotype within weeks of immunization 44, suggesting that the rate of conversion to central memory cells may differ between species. In other human studies, we have also consistently observed predominant effector phenotypes of human mycobacteria-specific CD4+ T cells in infants 33, 45 and adults 20. Mycobacteria-specific CD4+ T cells from children with latent M.tb infection or active TB 16, and chronically HIV-infected adults with latent M.tb infection 46, Aloxistatin also display this phenotype. Long-lived central memory cells prevail when Ag is cleared after vaccination, e.g. after tetanus toxoid vaccination 42, whereas chronic CMV, EBV or HIV infection is associated with predominance of effector memory cells 47. One might hypothesize

that chronic exposure to mycobacterial Ag is responsible for our observed phenotype. Adolescents with latent M.tb infection, one potential source of such chronic exposure, were not enrolled Astemizole into our study. Additional studies are required to dissect this further. No serious adverse events were recorded, and mild local reactions at the vaccination site were Ceritinib chemical structure predominant. These reactions were commonly reported in the first week after vaccination, did not interfere with daily activities and did not persist. Systemic reactions were uncommon and included mild flu-like symptoms. Clinically, there were no major differences between the adolescents’ and the children’s experience in the trial with a slightly increased incidence of non-vaccine-related systemic events reflecting

this younger age group’s increased risk of transient viral illnesses. This complements the good safety profile of MVA85A found in healthy adults from the same region 25, the United Kingdom 36 and The Gambia 24, as well as other recombinant MVA being tested in clinical trials 40, 48. Together these small phase I/II trials demonstrate a very promising safety profile of MVA85A, which is now being assessed in larger groups of participants, in an infant, phase IIb safety and efficacy study. In conclusion, MVA85A was found to be safe and highly immunogenic in TB-naïve, HIV-uninfected adolescents and children. The vaccine induced durable, polyfunctional CD4+ T-cell responses with a CCR7− effector memory phenotype. These data support future studies to evaluate the efficacy of this vaccine to prevent TB.

The up-regulation of TLR-2 and/or TLR-4 has been shown in macroph

The up-regulation of TLR-2 and/or TLR-4 has been shown in macrophages and gingival fibroblasts of inflamed periodontal tissue [15], which suggests that innate immune responses involving the TLRs as signalling receptors contribute to the inflammatory or immune response of periodontal tissue. Sirtuin 1 (SIRT1) is the human orthologue of the yeast Sir2 protein, the prototypic class III histone deacetylase. SIRT1 has been shown to play a central role in a variety of cellular processes such as stress resistance, metabolism, differentiation and ageing [16]. We have demonstrated previously that SIRT1 exerts anti-inflammatory

effects through Selleckchem Bortezomib the modulation of osteoclastogenic cytokine levels in human PDL cells [17]. Furthermore, SIRT1 has been implicated in the regulation of immune function, as it is expressed at high levels in the thymus, Silmitasertib research buy including in CD4+ and CD8+ thymocytes, and knocking out SIRT1 increases sensitivity to ionizing radiation-induced apoptosis [18]. Moreover, treatment of T cells with resveratrol, a SIRT1 activator, suppresses proliferation and cytokine production

in vitro[19]. Resveratrol also suppresses immune functions by inducing lymphocyte apoptosis [20]. These results suggest that SIRT1 may be involved in the production of immune defence genes in MS-stimulated PDL cells. We have reported previously that MS induces inflammatory cytokines including IL-1β, TNF-α and IL-6, as well as defence genes such as haem oxygenase-1 (HO-1), in human dental pulp cells [21]. Recently, we demonstrated that MS modulates odontoblastic/osteoblastic differentiation via modulation of the HO-1 pathway in dental pulp and PDL cells [22,23]. Although the activation of TLRs and production of anti-microbial peptides, cytokines and chemokines, as well as their receptors, are implicated in innate and adaptive immunity [24], there is little information on the involvement of SIRT1 in MS-induced immune genes of PDL cells. The aim of the present study was to investigate

the role of SIRT1 in the effects of MS on the expression Dolichyl-phosphate-mannose-protein mannosyltransferase of immune response genes in human PDL cells and to identify the underlying mechanisms involved. Dulbecco’s modified Eagle’s medium (DMEM), fetal bovine serum (FBS) and other tissue culture reagents were purchased from Gibco BRL (Grand Island, NY, USA). Resveratrol and sirtinol were purchased from Sigma-Aldrich (St Louis, MO, USA). Affinity purified polyclonal antibodies against mouse TLR-2, TLR-4, I-κBα, nuclear factor (NF)-κB p65 and β-actin monoclonal antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibodies against phospho-extracellular-regulated kinase (p-ERK), ERK, phospho-p38 (p-p38), p38, phospho- c-Jun N-terminal kinase (p-JNK) and JNK were purchased from Cell Signaling Inc. (Beverly, MA, USA).

Conclusion:  EETs are beneficial in

Conclusion:  EETs are beneficial in the setting of lung ischemia–reperfusion, when administered at reperfusion. However, further study will be needed to elucidate the mechanism of action. “
“Please cite this paper as: McGahon MK, McKee J, Dash DP, Brown E, Simpson DA, Curtis TM, McGeown JG, Scholfield CN. Pharmacological profiling of store-operated Ca2+ entry in retinal arteriolar smooth muscle. Microcirculation 19:

586–597, 2012. Objective:  Pharmacological profiling of SOCE and molecular profiling of ORAI and TRPC expression in arterioles. Methods:  Fura-2-based microfluorimetry was used to assess CPA-induced SOCE in rat retinal arteriolar myocytes. Arteriolar ORAI and TRP transcript expression was screened using RT-PCR. Results:  The SKF96365 and LOE908 blocked SOCE (IC50s of 1.2 and 1.4 μm, respectively). Gd3+ and La3+ potently inhibited SOCE (IC50s of 21 and 42 nm, respectively), but Ni2+ showed lower potency (IC50 = 11.6 μm). 2APB inhibited SOCE (IC50 = 3.7 μm) Selleckchem Kinase Inhibitor Library but enhanced

basal influx (>100 μm). Verapamil and nifedipine had no effect at concentrations that inhibit L-type Ca2+ channels, but diltiazem inhibited SOCE by approximately 40% (≥0.1 μm). The RT-PCR demonstrated transcript expression for ORAI 1, 2, and 3, and TRPC1, 3, 4, and 7. Transcripts for TRPV1 and 2, which are activated by 2APB, were also expressed. Conclusions:  The pharmacological profile of SOCE in retinal arteriolar smooth muscle appears unique when compared with other vascular Sodium butyrate tissues.

This suggests that the molecular mechanisms underlying SOCE can differ, even in closely related tissues. Taken together, the pharmacological and molecular data are most consistent with involvement of TRPC1 in SOCE, although involvement of ORAI or other TRPC channels cannot be excluded. “
“Microcirculation (2010) 17, 69–78. doi: 10.1111/j.1549-8719.2010.00002.x Background:  This study was designed to explore the effect of transient inducible nitric oxide synthase (iNOS) overexpression via cationic liposome-mediated gene transfer on cardiac function, fibrosis, and microvascular perfusion in a porcine model of chronic ischemia. Methods and Results:  Chronic myocardial ischemia was induced using a minimally invasive model in 23 landrace pigs. Upon demonstration of heart failure, 10 animals were treated with liposome-mediated iNOS-gene-transfer by local intramyocardial injection and 13 animals received a sham procedure to serve as control. The efficacy of this iNOS-gene-transfer was demonstrated for up to 7 days by reverse transcriptase–polymerase chain reaction in preliminary studies. Four weeks after iNOS transfer, magnetic resonance imaging showed no effect of iNOS overexpression on cardiac contractility at rest and during dobutamine stress (resting ejection fraction: control 27%, iNOS 26%; P = ns). Late enhancement, infarct size, and the amount of fibrosis were similar between groups.

Recently we have developed a novel method to induce IL-17 product

Recently we have developed a novel method to induce IL-17 production and generate Th17 cells using exclusively microbial stimulation [18], a method that

mimics much more closely the in vivo conditions during infection. Although we can confirm defective Th17 generation and IL-17 production by cells isolated check details from patients with HIES [9–11], several important aspects are now apparent when using this improved methodology. First, defective IL-17 induction differs between stimulation with S. aureus or C. albicans. When Th17 responses were assessed both these microorganisms, which are the most important in HIES patients, were equally defective in generating CD4+ IL-17+ cells. Surprisingly, however, C. albicans

was still capable of stimulating approximately 20–30% of normal IL-17 production, while S. aureus was completely defective as an IL-17 stimulus in HIES patients (Fig. 1c). This finding is important as it may explain why it is mainly mucosal; nailbed infection is the most common Candida complication in HIES patients (83% in one large study), while systemic candidiasis is relatively rare [3]. Notably, patients with chronic mucocutaneous candidiasis who have the same clinical spectrum of Candida infection [19] have also been reported to have a specific defect in Candida-induced GSK2126458 IL-17 production [20]. This supports the conclusion that IL-17 is important in mucosal anti-Candida host defence and that the lower IL-17 found in our patients is indeed clinically relevant. Secondly, an important observation of our study is represented by

stiripentol indistinguishable immunological responses in patients with the ‘classical’ clinical form of HIES, independent of the presence or absence of STAT3 mutations. All the patients who had a strong phenotype of the disease displayed similar defects in IL-17 production and Th17 generation. Our data are supported by the report of one HIES patient without STAT3 mutation and defective Th17 responses [21], and suggests strongly that in patients with the ‘classical’ presentation of HIES, but in which no STAT3 mutation is found, defects in the same immunological pathways are the most probable cause of the disease. This may also imply that defective Th17 responses are a more sensitive diagnostic tool for HIES. Thirdly, one of the most interesting findings of our study is the description of a clear association of a milder phenotype of the disease in a Dutch family with a less severe defect in IL-17 production, due probably to the linker domain triplet that did not lead to a frameshift [13]. Patients from this family suffer from skin infections with S. aureus, candidiasis of the nailbeds (but not of the mucosae), dermatitis, hyper-IgE and eosinophilia, but they lack any respiratory infections (either with S. aureus or other pathogens).

We believe that in some circumstances, small expression differenc

We believe that in some circumstances, small expression differences in multiple genes acting in the same signalling pathway could serve as a valuable biomarker

of diabetogenic process. Unexpectedly, the biggest differences in gene expression GSI-IX ic50 profile were found between the group of healthy relatives (DRLN) and the control group. Several of those differentially expressed immunorelevant genes are those regulating inflammation and innate immune responses. Data presented in this study suggest that predisposition to T1D can be generated by the action of myriad of genes with only a slightly altered gene expression levels. Thus, healthy, autoantibody-negative first-degree relatives of patients with T1D are predisposed to react inadequately to certain environmental and/or endogenous stimuli owing to their genetically controlled bias towards enhanced proinflammatory responses. However, in normal circumstances, the propensity for such responses in these subjects seems to be counterbalanced by the opposing action of the regulatory

T cells [14] or by other mechanisms [45], keeping chronic inflammatory responses on low levels. For this reason, vast majority of genetically predisposed people to autoimmune diabetes can stay healthy for entire duration of his/her life. However, in some cases, when the inflammatory responses are exacerbated and/or the regulation of negatively acting circuit is insufficient, the initiation of autoimmune processes leads to the production of

autoantibodies and insulitis. As this process might employ distinct and much smaller set of genes, the whole-genome expression profile stabilizes, resembling rather a ‘normal’ landscape of expression profile. The other possibility is that once beta-islet autoimmunity is initiated and the pancreas becomes a target for lymphocyte infiltration, PMBCs with proinflammatory attributes are depleted from the circulation and/or home to the pancreas and pancreatic draining lymph nodes, thus becoming invisible for their detection in the peripheral blood. This scenario could explain why significant differences in gene expression profile are observed between DRLN and DRLP PRKACG groups. From this point of view, DRLN seems to be a suitable target for discerning vital information about genes with immune and/or non-immune importance and their potential role in the initiation of molecular processes leading to the development of T1D. Once DRLN subjects became autoantibody positive (DRLP), most gene expression–related differences disappear. Results of this study and in particular the conclusion that non-specific immune processes and proinflammatory milieu are essential for the establishment of destructive insulitis are in agreement with conclusions from previous reports that provided an analogous insight into T1D pathogenesis [10, 12–14].

9±5 5 (P=0 003) To determine whether the advantage of B paraper

9±5.5 (P=0.003). To determine whether the advantage of B. parapertussis was manifest at earlier stages of infection, mice (four per group) were inoculated as in the previous experiment with a mixed 1 : 1 inoculum of B. pertussis and B. parapertussis and euthanized at days 1, 2, 4 and 7 postinoculation. buy Galunisertib The competitive advantage of B. parapertussis was observed as early as 24 h postinoculation (mean CI=7) and was maintained through the peak of infection (Fig. 1b). Together, these data indicate that B. parapertussis not only outcompetes B. pertussis in a mixed infection, but also that it benefits

from the presence of B. pertussis in the infection. To further explore the competition between these two organisms in a mixed infection, mice (four per group) were infected with mixed inocula selleck at ratios (B. pertussis to B. parapertussis) of 10 : 1, 3 : 1, 1 : 3 and 1 : 10 (106 total CFU). Mice were euthanized 7 days postinoculation, and the bacterial load and ratio of the two organisms were determined as before. Bordetella parapertussis outcompeted B. pertussis

at all inoculum ratios (Fig. 2). In mice where the inoculum contained B. parapertussis as the predominant strain (1 : 3 and 1 : 10), B. pertussis was at a significant disadvantage, with relatively low CFU recovered from the mice (no B. pertussis was recovered from two mice in the 1 : 10 group). Remarkably, even when the inoculum contained 10-fold excess of B. pertussis (10 : 1), greater CFU of B. parapertussis were recovered from the host, with a mean CI of 31 (Fig. 2). Overall, these data show that B. parapertussis is able to outcompete B. pertussis in a mixed infection over a range of input ratios and apparently gains a greater advantage (higher CI) when the initial inoculum contains higher numbers of B. pertussis. To determine

whether the advantage to B. parapertussis occurs early in the process of infection, experiments were conducted staggering the inoculation with the two organisms. Two different staggered inoculations were tested: (1) mice were inoculated first with 2.5 × 105 CFU B. pertussis (t0) and were inoculated HSP90 3 h later with 2.5 × 105 CFU B. parapertussis (t3), or (2) mice were inoculated initially with 2.5 × 105 CFU B. pertussis (d0) and inoculated 24 h later with 2.5 × 105 CFU B. parapertussis (d1). Mice from each group were euthanized on day 2 post-B. pertussis inoculation, and the bacterial loads and the ratio of the two organisms recovered were determined. In mice inoculated at t0 and t3, greater numbers of B. parapertussis than B. pertussis were recovered (CI=1.82, P<0.05, data not shown). When the inoculations were staggered by 24 h, neither strain had a significant advantage (CI=0.9, data not shown), even though B. pertussis colonization began 24 h earlier than that of B. parapertussis. These data indicate that B. parapertussis can outcompete B. pertussis during early infection even when B.