22 research outputs found
Detection of amyloid beta aggregates in the brain of BALB/c mice after Chlamydia pneumoniae infection
Neuroinflammation, initiated by cerebral infection, is increasingly postulated as an aetiological factor in neurodegenerative diseases such as Alzheimer’s disease (AD). We investigated whether Chlamydia pneumoniae (Cpn) infection results in extracellular aggregation of amyloid beta (Aβ) in BALB/c mice. At 1 week post intranasal infection (p.i.), Cpn DNA was detected predominantly in the olfactory bulbs by PCR, whereas brains at 1 and 3 months p.i. were Cpn negative. At 1 and 3 months p.i., extracellular Aβ immunoreactivity was detected in the brain of Cpn-infected mice but also in the brain of mock-infected mice and mice that were neither Cpn infected nor mock infected. However, these extracellular Aβ aggregates showed morphological differences compared to extracellular Aβ aggregates detected in the brain of transgenic APP751SL/PS1M146L mice. These data do not unequivocally support the hypothesis that Cpn infection induces the formation of AD-like Aβ plaques in the brain of BALB/c mice, as suggested before. However, future studies are required to resolve these differences and to investigate whether Cpn is indeed an etiological factor in AD pathogenesis
Qualitative and Quantitative Detection of Chlamydophila pneumoniae DNA in Cerebrospinal Fluid from Multiple Sclerosis Patients and Controls
A standardized molecular test for the detection of Chlamydophila pneumoniae DNA in cerebrospinal fluid (CSF) would assist the further assessment of the association of C. pneumoniae with multiple sclerosis (MS). We developed and validated a qualitative colorimetric microtiter plate-based PCR assay (PCR-EIA) and a real-time quantitative PCR assay (TaqMan) for detection of C. pneumoniae DNA in CSF specimens from MS patients and controls. Compared to a touchdown nested-PCR assay, the sensitivity, specificity, and concordance of the PCR-EIA assay were 88.5%, 93.2%, and 90.5%, respectively, on a total of 137 CSF specimens. PCR-EIA presented a significantly higher sensitivity in MS patients (p = 0.008) and a higher specificity in other neurological diseases (p = 0.018). Test reproducibility of the PCR-EIA assay was statistically related to the volumes of extract DNA included in the test (p = 0.033); a high volume, which was equivalent to 100 µl of CSF per reaction, yielded a concordance of 96.8% between two medical technologists running the test at different times. The TaqMan quantitative PCR assay detected 26 of 63 (41.3%) of positive CSF specimens that tested positive by both PCR-EIA and nested-PCR qualitative assays. None of the CSF specimens that were negative by the two qualitative PCR methods were detected by the TaqMan quantitative PCR. The PCR-EIA assay detected a minimum of 25 copies/ml C. pneumoniae DNA in plasmid-spiked CSF, which was at least 10 times more sensitive than TaqMan. These data indicated that the PCR-EIA assay possessed a sensitivity that was equal to the nested-PCR procedures for the detection of C. pneumoniae DNA in CSF. The TaqMan system may not be sensitive enough for diagnostic purposes due to the low C. pneumoniae copies existing in the majority of CSF specimens from MS patients
International Bordetella pertussis assay standardization and harmonization meeting report. Centers for Disease Control and Prevention, Atlanta, Georgia, United States, 19-20 July 2007.
Bactericidal Antibody Responses Elicited by a Meningococcal Outer Membrane Vesicle Vaccine with Overexpressed Factor H–Binding Protein and Genetically Attenuated Endotoxin
BACKGROUND: Outer membrane vesicle (OMV) vaccines from mutant N. meningitidis strains engineered to over-express factor H-binding protein (fHbp) elicited broadly protective serum antibody responses in mice. The vaccines investigated were not treated with detergents to avoid extracting fHbp, which is a lipoprotein. Because of their high endotoxin content, the vaccines would not be safe to administer to humans. METHODS: We prepared a native OMV vaccine from a strain engineered to over-express fHbp and in which the gene encoding LpxL1 was inactivated, which reportedly decreased endotoxin activity. RESULTS: The OMV vaccine from the mutant had similar or lower ability to induce proinflammatory cytokines by human peripheral blood mononuclear cells as a detergent-extracted wildtype OMV, and 1000- to 10,000-fold lower activity than a native wildtype OMV. In mice, the OMV vaccine from the mutant elicited higher serum bactericidal antibody responses against a panel of heterologous N. meningitidis strains than a control multicomponent recombinant protein vaccine, or a detergent-extracted OMV vaccine that previously had been demonstrated to confer protection against meningococcal disease in humans. CONCLUSIONS: The data illustrate the potential to develop a broadly immunogenic native OMV vaccine with decreased endotoxin activity that is potentially suitable for testing in humans
Prospective evaluation of an Australian pertussis toxin IgG and IgA enzyme immunoassay
Serological diagnosis of recent pertussis infection is an important part of both clinical assessment and epidemiological documentation of this disease. Standardization of serological testing and interpretation remains challenging despite international efforts to improve it. Currently, determining the anti-pertussis toxin (PT) IgG titer is recommended as the most accurate serological test in Europe and the United States, while Australia relies predominantly on measurement of Bordetella pertussis IgA antibody responses. Using B. pertussis PCR and the WHO clinical case definition as reference standards, the diagnostic utility of in-house anti-PT IgG and anti-PT IgA assays was evaluated prospectively in an Australian community-based cohort (n = 327). Patients provided up to four consecutive serum samples to document the kinetics of antibody response and decay. Previously validated cutoffs for positivity were converted to international units by using WHO-approved reference sera. At currently used cutoffs, both anti-PT IgG (>94 IU/ml) and anti-PT IgA (>20 IU/ml) assays had good specificity (80% [95% confidence interval {95% CI}, 68 to 88%] and 87% [95% CI, 77 to 94%]), but anti-PT IgG assay was consistently more sensitive than anti-PT IgA assay across a range of cutoffs (60 to 79% [95% CI, 53 to 84%] versus 41 to 62% [95% CI, 34 to 69%]). The combination of anti-PT IgG and anti-PT IgA assays performed no better than anti-PT IgG assay alone. The anti-PT IgA response in children under 12 years of age was poor. The accuracy of serology was optimal between 2 and 8 weeks after symptom onset. Cutoffs of >94 IU/ml for anti-PT IgG and >20 IU/ml for anti-PT IgA correlated well with recent pertussis infection and were consistent with recent recommendations from the EU Pertstrain group. Anti-PT IgG assay was superior to anti-PT IgA assay as the test of choice for the diagnosis of pertussis from a single sample
Standardizing Chlamydia pneumoniae assays: Recommendations from the centers for disease control and prevention (USA) and the laboratory centre for disease control (Canada)
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International Bordetella pertussis assay standardization and harmonization meeting report. Centers for Disease Control and Prevention, Atlanta, Georgia, United States, 19-20 July 2007.
An international meeting on Bordetella pertussis assay standardization and harmonization was held at the Centers for Disease Control and Prevention (CDC), Atlanta, GA, 19-20 July 2007. The goal of the meeting was to harmonize the immunoassays used for pertussis diagnostics and vaccine evaluation, as agreed upon by academic and government researchers, regulatory authorities, vaccine manufacturers, and the World Health Organization (WHO). The primary objectives were (1) to provide epidemiologic, laboratory, and statistical background for support of global harmonization; (2) to overview the current status of global epidemiology, pathogenesis and immunology of pertussis; (3) to develop a consensus opinion on existing gaps in understanding standardization of pertussis assays used for serodiagnosis and vaccine evaluation; and (4) to search for a multicenter process for addressing these priority gaps. Presentations and discussions by content experts addressed these objectives. A prioritized list of action items to improve standardization and harmonization of pertussis assays was identified during a group discussion at the end of the meeting. The major items included: (1) to identify a group that will organize, prepare, maintain, and distribute proficiency panels and key reagents such as reference and control sera; (2) to encourage the development and identification of one or more reference laboratories that can serve as an anchor and resource for other laboratories; (3) to define a performance-based assay method that can serve as a reference point for evaluating laboratory differences; (4) to develop guidance on quality of other reagents, e.g., pertussis toxin and other antigens, and methods to demonstrate their suitability; (5) to establish an international working group to harmonize the criteria to evaluate the results obtained on reference and proficiency panel sera; (6) to create an inventory to determine the amount of appropriate and well-characterized sera that are available globally to be used as bridging reagents for vaccine licensure; and (7) to seek specific guidance from regulatory authorities regarding the expectations and requirements for the licensure of new multicomponent pertussis vaccines