Profiling of humoral response to influenza A(H1N1)pdm09 infection and vaccination measured by a protein microarray in persons with and without history of seasonal vaccination.

BACKGROUND: The influence of prior seasonal influenza vaccination on the antibody response produced by natural infection or vaccination is not well understood. METHODS: We compared the profiles of antibody responses of 32 naturally infected subjects and 98 subjects vaccinated with a 2009 influenza A(H1N1) monovalent MF59-adjuvanted vaccine (Focetria, Novartis), with and without a history of seasonal influenza vaccination. Antibodies were measured by hemagglutination inhibition (HI) assay for influenza A(H1N1)pdm09 and by protein microarray (PA) using the HA1 subunit for seven recent and historic H1, H2 and H3 influenza viruses, and three avian influenza viruses. Serum samples for the infection group were taken at the moment of collection of the diagnostic sample, 10 days and 30 days after onset of influenza symptoms. For the vaccination group, samples were drawn at baseline, 3 weeks after the first vaccination and 5 weeks after the second vaccination. RESULTS: We showed that subjects with a history of seasonal vaccination generally exhibited higher baseline titers for the various HA1 antigens than subjects without a seasonal vaccination history. Infection and pandemic influenza vaccination responses in persons with a history of seasonal vaccination were skewed towards historic antigens. CONCLUSIONS: Seasonal vaccination is of significant influence on the antibody response to subsequent infection and vaccination, and further research is needed to understand the effect of annual vaccination on protective immunity.

A clinical evaluation of statin pleiotropy: statins selectively and dose-dependently reduce vascular inflammation.

Statins are thought to reduce vascular inflammation through lipid independent mechanisms. Evaluation of such an effect in atherosclerotic disease is complicated by simultaneous effects on lipid metabolism. Abdominal aortic aneurysms (AAA) are part of the atherosclerotic spectrum of diseases. Unlike atherosclerotic occlusive disease, AAA is not lipid driven, thus allowing direct evaluation of putative anti-inflammatory effects. The anti-inflammatory potency of increasing doses (0, 20 or 40 mg/day) simvastatin or atorvastatin was evaluated in 63 patients that were at least 6 weeks on statin therapy and who underwent open AAA repair. A comprehensive analysis using immunohistochemistry, mRNA and protein analyses was applied on aortic wall samples collected during surgery. The effect of statins on AAA growth was analyzed in a separate prospective study in incorporating 142 patients. Both statins equally effectively and dose-dependently reduced aortic wall expression of NFκB regulated mediators (i.e. IL-6 (P<0.001) and MCP-1 (P<0.001)); shifted macrophage polarization towards a M2 phenotype (P<0.0003); selectively reduced macrophage-related markers such as cathepsin K and S (P<0.009 and 0.0027 respectively), and ALOX5 (P<0.0009), and reduced vascular wall NFκB activity (40 mg/day group, P<0.016). No effect was found on other cell types. Evaluation of the clinical efficacy of statins to reduce AAA progression did not indicate an effect of statins on aneurysm growth (P<0.337). Hence, in the context of AAA the clinical relevance of statins pleiotropy appears minimal.

Characteristics of pediatric patients with enterovirus meningitis and no cerebral fluid pleocytosis.

UNLABELLED: Human non-polio enterovirus (EV) is the most important cause of aseptic meningitis in children. Only a few studies report the lack of cerobrospinal fluid (CSF) pleocytosis in children with confirmed EV meningitis; however, the characteristics of these children have not been well defined. This paper describes the clinical and laboratory features of EV meningitis in children with no CSF pleocytosis. Clinical, laboratory, and virological data of Dutch patients <16 years diagnosed with EV meningitis, between 2003 and 2008, were analyzed retrospectively. Data of children with and without CSF pleocytosis were compared. A total of 149 children were infected with EV. Patients presented mainly with fever (n = 113), malaise (n = 43), abdominal pain (n = 47), and irritability (n = 61). Of the 60 patients with EV meningitis, 23 had no pleocytosis. Those who lacked CSF pleocytosis were younger [odds ratio (OR) 1.00; 95% confidence interval (CI) 1.000-1.002; p = 0.001], had experienced drowsiness more (OR 9.60; 95% CI 2.24-41.15; p = 0.002), had lower white blood cell counts (OR 0.73; 95% CI 0.61-0.89; p = 0.001), and had higher C-reactive protein (OR 1.13; 95% CI 1.03-1.23; p = 0.006) than those with pleocytosis. CONCLUSION: These findings show that EV meningitis occurs in the absence of CSF pleocytosis, particularly in young infants, meaning that EV meningitis in this age group cannot be solely excluded by the absence of CSF pleocytosis. They also confirm the importance of genome detection in the diagnosis of EV meningitis in young infants.

Identification by genotyping of a commercial antigen preparation as the source of a laboratory contamination with Coxiella burnetii and as an unexpected rich source of control DNA.

By performing genotyping, a laboratory contamination involving Q fever was traced back to the antigen preparation used in a commercially available complement fixation test. It was established that such antigen preparations contain relatively high loads of DNA/RNA, making them potential sources of contamination but also convenient preparations for control material.

[Laboratory diagnosis of acute Q fever]. / Laboratoriumdiagnostiek van acute Q-koorts.

In the Netherlands an increasing number of laboratories are involved in diagnosing acute Q-fever. More uniformity in diagnostics and interpretation is desirable. To enable this, a working group on diagnostics of acute Q-fever was created on the initiative of the National Institute for Public Health and the Environment (RIVM) and the Dutch Association for Medical Microbiology (NVMM). The diagnostics of acute Q-fever includes a diagnostic flow chart (algorithm) consisting of tests for DNA and for antibodies against the antigens that appear in the successive stages of the disease. Reporting of both confirmed and suspected cases of acute Q-fever is obligatory.

Detection of respiratory viruses and Legionella spp. by real-time polymerase chain reaction in patients with community acquired pneumonia.

We conducted a study on throat swabs obtained from a group of hospitalized patients with community acquired pneumonia (CAP). Throat swab specimens from 242 adults admitted to hospital with CAP were tested. In total, 1 or more aetiological agents were identified by real-time PCR in 55 (23%) patients. The most frequently detected pathogens were coronavirus (17%), parainfluenza virus (6%) and influenza virus (4%). Overall, viral pathogens were identified by conventional techniques in 7 (2%) patients, and real-time PCR in 50 (21%) patients (p<0.0001). The diagnostic yield increased from 137 cases (57% of patients using conventional microbiological assays) to 158 cases (65% of patients using real-time PCR assays and conventional microbiological assays; p=0.06). A significantly higher percentage of mortality was present in patients with a mixed bacterial and viral infection. L. pneumophila PCR was positive in only 3 out of 11 cases (27%) of Legionnaires' disease (LD). This study demonstrates that real-time PCR can increase the number of microbiological detections of respiratory pathogens, mainly as a result of detection of respiratory viruses.

Utility of real-time PCR for diagnosis of Legionnaires' disease in routine clinical practice.

The main aim of our study was to determine the added value of PCR for the diagnosis of Legionnaires' disease (LD) in routine clinical practice. The specimens were samples submitted for routine diagnosis of pneumonia from December 2002 to November 2005. Patients were evaluated if, in addition to PCR, the results of at least one of the following diagnostic tests were available: (i) culture for Legionella spp. on buffered charcoal yeast extract agar or (ii) detection of Legionella pneumophila antigen in urine specimens. Of the 151 evaluated patients, 37 (25%) fulfilled the European Working Group on Legionella Infections criteria for a confirmed case of LD (the "gold standard"). An estimated sensitivity, specificity, and overall percent agreement of 86% (32 of 37; 95% confidence interval [CI] = 72 to 95%), 95% (107 of 112; 95% CI = 90 to 98%), and 93% (139 of 149), respectively, were found for 16S rRNA-based PCR, and corresponding values of 92% (34 of 37; 95% CI = 78 to 98%), 98% (110 of 112; 95% CI = 93 to 100%), and 97% (144 of 149), respectively, were found for the mip gene-based PCR. A total of 35 patients were diagnosed by using the urinary antigen test, and 34 were diagnosed by the 16S rRNA-based PCR. With the mip gene PCR one more case of LD (n = 36; not significant) was detected. By combining urinary antigen test and the mip gene PCR, LD was diagnosed in an additional 4 (11%) patients versus the use of the urinary antigen test alone. The addition of a L. pneumophila-specific mip gene PCR to a urinary antigen test is useful in patients with suspected LD who produce sputum and might allow the early detection of a significant number of additional patients.

Molecular evidence for the ubiquitous presence of Legionella species in Dutch tap water installations.

Our aim was to investigate the occurrence and identity of Legionella spp. in Dutch tap water installations using culture, real-time PCR and sequence analysis. The PCR assays used were a 16S rRNA gene based PCR with both a Legionella species specific probe and a L. pneumophila specific probe and a L. pneumophila-specific PCR based on the sequence of the mip gene. A total of 357 water samples from 250 locations in The Netherlands was investigated. The detection rates of Legionella spp. were 2,2% (8 of 357) by culture, and 87,1% (311 of 357) by PCR. The majority of samples was found to contain Legionella species other than L. pneumophila. These comprised of Legionella Like Amoebal Pathogens (LLAPs), L. busanensis, L. worsliensis and others. Fourteen (3,9%) samples were positive for L. pneumophila by either culture, 16S rRNA based PCR and/or mip based PCR. It is apparent from this study that Legionella spp. DNA is ubiquitous in Dutch potable water samples. Our findings further suggest that LLAPs and viable but nonculturable (VBNC) Legionella represent a large proportion of the population in man-made environments.

Risk factors for acute respiratory tract infections in general practitioner patients in The Netherlands: a case-control study.

BACKGROUND: Acute respiratory tract infections (ARTI) are an important public health problem. Improved identification of risk factors might enable targeted intervention. Therefore we carried out a case-control study with the aim of identifying environmental risk factors for ARTI consultations in the Dutch general population. METHODS: A subset of patients visiting their GP in the period of 2000-2003 with an ARTI (cases) and age-matched controls (visiting for other complaints) were included in a case-control study. They were asked to complete a questionnaire about potential risk factors. Conditional logistic regression was used to calculate odds ratio's (OR) and 95% confidence intervals (CI) to estimate the independent effect of potential risk factors. RESULTS: A total of 493 matched pairs of case and control subjects were enrolled. Exposure to persons with respiratory complaints, both inside and outside the household, was found to be an independent risk factor for visiting a GP with an ARTI (respectively ORadj = 1.9 and ORadj = 3.7). Participants exposed to dampness or mould at home (ORadj=0.5) were significantly less likely to visit their GP. In accordance with the general risk of consultations for ARTI, participants with a laboratory-confirmed ARTI who were exposed to persons with respiratory complaints outside the household were also significantly more likely to visit their GP (ORadj=2.5). CONCLUSION: This study confirmed that heterogeneity in the general population as well as in pathogens causing ARTI makes it complicated to detect associations between potential risk factors and respiratory infections. Whereas it may be difficult to intervene on the risk posed by exposure to persons with respiratory complaints, transmission of ARTI in the general population might be reduced by improved hygienic conditions.

Evaluation of real-time PCR for the early detection of Legionella pneumophila DNA in serum samples.

Legionella pneumonia can be difficult to diagnose. Existing laboratory tests all have shortcomings, especially in the ability to diagnose Legionnaires' disease (LD) at an early stage of the disease in a specimen that is readily obtainable. The aim of this study was to assess the performance of PCR as a rapid diagnostic method and to compare the results of different PCR assays of serum samples from patients with LD. Samples included 151 serum samples from 68 patients with proven LD and 60 serum samples from 36 patients with respiratory tract infections other than Legionella. PCR assays were based on the 5S rRNA gene, 16S rRNA gene and the mip gene. The samples from patients with infections caused by pathogens other than Legionella all tested negative in PCR. Among the patients with proven LD 54.4 % (37/68) tested positive in 5S rRNA PCR, 52.9 % (36/68) in mip gene PCR and 30.9 % (21/68) in 16S rRNA PCR in the first available serum sample. The association between threshold cycle value in 5S PCR positive serum samples (n=49) and C-reactive protein value was determined, and showed a strong negative correlation (Pearson correlation coefficient r=-0.63, P<0.0001). In addition to existing tests for the diagnosis of LD, detection of Legionella DNA in serum could be a useful tool for early diagnosis of LD caused by any Legionella species and serogroup, and has the potential to provide a diagnosis in a time frame that could affect initial infection management.

Transient paresis associated with cat-scratch disease: case report and literature review of vertebral osteomyelitis caused by Bartonella henselae.

Cat-scratch disease (CSD) rarely presents as vertebral osteomyelitis. We describe a case with paresis of the arm with total recovery after antibiotic and neurosurgical therapy. We reviewed 20 other cases of CSD vertebral osteomyelitis in the literature. This diagnosis should be considered in patients with systemic symptoms, back pain, and cat contact. The prognosis is generally good.