Low neonatal Toll-like receptor 4-mediated interleukin-10 production is associated with subsequent atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) and respiratory syncytial virus lower respiratory tract infection (RSV LRTI) are common diseases during early life. Impaired Th1-cell polarizing Toll-like receptor (TLR) responses play an important role in the pathogenesis of both diseases. Neonatal TLR-mediated production of Th1-type cytokines is decreased at birth, but rapidly increases during the first month of life. OBJECTIVE: To determine whether decreased TLR-mediated production of Th1-polarizing cytokines, at the age of 1 month is associated with subsequent AD or RSV LRTI. METHODS: A prospective healthy birth cohort study was performed. Whole blood concentrations of innate immune cells and TLR-mediated cytokine responses were measured at the age of 1 month in 291 neonates. AD was determined by a physician questionnaire at the age of 1 year and RSV LRTI was defined as parent-reported respiratory symptoms and presence of RSV RNA in a nose-throat specimen. RESULTS: Of participating neonates, 45 (15%) developed AD and 41 (14%) developed RSV LRTI. Risks of AD and RSV LRTI were not associated (χ(2) , P = 1.00). AD was associated with decreased concentrations of basophils (7.6 vs. 14.0 × 10(6) /mL, P = 0.002) and plasmacytoid dendritic cells (17.0 vs. 20.5 × 10(6) /mL, P = 0.04), increased concentrations of NK-cells (79.7 vs. 45.1 × 10(6) /mL, P = 0.03), and twofold lower TLR4-mediated IL-10 production (P = 0.001). In contrast, RSV LRTI was associated neither with neonatal concentrations of innate immune cells, nor with TLR-mediated TNF-α, IL-12p70, IL-10 or IFN-α production. CONCLUSIONS AND CLINICAL RELEVANCE: Atopic dermatitis, but not RSV LRTI, is associated with distinct pre-symptomatic differences in the innate immune system. We hypothesize that decreased neonatal IL-10-mediated immune regulation during early life might play a causal role in the initiation of AD.

Long time trends in influenza-like illness and associated determinants in The Netherlands.

We analysed long-term epidemiological trends in influenza-like illness (ILI) in The Netherlands and used an ecological analysis to estimate its relationship with age, influenza vaccination, and virological aspects. This study used data from weekly ILI consultation reports from sentinel general practitioners (1986/1987 to 2006/2007), virological data from sentinel ILI patients (1992/1993 to 2006/2007), and data for influenza vaccine uptake (1991-2005). The incidence of ILI consultations, although varying during the study period, was estimated to decrease in the total population by 12.2/10 000 persons each season (95% CI 8.6-15.9). Uptake of influenza vaccination in people aged > or = 65 years (elderly) increased from 28% in 1991 to >70% since 1997. ILI incidence in the elderly declined by 1.7/10 000 persons (P=0.05) per percentage vaccine uptake per season. The decline in ILI incidence over the last 20 years could be related to the increased vaccine uptake. However, insufficient data were present to assess the impact of other potential contributing factors, such as diminished fitness of influenza viruses and changes in consulting behaviour.

Community-acquired pathogens associated with prolonged coughing in children: a prospective cohort study.

A 2-year prospective study was performed of children with prolonged coughing to investigate the frequency of different respiratory pathogens, the rate of mixed infections, and possible differences in severity of disease between single and mixed infections. Sera from 135 children (136 episodes of prolonged coughing lasting 1-6 weeks) were tested for antibodies to different viruses and bacteria. Swabs were taken for culture and PCR to detect different viral and bacterial pathogens. One or more pathogens were found in 91 (67%) patients. One infectious agent was found in 49 (36%) patients, two agents in 35 (26%) patients, and more than two agents in seven (5%) patients. The most frequent pathogens encountered were rhinovirus (n = 43; 32%), Bordetella pertussis (n = 23; 17%) and respiratory syncytial virus (n = 15; 11%). The most frequent mixed infection was B. pertussis and rhinovirus (n = 14; 10%). No significant differences in clinical symptoms were observed between patients with or without pathogens; however, patients with mixed infections were significantly older. There was a strong seasonal influence on the number of infections, but not on the number of mixed infections. In children with prolonged coughing, there was a high frequency of mixed infections regardless of the season. However, mixed infection was not associated with increased disease severity. No clinical symptoms were found that allowed discrimination between specific pathogens.

No evidence of Legionella infection in general practice patients presenting with acute respiratory infections in The Netherlands.

The role of Legionella spp. in the aetiology of acute respiratory infections (ARIs) is largely unknown. In this case-control study, conducted in a general practitioner setting during 2000 and 2001, nose and throat samples from patients presenting with ARIs (n = 230) and controls (n = 200) were analysed for the presence of Legionella spp. by real-time PCR. Legionella DNA was not detected in any of the cases or controls. Thus, Legionella spp. do not seem to play a role in patients presenting with ARIs, nor were they present in patients who visited their general practitioner for complaints other than ARIs.

Influenza-associated encephalopathy: no evidence for neuroinvasion by influenza virus nor for reactivation of human herpesvirus 6 or 7.

During 2 consecutive influenza seasons we investigated the presence of influenza virus, human herpesvirus (HHV) type 6, and HHV-7 in cerebrospinal fluid samples from 9 white children suffering from influenza-associated encephalopathy. We conclude that it is unlikely that neuroinvasion by influenza virus or reactivation of either HHV-6 or HHV-7 is involved.

[The 2003/2004 influenza season in the Netherlands with a limited epidemic of the virus variant A/Fujian, and the vaccine composition for the 2004/2005 season]. / Het influenzaseizoen 2003/'04 in Nederland met een beperkte epidemie door de virusvariant A/Fujian, en de vaccinsamenstelling voor het seizoen 2004/'05.

In contrast to the three previous influenza seasons, the influenza epidemic of the 2003/2004 season started early in week 49 of 2003. The epidemic was predominantly caused by influenza-A viruses of the H3N2 subtype. All isolated influenza-A viruses were antigenically related to influenza virus A/Fujian/411/02, which was already detected in the influenza season 2002/2003 and that deviated from the vaccine-reference strain A/Moscow/10/99 to a certain extent. The magnitude of the epidemic was limited despite the fact that it was caused by influenza-A H3N2-virus-drift variants. Immunity caused by natural infection with influenza viruses during previous seasons or vaccination has possibly provided sufficient cross protection against these new H3N2-drift variant. No influenza-A viruses of the H1N1 or H1N2 subtypes were detected in the influenza season 2003/2004. Only a small number of influenza-B viruses were isolated, which all belonged to the B/Yamagata/16/88 lineage, which was temporarily replaced by the B/Victoria2/87 lineage in the previous influenza season. On the basis of epidemiological and serological data the World Health Organization has recommended the following vaccine composition for the 2004/2005 influenza season: A/Fujian/411/02 (H3N2), A/New Caledonia/20/99 (H1N1) and B/Shanghai/361/02.

[The 2002/2003 influenza season in the Netherlands and the vaccine composition for the 2003/2004 season]. / Het influenzaseizoen 2002/'03 in Nederland en de vaccinsamenstelling voor het seizoen 2003/'04.

As in the 2000/2001 and 2001/2002 seasons, the influenza epidemic in the 2002/2003 season started late (week 7 of 2003) and was only moderate in size. Influenza A (H3N2) and B viruses were detected in equal numbers among patients of general practitioners and these two viruses were therefore equally responsible for the epidemic. However, H3N2 viruses dominated isolates taken from hospitals. In haemagglutination-inhibition (HI) assays most of the H3N2 viruses proved highly reactive with antiserum to the vaccine-reference strain A/Moscow/10/99. This was also true for a number of isolates, including those obtained from nursing home residents, closely related to the reference strain A/Finland/170/03. However, an estimated 4% of the H3N2 isolates belonged to the variant A/Fujian/411/02 from China, which constituted the majority of the H3N2 viruses isolated in Europe in the later phase of the season. This variant reacted poorly with antiserum to A/Moscow/10/99. In H1 tests all influenza A(H1N1)-virus isolates and all B-virus isolates were closelyrelated to the corresponding vaccine-reference strains. Taking this data into consideration, the World Health Organization has advised the same vaccine composition for the 2003/2004 season as for the 2002/2003 season, namely: A/Moscow/10/99 (H3N2), A/New Caledonia/20/99 (H1N1) and B/Hong Kong/330/01. There is the possibility of a mismatch occurring between the H3N2-vaccine strain and the circulating H3N2 viruses in the coming influenza season. In March and April 2003 there was an outbreak of influenza-A (H7N7) fowl plague in the Netherlands. A special monitoring survey revealed that 91 people who had handled infected poultry became infected with the H7N7 virus. One of these later died as a result of this. None of the avian and human H7N7-virus isolates examined contained human or porcine influenza-A virus genes.

[The 2001/2002 influenza season and the vaccine composition for the 2002/2003 season]. / Het influenzaseizoen 2001/'02 en de vaccinsamenstelling voor het seizoen 2002/'03.

The epidemic in the influenza season 2001/2002 was of moderate activity just like in 2000/2001. The influenza epidemic started in week 2 of 2002 when the clinical influenza activity reported by the general practitioner network of the Netherlands Institute of Primary Health Care (NIVEL) increased. This was caused by influenza A viruses of the H3N2 subtype in particular. All influenza A viruses of this subtype were closely related to the vaccine strain for this subtype, A/Moscow/10/99. Influenza B viruses and influenza A/H1 viruses isolated this season had surprising features. The influenza B viruses originated from two lineages. Viruses of the B/Yamagata/16/88 lineage have been circulating for more than twelve years. The vaccine reference strain B/Sichuan/379/99 belongs to this lineage. The B/Victoria/2/87 lineage reappeared again after an absence in Europe of more than ten years and accounted for 50% of the influenza B viruses that were isolated in the Netherlands. Therefore the vaccine will have provided only partial protection against influenza B. The only influenza A/H1 virus that was isolated appeared to be of a new subtype H1N2. The H1 hemagglutinin of this virus was closely related to that of the vaccine strain A/New Caledonia/20/99. The N2 neuraminidase originated from recent human influenza A/H3N2 viruses. Therefore the vaccine probably provided good protection against the new H1N2 subtype. Based in part on these data, the World Health Organization has advised that the vaccines for the season 2002/2003 should contain the following or comparable influenza-virus strains: A/Moscow/10/99 (H3N2), A/New Caledonia/20/99 (H1N1) and B/Hong Kong/330/01, the latter being an influenza B virus of the B/Victoria/2/87 lineage.

[2000/01 influenza season and the vaccine composition for the season 2001/'02]. / Het influenzaseizoen 2000/'01 en de vaccinsamenstelling voor het seizoen 2001/'02.

In the 2000/01 season, the size of the influenza epidemic in the Netherlands was exceptionally small. Since the start of the Continuous Morbidity Registration of the Netherlands Institute of Primary Health Care (NIVEL) in 1970, the peak incidence of influenza-like illnesses has never been so low. The aetiology of the epidemic was also unusual. Most remarkable was the relatively extensive circulation of subtype H1N1 and the low activity of subtype H3N2. The epidemic started in week 1 of 2001 and ended in week 8. The antigenic properties of the influenza A (H1N1) viruses closely resembled those of the vaccine strain A/New Caledonia/20/99. This new variant of subtype H1N1 was first isolated in Asia in 1995 and was only (sporadically) detected in the Netherlands in the 1999/2000 season. Phylogenetic analysis showed that these viruses represent a new line of subtype H1N1. Following the influenza-activity caused by H1N1 viruses in the 2000/01 season, a small number of B and H3N2 viruses were also isolated up to week 19. Antigenically, these viruses were identical to those obtained in the previous years. On the basis of the antigenetic analyses presented, it can be concluded that the vaccine provided good protection against the circulating influenza viruses in the 2000/01 season. The World Health Organization recommends that influenza vaccines intended for use in the 2001/02 season of the northern hemisphere should contain the following, or antigenically similar, strains: A/Moscow/10/99 (H3N2), A/New Caledonia/20/99 (H1N1), and B/Sichuan/379/99.

Mycoplasma pneumoniae P1 type 1- and type 2-specific sequences within the P1 cytadhesin gene of individual strains.

Mycoplasma pneumoniae strains traditionally are divided into two types, based on sequence variation in the P1 gene. Recently, however, we have identified 8 P1 subtypes by restriction fragment length polymorphism analysis. In the present study the P1 gene sequences of three P1 type 1 and two P1 type 2 M. pneumoniae strains were analyzed. A new P1 gene sequence in a type 1 strain with partial similarity to a recently reported variable region in the P1 gene of an M. pneumoniae type 2 strain (T. Kenri, R. Taniguchi, Y. Sasaki, N. Okazaki, M. Narita, K. Izumikawa, M. Umetsu, and T.Sasaki, Infect. Immun. 67:4557-4562, 1999) was identified. In addition, the P1 gene of the type 1 strain contained another region with nucleotide polymorphisms identical to a stretch in the P1 gene of one of our type 2 strains. These findings indicate that recombination between sequences specific for P1 type 1 and type 2 had occurred and that P1 type 1 and type 2 hybrid sequences can be present within the P1 gene of an individual strain. Identical or nearly identical variable P1 gene sequences were present in several repetitive regions outside the P1 gene locus in the genome of M. pneumoniae strain M129, implying recombination as a mechanism for generation of the P1 gene variation. Additionally, in the P1 gene sequences of four of the five strains studied, single-nucleotide polymorphisms different from the previously reported P1 type 1 and 2 characteristic sequences were identified. The polymorphic sites are candidate targets for genotyping of M. pneumoniae by direct sequencing of amplicons from clinical specimens.

Results of molecular detection of Mycoplasma pneumoniae among patients with acute respiratory infection and in their household contacts reveals children as human reservoirs.

During a 30-month prospective study in The Netherlands, the distribution of Mycoplasma pneumoniae and respiratory viruses among 1172 patients with acute respiratory infection (ARI) who were treated in the outpatient general practitioner setting was studied. M. pneumoniae, as detected by polymerase chain reaction analysis, was present in 39 (3.3%) patients. The infection rate was similar in all age groups. Nose and throat samples collected from 79 household contacts of M. pneumoniae-positive index patients revealed M. pneumoniae in 12 (15%) cases. The frequency of M. pneumoniae among household contacts of index patients treated with appropriate antibiotics and untreated index patients was similar. Nine of the 12 M. pneumoniae-positive household contacts were <16 years old (P=.02), and 4 (44%) of them did not develop ARI. Apparently, children are a relevant reservoir for M. pneumoniae.

Detection of multiple viral DNA species in synovial tissue and fluid of patients with early arthritis.

OBJECTIVE: Viruses have a role in the pathogenesis of various forms of arthritis. This study aimed at determining whether viral DNA can be detected in joint samples in the early stages of idiopathic arthritides. METHODS: Synovial fluid (SF) and synovial tissue (ST) samples were obtained from 73 patients, with undifferentiated arthritis (n=22), rheumatoid arthritis (n=13), spondyloarthropathy (n=17), crystal arthropathy (n=8), osteoarthritis (n=7), septic arthritis (n=5), and trauma (n=1). The presence of viral DNA was investigated by polymerase chain reaction analysis. RESULTS: Cytomegalovirus was present in 25 patients, parvovirus B19 in 15 patients, Epstein-Barr virus in 12 patients, and herpes simplex virus in 16 patients (in ST, SF, or both), respectively. The joint samples were negative for viral DNA from adenovirus and varicella-zoster virus. In ST, eight patients were double positive for parvovirus B19 and another viral DNA, with herpes simplex virus being the most prevalent. Seven patients were double positive for other viruses (cytomegalovirus, herpes simplex virus, Epstein-Barr virus). In SF, four patients were double or triple positive for viral DNA. Paired samples were available in 56 patients. In these, viral DNA was detected in 37 patients in ST, as compared with 19 in SF. CONCLUSION: These data show that one or more viruses can be detected in the synovial specimens of patients with early arthritis, irrespective of the clinical diagnosis. This observation might be explained by migration of inflammatory cells harbouring viral DNA into the inflamed joints.

Presence of bacterial DNA and bacterial peptidoglycans in joints of patients with rheumatoid arthritis and other arthritides.

OBJECTIVE: The continuous presence of bacteria or their degraded antigens in the synovium may be involved in the pathogenesis of rheumatoid arthritis (RA). The aim of this study was to determine the presence of bacterial nucleic acids and bacterial cell wall constituents in the joints of patients with RA and other forms of arthritis. METHODS: Joint samples were obtained from patients with RA (n = 26), septic arthritis (n = 2), inflammatory osteoarthritis (n = 5), and gout (n = 6), and joint trauma (n = 1). Universal 16S-ribosomal RNA primers were used to detect the presence of bacterial DNA in these samples, using stringent regimens for sample collection and molecular microbiologic analysis. Automated sequencing and comparative data analysis were performed to identify the species. The presence of bacterial peptidoglycan-polysaccharide complexes in synovial tissue was detected by immunohistologic analysis with a specific antibody. RESULTS: The bacterial species cultured from the synovium could be identified in both of the patients with septic arthritis. DNA amplicons were also detected in the synovial fluid and/or tissue samples from 5 patients with RA and 2 patients with crystal-induced arthritis; these originated from multiple bacterial species. Staining for peptidoglycan-polysaccharide complexes was positive in the synovial tissue of both patients with septic arthritis, 16 with RA, 4 with inflammatory osteoarthritis, 4 with crystal-induced arthropathy, and 1 with joint trauma. The staining was mainly found in cells in the synovial sublining, including macrophages. CONCLUSION: The results indicate that bacterial DNA and bacterial cell wall constituents are retained in the joints of some patients with arthritis, where they might enhance synovial inflammation.

Detection of bacterial DNA in serial synovial samples obtained during antibiotic treatment from patients with septic arthritis.

OBJECTIVE: The management of septic arthritis could benefit from sensitive tests that detect the persistence of microorganisms in the joint. The aim of this study was to determine the feasibility of monitoring the presence of bacterial DNA in synovial samples from septic arthritis patients during antibiotic treatment. METHODS: Synovial fluid (SF) and synovial tissue (ST) samples were collected serially from 6 patients with septic arthritis before and during antibiotic therapy. In addition, peripheral blood (PB) samples were available for polymerase chain reaction (PCR) analysis from 5 of the 6 patients before treatment. All samples were analyzed for the presence of bacterial DNA with the use of a PCR with universal 16S ribosomal RNA primers. Automated sequencing and comparative data analysis were performed to identify the species. These data were compared with Gram staining and culture results. RESULTS: The bacterial species cultured from the synovium could be identified in all 6 patients using PCR and subsequent sequence analysis of the amplicons. In virtually all cases, positive Gram stain and culture findings in the synovial samples became negative after 2-3 days of antibiotic treatment. Bacterial DNA persisted in the SF and/or ST after culture conversion; in 2 patients, bacterial DNA was still detected at day 10, in 1 patient, at day 20, and in another patient, at day 22 after the initiation of treatment. Synovial samples were available for PCR analysis from 2 patients at day 26. At this time point, bacterial DNA could not be detected anymore. All PB samples were negative by both culture and PCR analysis. CONCLUSION: PCR analysis can be used to monitor the presence of bacterial DNA in synovial samples from patients with septic arthritis during antibiotic treatment. The absence of bacterial DNA could help in the decision to discontinue antibiotic treatment.

Detection of mycobacteria in joint samples from patients with arthritis using a genus-specific polymerase chain reaction and sequence analysis.

OBJECTIVE: Mycobacteria have been implicated in the pathogenesis of various forms of arthritis. The aim of this study was to examine the diagnostic potential of molecular biological techniques as well as to investigate the pathogenetic role of mycobacteria in chronic arthritis. PATIENTS AND METHODS: DNA, extracted from synovial fluid and synovial tissue samples from patients with mycobacterial septic arthritis (n = 2), seronegative spondyloarthropathies (SpA) (n = 18), undifferentiated arthritis (UA) (n = 21) and rheumatoid arthritis (RA) (n = 40), was analysed using a mycobacterial genus-specific polymerase chain reaction (PCR) applied to amplify mycobacterial DNA. Subsequently, automated sequencing was performed for speciation. Samples from patients with either non-mycobacterial septic arthritis, osteoarthritis (OA), crystal arthritis or joint trauma served as negative controls (n = 19). RESULTS: Mycobacterium tuberculosis complex and Mycobacterium marinum were detected in the two patients with mycobacterial septic arthritis. The other species identified were Mycobacterium hodleri (in one RA patient), Mycobacterium smegmatis (in one OA patient and two RA patients) and Mycobacterium austroafricanum (in one crystal arthritis patient). All other samples were negative. CONCLUSIONS: The results suggest that the mycobacterial genus-specific PCR applied on DNA extracts isolated directly from joint samples may be employed as an additional diagnostic tool in the case of clinical suspicion of a mycobacterial infection. No evidence was obtained for a pathogenetic role of mycobacteria in SpA, UA or RA.

Detection of Borrelia burgdorferi sensu stricto by reverse line blot in the joints of Dutch patients with Lyme arthritis.

OBJECTIVE: To analyze the presence of Borrelia burgdorferi sensu lato in synovial samples from the knee joint of patients with Lyme arthritis by polymerase chain reaction, and to differentiate the species by reverse line blot (RLB). METHODS: Synovial fluid (SF) and synovial tissue (ST) samples were obtained from patients with Lyme arthritis (n = 4) and from patients with various other forms of arthritis (n = 9). DNA extracted from synovial samples was amplified by using, as a target, the spacer region between the 5S and 23S ribosomal RNA genes of B. burgdorferi sensu lato. Subsequently, 4 species-specific DNA probes were used in the RLB for specific hybridization. RESULTS: DNA from B. burgdorferi sensu stricto DNA was detected in the SF and ST from 3 patients with Lyme arthritis. B. burgdorferi sensu lato DNA was not detected in the synovial samples from 9 control patients. CONCLUSION: The relationship between different species of B. burgdorferi sensu lato and arthritis can be studied using direct analysis of extracted DNA from joint samples. This method can be used to study the association between particular clinical manifestations of Lyme disease and different species of B. burgdorferi sensu lato.

Surveillance of respiratory pathogens and influenza-like illnesses in general practices - The Netherlands, winter 1997-98.

The Netherlands Institute of Primary Health Care (NIVEL) has coordinated the activities of a sentinel surveillance network of 43 general practices since 1970. These practices care for 1% of the Dutch population, a sample representative of the national pop

Detection of bacterial DNA in joint samples from patients with undifferentiated arthritis and reactive arthritis, using polymerase chain reaction with universal 16S ribosomal RNA primers.

OBJECTIVE: Bacteria are considered to be important in the pathogenesis of several forms of arthritis. The goal of this study was to apply the 16S ribosomal RNA (rRNA)-polymerase chain reaction method for the detection of bacterial DNA in synovial fluid (SF) and synovial tissue (ST) from inflamed joints. METHODS: Samples from 5 patients with septic arthritis and from 7 with osteoarthritis or arthritis secondary to joint trauma were used as controls. Samples from 6 patients with spondylarthropathy (SpA) and from 20 with undifferentiated arthritis (UA) were analyzed for the presence of bacterial DNA using universal 16S rRNA primers. Automated sequencing and comparative data analysis were performed to identify the species. RESULTS: In the positive control group, the bacterial species cultured from the synovium could be identified in all cases. No bacterial DNA was detected in the SF and ST from patients in the negative control group. In 4 of 6 patients with SpA and 7 of 20 with UA, the analysis of joint samples revealed the presence of bacterial DNA. Sequence analysis indicated the presence of multiple species, which was confirmed by sequencing of cloned products. CONCLUSION: When the the above techniques were used with a stringent regimen, we were able to demonstrate that it is possible to collect and analyze joint samples without contaminating bacterial DNA. The accumulation of phagocytic cells that contain bacterial DNA of various species could play a role in the pathogenesis of both SpA and UA.

Yersinia enterocolitica: a cause of chronic polyarthritis.

To investigate the role of Yersinia persistence in chronic undifferentiated arthritis, two patients who had chronic undifferentiated polyarthritis and circulating IgA and IgG antibodies to Yersinia outer proteins were studied. Immunofluorescence using antibodies directed against Yersinia adhesin A was performed on colonic and synovial tissue. Synovial tissue T cells were cloned aspecifically and screened for their proliferative responses to Yersinia enterocolitica. Furthermore, a Yersinia-specific polymerase chain reaction (PCR) was performed on synovial tissue. Both patients were found to have Yersinia antigens in colonic and synovial tissue. Y. enterocolitica-positive T-cell clones were grown from the synovial tissue: 4 CD4+ clones of 37 clones from patient 1 and 6 CD4+ clones of 53 clones from patient 2. Yersinia-specific PCR products were not detected in the synovial tissue specimens. The results support the hypothesis that an immune-mediated response to Yersinia antigens may play an important role in the pathogenesis of chronic undifferentiated arthritis.