Chlamydophila pneumoniae diagnostics: importance of methodology in relation to timing of sampling.

The diagnostic impact of PCR-based detection was compared to single-serum IgM antibody measurement and IgG antibody seroconversion during an outbreak of Chlamydophila pneumoniae in a military community. Nasopharyngeal swabs for PCR-based detection, and serum, were obtained from 127 conscripts during the outbreak. Serum, drawn many months before the outbreak, provided the baseline antibody status. C. pneumoniae IgM and IgG antibodies were assayed using microimmunofluorescence (MIF), enzyme immunoassay (EIA) and recombinant ELISA (rELISA). Two reference standard tests were applied: (i) C. pneumoniae PCR; and (ii) assay of C. pneumoniae IgM antibodies, defined as positive if >or=2 IgM antibody assays (i.e. rELISA with MIF and/or EIA) were positive. In 33 subjects, of whom two tested negative according to IgM antibody assays and IgG seroconversion, C. pneumoniae DNA was detected by PCR. The sensitivities were 79%, 85%, 88% and 68%, respectively, and the specificities were 86%, 84%, 78% and 93%, respectively, for MIF IgM, EIA IgM, rELISA IgM and PCR. In two subjects, acute infection was diagnosed on the basis of IgG antibody seroconversion alone. The sensitivity of PCR detection was lower than that of any IgM antibody assay. This may be explained by the late sampling, or clearance of the organism following antibiotic treatment. The results of assay evaluation studies are affected not only by the choice of reference standard tests, but also by the timing of sampling for the different test principles used. On the basis of these findings, a combination of nasopharyngeal swabbing for PCR detection and specific single-serum IgM measurement is recommended in cases of acute respiratory C. pneumoniae infection.

Field detection of Francisella tularensis.

A field investigation was undertaken following an outbreak of water-borne tularemia in Northern Norway. Francisella tularensis bacterial cellular components were analysed by rapid immunochromatography (RI)-testing, enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR). Water from 1 reservoir, fed from a rapid stream, tested negative. From another reservoir, 2 of a chain of 3 wells tested negative. The third well, at the end of the chain, contained lemming (Lemmus lemmus) carcasses and gave ample proof of F. tularensis contamination. We concluded that the origin of the outbreak was dead, infective lemming carcasses in the water sources. For the various sampling materials, the RI-test proved itself particularly handy and versatile, compared with the ELISA and the PCR.

Ground penetrating radar surveys to locate 1918 Spanish flu victims in permafrost.

The "Spanish Flu" killed over 40 million people worldwide in 1918. Archival records helped us identify seven men who died of influenza in 1918 and were interred in Longyearbyen, Svalbard, Norway, 1,300 km from the North Pole. Ground Penetrating Radar (GPR) was used successfully, in a high-resolution field survey mode, to locate a large excavation with seven coffins, near the existing seven grave markers. The GPR indicated that the ground was disturbed to 2 m depth and was frozen below 1 m. Subsequent excavation showed that: a) the GPR located the position of the graves accurately, b) the coffins were buried less than 1 m deep, and c) that the frozen ground was 1.2 m deep where the coffins were located. The GPR assisted in planning the exhumation, safely and economically, under the high degree of containment required. Virologic and bacteriologic investigations on recovered tissues may give us an opportunity to isolate and identify the micro-organisms involved in the 1918 influenza and expand our knowledge on the pathogenesis of influenza.

Organobismuth compounds: activity against Helicobacter pylori.

BACKGROUND AND METHODS: The antibacterial activity of originally devised and synthesized organobismuth compounds was tested against fresh clinical isolates of Helicobacter pylori and compared with clinically well-established inorganic bismuth salts currently used in triple antibacterial treatment to eradicate H. pylori. The test conditions in vitro were standard for determination of minimum inhibitory concentrations (MICs). RESULTS: Organic compounds with covalently bound bismuth showed stronger and more consistent antibacterial activity than inorganic bismuth salts. Whereas the most active among the standard therapeutic inorganic compounds showed MICs against the test organisms of 4-8 mg/l (bismuth salicylate) and 0.5-64 mg/l, the most active neosynthesized organic substance, tris(2.6-dimethylphenyl)-bismuthine, consistently showed an MIC of 4 mg/l against all bacterial strains. CONCLUSIONS: The new line of organobismuth compounds might offer a therapeutic potential against the bacteria causing peptic ulcer disease.

Antibiotic resistance of pneumococci in Norway.

A collection of 178 pneumococcal isolates found in Norway during the period 1987-1994 were tested for their susceptibility to benzylpenicillin, macrolides (azithromycin, clarithromycin, dirithromycin, erythromycin, roxithromycin, spiramycin), fluoroquinolones (ciprofloxacin, sparfloxacin), imipenem, chloramphenicol, and vancomycin by a standard agar dilution procedure. To benzylpenicillin, two strains (1%) showed resistance and 14 strains (8%) intermediate susceptibility. Towards erythromycin, eight strains (4%) showed resistance and four strains (2%) intermediate susceptibility. Cross-resistance was demonstrated among the macrolides. Among the fluoroquinolones, intermediate susceptibility occurred with 42% of the isolates for sparfioxacin and 90% for ciprofloxacin; to the latter 5.1% proved resistant. The sum of intermediate and highly resistant isolates was 53% for chloramphenicol. Both penicillin-resistant strains were isolated during the last 2 years of collection and came from patients of non-Norwegian ethnic background. Imported strains appeared over represented among the strains resistant to penicillin and macrolides. Only imipenem and vancomycin showed full susceptibility for all pneumococci tested. An over representation of serogroup 6 strains was apparent among the strains with intermediate susceptibility and high resistance to benzylpenicillin. It is apparent that high-level resistance has, not so far, become a difficult problem in Norway. Nevertheless, the situation requires monitoring of the resistance level, particularly in meningitis and septic patients, and certainly in patients who cntail a higher than usual possibility of acquiring pneumococci from pools of resistant strains outside Norway (visitors, immigrants and recent returness from abroad).

[Chlamydia pneumoniae infections in Norway]. / Chlamydia pneumoniae-infeksjoner i Norge.

During the last 5-6 years our understanding of Chlamydia pneumoniae has changed radically. C. pneumoniae is no longer considered a dangerous, obligatory pathogen. Rather, it is a common, highly contagious intracellular opportunist, inducing poor immunity and with a tendency to repeated reinfections. At present, a possible role in the formation of atheromatous plaques is being discussed. There is a significantly higher prevalence of antibodies against C. pneumoniae in coronary heart disease patients than in controls. Another unsolved problem is that of therapy, since chronic lung infection resists long-term macrolide antibiotic treatment. Should additional treatment with cortisone be given? Here we clearly need clinical trials before we move in a totally new direction.

Measles antibodies and herd immunity in 20- and 40-year-old Norwegians.

The introduction of a measles vaccination programme in Norway in 1969 using one dose of vaccine, and since 1983 two doses, was followed by a substantial decrease in the incidence of the disease. Since 1992, the annual incidence has been less than 20 cases. Small clusters and outbreaks have occasionally been observed among military personnel and unvaccinated children. This paper describes a seroepidemiological investigation of the level of immunity among 1,188 military conscripts, aged 18-28 years (mean 20.7) compared with 695 healthy 40-year-olds. The conscripts had been offered measles vaccine in infancy, in some cases also at 12-13 years of age, but they had also been exposed to wild measles virus, since the virus continued to circulate many years after the vaccination had started. The measles immunity in this group is considered to indicate the immunity level among the first 5 cohorts offered measles vaccine in Norway. The 40-year-olds had grown up in a community with no measles vaccination. Their level of immunity gives an indication of the level finally obtained when there are no vaccinations, and thus of the level that would induce herd immunity against measles in the Norwegian population. The aims of the vaccination programme must be to obtain a corresponding immunity. The results of the investigation show that the percentages with measles antibodies in the respective groups were 92.3 and 98.1. The observation of measles outbreaks among young Norwegian conscripts, as well as reports from several countries on outbreaks in university and college settings with levels of seropositivity of even more than 95%, indicate that the seropositivity in the 20-year-old group may be too low to afford protection, especially when this group is living under close conditions. Consideration should be given to the need for an intensification of the existing vaccination programme to ensure that the protection level needed for herd immunity is reached.

Epidemic patterns and carriage of Chlamydia pneumoniae in Norway.

Chlamydia pneumoniae infection, in earlier days misdiagnosed as ornithosis, is very common in Norway. The disease develops slowly, a feature that may account for the very large number of subclinical cases, which may be seven fold more common than clinical cases. Subclinical cases produce an antibody response similar to that seen in overt clinical disease. Silent carriage of C. pneumoniae in healthy individuals may be frequent. Therapy based on positive cell culture or polymerase chain reaction (PCR) in the absence of pneumonic symptoms may be questionable. PCR has, however, given the slow development of disease, revealed itself as a handy epidemiological technique useful for the survey of healthy populations.

Field investigations of tularemia in Norway.

In Norway, tularemia is a common disease in small rodent and hare populations, where large outbreaks can be observed. In humans, the yearly number of cases is low, usually less than ten, with peaks up to 44 recorded in recent years. Serological investigations on hunters and healthy school children nevertheless indicate, with up to 4.7% positivity in the latter group, that Francisella tularensis low-grade infection is widespread. F. tularensis in co-culture with amoebae, e.g. Achantamoeba castellanii, may grow after internalization and kill the amoeba. As with Legionella, Francisella virulence may be enhanced after protozoan ingestion. This suggests a mechanism that can explain the pattern of dissemination and infection in our region.

[Bacteremia in the county of Ribe]. / Bakteriaemi i Ribe Amt.

The Department of Microbiology at the Central Hospital of Esbjerg, established in 1987, serves the five hospitals in Ribe county, Denmark. From early on, the department has endeavoured to guide the hospital's antimicrobial policy. In order to investigate whether this involvement had any measurable impact on the antimicrobial resistance pattern in our region, we compared the resistance patterns of 212 strains isolated from the blood of bacteraemic patients in 1988 to those of 317 strains isolated in 1992. No increase in antibiotic resistance was revealed. This is noteworthy since new specialties have been established at the Central Hospital during this period, with an increased number of patients requiring antimicrobial therapy. It is important to survey the antibiotic resistance pattern closely, and that this is done locally.

Screening urine samples by leukocyte esterase test and ligase chain reaction for chlamydial infections among asymptomatic men.

Urine samples from 358 asymptomatic males were screened for urethral inflammation by the leukocyte esterase (LE) test and for Chlamydia trachomatis by the ligase chain reaction (LCR). LE and LCR positivity rates were 7.5% (27 of 358 samples) and 2.8% (10 of 358 samples), respectively. Eight of the 10 LCR-positive samples were detected by the LE screening test. The urine LE prescreening test in combination with the LCR assay may be a reasonable approach for genitourinary chlamydial disease control.

Detection of Chlamydia pneumoniae in clinical specimens by polymerase chain reaction using nested primers.

A nested primers strategy was used to develop a two-step PCR test for the direct species-specific detection of the 16s rRNA gene of Chlamydia pneumoniae. This test was applied to 58 nasopharyngeal or oropharyngeal swab specimens collected from patients in studies of community-acquired pneumonia and in a local outbreak of respiratory disease. Twelve patients (21%) showed evidence of Chlamydia pneumoniae infection in serological tests (7/56; 13%), culture (8/58; 14%) or PCR (10/58; 17%). Nested PCR but not single-step PCR was found to be as sensitive as culture or serology for detection of infection with this organism. In summary, nested PCR can be useful in direct testing of clinical specimens for Chlamydia pneumoniae, making additional DNA purification steps unnecessary.

[Acute bronchitis in adults. Clinical findings, microorganisms and use of antibiotics]. / Akutt bronkitt hos voksne. Kliniske funn, mikroorganismer og bruk av antibiotika.

Among 72 adult patients with a diagnosis of acute bronchitis, serological investigation established the presence of an aetiologic agent in 29 (40%). Influenza virus was the most common pathogen. Seven patients had bacterial infection, caused by pneumococci in four patients and Mycoplasma pneumoniae in three. Five of the patients had pneumonia as diagnosed by radiography, and mycoplasmal aetiology was established in one of these. Altogether, 11 patients either had bacterial infection or radiographic pneumonia. Although the doctors' recording of wheezes was strongly associated with prescription of antibiotics (p < 0.0001), wheezes were heard only in two of the 11 patients with pneumonia or bacterial infection, compared with 30 of the 61 patients with viral or unspecified bronchitis. The median value of C-reactive protein (CRP) was 52 mg/l in the 11 patients, significantly higher than < 11 mg/l in the 61 other patients (p < 0.0001). The corresponding values for erythrocyte sedimentation rate were 45 and 14 mm/h (p < 0.0005). The results indicate that certain patients with acute bronchitis should be treated with antibiotics, and that the erythrocyte sedimentation rate and the CRP-test may be useful in detecting which patients this applies to.

Detection of Chlamydia trachomatis in the urine of young Norwegian males by enzyme immunoassay.

First-void urine samples from 392 Norwegian military conscripts were investigated for the presence of Chlamydia trachomatis by enzyme immunoassay (EIA) on day 1 and day 5 after collection. Positive samples were subsequently investigated by direct immunofluorescence (IF) microscopy for the presence of chlamydial elementary bodies (EBs) in the urine pellet, and urethral swab material taken from the EIA-positive individuals was cultured. 4.8% (19/392) of the urine samples were EIA-positive on day 1, and 5.4% (21/392) were positive on day 5, with a combined total of 6.6% (26/392). Twenty-four of the 26 urine samples were confirmed as positive on IF microscopy. Urethral swabs were taken from 21 EIA-positive individuals. Six of the swabs were positive on cell culture, whereas nine were positive on IF microscopy of swab material, suggesting that these techniques perform better in symptomatic cases than in male Chlamydia trachomatis carriers. In the urine samples a notable discrepancy in EIA results was seen when the same refrigerated samples were retested on day 5 compared to day 1. This discrepancy was probably due to storage-related factors.

[Chlamydia pneumoniae--pathogenesis and perspectives]. / Chlamydia pneumoniae--patogenese og perspektiver.

Chlamydia pneumoniae, a Gram-negative bacterium, formerly named TWAR but identified as a distinct species since 1988, is now considered to be the most common agent of chlamydial infection in Scandinavia. C pneumoniae has a different tissue trophism from that of Chlamydia trachomatis, since C pneumoniae may infect bronchi and lungs, macrophages, monocytes, and endothelial cells. C pneumoniae, like other chlamydiae, has a slow, intracellular life cycle. An absence of reaction from the host cells, combined with scant tissual reaction owing to the low endotoxic activity of chlamydial lipopolysaccharide, may help to explain the usually discreet clinical picture. Atherosclerosis and coronary heart disease may follow chronic lung infection, and acute pneumonic episodes can trigger myocardial infarct. Asymptomatic infection with C pneumoniae is widespread. Intriguing diagnostic questions are the possible existence of a non-pathogenic carrier state, and the conceivable sensitization of the host with respect to a heterotypic, secondary chlamydial infection by, for example, C trachomatis, giving rise to an aggravated clinical picture. Early antibiotics are indicated to avoid the development of chronic disease.

Susceptibility pattern of Scandinavian Francisella tularensis isolates with regard to oral and parenteral antimicrobial agents.

Some recently introduced antimicrobial agents have only been incompletely evaluated for use in Francisella tularensis infections. The present study evaluated the susceptibility pattern of Scandinavian human, rodent, and hare F. tularensis isolates with respect to a selection of traditional as well as recently introduced antimicrobial agents. All strains were resistant to the following beta-lactams: penicillin, cephalexin, cefuroxime, ceftazidime, aztreonam, imipenem, and meropenem with minimal inhibitory concentrations > 32 mg/l. Against macrolides, a mixed susceptibility/resistance pattern appeared. All strains were susceptible to gentamicin, chloramphenicol, doxycycline, and four quinolones. Since the quinolones showed the lowest MIC values, and in addition give a good intracellular penetration, we conclude that future drugs to consider against tularemia should definitely include this group of antibiotics. The outpatient mode of antibiotic treatment is especially relevant as the Scandinavian variant of F. tularensis infection is nonlethal, usually pustuloglandular, and not septicemic. Therefore, oral drugs must be sought, and the quinolone group also satisfies this requirement.