Targeted screening as a tool for the early detection of chronic Q fever patients after a large outbreak

In the aftermath of the Dutch Q fever outbreak, an increasing number of patients are being diagnosed with chronic Q fever. Most of these patients are unaware of being infected with Coxiella burnetii, the causative agent of Q fever. To find patients in an earlier, asymptomatic stage, a targeted screening strategy (TSS) for patients with risk factors for chronic Q fever was started in the southeast region of Noord-Brabant. In total, 763 patients were tested using an IgG phase II indirect fluorescent antibody test (IFAT), of which 52 (7 %) patients tested positive. Ten of these 52 patients displayed a chronic Q fever serological profile. All of these 10 patients had a heart valve(s) or (endo-)vascular prosthesis. All except one were asymptomatic. Suggestive signs for chronic infections on positron emission tomography–computed tomography (PET-CT) were demonstrated in 5 (50 %) of these patients. Forty-two out of the 52 patients with a positive screening test showed a past Q fever serological profile. After a year of follow-up (every 3 months), none of these patients showed elevation of antibody titres and no new chronic Q fever patients were found in this group. A targeted screening programme is a useful instrument for detecting patients at risk of developing chronic Q fever.     

Serology in chronic Q fever is still surrounded by question marks

Detection of antibodies using immunofluoresence tests (IFAT) is recommended for diagnosis of chronic Q fever, but other commercial antibody assays are also available. We compared an enzyme-linked immunosorbent assay (ELISA) (Virion/Serion) and a complement fixation test (CFT) (Virion/Serion) for the detection of Coxiella burnetii IgG phase I and IgA phase I in early- and follow-up serum samples from patients with chronic Q fever, diagnosed according to an algorithm that involves IFAT. For this, we tested sera of 49 patients, including 30 proven, 14 probable and five possible chronic Q fever cases. Sensitivity of CFT for diagnosis of chronic Q fever was suboptimal (85 %), as eight patients, including five with chronic Q fever, tested negative at time of diagnosis, whereas IgG phase I antibodies were detected in these five patients by ELISA. Sensitivity of ELISA was higher, although three probable patients were missed. No differences in ELISA IgA phase I detection between proven chronic Q fever and probable were observed; instead possible patients were in majority IgA negative (60 %). Serological examination using ELISA and CFT in follow-up sera from 26 patients on treatment was unsatisfactory. Like IFAT, all kinetic options were possible: decreasing, remaining stable or even increase during time. This study demonstrated that the sensitivity of CFT-based phase I antibody detection is low and therefore not recommended for diagnosis of chronic Q fever. Based on our results, serological follow-up to guide treatment decisions was of limited value.     

Long-term serological follow-up after primary Coxiella burnetii infection in patients with vascular risk factors for chronic Q fever

We evaluated the long-term serological follow-up of patients with vascular risk factors for chronic Q fever that were previously Coxiella burnetii seropositive. C. burnetii phase I IgG titers were reevaluated in patients that gave informed consent or retrospectively collected in patients already deceased or lost to follow-up. Of 107 patients, 25 (23.4%) became seronegative, 77 (72.0%) retained a profile of past resolved Q fever infection, and five (4.7%) developed chronic Q fever. We urge clinicians to stay vigilant for chronic Q fever beyond two years after primary infection and perform serological testing based on clinical presentation.

     

Evaluation of Commonly Used Serological Tests for Detection of Coxiella burnetii Antibodies in Well-Defined Acute and Follow-Up Sera

In this study, we compared Coxiella burnetii IgG phase I, IgG phase II, and IgM phase II detection among a commercially available enzyme-linked immunosorbent assay (ELISA) (Virion/Serion), an indirect fluorescent antibody test (IFAT) (Focus Diagnostics), and a complement fixation test (CFT) (Virion/Serion). For this, we used a unique collection of acute- and convalescent-phase sera from 126 patients with acute Q fever diagnosed by positive Coxiella burnetii PCR of blood. We were able to establish a reliable date of onset of disease, since DNA is detectable within 2 weeks after the start of symptoms. In acute samples, at t = 0, IFAT demonstrated IgM phase II antibodies in significantly more sera than did ELISA (31.8% versus 19.7%), although the portion of solitary IgM phase II was equal for IFAT and for ELISA (18.2% and 16.7%, respectively). Twelve months after the diagnosis of acute Q fever, 83.5% and 62.2% of the sera were still positive for IgM phase II with IFAT and ELISA, respectively. At 12 months IFAT IgG phase II showed the slowest decline. Therefore, definitive serological evidence of acute Q fever cannot be based on a single serum sample in areas of epidemicity and should involve measurement of both IgM and IgG antibodies in paired serum. Based on IgG phase II antibody detection in paired samples (at 0 and 3 months) from 62 patients, IFAT confirmed more cases than ELISA and CFT, but the differences were not statically significant (100% for IFAT, 95.2% for ELISA, and 96.8% for CFT). This study demonstrated that the three serological tests are equally effective in diagnosing acute Q fever within 3 months of start of symptoms. In follow-up sera, more IgG antibodies were detected by IFAT than by ELISA or CFT, making IFAT more suitable for prevaccination screening programs.     

Q fever serology: cutoff determination for microimmunofluorescence.

Q fever, a worldwide zoonosis caused by Coxiella burnetii, lacks clinical specificity and may present as acute or chronic disease. Because of this polymorphism, serological confirmation is necessary to assess the diagnosis. Although microimmunofluorescence is our reference technique, the cutoff titers that are currently used to make a diagnosis of active or chronic Q fever were determined years ago with limited series of patients and sera. We determined the titers of immunoglobulin G (IgG), IgM, and IgA against both phases (I and II) of Coxiella burnetii. Rheumatoid factor was removed before testing IgM and IgA. We report here the various cutoff titers and the kinetics of antibody development from 2,218 first serum samples of patients, among whom 208 suffered from acute Q fever and 53 had chronic Q fever. In active Q fever, we have defined a low cutoff (phase II IgG titer < or = 100) below which the diagnosis cannot be made and would need further confirmation and confirmed a high cutoff (phase II IgG titer > or = 200 and phase II IgM titer > or = 50) over which the diagnosis can be made. For chronic Q fever diagnosis, phase I IgA titers are not contributive despite previous works claiming their usefulness; a phase I IgG titer of > or = 800 is highly predictive (98%) and sensitive (100%). We have also studied the possibility of rejecting or evoking the diagnosis of chronic Q fever by phase II IgG and IgA titers. This method is useful when phase I testing is not available, but the sensitivity remains low (57%).     

Q fever and spontaneous abortion

Q fever, caused by Coxiella burnetii, may result in abortions in infected animals and pregnant women. However, the role that Q fever plays in spontaneous abortions is still unknown. This study examined the association between Q fever serology and abortion in a region where Q fever is endemic. A case–control population-based study was conducted in General Yagüe Hospital (Burgos area, Spain) between June 2009 and July 2010. A total of 801 samples from 500 pregnant women were tested, of whom 273 had a spontaneous abortion and 227 gave birth. IgG and IgM antibody titres against Q fever were determined in their two phases (I and II) by immunofluorescence assay. Seropositivity (phase I IgG ≥1:16 or phase II IgG ≥1:80) was detected in 88/273 (32.2%) cases and 53/227 (23.3%) controls; p <0.01, OR 1.5, 95% CI 1.0–2.3. Seropositivity for both phases of IgG, compatible with recent or persistent infection, was detected in 55 (20.1%) vs 22 (9.7%); p <0.001, OR 2.3, 95% CI 1.3–3.9. High phase II IgG antibodies compatible with active or recent infection (titres ≥1:160) were detected in 27 (9.6%) vs 7 (3.1%); p <0.002, OR 3.4, 95% CI 1.4–8.0, respectively. Q fever was diagnosed in 14 (5.1%) cases. The risk of abortion associated with serological markers of active or recent Q fever in pregnant women was measurable and noticeable in this population, and accounted for 12% (95% CI 4–21%).     

Microbiological Challenges in the Diagnosis of Chronic Q Fever

Diagnosis of chronic Q fever is difficult. PCR and culture lack sensitivity; hence, diagnosis relies mainly on serologic tests using an immunofluorescence assay (IFA). Optimal phase I IgG cutoff titers are debated but are estimated to be between 1:800 and 1:1,600. In patients with proven, probable, or possible chronic Q fever, we studied phase I IgG antibody titers at the time of positive blood PCR, at diagnosis, and at peak levels during chronic Q fever. We evaluated 200 patients, of whom 93 (46.5%) had proven, 51 (25.5%) had probable, and 56 (28.0%) had possible chronic Q fever. Sixty-five percent of proven cases had positive Coxiella burnetii PCR results for blood, which was associated with high phase I IgG. Median phase I IgG titers at diagnosis and peak titers in patients with proven chronic Q fever were significantly higher than those for patients with probable and possible chronic Q fever. The positive predictive values for proven chronic Q fever, compared to possible chronic Q fever, at titers 1:1,024, 1:2,048, 1:4,096, and ≥1:8,192 were 62.2%, 66.7%, 76.5%, and ≥86.2%, respectively. However, sensitivity dropped to <60% when cutoff titers of ≥1:8,192 were used. Although our study demonstrated a strong association between high phase I IgG titers and proven chronic Q fever, increasing the current diagnostic phase I IgG cutoff to >1:1,024 is not recommended due to increased false-negative findings (sensitivity < 60%) and the high morbidity and mortality of untreated chronic Q fever. Our study emphasizes that serologic results are not diagnostic on their own but should always be interpreted in combination with clinical parameters.     

Real-Time PCR with Serum Samples Is Indispensable for Early Diagnosis of Acute Q Fever

The world''s largest Q fever outbreak is ongoing in The Netherlands with around 3,000 confirmed cases since the first half of 2007. Increased awareness has resulted in early referral of patients for diagnostics. An important drawback to serological diagnosis of acute Q fever is the lag phase in antibody response. Therefore, we evaluated the performance of a real-time PCR for detection of Coxiella burnetii DNA using serum samples from patients with acute Q fever. PCR, targeting IS1111, was retrospectively performed on acute-phase and follow-up convalescent-phase serum samples from 65 patients with acute Q fever as diagnosed by immunofluorescence assay. The results obtained by PCR were related to disease stage as defined by subsequent appearance of phase II IgM, phase II IgG, phase I IgM, and phase I IgG (IgM-II, IgG-II, IgM-I, and IgG-I, respectively) antibodies and time since onset of disease. In addition, we analyzed seronegative acute-phase serum samples from patients with inconclusive Q fever serology, because no convalescent-phase serum samples were available. PCR was scored positive in 49/50 (98%) seronegative sera, 9/10 (90%) sera with isolated IgM-II antibodies, 3/13 (23%) sera with IgM-II/IgG-II antibodies, 2/41 (5%) sera with IgM-II/IgG-II/IgM-I antibodies, 0/15 (0%) sera with IgM-II/IgG-II/IgM-I/IgG-I antibodies, and 0/1 (0%) serum sample with IgM-II/IgG-II/IgG-I antibodies. The latest time point after onset of disease in which C. burnetii DNA could be detected was at day 17. In patients with inconclusive Q fever serology, PCR was positive in 5/50 (10%) cases. We conclude that real-time PCR with serum samples is indispensable for early diagnosis of acute Q fever. C. burnetii DNA becomes undetectable in serum as the serological response develops.Q fever, an infection caused by the bacterium Coxiella burnetii, results in a self-limiting disease in 40 to 50% of infected cases. Pneumonia is the predominant presenting symptom in acute Q fever, although fever and hepatitis are also frequently observed (9, 10). Failure to diagnose acute Q fever and delay in treatment may lead to prolonged morbidity and increased hospital admission rates (4, 7, 11, 14).During three consecutive years, large Q fever epidemics occurred in an area in the south of The Netherlands where the disease was formerly not prevalent (11). In 2007 there were a total of 191 confirmed cases reported, in 2008 a total of 998, and in 2009 more than 2,000 confirmed cases were reported, which ranks the outbreak as the largest Q fever epidemic recorded to date. The affected area has a large density of dairy goats, of which a number have tested positive for Q fever. Next to the differences in sizes of the epidemics, the interval between onset of disease and date of diagnosis decreased from a median of 77 days in 2007 to 29 days in 2008 and 17 days in 2009 (12). Moreover, the hospital admission rates were reduced from 40% in 2007 to 20% in 2008 (11). Both observations are most likely due to increased awareness among physicians in the affected area resulting in early submission of clinical samples to the laboratory, subsequent earlier diagnosis, and probably fewer undiagnosed cases. The majority of diagnostic samples from both epidemics were submitted to our laboratory, which lies in the center of the epidemic area and serves a catchment area of roughly 500,000 persons in a semirural district supporting two hospitals and surrounding general practitioners.The gold standard for serological diagnosis of an infectious disease is either a seroconversion or a 4-fold rise in antibody titer. The reference test for serological diagnosis of Q fever is the immunofluorescence assay (IFA) (8). Antibodies are expressed against phase II antigens during the acute infection and against phase I antigens in the established infection. For both antigens, IgM antibody production precedes IgG production, and thus three phases can be distinguished in acute Q fever: a seronegative phase followed by IgM/IgG phase II seroconversion during the acute infection and subsequent IgM/IgG phase I seroconversion in the established infection. However, an important drawback to serological diagnosis of acute Q fever is the lag phase in antibody response of 7 to 15 days after onset of clinical symptoms (8).Apart from serology, C. burnetii-specific PCR of serum samples can be an additional tool to diagnose Q fever in the early acute phase, but conflicting sensitivities have been reported (3, 13). Here, we evaluated the performance of an in-house-developed real-time PCR assay for detection of C. burnetii DNA in serum samples from patients with acute Q fever.     

Association between serological evidence of past Coxiella burnetii infection and atherosclerotic cardiovascular disease in elderly patients

Q fever, caused by Coxiella burnetii, may cause vascular complications, but the role that this infection may play in the development of atherosclerotic cardiovascular disease remains unknown. This study examined the association between Q fever serology and cardiovascular disease in a region where Q fever is endemic. A case-control study was conducted in the Hospital Universitario de Burgos (Spain) between February 2011 and June 2012. A total of 513 samples were tested, from 454 hospitalized patients ≥65 years old, of whom 164 were cases (patients with prevalent or incident coronary heart, cerebrovascular or peripheral artery, disease) and 290 controls (patients without cardiovascular disease). Serum IgG antibody phase II titres against Q fever were determined by immunofluorescence assay. Seropositivity (titres ≥1:256) was detected in 84/164 (51.2%) cases and in 109/290 (37.6%) controls (p = 0.005; OR, 1.7; 95% CI, 1.1–2.5). This ratio increases when adjusted for sex, hypertension, dyslipidaemia, smoking, diabetes and atrial fibrillation (OR, 2.6; 95% CI, 1.5–4.7). The geometric mean titre (GMT) for C. burnetii phase II assay was higher in cases than in controls (p = 0.004). We found no significant relationship between cardiovascular disease and C. pneumoniae, and Cytomegalovirus seropositivity (both determined by the IgG ELISA method). In conclusion, serological evidence of past Q fever is associated with atherosclerotic cardiovascular disease in elderly patients in an endemic region.     

Early Diagnosis and Treatment of Patients with Symptomatic Acute Q Fever Do Not Prohibit IgG Antibody Responses to Coxiella burnetii

Little is known about the effect of timing of antibiotic treatment on development of IgG antibodies following acute Q fever. We studied IgG antibody responses in symptomatic patients diagnosed either before or during development of the serologic response to Coxiella burnetii. Between 15 and 31 May 2009, 186 patients presented with acute Q fever, of which 181 were included in this retrospective study: 91 early-diagnosed (ED) acute Q fever patients, defined as negative IgM phase II enzyme-linked immunosorbent assay (ELISA) and positive PCR, and 90 late-diagnosed (LD) acute Q fever patients, defined as positive/dubious IgM phase II ELISA and positive immunofluorescence assay (IFA). Follow-up serology at 3, 6, and 12 months was performed using IFA (IgG phase I and II). High IgG antibody titers were defined as IgG phase II titers of ≥1:1,024 together with IgG phase I titers of ≥1:256. At 12 months, 28.6% of ED patients and 19.5% of LD patients had high IgG antibody titers (P = 0.17). No statistically significant differences were found in frequencies of IgG phase I and IgG phase II antibody titers at all follow-up appointments for adequately and inadequately treated patients overall, as well as for ED and LD patients analyzed separately. Additionally, no significant difference was found in frequencies of high antibody titers and between early (treatment started within 7 days after seeking medical attention) and late timing of treatment. This study indicates that early diagnosis and antibiotic treatment of acute Q fever do not prohibit development of the IgG antibody response.     

The prognostic value of serological titres for clinical outcomes during treatment and follow-up of patients with chronic Q fever

ObjectivesWe assessed the prognostic value of phase I IgG titres during treatment and follow-up of chronic Q fever.MethodsWe performed a retrospective cohort study to analyse the course of phase I IgG titres in chronic Q fever. We used a multivariable time-varying Cox regression to assess our primary (first disease-related event) and secondary (therapy failure) outcomes. In a second analysis, we evaluated serological characteristics after 1 year of therapy (fourfold decrease in phase I IgG titre, absence of phase II IgM and reaching phase I IgG titre of ≤1:1024) with multivariable Cox regression.ResultsIn total, 337 patients that were treated for proven (n = 284, 84.3%) or probable (n = 53, 15.7%) chronic Q fever were included. Complications occurred in 190 (56.4%), disease-related mortality in 71 (21.1%) and therapy failure in 142 (42.1%) patients. The course of phase I IgG titres was not associated with first disease-related event (HR 1.00, 95% CI 0.86–1.15) or therapy failure (HR 1.02, 95% CI 0.91–1.15). Similar results were found for the serological characteristics for the primary (HR 0.97, 95% CI 0.62–1.51; HR 1.12, 95% CI 0.66–1.90; HR 0.99, 95% CI 0.57–1.69, respectively) and secondary outcomes (HR 0.86, 95% CI 0.57–1.29; HR 1.37, 95% CI 0.86–2.18; HR 0.80, 95% CI 0.48–1.34, respectively).DiscussionCoxiella burnetii serology does not reliably predict disease-related events or therapy failure during treatment and follow-up of chronic Q fever. Alternative markers for disease management are needed, but, for now, management should be based on clinical factors, PCR results, and imaging results.     

Prevalence of chronic Q fever in patients with a history of cardiac valve surgery in an area where Coxiella burnetii is epidemic

Chronic Q fever develops in 1 to 5% of patients infected with Coxiella burnetii. The risk for chronic Q fever endocarditis has been estimated to be ≈ 39% in case of preexisting valvulopathy and is potentially even higher for valvular prostheses. Since 2007, The Netherlands has faced the largest Q fever outbreak ever reported, allowing a more precise risk estimate of chronic Q fever in high-risk groups. Patients with a history of cardiac valve surgery were selected for microbiological screening through a cardiology outpatient clinic in the area where Q fever is epidemic. Blood samples were analyzed for phase I and II IgG against C. burnetii, and if titers were above a defined cutoff level, C. burnetii PCR was performed. Chronic Q fever was considered proven if C. burnetii PCR was positive and probable if the phase I IgG titer was ≥ 1:1,024. Among 568 patients, the seroprevalence of C. burnetii antibodies (IgG titer greater than or equal to 1:32) was 20.4% (n = 116). Proven or probable chronic Q fever was identified among 7.8% of seropositive patients (n = 9). Valve characteristics did not influence the risk for chronic Q fever. Patients with chronic Q fever were significantly older than patients with past Q fever. In conclusion, screening of high-risk groups is a proper instrument for early detection of chronic Q fever cases. The estimated prevalence of chronic Q fever is 7.8% among seropositive patients with a history of cardiac valve surgery, which is substantially higher than that in nonselected populations but lower than that previously reported. Older age seems to increase vulnerability to chronic Q fever in this population.     

The sexual dimorphism of anticardiolipin autoantibodies in acute Q fever patients

ObjectivesQ fever is a zoonotic disease caused by Coxiella burnetii which affects men more than women (sex ratio men/women: 2.2). Acute Q fever complications are associated with elevation of anticardiolipin (aCL) antibodies. Here, we investigate the sexual dimorphism of aCL antibodies during acute C. burnetii infection.MethodsIgG aCL antibodies were evaluated at the time of Q fever serological diagnosis with enzyme-linked immunosorbent assay. Results were analysed according to sex.ResultsAmong the 1323 patients with Q fever tested for aCL, 1013 had acute Q fever (692 men/321 women) and 310 had persistent focalized infection (226 men/84 women). In cases of acute Q fever, men presented a significantly higher proportion of positive aCL antibodies (351/692, 50.7%) than women (113/321, 35.2%) (p <0.05). In addition, men had significantly higher aCL antibodies levels than women (p <0.001).ConclusionsWe highlight a relationship between sex and markers of autoimmunity during Q fever. Further investigations are necessary to better understand the mechanisms of this sexual dimorphism.     

Serological differentiation between acute (late control) and endocarditis Q fever.

AIMS--To differentiate the serological profiles of chronic (endocarditis) Q fever from the late follow up of acute cases. METHODS--Twenty patients (10 diagnosed with acute and 10 with endocarditis Q fever) were studied. Those diagnosed with acute infection were followed up from 2.5 to 88 months (mean 35.8 months). Serological variables included indirect immunofluorescence against phase I and II of Coxiella burnetii (IgM, IgG, and IgA), complement fixation and rheumatoid factor (RF). RESULTS--All patients with titres of IgA against phase I, after IgG removal, equal to or above 320 and a complement fixation value equal to or above 128 had endocarditis. No patient with acute Q fever had such a serological profile. CONCLUSIONS--The combination of IgA against phase I and complement fixation values may be sufficient to differentiate the serological profile of chronic (endocarditis) Q fever from the late follow up of acute cases.     

Detection of phase I IgG antibodies to Coxiella burnetii with EIA as a screening test for blood donations

The presence of a high phase I IgG antibody titre may indicate chronic infection and a risk for the transmission of Coxiella burnetii through blood transfusion. The outbreak of Q fever in the Netherlands allowed for the comparison of an enzyme immunoassay (EIA) with the reference immunofluorescence assay (IFA) in a large group of individuals one year after acute Q fever. EIA is 100?% sensitive in detecting high (≥1:1,024) phase I IgG antibody titres. The cost of screening with EIA and confirming all EIA-positive results with IFA is much lower than screening all donations with IFA. This should be taken into account in cost-effectiveness analyses of screening programmes.     

Predominant Immunoglobulin A Response to Phase II Antigen of Coxiella burnetii in Acute Q Fever

Diagnosis of acute Q fever is usually confirmed by serology, on the basis of anti-phase II antigen immunoglobulin M (IgM) titers of ≥1:50 and IgG titers of ≥1:200. Phase I antibodies, especially IgG and IgA, are predominant in chronic forms of the disease. However, between January 1982 and June 1998, we observed anti-phase II antigen IgA titers of ≥1:200 as the sole or main antibody response in 10 of 1,034 (0.96%) patients with acute Q fever for whom information was available. In order to determine whether specific epidemiological or clinical factors were associated with these serological profiles, we conducted a retrospective case-control study that included completion of a standardized questionnaire, which was given to 40 matched controls who also suffered from acute Q fever. The mean age of patients with elevated phase II IgA titers was significantly higher than that usually observed for patients with acute Q fever (P = 0.026); the patients were also more likely than controls to live in rural areas (P = 0.026) and to have increased levels of transaminase in blood (P = 0.03). Elevated IgA titers are usually associated with chronic Q fever and are directed mainly at phase I antigens. Although the significance of our findings is unexplained, we herein emphasize the fact that IgA antibodies are not specific for chronic forms of Q fever and that they may occasionally be observed in patients with acute disease. Moreover, as such antibody profiles may not be determined by most laboratories, which test only for total antibody titers to phase I and II antigens, the three isotype-specific Ig titers should be determined as the first step in diagnosing Q fever.     

Molecular detection of Coxiella burnetii in the sera of patients with Q fever endocarditis or vascular infection

In the absence of a specific diagnosis based on serology, chronic Q fever is inevitably fatal. However, diagnosis is often delayed because the test is not widely available. To shorten the diagnostic delay, we adapted a nested-PCR assay with serum as a template and the LightCycler as a thermal cycler, termed LCN-PCR. We retrospectively and prospectively applied this method to samples from 48 patients diagnosed with Q fever endocarditis or vascular infection and to samples from 100 controls with endocarditis caused by other microorganisms. We also prospectively applied this technique to samples from 30 patients treated for a Q fever endocarditis and to samples from 13 patients with a convalescent acute Q fever with ambiguous immunoglobulin G (IgG) phase I titer. LCN-PCR had a specificity of 100%. It was positive only in samples from patients with evolutive Q fever, as none of the samples from patients with a treated chronic Q fever or with a convalescent acute Q fever presented positive results. When performed prospectively on recently stored sera, the sensitivity of LCN-PCR is 64% (7 of 11 samples; P = 0.004), but the efficiency of LCN-PCR was dramatically altered by the storage of specimens at -20 degrees C. High IgG phase I titers decreased the sensitivity of LCN-PCR. A significant difference was observed among LCN-PCR results for sera with IgG phase I titers of > or =1:25,600 compared to sera with IgG phase I titers of <1:25,600 (0 of 15 samples versus 13 of 33 samples; P = 0.004). In patient samples with titers below 1:25,600 tested prospectively, sensitivity was 100% (7 of 7). The LCN-PCR assay may be helpful in establishing an early diagnosis of chronic Q fever.     

Enzyme-linked immunosorbent assay for diagnosis of chronic Q fever.

From 1982 through 1987 we diagnosed 13 chronic Q fever cases. Clinically these patients presented a culture-negative endocarditis, and all but two had high complement-fixing antibody titers to Coxiella burnetii phase I (reciprocal titer above 200). With the enzyme-linked immunosorbent assay (ELISA), titers of immunoglobulin G (IgG) to phases I and II of C. burnetii averaged 158,000 and 69,900, respectively, whereas they reached 300 and 3,200 in acute Q fever cases. Similarly, IgA to both phases of C. burnetii and IgM to phase I were consistently higher during chronic than acute Q fever. The serological follow-up of one patient with chronic Q fever over a 4-year period showed a good correlation between the titers of IgG and IgM antibody titers detected by ELISA and indirect fluorescent-antibody test (IFA) to both phases of C. burnetii. Few discrepancies appeared with IgA. Shortly after initiation of antibiotic treatment, a slow and steady decrease of the antibody titers to C. burnetii phases I and II was observed. The complement fixation, IFA, and ELISA tests showed the same type of antibody response. The ELISA proved to be an excellent diagnostic test for chronic Q fever. It distinguished negative from positive reactions clearly, and results were highly reproducible. The reading is objective, and the test is simple to perform and more sensitive than the IFA and complement fixation tests. The ELISA is recommended for serologic evaluation of patients with chronic Q fever.     

Emerging evidence for Q fever in humans in Denmark: role of contact with dairy cattle

Until recently, Q fever was notified in very low numbers annually in Denmark and it was always considered to be acquired abroad. Preliminary reports now describe Coxiella burnetii in milk samples from Danish dairy cattle. Serum samples of a large cohort of farmers, veterinarians, inseminators and hoof trimmers, all having occupational contact with dairy cattle, were tested for the presence of IgG to phase I and phase II antigens of C. burnetii. In 39 of 359 individuals studied (11%), the presence of antibodies to C. burnetii was found. Veterinarians had the highest seropositivity rate (36%). This survey suggests that C. burnetii is a recently recognized domestic infection in Denmark and that risk of infection is associated with occupation.