Q fever and lymphadenopathy: report of four new cases and review

Coxiella burnetii, the causative agent of Q fever, is responsible for various clinical syndromes, but lymphadenitis has been described during Q fever in only three recent case reports. Four new cases of acute Q fever associated with lymphadenopathy are reported here, and these cases are discussed along with the three previously reported cases. Coxiella burnetii was isolated for the first time from a lymph node. Q fever should be considered an etiologic agent of lymphadenitis.     

Identification of protein candidates for the serodiagnosis of Q fever endocarditis by an immunoproteomic approach

Q fever is a worldwide zoonosis caused by Coxiella burnetii bacterium. Two clinical forms are present: acute Q fever and chronic disease, including endocarditis. Currently, the diagnosis of Q fever endocarditis is based on the detection of anti-phase I antibodies. The objective of the study was to identify candidate proteins for the serological diagnosis of endocarditis due to C. burnetii. The immunoreactivities of sera from 12 patients with C. burnetii infections, including the sera from patients with endocarditis and with the acute clinical form of Q fever, were compared with those of three control subjects who did not have Q fever. We identified 29 candidate antigenic proteins by mass spectrometry. Two proteins, arginine repressor and OmpH, were recognised exclusively by the sera of patients with Q fever endocarditis. These proteins are promising candidates for the development of serodiagnostic assays for Q fever endocarditis.     

Immunological arousal during acute Q fever infection

Physicians often encounter patients who present with a vague clinical syndrome. A wide serological workup is often ordered, which may include tests for Coxiella burnetii in endemic areas. Often, the results of these tests pose new dilemma, with overlapping positive laboratory assays. The objective of this investigation was to characterise the serological overlap between acute Q fever and other infectious and immunological diseases. We retrospectively scanned the files of patients with a positive or equivocal immunoglobulin (Ig) M for C. burnetii phase II over a period of 8 years in a general hospital. Clinical and laboratory data, including antibodies to infectious agents and antibodies related to immunological states, were recorded. Anti-nuclear antibody (ANA), smooth muscle antibody (SMA) and rheumatoid factor were positive in 38%, 33.3% and 22.2% of the cases, respectively. In patients with acute Q fever, elevated IgM levels for Epstein–Barr Virus (EBV), cytomegalovirus (CMV), Mycoplasma pneumoniae, parvovirus, Bordetella pertussis, Rickettsia conorii and R. typhi were noted in 13.8%, 8.3%, 12.12%, 22.2%, 25%, 13% and 21.7% of cases, respectively. Acute Q fever induces a non-specific immunological arousal in a significant number of patients. This may interfere with diagnosis and delay treatment. Caution, clinical judgment and serological follow-up is warranted in such conditions.     

Coxiella burnetii infection among subjects infected with HIV type 1 in the Central African Republic

Sixty-six sera from HIV-1-seropositive adult African subjects and 49 sera from HIV-seronegative age and sex matched healthy African controls living in Bangui, Central African Republic, were screened forCoxiella burnetii antibody by an indirect immunofluorescent antibody test. 16.7 % of HIV-infected patients and 16.3 % of the HIV-negative controls had positive IgG titres, with no significant difference between the two groups. Two of the seven HIV-infected patients seropositive forCoxiella burnetii for whom clinical data was available had a medical history compatible with symptomatic Q fever. These findings indicate that there is a high degree of exposure toCoxiella burnetii infection in Bangui. In individuals co-infected with HIV andCoxiella burnetii, cellular immunosuppression could favour symptomatic Q fever. Physicians should be aware of the possibility of symptomaticCoxiella burnetii infection among HIV-infected people, particularly in endemic regions for both infections such as in sub-saharan Africa.     

Reactivation of Q fever following cardiac surgery

To determine whether reactivation of Q fever occurs following cardiac surgery 43 patients who had antibodies toCoxiella burnetii were identified pre-operatively. Serum samples were collected 1, 4 to 6, and 24 weeks post-operatively. One patient had active Q fever endocarditis. Seven of the remaining 42 (17 %) had a fourfold rise in titres for antibodies toCoxiella burnetii post-operatively. Only two of these seven had an increase in antibody titres for other agents suggesting that the fourfold titre rise forCoxiella burnetii was not part of a polyclonal response. It is concluded that self-limited reactivation of infection may occur amongCoxiella burnetii seropositive patients undergoing open heart surgery.     

Dysregulation of Serum Gamma Interferon Levels in Vascular Chronic Q Fever Patients Provides Insights into Disease Pathogenesis

A large community outbreak of Q fever occurred in the Netherlands in the period 2007 to 2010. Some of the infected patients developed chronic Q fever, which typically includes pathogen dissemination to predisposed cardiovascular sites, with potentially fatal consequences. To identify the immune mechanisms responsible for ineffective clearance of Coxiella burnetii in patients who developed chronic Q fever, we compared serum concentrations of 47 inflammation-associated markers among patients with acute Q fever, vascular chronic Q fever, and past resolved Q fever. Serum levels of gamma interferon were strongly increased in acute but not in vascular chronic Q fever patients, compared to past resolved Q fever patients. Interleukin-18 levels showed a comparable increase in acute as well as vascular chronic Q fever patients. Additionally, vascular chronic Q fever patients had lower serum levels of gamma interferon-inducible protein 10 (IP-10) and transforming growth factor β (TGF-β) than did acute Q fever patients. Serum responses for these and other markers indicate that type I immune responses to C. burnetii are affected in chronic Q fever patients. This may be attributed to an affected immune system in cardiovascular patients, which enables local C. burnetii replication at affected cardiovascular sites.     

Emergence of Q Fever Arthritis in France

Osteoarticular infection is an uncommon presentation of Q fever. Positron emission tomography (PET) scanning is a valuable tool for the diagnosis of Coxiella burnetii graft prosthesis infection and endocarditis. Our objective was to test a series of culture-negative osteoarticular samples using molecular assays for Coxiella burnetii. We tested for C. burnetii by molecular assays targeting the IS1111 and the IS30A spacer regions, using culture-negative osteoarticular samples obtained in our laboratory between January 2011and December 2012. We examine a total of 1,410 osteoarticular samples, and we observed two cases of arthritis and subacromial bursitis caused by C. burnetii. The infections were localized using PET scanning, and the diagnosis was confirmed through serology. For one, a C. burnetii strain with a multispacer sequence type 8 genotype was isolated from synovial fluid culture. Q fever articular infections could be undiagnosed because of the long evolution of articular attack, and patients with high antibody titers against C. burnetii should be tested using PET scanning to localize the site of infection.     

Comparison of ELISA and indirect immunofluorescent antibody assay detecting <Emphasis Type="Italic">Coxiella burnetii</Emphasis> IgM phase II for the diagnosis of acute Q fever

A commercially available enzyme-linked immunosorbent assay (ELISA) detecting Coxiella burnetii phase II-specific IgM for the diagnosis of acute Q fever was compared with indirect immunofluorescent antibody assay (IFA). IFA is the current reference method for the detection of antibodies against C. burnetii, but has disadvantages because the judgment of fluorescence is subjective and tiring, and the test is expensive and automation is not possible. To examine whether phase II IgM ELISA could be used as a screening assay for acute Q fever, we compared the sensitivity and specificity of IFA and ELISA. The sensitivity of the IFA and ELISA tests were 100 and 85.7%, respectively, with a specificity of 95.3 and 97.6%, respectively. Because of the high sensitivity and specificity of the ELISA in combination with the practical disadvantages of the IFA, we introduced a new algorithm to screen samples of patients with symptoms of acute Q fever infection.     

Validation of an enzyme immunoassay for serodiagnosis of acute Q fever

An enzyme immunoassay was validated for the serodiagnosis of acute Q fever. Minimum positive tests were determined for both serial dilutions and a single dilution of patient sera. To establish the specificity of the test, 152 serum samples were tested from individuals with no evidence of pastCoxiella burnetii infection. Diagnostic titers were set at 128 for the IgM and IgG responses to phase I, at 512 for the IgM response to phase II and at 1,024 for the IgG response to phase IICoxiella burnetii. These titers gave a falsepositive rate of 1 %. Alternatively, testing a single dilution of sera (1:128) gave specificities ranging from 97.3 to 98.7 %. Tests with the greatest sensitivities, using serially diluted early convalescent-phase sera, were the IgM (84 %) and IgG (80 %) responses to phase IICoxiella burnetii. At a single serum dilution, 92 % of early convalescent sera had a positive IgG response to phase IICoxiella burnetii. With a high specificity and good sensitivity, the EIA can be used to diagnose acute Q fever with a single convalescent serum specimen. The duration of a positive response was greater than five years.     

Conventional viral cultures and shell vial assay for diagnosis of apparently culture-negativeCoxiella burnetii endocarditis

A patient with culture-negative endocarditis was diagnosed with Q fever endocarditis based on the results of serological tests and positive leukocyte cultures obtained using conventional viral cultures and the shell vial technique. This case report suggests that isolation ofCoxiella burnetii from blood may allow better diagnostic and therapeutical evaluation of patients with Q fever endocarditis. The use of both conventional and shell vial viral cultures is recommended for the isolation ofCoxiella burnetii from the blood of patients with apparently culture-negative endocarditis.     

Q Fever in the Greek Island of Crete: Detection, Isolation, and Molecular Identification of Eight Strains of Coxiella burnetii from Clinical Samples

Over a period of 6 years (1989 to 1995), serum samples from 3,300 patients suspected to be infected by Coxiella burnetii were assayed for the presence of antibodies against antigen phase II of the microorganism by the indirect immunofluorescence antibody technique (IFAT). One hundred fifty-two cases were recorded, and blood samples from 17 patients were cultured for the isolation of the pathogen. By a centrifugation shell vial technique, eight strains were isolated from patients suffering from acute Q fever. The microorganism was detected in the cultures by IFAT, by Gimenez staining, and by the cytopathogenic effect on Vero and human embryonic lung (HEL) cells. PCR followed by restriction fragment length polymorphism analysis was used to confirm the diagnosis and identify the Coxiella burnetii strains within the cell cultures as well as to compare them with reference strains. In order to avoid time-consuming cultures, to achieve direct detection of Coxiella burnetii in clinical samples (blood, buffy coat, etc.), and to increase the specificity and sensitivity of the detection, nested PCR was performed. The first step of DNA extraction was performed with the QIAamp blood kit 250. For the second step of the PCR assays, the conditions of temperature and times of recycling were properly modified, and the microorganism was detected within 4 h. Our study demonstrates that Q fever is an endemic disease in Crete and that the diagnosis of Coxiella burnetii infection can be rapidly achieved by the detection of the microorganism in buffy coat samples by nested PCR. Although the presenting symptoms of the disease in this study differed from those in other studies, the Cretan strains do not differ genotypically from the reference strains (Nine Mile and Q212).     

Q fever endocarditis: diagnostic approaches and monitoring of therapeutic effects

Summary The scope of current diagnostic methods for Q fever endocarditis includes serology, direct demonstration of Coxiella burnetii in the resected heart valve tissue, and animal inoculation studies. Illustrated by a clinical case report, the different methods are presented and discussed. Serology represents the primary method, using the techniques of complement fixation, indirect immunofluorescence, and enzyme-linked immunosorbent assay (ELISA). The latter two techniques allow the detection of immunoglobulins G, M, and A to the phase I and II antigens of C. burnetii. After cardiac surgery, we visualized C. burnetii on smears and specifically stained it on histologic sections of the resected heart valve by light and electron microscopic immunohistochemistry. In addition, seroconversion in animals after inoculation with valve specimens confirmed the presence of C. burnetii in the heart valve. The antibody titers determined by ELISA correlated well with the patient's clinical course during the treatment period. Therefore it is suggested that its usefulness for monitoring the efficacy of antimicrobial agents in patients with Q fever endocarditis should be further evaluated.Abbreviations IFA indirect fluorescent antibody test - ELISA enzyme-linked immunosorbent assay - CF complement fixation - TMP/SMX trimethoprim/sulfamethoxazole - CDC Center for Disease Control - PAP peroxidase-antiperoxidase - PBS phosphate-buffered saline     

Comparison of immunofluorescence with enzyme immunoassay for detection of Q fever

To evaluate enzyme immunoassay (EIA) as an alternative to indirect immunofluorescence assay (IFA) to screen for Q fever in humans, 157 serum samples from patients suspected of having the disease were tested for immunoglobulin G antibodies toCoxiella burnetii. The agreement between the tests and the sensitivity of EIA were excellent (96.8% and 98.4%, respectively) when an IFAtiter of > 1/160 was considered positive. All serum samples with a titer of > 1/320 in the IFA were also positive by the EIA. The EIA seems to be an acceptable alternative to IFA for screening for Q fever.     

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.     

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.     

Chronic Q Fever in the Netherlands 5 Years after the Start of the Q Fever Epidemic: Results from the Dutch Chronic Q Fever Database

Coxiella burnetii causes Q fever, a zoonosis, which has acute and chronic manifestations. From 2007 to 2010, the Netherlands experienced a large Q fever outbreak, which has offered a unique opportunity to analyze chronic Q fever cases. In an observational cohort study, baseline characteristics and clinical characteristics, as well as mortality, of patients with proven, probable, or possible chronic Q fever in the Netherlands, were analyzed. In total, 284 chronic Q fever patients were identified, of which 151 (53.7%) had proven, 64 (22.5%) probable, and 69 (24.3%) possible chronic Q fever. Among proven and probable chronic Q fever patients, vascular infection focus (56.7%) was more prevalent than endocarditis (34.9%). An acute Q fever episode was recalled by 27.0% of the patients. The all-cause mortality rate was 19.1%, while the chronic Q fever-related mortality rate was 13.0%, with mortality rates of 9.3% among endocarditis patients and 18% among patients with a vascular focus of infection. Increasing age (P = 0.004 and 0.010), proven chronic Q fever (P = 0.020 and 0.002), vascular chronic Q fever (P = 0.024 and 0.005), acute presentation with chronic Q fever (P = 0.002 and P < 0.001), and surgical treatment of chronic Q fever (P = 0.025 and P < 0.001) were significantly associated with all-cause mortality and chronic Q fever-related mortality, respectively.     

Correlations between Peripheral Blood Coxiella burnetii DNA Load,Interleukin-6 Levels,and C-Reactive Protein Levels in Patients with Acute Q Fever

From 2007 to 2010, the Netherlands experienced the largest reported Q fever outbreak, with >4,000 notified cases. We showed previously that C-reactive protein is the only traditional infection marker reflecting disease activity in acute Q fever. Interleukin-6 is the principal inducer of C-reactive protein. We questioned whether increased C-reactive protein levels in acute Q fever patients coincide with increased interleukin-6 levels and if these levels correlate with the Coxiella burnetii DNA load in serum. In addition, we studied their correlation with disease severity, expressed by hospital admission and the development of fatigue. Interleukin-6 and C-reactive protein levels were analyzed in sera from 102 patients diagnosed with seronegative PCR-positive acute Q fever. Significant but weak negative correlations were observed between bacterial DNA loads expressed as cycle threshold values and interleukin-6 and C-reactive protein levels, while a significant moderate-strong positive correlation was present between interleukin-6 and C-reactive protein levels. Furthermore, significantly higher interleukin-6 and C-reactive protein levels were observed in hospitalized acute Q fever patients in comparison to those in nonhospitalized patients, while bacterial DNA loads were the same in the two groups. No marker was prognostic for the development of fatigue. In conclusion, the correlation between interleukin-6 and C-reactive protein levels in acute Q fever patients points to an immune activation pathway in which interleukin-6 induces the production of C-reactive protein. Significant differences in interleukin-6 and C-reactive protein levels between hospitalized and nonhospitalized patients despite identical bacterial DNA loads suggest an important role for host factors in disease presentation. Higher interleukin-6 and C-reactive protein levels seem predictive of more severe disease.     

High Coxiella burnetii DNA Load in Serum during Acute Q Fever Is Associated with Progression to a Serologic Profile Indicative of Chronic Q Fever

PCR is very effective in diagnosing acute Q fever in the early stages of infection, when bacterial DNA is present in the bloodstream but antibodies have not yet developed. The objective of this study was to further analyze the diagnostic value of semiquantitative real-time PCR (qPCR) in diagnosing acute Q fever in an outbreak situation. At the Jeroen Bosch Hospital, in 2009, qPCR testing for Coxiella burnetii DNA was performed for 2,715 patients suspected of having acute Q fever (positive, n = 385; negative, n = 2,330). The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the qPCR assay were calculated for patients with negative qPCR results with a follow-up sample obtained within 14 days (n = 305) and qPCR-positive patients with at least one follow-up sample (n = 369). The correctness of the qPCR result was based on immunofluorescence assay results for samples submitted for qPCR and follow-up testing. The sensitivity of the Q fever qPCR assay was 92.2%, specificity 98.9%, PPV 99.2%, and NPV 89.8%. Patients who later developed serologic profiles indicative of chronic Q fever infection had significantly higher C. burnetii DNA loads during the acute phase than did patients who did not (P < 0.001). qPCR testing is a valuable tool for the diagnosis of acute Q fever and should be used in outbreak situations when the onset of symptoms is <15 days earlier. Special attention is needed in the follow-up monitoring of patients with high C. burnetii DNA loads during the acute phase, as this might be an indicator for the development of a serologic profile indicative of chronic infection.     

Lack of pathotype specific gene in human Coxiella burnetii isolates

Human Q fever, due to the obligate intracellular bacterium Coxiella burnetii may be acute or chronic. The acute form is rarely fatal, although the chronic form, mostly represented by chronic Q fever endocarditis, is severe and usually fatal in lack of appropriate treatment. A correlation between the human disease state and plasmid types has been described, and specific DNA sequences unique to each plasmid type and which would code for specific pathotypes have been characterized. Because this gene specificity of the pathogenesis was hypothesized only on a small number of isolates, we evaluated seven reference isolates and 30 recent C. burnetii isolates from France for which all clinical data were available using the polymerase chain reaction by employing two specific primer sets. Our studies indicate that the previously described CbhE′ plasmid-gene is not unique to C. burnetii isolates associated with acute disease, which would mean that the hypothesis of the correlation between gene specificity and pathotype has to be revised.     

Q fever osteoarticular infection: four new cases and a review of the literature

Q fever is a worldwide-occurring zoonosis caused by Coxiella burnetii. Better knowledge of the disease and of evolving diagnostics can enable recognition of unusual manifestations. Reported here are four cases of Q fever osteoarticular infections in adults: two cases of Q fever tenosynovitis, which represent the first two reports of this infection, and two cases of Q fever spondylodiscitis complicated by paravertebral abscess. In addition, the literature is reviewed on the 15 previously reported cases of Q fever osteoarticular infection, six of which were vertebral infections. Osteomyelitis is the usual manifestation Q fever osteoarticular infection. Because its onset is frequently insidious, diagnosis is usually delayed. The main differential diagnosis is mycobacterial infection, based on the histological granulomatous presentation of lesions. Whereas serology is the reference diagnostic method for Q fever, detection of C. burnetii in tissue specimens by PCR and cell culture provides useful additional evidence of infection. Culture-negative osteoarticular samples with granulomatous presentation upon histological examination should raise suspicion of Q fever.