case - patients and controls were recruited from jeroen bosch hospital in s - hertogenbosch and bernhoven hospital in oss and veghel ; both are regional hospitals located in the center of the q fever epidemic area in the netherlands .
the study design was approved by the medical research ethics committee of the university medical centre utrecht .
we used existing datasets and spontaneous notifications from the 2 hospitals to identify all chronic q fever diagnoses among patients > 18 years of age during january 1 , 2007may 1 , 2011 . in the past ,
chronic q fever is considered proven if c. burnetii is detected by pcr in blood or tissue in the absence of acute infection , but sensitivity of this technique is only 50% ( 15,16 ) . persisting high levels of igg to phase i antigens ( phase i igg ) and , to a lesser extent , phase ii antigens ( phase ii igg ) are also indicative of chronic q fever ( 1 ) .
the optimal immunofluorescence assay ( ifa ) cutoff value for phase i igg titer is still matter of debate and is dependent on the test used but is probably within the range of 8001,600 ( 7,1719 ) .
recently , the dutch q fever consensus group proposed a new diagnostic approach that combines pcr , serologic testing , and clinical data and categorizes cases into proven , probable , or possible chronic q fever ( 20 ) .
proven cases are those among patients with positive pcr results for c. burnetii in blood or tissue or a phase i igg titer of > 1,024 in combination with a vascular infection proven by positron emission tomography ( pet ) , computed tomography ( ct ) , magnetic resonance imaging ( mri ) , or endocardial involvement according to the major criteria of the modified duke criteria on echocardiogram ( 21 ) .
probable cases are those among patients with phase i igg titers of > 1,024 and known risk factors : nonmajor valvulopathy according to the modified duke criteria ( 21 ) , suspected nonvascular or noncardial localization of chronic q fever infection , or aspecific signs of chronic infection .
possible cases are those among patients with phase i igg titers of > 1,024 without other risk factors as listed for probable or proven chronic q fever . in contrast to the other 2 subgroups , in general
, possible chronic q fever patients do not receive long - term antimicrobial drug treatment but instead enter a follow - up program ; many demonstrate spontaneous decline in phase i igg titers .
we defined cases according to these definitions ( 20 ) ( table 1 ) . *
ifa , immunofluorescence assay ; pet , positron emission tomography ; ct , computed tomography ; mri , magnetic resonance imaging .
controls were selected from an existing cohort of patients with acute q fever , seen by general practitioners in 2009 , who had positive pcr results for c. burnetii in serum samples .
controls were included if they were > 18 years of age at the time of acute q fever and if the serologic profile was not suggestive of chronic q fever during > 1 year of follow - up ( i.e. , decreasing antibody titers and phase i igg titer < 1,024 ) .
patients with serologic follow - up of < 1 year after the episode of acute q fever were excluded from analysis .
all case - patients except 1 and all controls lived in the same postal code area ( 50005400 ) in the netherlands .
microbiological diagnostics for chronic q fever case - patients consisted of ifa ( focus diagnostics , inc . , cypress , ca , usa ) of serum samples and pcr for c. burnetii dna in serum , plasma , and tissue samples .
the diagnostic workup to evaluate c. burnetii infection in control patients with documented acute q fever had been performed according to a diagnostic algorithm for acute q fever introduced in may 2009 . in brief
, serum samples were screened with elisa for igm against c. burnetii phase ii antigens ( mii - screen ; institut virion serion gmbh , wrzburg , germany ) .
depending on date of onset of disease and inpatient or outpatient setting , pcr for c. burnetii dna was performed if the mii - screen result was negative ( 2325 ) . in patients with confirmed acute q fever , serologic follow - up was performed at 3 , 6 , and 12 months , consisting of ifa for igm and igg against c. burnetii phase i and phase ii antigens .
we collected patient characteristics including demographic variables , medical history , medication , pathology and microbiology results , imaging records , therapy , and outcome for case - patients and controls .
case - patient information was already available in the hospital registration systems and was interpreted by 2 researchers ( l.k . and s.d . ) .
all controls were sent a questionnaire and an informed consent form that asked for permission to request patient s data from the general practitioner and from the hospital registration system .
although debatable , routine echocardiographic screening after diagnosis of acute q fever is not the standard of care in the netherlands because no benefit was found in an earlier evaluation ( 26,27 ) .
therefore , for chronic q fever case - patients and acute q fever controls , details about cardiac valvulopathy were retrieved by review of medical records .
spss version 18.0 was used for storage and analysis of the collected data ( spss inc . ,
, we conducted 3 analyses : 1 ) an overall analysis of all chronic q fever cases ( i.e. , proven , probable , and possible ) ; 2 ) an analysis of proven and probable chronic q fever cases ; and 3 ) an analysis of proven chronic q fever cases only .
we performed these analyses to determine whether exclusion of possible chronic q fever and , to a lesser extent , probable chronic q fever ( the groups in which disease status is doubtful ) influenced the overall results .
univariate and subsequent multivariate logistic regression analyses were performed to calculate odds ratios ( ors ) , corresponding 95% cis , and p values for the development of chronic q fever . in univariate analysis ,
variables with no observations among case - patients and < 2 observations in the control group ( or vice versa ) were excluded ( i.e. , hematologic malignancies , bone marrow transplantation , dialysis , renal transplant , nonrenal organ transplant , congenital cardiac deviation , pulmonary diseases , and autoimmune disorder ) . for potential dichotomous risk factors ,
i.e. , those that had 0 observations among either the case - patients or controls but > 2 observations in the other , we applied a fisher exact test to calculate p values .
variables with > 1 observations and < 25% missing values in case - patients and controls , a p value of < 0.10 in univariate analysis , or known association in previous reports with the development of chronic q fever were subsequently analyzed in a multivariate model . the variables vascular history and valvulopathy were not included in multivariate analysis because they were included in variables that were listed separately ( i.e. , vascular prosthesis , aneurysm , other vascular surgery , peripheral arterial disease , cerebrovascular disease , valvular surgery , and nonsurgical valvular disease ) . eighteen case - patients and 0 controls had a history of valvular surgery .
because of the expected importance of this risk factor and the high incidence among case - patients , we considered its inclusion in the multivariate analysis critical ; moreover , the logistic regression model could not be fitted with this variable excluded .
therefore , we randomly changed one of the observations of the control group from 0 to 1 , which artificially reduced the association but enabled us to fit the regression model . the variables age , vascular history , vascular prosthesis , aneurysm , other vascular surgery , cerebrovascular disease , peripheral vascular disease , valvulopathy , valvular surgery , valvular deviation , ischemic heart disease , other cardiovascular diseases , hypertension , dyslipidemia , diabetes , nonhematologic malignancy , and renal insufficiency could be included in multivariate analysis of all groups . the variable immune disorder was also included in multivariate analysis for the probable and proven and the proven subgroups .
differences between case - patients and controls were shown in use of statins , clopidogrel , acenocoumarol , and proton pump inhibitors and hospitalization and adequate treatment during acute q fever ( p<0.10 ) .
however , these variables could not be included in the multivariate analysis because > 25% of values were missing , most among case - patients in whom an acute q fever episode had gone unrecognized . after selecting predictors for our final multivariate model
interactions were not significant and therefore not included in the model . to assess the goodness - of - fit of the final model , we plotted sensitivity and specificity by using a receiver operating characteristic curve and estimated the area under the curve ( c - statistic ) .
case - patients and controls were recruited from jeroen bosch hospital in s - hertogenbosch and bernhoven hospital in oss and veghel ; both are regional hospitals located in the center of the q fever epidemic area in the netherlands .
the study design was approved by the medical research ethics committee of the university medical centre utrecht .
we used existing datasets and spontaneous notifications from the 2 hospitals to identify all chronic q fever diagnoses among patients > 18 years of age during january 1 , 2007may 1 , 2011 . in the past ,
chronic q fever is considered proven if c. burnetii is detected by pcr in blood or tissue in the absence of acute infection , but sensitivity of this technique is only 50% ( 15,16 ) . persisting high levels of igg to phase i antigens ( phase i igg ) and , to a lesser extent , phase ii antigens ( phase ii igg ) are also indicative of chronic q fever ( 1 ) .
the optimal immunofluorescence assay ( ifa ) cutoff value for phase i igg titer is still matter of debate and is dependent on the test used but is probably within the range of 8001,600 ( 7,1719 ) .
recently , the dutch q fever consensus group proposed a new diagnostic approach that combines pcr , serologic testing , and clinical data and categorizes cases into proven , probable , or possible chronic q fever ( 20 ) . proven cases are those among patients with positive pcr results for c. burnetii in blood or tissue or a phase i igg titer of > 1,024 in combination with a vascular infection proven by positron emission tomography ( pet ) , computed tomography ( ct ) , magnetic resonance imaging ( mri ) , or endocardial involvement according to the major criteria of the modified duke criteria on echocardiogram ( 21 ) .
probable cases are those among patients with phase i igg titers of > 1,024 and known risk factors : nonmajor valvulopathy according to the modified duke criteria ( 21 ) , suspected nonvascular or noncardial localization of chronic q fever infection , or aspecific signs of chronic infection .
possible cases are those among patients with phase i igg titers of > 1,024 without other risk factors as listed for probable or proven chronic q fever .
in contrast to the other 2 subgroups , in general , possible chronic q fever patients do not receive long - term antimicrobial drug treatment but instead enter a follow - up program ; many demonstrate spontaneous decline in phase i igg titers .
we defined cases according to these definitions ( 20 ) ( table 1 ) . * described in ( 20 ) .
ifa , immunofluorescence assay ; pet , positron emission tomography ; ct , computed tomography ; mri , magnetic resonance imaging .
controls were selected from an existing cohort of patients with acute q fever , seen by general practitioners in 2009 , who had positive pcr results for c. burnetii in serum samples .
controls were included if they were > 18 years of age at the time of acute q fever and if the serologic profile was not suggestive of chronic q fever during > 1 year of follow - up ( i.e. , decreasing antibody titers and phase i igg titer < 1,024 ) .
patients with serologic follow - up of < 1 year after the episode of acute q fever were excluded from analysis .
all case - patients except 1 and all controls lived in the same postal code area ( 50005400 ) in the netherlands .
microbiological diagnostics for chronic q fever case - patients consisted of ifa ( focus diagnostics , inc . , cypress , ca , usa ) of serum samples and pcr for c. burnetii dna in serum , plasma , and tissue samples .
the diagnostic workup to evaluate c. burnetii infection in control patients with documented acute q fever had been performed according to a diagnostic algorithm for acute q fever introduced in may 2009 . in brief
, serum samples were screened with elisa for igm against c. burnetii phase ii antigens ( mii - screen ; institut virion serion gmbh , wrzburg , germany ) .
depending on date of onset of disease and inpatient or outpatient setting , pcr for c. burnetii dna was performed if the mii - screen result was negative ( 2325 ) . in patients with confirmed acute q fever , serologic follow - up was performed at 3 , 6 , and 12 months , consisting of ifa for igm and igg against c. burnetii phase i and phase ii antigens .
we collected patient characteristics including demographic variables , medical history , medication , pathology and microbiology results , imaging records , therapy , and outcome for case - patients and controls .
case - patient information was already available in the hospital registration systems and was interpreted by 2 researchers ( l.k . and s.d . ) .
all controls were sent a questionnaire and an informed consent form that asked for permission to request patient s data from the general practitioner and from the hospital registration system .
although debatable , routine echocardiographic screening after diagnosis of acute q fever is not the standard of care in the netherlands because no benefit was found in an earlier evaluation ( 26,27 ) .
therefore , for chronic q fever case - patients and acute q fever controls , details about cardiac valvulopathy were retrieved by review of medical records .
spss version 18.0 was used for storage and analysis of the collected data ( spss inc . ,
within this study , we conducted 3 analyses : 1 ) an overall analysis of all chronic q fever cases ( i.e. , proven , probable , and possible ) ; 2 ) an analysis of proven and probable chronic q fever cases ; and 3 ) an analysis of proven chronic q fever cases only .
we performed these analyses to determine whether exclusion of possible chronic q fever and , to a lesser extent , probable chronic q fever ( the groups in which disease status is doubtful ) influenced the overall results .
univariate and subsequent multivariate logistic regression analyses were performed to calculate odds ratios ( ors ) , corresponding 95% cis , and p values for the development of chronic q fever . in univariate analysis ,
variables with no observations among case - patients and < 2 observations in the control group ( or vice versa ) were excluded ( i.e. , hematologic malignancies , bone marrow transplantation , dialysis , renal transplant , nonrenal organ transplant , congenital cardiac deviation , pulmonary diseases , and autoimmune disorder ) . for potential dichotomous risk factors ,
i.e. , those that had 0 observations among either the case - patients or controls but > 2 observations in the other , we applied a fisher exact test to calculate p values .
variables with > 1 observations and < 25% missing values in case - patients and controls , a p value of < 0.10 in univariate analysis , or known association in previous reports with the development of chronic q fever were subsequently analyzed in a multivariate model .
the variables vascular history and valvulopathy were not included in multivariate analysis because they were included in variables that were listed separately ( i.e. , vascular prosthesis , aneurysm , other vascular surgery , peripheral arterial disease , cerebrovascular disease , valvular surgery , and nonsurgical valvular disease ) .
because of the expected importance of this risk factor and the high incidence among case - patients , we considered its inclusion in the multivariate analysis critical ; moreover , the logistic regression model could not be fitted with this variable excluded .
therefore , we randomly changed one of the observations of the control group from 0 to 1 , which artificially reduced the association but enabled us to fit the regression model .
the variables age , vascular history , vascular prosthesis , aneurysm , other vascular surgery , cerebrovascular disease , peripheral vascular disease , valvulopathy , valvular surgery , valvular deviation , ischemic heart disease , other cardiovascular diseases , hypertension , dyslipidemia , diabetes , nonhematologic malignancy , and renal insufficiency could be included in multivariate analysis of all groups . the variable immune disorder was also included in multivariate analysis for the probable and proven and the proven subgroups .
the variable pacemaker was also included in multivariate analysis for the proven group . differences between case - patients and controls were shown in use of statins , clopidogrel , acenocoumarol , and proton pump inhibitors and hospitalization and adequate treatment during acute q fever ( p<0.10 ) .
however , these variables could not be included in the multivariate analysis because > 25% of values were missing , most among case - patients in whom an acute q fever episode had gone unrecognized . after selecting predictors for our final multivariate model
interactions were not significant and therefore not included in the model . to assess the goodness - of - fit of the final model , we plotted sensitivity and specificity by using a receiver operating characteristic curve and estimated the area under the curve ( c - statistic ) .
we identified 105 case - patients with proven , probable , or possible chronic q fever ; 44 ( 42% ) had proven , 28 ( 27% ) probable , and 33 ( 31% ) possible disease .
of the case - patients with proven chronic q fever , 27 ( 61% ) had positive pcr results for c. burnetii in blood only , 5 ( 11% ) in tissue only , 8 ( 18% ) in tissue and blood , and 4 ( 9% ) in neither blood nor tissue .
the focus of infection in cases of proven chronic q fever was endocarditis for 12 case - patients ( 27% ) and endovascular infection for 26 ( 59% ) ; 6 ( 14% ) had no clear infection focus .
of the case - patients with probable chronic q fever , suspected foci were cardiac valves in 12 ( 43% ) , endovascular lesions in 1 ( 4% ) , and another focus ( e.g. , pregnancy or clinical symptoms of infection such as weight loss , night sweats , and fever ) in 15 ( 54% ) .
long - term antimicrobial drug treatment was started for 40/44 case - patients ( 91% ) with proven chronic q fever , 18/28 case - patients ( 64% ) with probable chronic q fever , and 5/32 case - patients ( 15% ) with possible chronic q fever .
three patients with proven chronic q fever patients died before diagnosis of chronic q fever ; 1 refused therapy . in all , 289 controls who had pcr - proven acute q fever in 2009 were sent a questionnaire .
of these , 201 ( 69.6% ) responded , signed the informed consent form , and fulfilled the inclusion criteria ( figure 1 ) .
enrollment , selection , and inclusion criteria forcase - patients and controls for case  control study to identify risk factors for chronic q fever , the netherlands . results of the univariate analysis are listed in table 2 .
comparisons for age , vascular history , vascular prosthesis , aneurysm , other vascular surgery , cerebrovascular disease , peripheral vascular disease , valvulopathy , valvular surgery , valvular deviation , ischemic heart disease , other cardiovascular diseases , hypertension , dyslipidemia , diabetes , nonhematologic malignancy ( defined as several kinds of solid tumors ) , renal insufficiency , and pregnancy showed significant differences between case - patients and controls . *
patients wth information available for that category . or , odds ratio ; ns , nonsurgical ; na , not applicable ; copd , chronic obstructive pulmonary disease . 
 case - patients : aortic valve defects , 10 ( no bicuspid valves ) ; mitral valve defects , 9 ( no prolapse ) ; tricuspid valve defects , 4 .
controls : aortic valve defects , 6 ( no bicuspid valves ) ; mitral valve defects , 3 ( 1 prolapse ) . 
* atrial fibrillation , congestive heart failure , pericarditis , bradycardia , ischemic cardiomyopathy , and left ventricular hypertrophy . 
prednisone cumulative dose > 750 mg ; use of tumor necrosis factor blocker , methotrexate , mycofenolate mofetil ; splenectomy .  
# # defined as 1014 d of doxycycline treatment results of the multivariate analyses are shown in table 3 .
valvular surgery ( or 31.5 , 95% ci 3.99249 ) , vascular prosthesis ( or 10.4 , 95% ci 2.1750.0 ) , aneurysm ( or 8.65 , 95% ci 1.7442.9 ) , nonhematologic malignancy ( or 3.90 , 95% ci 1.3311.5 ) , and age ( or 1.03 , 95% ci 1.011.06 ) were independently associated with the development of chronic q fever .
the final discriminative performance was good , with a c - statistic of 0.71 ( 95% ci 0.710.83 ) ( figure 2 ) . * or ,
possible risk factors entered in all analyses : age , vascular prosthesis , aortic aneurysm , other vascular surgeries , peripheral arterial disease , cerebrovascular disease , valvular surgery , valvular disease ( nonsurgical ) , ischemic cardiac disease , other cardiac history , hypertension , dyslipidemia , diabetes , nonhematologic malignancy , renal insufficiency .
immune disorder was also entered in the analyses of proven and probable chronic q fever and of proven chronic q fever .
valvular surgeries in the proven group are subdivided into biological valve ( n = 6 ) , prosthetic valve ( n
= 3 ) , and valve repair ( n = 1 ) all located in the aortic valve ( n = 10 ) . within the controls there were no patients with history of valvular surgery . 
 locations of vascular prostheses in proven group were infrarenal and iliac ( n = 6 ) , infrarenal ( n = 4 ) , thoracic ( n = 2 ) , and unknown ( n = 2 ) . types of vascular prosthesis were y - prosthesis ( n = 7 ) , endovascular aneurysm repair ( n = 2 ) , stent graft ( n = 2 ) , bentall ( n = 1 ) , and unknown ( n = 2 ) .
locations of aneurysms in proven group were infrarenal ( n = 6 ) , infrarenal and iliac ( n = 2 ) , and suprarenal , infrarenal , and iliac ( n = 1 ) . within the control group ,
# observed stages of chronic kidney disease according to the kidney disease outcome quality initiative guidelines ( 28 ) in the proven group were stage 3 ( n = 6 ) , stage 4 ( n = 2 ) , and stage 5 ( n = 1 ) and in the controls solely stage 3 ( n = 2 ) . 
goodness - of - fit models for case  control study to identify risk factors for chronic q fever , the netherlands .
a ) all chronic q fever cases ( n = 105 ) ; area under the curve ( c - statistic ) 0.77 ( 95% ci 0.710.83 ) ; p<0.001 .
b ) proven and probable chronic q fever cases ( n = 72 ) ; c - statistic 0.86 ( 95% ci 0.810.92 ) ; p<0.001 .
c ) proven chronic q fever cases ( n = 44 ) ; c - statistic 0.91 ( 95% ci 0.850.97 ) ; p<0.001 .
patient risk factors included in the model ( no . observations ) : a ) valvular surgery ( 18 ) ; vascular prosthesis ( 15 ) ; aneurysm ( 12 ) ; nonhematologic malignancy ( 16 ) ; age , continuous , mean 63.9 y ; b ) valvular surgery ( 18 ) ; vascular prosthesis ( 15 ) ; aneurysm ( 12 ) ; renal insufficiency ( 12 ) ; age , continuous , mean 67.3 y ; c ) valvular surgery ( 10 ) ; vascular prosthesis ( 14 ) ; aneurysm ( 9 ) ; renal insufficiency ( 9 ) ; age , continuous , mean 68.4 .
patient risk factors identified in the analysis of the proven cases , representing the most definite chronic q fever cases , were valvular surgery ( or 43.6 , 95% ci 4.70405 ) , vascular prosthesis ( or 26.8 , 95% ci 4.88147 ) , aneurysm ( or 25.9 , 95% ci 4.55147 ) , renal insufficiency ( or 16.0 , 95% ci 2.06123 ) , and age ( or 1.06 , 95% ci 1.021.11 ) .
the final discriminative performance was good , with a c - statistic of 0.91 ( 95% ci 0.850.97 ) ( figure 2 ) .
to our knowledge , this is the first study that analyzed a large number of potential risk factors for chronic q fever in a large number of patients .
most former studies have been limited by a low number of cases and evaluation of few risk factors . moreover
, quantification of these risk factors was lacking ( 69,11,12 ) . in our study , we focused mainly on case - patients with proven chronic q fever becausethis group included patients with the most definite form of chronic q fever
. proven chronic q fever also showed the strongest correlation with the identified risk factors . in multivariate analysis ,
valvular surgery , vascular prosthesis , aneurysms , renal insufficiency , and age were significant risk factors for the development of chronic q fever in patients with proven cases . in the analysis of all patients with chronic q fever cases , nonhematologic malignancy also seemed to be a risk factor ; however , this could not be reproduced in the subanalyses of the more definite cases ( e.g. , proven and probable cases ) .
valvular surgery , vascular prostheses , and aneurysms were the strongest predictors in this study , which confirms observational findings from earlier studies .
a novel finding is the association between mild renal insufficiency and chronic q fever . the majority of patients with chronic q fever and renal disease in our study had stage 3 renal insufficiency according to kidney disease outcome quality initiative guidelines ( 28 ) .
although terminal renal insufficiency can decrease the immune response , this association was not found for mild renal disease ( 29 ) .
renal insufficiency is associated with vascular disease , which may explain the elevated incidence of chronic q fever in these patients ( 30 ) .
increasing age also predisposes for the development of chronic q fever ; this predisposition was also illustrated in a recent report of van der hoek et al .
the explanation probably lies in the increased prevalence of cardiovascular diseases and the decreased cellular immunity during aging ( 31,32 ) .
age > 60 years appeared the best cutoff above which the risk for chronic q fever increases significantly .
preexisting cardiac valvulopathy has been found to give an estimated risk of 39% for the development of chronic q fever after infection with c. burnetii ( 6,33,34 ) .
in contrast , recent reports showed no elevated risk for patients with mild valvulopathy in the ongoing outbreak in the netherlands ( 26,27 ) .
although our univariate analyses showed that nonsurgical cardiac valvulopathy increased the risk for the development of chronic q fever , this finding was not confirmed in the multivariate analysis .
this finding can be explained by the fact that 9/17 ( 53% ) case - patients with nonsurgical valvulopathy also had a history of valvular surgery of one of the other valves .
the location and type of valvular defects did not differ significantly between case - patients and controls ( table 2 ) .
a possible explanation for the discrepancy with previous observations lies in the fact that our study was conducted 4 years after start of the q fever epidemic , but chronic q fever endocarditis in patients with nonsurgical cardiac valvulopathy might become evident later ( 6,8 ) .
furthermore , strain - specific differences in clinical signs and symptoms might also be of importance ( 26 ) .
presence of valvulopathy in case - patients and controls could have been missed becausethis was assessed only through review of medical records . however ,
echocardiography , which was standard care for all patients with suspected cases of chronic q fever , revealed no additional congenital or bicuspid valve defects , in comparison to assessment of valvulopathy through review of medical records . from other than the above - mentioned defects
, it could not be determined by these echocardiograms if these were preexisting or caused by chronic q fever .
immunosuppression , although not well defined , has been indicated as a risk factor in former reports , but clear definition and statistical empowerment is lacking ( 8) . although our univariate analysis did show an elevated risk for immunosuppression , especially for patients with proven chronic q fever cases , this elevated risk was not confirmed in multivariate analysis .
immunocompromised patients may be underrepresented in our study becauseit was conducted in a peripheral hospital setting .
pregnancy , another formerly reported risk factor , showed an association with the development of chronic q fever in univariate analysis .
however , because there were no pregnant women in the control group and only 3 pregnant women in all case groups , evaluation of pregnancy in multivariate analyses was not possible .
a study specifically designed to evaluate associations between pregnancy and q fever is ongoing in the netherlands ( 35 ) . in our opinion
, our data were representative for this large q fever outbreak because they were well documented data and patients were willing to participate .
the fact that case - patients and controls were living in the same area increases the comparability of these groups and strengthens the results .
first , all controls had an acute episode in 2009 , but information about their signs and symptoms was obtained in 2011 , introducing possible recall bias .
we tried to reduce this bias by requesting additional information from the general practitioners and by reviewing clinical test results and physicians reports in the hospital registration systems . because information bias could also have been introduced by the subjective interpretation of physicians reports for case - patients and controls , 2 of our researchers interpreted the results independently .
serologic follow - up of the controls after the acute q fever episode lasted only 1 year , which is the normal follow - up period in the netherlands .
however , because chronic q fever can become manifest years after initial infection , development of chronic q fever after this follow - up period is still possible ( 6,22 ) .
still , 75% of chronic q fever cases develop within 6 months after primary infection ( 22 ) .
moreover , according to the observed decrease in antibody titers of these patients , progression to chronic q fever is not likely .
in addition , as a consequence of the inclusion of patients with symptomatic acute q fever as a control group , the results can only be generalized to patients with symptomatic acute q fever , although the results probably provide an adequate indication of risks factors for patients with mild or asymptomatic primary q fever .
notably , almost all controls received antimicrobial drug treatment at time of acute q fever , in contrast to the case - patients , among which only a minority had symptomatic acute q fever .
thus , antimicrobial drug treatment might influence the chance of chronic q fever development , although there is no quantitative evidence that treatment for acute q fever reduces the chance for chronic q fever ( 4 ) .
chronic q fever cases were selected and classified according to the definitions of the dutch q fever consensus group ( 20 ) , which still need confirmation .
the definition of probable chronic q fever contains several patient criteria that we also included as potential risk factors in our study ( e.g. , valvular disease , vascular prosthesis , aneurysm , and immunosuppressive state ) .
nevertheless , proven chronic q fever , for which these criteria are not part of the definition , was also predicted with the identified risk factors in multivariate analysis , thereby confirming the independent risk association of these variables .
some chronic q fever cases were identified during screening programs for patients who had valvular surgery , aneurysms , or vascular prostheses .
patients with these risk factors may therefore be overrepresented within our study , although all proven case - patients had symptomatic disease .
last , the results of this study have to be considered in view of a predominant c. burnetii strain that is responsible for the majority of q fever cases in humans in the netherlands ( 36 ) . worldwide ,
q fever manifestations differ geographically , which might result from differences in c. burnetii strains ( 4 ) . in conclusion , previous valvular surgery , vascular prosthesis , aneurysms , renal insufficiency , and age were identified as major risk factors for the development of chronic q fever among persons infected with c. burnetii .
because untreated chronic q fever comes with serious risk for illness and death , awareness is required in people with acute q fever possessing the identified risk factors .
this may require close follow - up or even prophylactic treatment in high - risk groups .
moreover , in case of large q fever outbreaks , screening is advisable for patients with these identified risk factors .
 previous cardiac valvular surgery , vascular prosthesis , aortic aneurysm , renal insufficiency , and older age increased risk . 