are frequently present in the 
 water systems of large buildings , and exposure to these bacteria occurs therefore 
 regularly .
nonetheless , legionnaires ' diseases ( lds ) , the most severe form of 
 illness due to legionella spp . , seem 
 to be a rare outcome of exposure . this has been underpinned by outbreak 
 investigations suggesting that only 0.15% of persons exposed to legionella develops ld .
most legionella infections may be subclinical 
 or result in an influenza - like illness ( pontiac 
  fever )
. in particular , subclinical infections may be common among individuals 
 with regular exposure to legionella [ 1 , 2 ] . in an outbreak of ld at a floral show , antibody levels were higher in 
 exposed but asymptomatic exhibitors than in the general population .
health 
 complaints differed by the workplace locations of the exhibitors but were 
 largely independent of antibody levels . although legionella has been 
 detected by culture in up to 70% of water samples from hospitals ' water 
 distribution systems [ 48 ] , and nosocomial ld is a well - known problem , 
 little is known about rates of legionella infections in communities and workplaces .
the aim of the present cross - sectional study was to analyse antibody 
 levels among hospital workers with known exposure to legionella and to determine the correlation between antibodies to legionella and self - reported symptoms 
 compatible with legionella infection . 

furthermore , we examined domestic and other environmental risk factors for 
 seropositivity among the hospital workers .
the study was undertaken at a 643-bed acute - care hospital providing both 
 general and specialised hospital care .
the hospital blocks include both new and 
 old buildings up to a hundred years old .
there have been no cooling towers functioning 
 in the hospital area since 2001 . before 2003
there were 21 separate hot water 
 systems with blind ends in every system . from 1998 to 2003 all hot water tanks 

were removed and replaced by heat exchangers . as part of measures for reducing 
 the risk of legionella infection at the 
 hospital , the temperature of the outgoing hot water
is maintained at least 60c ; whereas the circulating temperature and the temperatures at the most remote points - of - use 
 are at least 50c .  
once a week , the 
 temperature is increased to 6770c in about three hours .
there is no routine monitoring 
 of the temperatures of the water in the pipes or at the points - of - use . in spite 
 of these precautions ,
six nosocomial ld cases from five departments were reported 
 at the hospital between 1999 and 2005 .
the hospital has guidelines for the 
 prevention of ld among susceptible patients , including recommendations to avoid 
 exposure to aerosols and to use sterile water for drinking purposes , and so forth .
water samples from the hospital were analysed for viable legionella at statens serum institut 
 within two days of sampling .
the results were recorded as the highest number of 
 colonies confirmed as legionella ( cfu / litre ) . 
 from each water sample with growth of legionella , one to five colonies were selected and tested by legionella latex test ( oxoid dr0800 , basingstoke , uk ) , 
 by this method the isolates were divided into l. pneumophila serogroup 1 , l. 
 pneumophila serogroup 214 , and legionella spp .
the lowest 
 count of legionella that reliably can 
 be detected by this method is 100 cfu / litre . in the period 1999 to 2005
, 230 waters samples were analysed , and 214 
 ( 93% ) were positive for legionella spp . 
 with counts up to 28 0000 cfu / litre .
all departments included had positive 
 water tests for l. pneumophila , and l. pneumophila sg 1 were found in all 
 departments but one .
the samples ( 74 ) taken in the year of the study , 2005 , 
 showed that all water distribution systems of the selected departments were 
 positive for legionella with counts 
 up to 18 000 cfu / litre .
l. pneumophila sg 1 was present in 14% of the samples , sg 214 in 60% ( l. pneumophila sg 3 in 19% ) , and in 1% of the samples legionella spp .
a 
 month before our study we tested representative showers at the departments and 
 at the staff changing - rooms ; three of four showers at the staff changing - rooms 
 showed low levels of legionella spp .
a total of 675 employees from nine different hospital departments were 
 invited to participate in the study .
the eligible employees had various risks of 
 exposure , including showering patients , performing surgical hand wash , or using 
 the shower of the hospital for personal purpose .
all participants were asked to complete a questionnaire about 
 self - reported health and relevant exposures during the past year .
the questions 
 about health status included a history of ailments such as 
 influenza - like illness , pneumonia , common cold ; health care seeking including 
 hospitalisations and visits to general practitioners ; absences from work due to 
 illness ; specific symptoms ( cough , fever , malaise , stomach pain , shiver , 
 diarrhoea , headache , myalgia , cold ) .
participants were requested to report 
 symptoms only if they had persisted for at least two consecutive days in the 
 previous year .
the questions on occupational exposures included frequency and duration 
 of showering patients , using a shower at the hospital for personal purpose , and 
 frequency of performing surgical hand wash .
combined hospital exposure included 
 any frequency of showering patients , self - showering , or surgical hand wash .
the questions on nonoccupational exposures ( reflecting potential environmental 
 risk factors ) included type of residence ; residence built before 1970 ; district 
 heating ; presence of hot water tank ; hot water tap - time ( considered to be slow 
 if not hot in 1/21 minute ) ; 
 closure of water distribution ; closure of home ; use of spa - bath ; shower 
 elsewhere than home ; swimming pool ; travel abroad ; hotel stay in denmark ; visit 
 to a danish summer cottage ; air - condition in private car .
socioeconomic variables were school - education , job skills , and family 
 income . all questions concerned exposures during the previous year .
blood samples from hospital workers were analysed for antibodies to legionella by indirect immunofluorescence antibody 
 test ( ifat ) with plate grown and heat inactivated l. pneumophila serogroup ( sg ) 1 to 6 , l. micdadei and l. bozemanii as antigens .
the assay is based on the well - characterised assay 
 described by wilkinson et al . , which follows the guidelines from the 
 centers for disease control and prevention ( cdc ) .
the in - house if test has recently been compared with 
 commercial kits for detection of antibodies to l. pneumophila , and it was found that the in house test was at 
 least as specific and sensitive as the commercial kits .the serum samples were titrated from 1 : 64 and upwards to end - point 
 titre .
a titre of    1 : 128 was used to 
 define a positive antibody response 
 to legionella .
national laboratory test criteria for a confirmed diagnosis of legionella infection include a four - fold 
 or greater rise in antibody titre to   1 : 128 in ifat ( seroconversion ) to l. 
 pneumophila sg 1 , 3 , or 6 .  
seroconversion to other legionella antigens 
 and positive titres ( 1 : 256 ) to 
 any legionella antigen are considered indicative of a recent or previous legionella infection . to compare the results obtained for health care workers with the general 
 population , we analysed blood samples collected from 308 and 400 healthy blood 
 donors living in the two towns of randers 
  and vejle , as described previously .
these towns are situated in the 
 neighbour regions of the catchment area of the study hospital . in 2004 ,
the 
 incidences of notified ld in the two towns were 48 and 19 per million 
 inhabitants , respectively . in 2004 ,
the incidence of ld in the town of the 
 study hospital was 17 per million which is within average incidence of ld in denmark .
there was no difference in age , gender , or overall antibody distribution 
 between the towns .
epi data ( ver 3 , odense , 
  denmark ) was 
 used for data entry .
univariable analyses were performed with antibody status 
 as the dependent variable ; variables with p < .2 were added to the model . 

based on this p - value , multiple logistic regression analyses were applied to 
 determine associations with health status and risk factors , respectively , 
 adjusted for age , gender , and current smoking .
variables of significance in the 
 multiple analysis were reported with odds ratio ( or ) and 95% confidence 
 interval ( ci ) .
the prevalence of seropositivity declined by 
 age in a log - linear fashion and therefore age ( in years ) was fitted as a 
 continuous variable .
the study was approved by the regional scientific ethical committee 
 ( vn2005/7 ) and the danish data protection agency .
the antibody titres   1 : 128 for all serogroups were significantly higher 
 in the hospital staff ( 45.1% ) than in the donor population ( 22.9% ) ( or 3.41 , 
 95% ci 2.444.77 ) .
there was no significant difference in antibody levels for l. pneumophila sg 1 ( or 1.43 , 95% ci 
 0.942.16 ) .
one person only from the hospital staff was positive to l. bozemanii , and none was positive to l. micdadei .
the hospital staff had a mean age of 44 years ( range 20 to 67 years ) 
 with a male / female ratio of 36/222 .
there was no 
 difference in mean age and range or smoking between the hospital staff and the 
 healthy blood donor population , although nearly a quarter ( 22.5% ) of the 
 donors was smokers .
male / female ratio was differently , with 56.9% of the donors 
 being males , however , among donors the antibody levels were independent of 
 gender .
in general , there were no marked differences in self - reported morbidity 
 between seropositive and seronegative individuals .
persons with legionella antibodies tended to report 
 more absence from work due to illness or having had a common cold than 
 seronegative persons reported ( table 1 ) .
furthermore , persons who reported symptoms 
 other than influenza - like symptoms had a lower risk of developing antibodies ( or 
 0.34 ; 95% ci 0.120.99 ) , but the numbers were small ( table 1 ) . based on a p - value of   < .2 , multiple regression analysis of seropositives 
 ( titre   1 : 128 ) revealed association with previous pneumonia ( or 0.29 ; 95% ci 
 0.090.94 ) and current smoking ( or 3.54 ; 95% 1.1011.49 ) .
seropositives for sg 
 1 ( titre   1 : 128 ) were not associated with any of the health - related variables 
 in multiple regression analyses .
furthermore , we constructed a symptom complex of influenza - like illness 
 ( cough , fever , malaise , stomach pain , shiver , diarrhoea , headache , myalgia , 
 cold ) .
there were no consistent association between legionella antibodies and a symptom complex of at least three ( or 
 1.95 ; 95% ci 1.003.78 ) , four ( or 1.08 ; 95% ci 0.901.29 ) , or five ( or 0.99 ; 95% 
 ci 0.841.17 ) symptoms of the complex of influenza - like illness with adjustment 
 for age , gender , and current smoking .
finally , there were no associations between symptoms and exposures at 
 the hospital ( data not shown ) .
individuals taking showers in other places than home or having 
 air - conditioning in private car had an increased risk of having antibodies 
 ( table 1 ) .
the multiple regression model ( seropositives with titre   1 : 128 ) of 
 risk variables with p < .2 and gender , age , current smoking showed 
 significant increase in antibodies with showering elsewhere than home ( or 1.89 ; 
 95% ci 1.083.31 ) , air - conditioning in car ( or 1.99 ; 95% ci 1.153.35 ) , and decreasing 
 age ( or 0.97 ; 95% ci ( 0.940.99 ) .
multiple regression analysis of seropositivity to sg 1(titre   1 : 128 ) 
 showed increased antibody levels to sg 1 when having a hot water tank at home ( or 
 4.49 ; 95% ci 1.5313.1 ) and by decreasing age ( or 0.95 ; 95% ci 0.901.00 ) and decreased 
 antibody levels when having had hotel stays in denmark 
  ( or 0.32 ; 95% ci 0.110.95 ) . 

further age analysis in age groups showed a lower prevalence in persons 50 
 years and above ( or 0.79 ; 95% ci 0.650.95 ) compared with individuals below 50 
 years .
antibody levels were independent of hospital department and type of occupational 
 exposure ( data not shown ) .
thus , there were no significant differences in 
 antibody level between staff members working with patients , showering patients , 
 taking personal showers or doing surgical hand wash .
there were no differences 
 according to frequency of the exposure ( almost daily to never ) .
we found a higher prevalence of legionella antibodies ( 1 : 128 ) in the hospital staff with continuous exposure from 
 the water system than in blood donors being representative of the general 
 health population .
antibody levels were not higher in members of the hospital 
 staff at risk of frequent aerosol exposures from showers or surgical hand 
 washing .
we found no association between an influenza - like symptom complex and 
 the presence of antibodies .
the epidemiology of subclinical legionella infections is largely unknown , especially beyond the outbreak setting .
however , 
 outbreak investigations indicate that the antibody response in the healthy 
 population declines with the distance to the source of the outbreak [ 13 , 14 ] .
an outbreak of pontiac 
  fever indicated coherence between 
 attack rate and distance to source too . compared with other studies we 
 found a high prevalence of seropositive individuals suggesting a high exposure and 
 probably ongoing exposure at the hospital .
this finding is consistent with an 
 italian study of a healthy hospital staff which found a high prevalence of 
 antibodies to l. pneumophila sg 114 , but only a prevalence of 3.0% for l. pneumophila sg 16 which are the 
 serogroups ( especially l. pneumophila sg 1 ) most frequently reported causing disease .
the distribution of the 
 levels of legionella antibody shifts 
 to the right ( higher levels ) with increasing exposure in an outbreak situation 
 ; a similar distribution may occur at the hospital setting with a probable 
 higher exposure of the staff compared with the donor population .
we found no specific 
 sources of exposure at the hospital nor subgroups being at higher risk .
this is 
 in contrast to a study from italy 
  where dental personnel had a higher risk of antibodies compared to other 
 hospital workers , possibly due to dental staff being close to aerosols .
aerosols 
 have been shown to be able to spread over a large area outdoors .
we do not 
 know if aerosols will disperse over large areas indoors , but this is 
 conceivable .
surprisingly , a small fountain without obvious aerosols - generating 
 capability was recently implicated as the source of an outbreak of ld . 

this corroborates that exposure to legionella arising from aerosol - generating sources at health care facilities may occur 
 relatively far from the source .
the hospital workers distance to aerosol 
 sources at the hospital or their number of contacts with the sources had no 
 influence on their antibody level .
the questions about exposure and health were all about conditions in the 
 previous year .
the health symptoms are common , frequent , and probably not easy 
 to remember .
this poses the problem of recall bias , but recall bias will affect 
 both groups equally as no one is aware of having antibodies .
self - reported 
 exposure time and frequencies of exposure at work seem to be valid and useful 
 .
the association between types of symptoms and high antibody levels in 
 some previous studies seems to be weak and inconsistent [ 1 , 3 , 20 ] .
we found no symptoms related to high antibody 
 level , even though single chance findings could be expected due to the large 
 number of tested symptoms .
our study was limited by being based on the 
 serological analysis of single serum samples ; it is well known that antibodies 
 to legionella can be detected months 
 after an infection .
reliable serological diagnosis of a recent or current legionella infection can best be done by 
 the detection of a seroconversion , which for our if test is defined as a fourfold 
 rise in titre to at least 1 : 128 .
in addition , it can take two to several weeks 
 before antibodies can be detected after the onset of symptoms or after exposure .
a 
 follow - up study of a staff cohort would have enabled us to detect both the 
 changes in antibodies and the related symptoms .
we do not know to what extent the seroprevalence varies in different populations , 
 though we found hardly any variation between our two blood donor populations in 
 the two different towns , one with an average incidence and one with an endemic high 
 incidence of ld , respectively .  
we 
 know that the incidence of ld in the study town was within the average 
 incidence of ld in denmark , 
 and we therefore assume that the overall prevalence of antibodies to legionella in the population in the town 
 of our hospital was at the same level as the donor populations in the reference 
 towns .
an inverse relation between age and seroprevalence has not been 
 demonstrated in other studies [ 14 , 16 ] .
we investigated the staff at a hospital with an ongoing high amount of legionella in the water system .
we found 
 that almost half of the staff had serological signs of legionella infection , but these antibodies could not be related to specific 
 occupational exposures or symptoms .
although this indicates that the health 
 implications for workers at health care facilities may be limited , we do not 
 know the health risks if a virulent legionella invades the distribution system .
treatment and 
 maintenance of water systems in healthcare to minimise the threat of legionella contamination following well - described methods should therefore be a standard procedure in order not only to minimise 
 the risk of nosocomial ld but also to reduce occupational risks .
 although legionnaires ' disease frequently is acquired in health care institutions , little is 
                   known about the occupational risk of legionella infection among health care workers . 
 the aim of the present cross - sectional study was to analyse antibody levels among exposed hospital workers and to determine the correlation between antibodies to legionella and self - reported symptoms . 
 the study included 258 hospital employees and a reference group of 708 healthy blood donors .  
 
 hospital workers had a higher prevalence of legionella antibody titres ( 1 : 128 ) than blood donors ( odds ratio 3.4 ; 95% ci 2.44.8 ) . 
 antibody levels were not higher among staff members at risk of frequent aerosol exposure than among less exposed employees . 
 there was no consistent association between a history of influenza - like symptom complex and the presence of antibodies . 
 
 the results indicate that hospital workers have a higher risk of legionella infections than the general population 
 . however , since no excess morbidity was associated with seropositivity , most legionella infections may be asymptomatic . 