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Evaluation of Hypersensitivity Pneumonitis Among Workers Exposed to Metal Removal Fluids



Hypersensitivity pneumonitis (HP) was identified among employees in an automobile parts manufacturing facility. Mycobacteria immunogenum (MI) was identified as a metal removal fluid (MRF) contaminant at this facility and had been identified as a contaminant in other facilities where HP had occurred. We therefore questioned whether measurement of MI-specific cell-mediated immunity would be associated with HP in this facility. We also questioned whether measures of cell-mediated immunity would be more informative about the presence of HP than evaluation of serum anti-MI antibody levels. Workers were categorized for exposure and disease status by questionnaire and review of medical records. Cell-mediated immunity to MI was assessed by measuring in vitro secretion of cytokines (interleukin 8, tumor necrosis factor alpha, and interferon-gamma) from peripheral blood mononuclear cells or anticoagulated whole blood induced by culture with MI antigen. Serum antibodies against MI were also measured. Six study participants met our survey definition for HP and 48 did not. As has been reported for various agents causing HP, serum antibody levels against MI were increased in both exposed workers and workers with HP. Serum antibodies did not distinguish between the two. When expressed as a percentage of secretion induced by lipopolysaccharide, MI induced a significant increase in interleukin-8 secretion in exposed participants' whole blood cultures. There were trends for increased MI-induced secretion of interferon-gamma by peripheral blood mononuclear cells from both exposed workers and workers with HP. However, these trends did not attain statistical significance. Thus, several measures of immunity to MI distinguished between exposed and unexposed workers but not between workers with and without HP. These evaluations of cell-mediated immunity were not more informative than measurement of serum antibodies. As was done at this facility, institution of a comprehensive safety and health plan for MRF is necessary to eliminate (or minimize) health effects related to occupational exposures in the machining environment.
Applied Occupational and Environmental Hygiene, 18: 953–960, 2003
Copyright c
Applied Industrial Hygiene
ISSN: 1047-322X print / 1521-0898 online
DOI: 10.1080/10473220390237683
Evaluation of Hypersensitivity Pneumonitis Among
Workers Exposed to Metal Removal Fluids
Douglas Trout,1David N. Weissman,2Daniel Lewis,2Rodney A. Brundage,2
Alfred Franzblau,3,4and Daniel Remick4,
1Division of Surveillance, Hazard Evaluations, and Field Studies, National Institute for Occupational
Safety and Health, Cincinnati, Ohio; 2Health Effects Laboratory Division, National Institute
for Occupational Safety and Health, Morgantown, West Virginia; 3University of Michigan School of Public
Health, Ann Arbor, Michigan; 4University of Michigan Medical School, Ann Arbor, Michigan
Hypersensitivity pneumonitis (HP) was identified among
employees in an automobile parts manufacturing facility.
Mycobacteria immunogenum (MI) was identified as a metal
removal fluid (MRF) contaminant at this facility and had
been identified as a contaminant in other facilities where HP
had occurred. We therefore questioned whether measure-
ment of MI-specific cell-mediated immunity would be asso-
ciated with HP in this facility. We also questioned whether
measures of cell-mediated immunity would be more infor-
mative about the presence of HP than evaluation of serum
anti-MI antibody levels. Workers were categorized for expo-
sure and disease status by questionnaire and review of med-
ical records. Cell-mediated immunity to MI was assessed
by measuring in vitro secretion of cytokines (interleukin
8, tumor necrosis factor alpha, and interferon-γ)from pe-
ripheral blood mononuclear cells or anticoagulated whole
blood induced by culture with MI antigen. Serum antibod-
ies against MI were also measured. Six study participants
met our survey definition for HP and 48 did not. As has
been reported for various agents causing HP, serum an-
tibody levels against MI were increased in both exposed
workers and workers with HP. Serum antibodies did not
distinguish between the two. When expressed as a percent-
age of secretion induced by lipopolysaccharide, MI induced
a significant increase in interleukin-8 secretion in exposed
participants’ whole blood cultures. There were trends for
increased MI-induced secretion of interferon-γby periph-
eral blood mononuclear cells from both exposed workers and
workers with HP. However, these trends did not attain sta-
tistical significance. Thus, several measures of immunity to
MI distinguished between exposed and unexposed workers
Supported in part by National Institutes for Health grant GM 50401.
but not between workers with and without HP. These eval-
uations of cell-mediated immunity were not more informa-
tive than measurement of serum antibodies. As was done at
this facility, institution of a comprehensive safety and health
plan for MRF is necessary to eliminate (or minimize) health
effects related to occupational exposures in the machining
Keywords Mycobacterium Immunogenum, Hypersensitivity Pneu-
monitis, Metalworking Fluids, Metal Removal Fluids,
Coolant, Machining
In 2002, the National Institute for Occupational Safety and
Health (NIOSH) received a request for a health hazard evalu-
ation (HHE) from an automobile parts manufacturing facility.
The request concerned respiratory problems and hypersensitiv-
ity pneumonitis (HP) thought to be associated with occupational
exposures to metal removal fluids (MRFs). In response to the
HHE request, NIOSH investigators performed a record review
and a questionnaire and serologic survey among employees in
selected departments at the facility. The goal of this evaluation
was to provide further understanding of potential causes of HP,
which had been diagnosed among some employees at the facil-
ity. This report summarizes the results of this evaluation.
At the time of the survey, the facility was three years old
and employed approximately 2000 workers in the manufacture
of automatic transmissions and transmission components. The
facility has a medical department with on-site physician and
nursing staff. Multiple central MRF systems supplied a semi-
synthetic MRF to machines performing a variety of types of
machining; the metals being machined were primarily steel and
aluminum. Machine tools at this facility are generally enclosed
and exhausted to the outdoors, and 55 percent (28 of 51) of
recent industrial hygiene samples of MRF aerosol (total partic-
ulate) were below 0.2 milligrams per cubic meter (mg/m3). Bulk
sampling of central MRF systems around the time the illnesses
were recognized revealed contamination of several systems with
Mycobacteria immunogenum, a microorganism that has been
found as a contaminant in other machining plants where there
have been outbreaks of HP.(1)
In September 2001, a 35-year-old male toolmaker at the facil-
ity developed a non-productive cough, low-grade fever, dyspnea
with minimal exertion, and myalgias. He smoked 1
2to 1 pack of
cigarettes per day, but stopped around the time these symptoms
began. He was initially treated with several courses of antibiotics
without improvement. Pulmonary consultation two months after
symptom onset was notable for continued symptoms, a normal
physical examination, and oxygen saturation with brief exertion
(walking) on room air of 81 percent. Spirometry revealed a mod-
erate reduction in forced vital capacity with a normal ratio of
forced expiratory volume in one second to forced vital capac-
ity. Diffusing capacity for carbon monoxide was within normal
limits. A chest radiograph was interpreted as no acute process.
The patient was begun on prednisone (60 milligrams per day)
due to concerns that he was possibly experiencing pulmonary
manifestations of an autoimmune disease. Approximately three
weeks after initial pulmonary consultation the patient was taken
off work. A ventilation/perfusion scan was performed and was
interpreted as low probability to indeterminatefor pulmonary
embolism and the patient was admitted to the hospital for anti-
coagulation (treatment for presumed thromboembolic disease).
During that hospitalization, physical examination was notable
for a few coarse interstitial crackles bilaterally. Computed to-
mography of the chest revealed diffuse interstitial inltrates
(ground glasschanges). Anticoagulation treatment was dis-
continued. Open lung biopsy approximately six weeks after
initial pulmonary consultation revealed chronic interstitial pneu-
monia with rare giant cells and vague non-necrotizing gran-
ulomas most consistent with HP. Follow-up examination 21
months after initial pulmonary consultation revealed resolution
of symptoms, oxygen saturation with brief exertion on room air
of 94 percent, and normal spirometry. The patient was main-
tained on a tapering dose of prednisone until it was discontin-
ued approximately three months after initial pulmonary consul-
tation. He returned to work, approximately ve months after
initial pulmonary consultation, to an area of the facility outside
the machining areas where he had formerly worked.
Occupational exposure to MRF aerosols causes a variety of
pulmonary symptoms and illnesses (including HP); these health
effects have been summarized previously.(14) HP (also called
extrinsic allergic alveolitis) is a granulomatous, interstitial lung
disease that occurs after repeated inhalation and sensitization to
a wide variety of microbial agents (bacteria, fungi, amoebae),
animal proteins, and low-molecular weight chemical antigens.
It is marked by a pneumonitis, which is reversible if exposure to
the antigen is stopped; continued exposure can lead to a chronic
interstitial brosis or scarring of the lungs. In general, HP is
marked by nonspecic symptoms. Suggestions for uniform cri-
teria for the diagnosis of HP have been published.(5,6)
HP is thought to involve both humoral and cell-mediated
immune responses.(7,8) Detection of antibody is associated with
exposure to a suspect causative antigen, but does not by itself
conrm the presence of an active disease.(9) With recognition
of the difculty of performing specic inhalation challenges
to evaluate the potential causative role of a specic antigen,
measurements of the secretion of substances playing roles in
cell-mediated immunity (as a surrogate for clinical illness) have
been described.(8)
One study found that peripheral blood cells from subjects
with pigeon breeders disease (a form of hypersensitivity pneu-
monitis) produced increased amounts of a cytokine (migration
inhibitory factor) after in vitro challenge compared with those
from asymptomatic persons.(10) Interleukin 8 (IL-8), tumor
necrosis factor alpha (TNF-α), and interferon gamma (IFN-γ)
have been suggested as playing a role in the pathogenesis of sev-
eral types of HP.(11) We therefore questioned whether measure-
ment of MI-specic cell-mediated immunity as MI-stimulated
secretion of IL-8, TNF-α, or IFN-γby peripheral blood cells
would be associated with HP in this facility. We also questioned
whether such measures would be more informative about the
presence of HP than evaluation of serum anti-MI antibody levels.
Medical Record Review and Questionnaire
Facility medical records concerning employees from machin-
ing areas who had presented to the facility medical department
within the previous several months with respiratory symptoms
and/or illnesses were reviewed. Subsequently, a brief question-
naire, which included questions concerning job duties, current
symptoms, and medical history was administered in June 2002
to employees in selected manufacturing departments. Included
among those departments were three machining departments
and one assembly department. Current cigarette smoking sta-
tus was determined by response to the question Do you now
smoke cigarettes?The goal of the questionnaire was to iden-
tify groups of employees to take part in the serological survey
described below. The protocol for this HHE, including ques-
tionnaire administration and serologic survey, was approved by
the NIOSH and University of Michigan human subjects review
boards. All participants provided informed consent.
Based on laboratory capabilities for the serological stud-
ies, we planned to recruit approximately 15 participants from
each of four groups (see below). Symptomaticemployees in-
cluded those employees who reported on the questionnaire in
the preceding year: a) chest u (fever, shivering, cough, tired,
weak, ache all over) or pneumonia, with symptoms related to
work; and/or b) episode of breathing difculty (such as wheez-
ing, shortness of breath, and/or cough) with symptoms related
to work. Asymptomaticemployees were dened as employ-
ees who reported neither anor babove. If more than 15
potential participants were available from any of the groups, 15
participants were chosen randomly from the total number.
Participants were categorized into one of four groups: Group 1
employees working in machining areas (exposedto MRF)
who were suspected or conrmed to have respiratory illness
(possibly including HP) by their physicians; Group 2 employ-
ees from the machining areas who were symptomatic but who
had not presented to the facility medical department; Group 3
employees from the machining areas who were asymptomatic;
and Group 4 employees from the assembly area (unexposed
to MRF) who were asymptomatic. HP was dened using a pub-
lished case denition previously used in the evaluation of res-
piratory illness among workers in a large machining plant in
19951996 (Table I).(6) We considered all persons with possi-
ble,”“probable,or deniteHP to meet our case denition for
Cytokine and Antibody Analyses
To evaluate whether M. immunogenum, which was cultured
from the MRF at the facility, may be related to the HP diag-
nosed among some workers at the plant, we performed cytokine
and antibody analyses. A nding of increased cytokine secretion
from workers with HP compared to those from workers without
HP would provide some evidence that the affected workers were
specically sensitized to M. immunogenum. Our cytokine anal-
yses measured the response of workerswhole blood (Assay 1)
and peripheral blood mononuclear cells (PBMC) (Assays 2 and
3) after in vitro exposure to a M. immunogenum antigen prepa-
ration. Blood was obtained by venipuncture and collected into
Diagnostic criteria for hypersensitivity pneumonitis(6)
Physician diagnosis of hypersensitivity pneumonitis
Onset of at least two pulmonary symptoms (cough, wheeze,
chest tightness, shortness of breath) and one systemic
symptom (fever, weight loss) after July 2001
Recurrence of pulmonary or systemic symptoms after three or
more day avoidance
Restrictive pattern on spirometry not due to obesity
Impairment of pulmonary diffusing capacity less than 80%
Interstitial or reticulonodular pattern on chest x-ray or
computed tomography
Biopsy evidence of non-caseating granulomas
Denite case of HP: Meeting six or seven criteria.
Probable case of HP: Meeting ve criteria.
Possible case of HP: Meeting four criteria.
heparinized tubes. Samples were stored on ice and taken di-
rectly for evaluation at the University of Michigan (Assay 1),
or shipped overnight for evaluation at NIOSH-Morgantown
M. immunogenum (MI) was obtained from the American
Type Culture Collection (ATCC), Manassas, VA (#700506).
This isolate has previously been implicated as being associated
with MRF-related HP.(12) Puried protein derivative from MI
(MI-PPD) was prepared in a fashion similar to that described by
Landi.(13) Briey, MI was cultured in Middlebrook 7H9 broth
supplemented with OADC for 3 weeks at 37C. At 3 weeks,
growth was heavy with a large amount of precipitated material
present in cultures. Cultures were sterilized by autoclaving for
15 minutes. Culture supernatants were obtained by centrifuga-
tion (20,000 ×g, 4C, 1 hour). Protein was precipitated from
culture supernatants using saturated ammonium sulfate. Precip-
itate was obtained by centrifugation (20,000 ×g, 4C, 1 hr) and
re-dissolved in a minimum volume of phosphate-buffered saline
(PBS). This material was dialyzed against four 20-fold volumes
of PBS at 4C over the next 48 hours. The dialysate was re-
moved, claried by centrifugation (20,000 ×g, 4C, 4 hours),
passed through a 0.45 µmlter, aliquoted, and stored frozen
at 70C. The resulting MI-PPD preparation was sterile, had
a protein concentration of 31 mg/ml by Lowry assay, and an
endotoxin concentration of 1.3 ng/ml by limulus amoebocyte
lysate (LAL) assay. The optimal dilution of the MI-PPD prepa-
ration for stimulating IFN-γsecretion by a tuberculin-positive
volunteers PBMC using Assay 2 (described below) was 1:20.
Assay 1
The PPD prepared from M. immunogenum (MI-PPD; see
above) antigen preparation was prepared at ten times concen-
tration, and 110 microliter (µl) were added to one ml of whole
blood. Enough stimulant was prepared for at least 40 partici-
pants. The blood and the MI-PPD were combined in the fol-
lowing order. The MI-PPD stock was 31 milligram (mg)/ml,
and stimulation was performed at 0.248 µg/ml. The stock was
diluted 1/100 (990 µl control vehicle [RPMI] +10 µl MI-PPD).
Forty µl of the above dilution was added to 5 ml of RPMI for a
nal concentration of 2.48 µg/ml. The lipopolysaccharide (LPS)
antigen was prepared as follows. The stock was 100 µg/ml and
we stimulated at 100 nanogram (ng)/ml (50 µl of stock was
added to 5 ml RPMI for a nal concentration of 1000 ng/ml.
After adding blood, the tubes were placed on the rotator in the
incubator (37C) for 20 hours. At the end of 20 hours, the tubes
were spun at 2000 ×g for three minutes, and the plasma was
drawn off and frozen at 70C. Enzyme-linked immunosor-
bent assay (ELISA) was used to analyze for TNF-α, IFN-γ,
and IL-8.
Assays 2 and 3
PBMC were prepared by centrifugation over sodium dia-
trizoate/Ficoll gradients (Isolymph, Gallard Schlesinger, Carle
Place, NY).
Cultures for Assay 2 (MI-PPD stimulated IFN-γsecretion by
PBMC) were performed in 96-well round-bottomed microtiter
plates using RPMI-1640 tissue culture medium supplemented
with 25 mM HEPES, 2 mM L-glutamine, 100 units/ml peni-
cillin, 0.1 mg/ml streptomycin, 5 percent fetal calf serum, and
5 percent human AB serum (CRPMI). 2×105PBMC in a volume
of 200 µl culture medium per well were cultured with CRPMI
alone (negative control), phytohemagglutinin-P (PHA) 5 µg/ml
(positive control), puried protein derivative (PPD) 20 µg/ml
(Accurate Chemical & Scientic Co., Westbury, NY), or MI-
PPD (see above) diluted to a nal concentration of 1:10, 1:20,
or 1:100. Cultures were performed at 37C and 5 percent carbon
dioxide (CO2) for 5 days, an interval previously documented to
be optimal for determination of PPD-induced IFN-γsecretion
by PBMC from tuberculin positive individuals (data not shown).
After 5 days, conditioned culture supernatants were obtained
and saved frozen at 70C until assayed for IFN-γcontent by
ELISA (Interferon Gamma Duoset,R&DSystems, Minneapo-
lis, MN). The data for the 1:20 dilution are presented here; these
data correlated with data for the 1:10 dilution (r =0.84) and less
well with data from the 1:100 dilution (r =0.37).
To perform ELISPOT assays (Assay 3), 96-well PVDF
membrane-bottomed plates (MAIPS4510, Millipore, Bedford,
MA) were coated overnight with 50 µl/well of 10 µg/ml solu-
tion of anti-human IFN-γmonoclonal antibody (clone 1-D1K;
Mabtech Inc., Cincinnati, OH) in carbonate buffer, pH 9.6. After
washing and blocking with CRPMI, cultures were performed
in CRPMI alone (negative control), PHA-P 5 µg/ml (positive
control), puried protein derivative (PPD) 20 µg/ml (Accurate
Chemical & Scientic Co., Westbury, NY), or MI-PPD diluted
toanal concentration of 1:20. 5 ×104cells were added to cul-
tures performed with PHA-P; all other cultures were performed
with 2 ×105cells. Plates were incubated at 37C, 5 percent
CO2overnight. To visualize spots, plates were incubated with
the following reagents with appropriate washing steps between
incubations: biotinylated anti-human IFN-γmonoclonal anti-
body 1 µg/ml (clone 7-B6-1, Mabtech Inc., Cincinnati, OH),
Vectastain AB reagent (Vectastain ABC Elite kit, Vector Labo-
ratories, Burlingame, CA), and Vector VIP peroxidase substrate
(Vector Laboratories). Plates were washed with water, dried, and
spots counted using a dissecting microscope. Data are expressed
as spots per 105PBMC.
Antibody Studies
An antigen extract was prepared from the same isolate of M.
immunogenum used for the MI-PPD preparations. The antigen
preparation, which contains both cell-associated and secreted
products, was made by collecting the bacterial cells by cen-
trifugation and re-suspending in saline as a 10 percent volume/
volume suspension. The bacterial suspensions were sonicated
for one minute using a Branson Model 350 sonier set at 40 per-
cent output, 50 percent duty cycle pulse. The sonicates were
claried by centrifugation at 3500 rpm for 20 minutes, and the
supernatant uid was recovered, stored at 20C, and used as
the source of antigen. The protein content of bacterial sonicate
was determined using a modied Lowry method (BioRad) ac-
cording to the manufacturers recommendations. The bacterial
sonicates were diluted with carbonate coating buffer (pH 9.6)
for use in the ELISA assay.
A direct ELISA for antibodies to M. immunogenum was
developed using the procedures described by Voller and Bid-
well (Manual of Clinical Immunology, 4th edition, ASM Press,
Washington, DC). ELISA plates were coated with the M. im-
mungenum extract (3 to 5 µg protein/ml) in carbonate coat-
ing buffer overnight at 4C, blocked with 1 percent human
serum albumin. The subject sera were initially tested in dupli-
cate at a 1:100 dilution, and antibody binding was detected using
peroxidase-labeled anti-human immunoglobulins (Sigma) and
developed with TMB substrate. Appropriate positive and neg-
ative human serum controls were performed with each plate,
including an antigen blank. Results were reported as optical
density at 450 nm.
Data Analysis
To control for the number of viable cytokine-producing cells,
for Assays 1 and 2 the secretion of cytokine was expressed as a
percentage of cytokine secretion after stimulation from a strong
stimulant (lipopolysaccharide [LPS] [Assay 1] or phytohemag-
glutinin [PHA] [Assay 2]). The equation(14) used to calculate the
% LPS(or %PHA) was:
[(cant ccon)/(cLPS ccon )] ×100 [1]
where cant =cytokine stimulated by antigen, ccon =cytokine
stimulated by control, and cLPS =cytokine stimulated by LPS (or
in Assay 2, by PHA). For the ELISPOT (Assay 3), the index used
in the evaluation was the difference between antigen-stimulated
result and the result from the negative control. Comparisons
of cytokine concentrations (Assays 1 and 2), ELISPOT results
(Assay 3), and antibody levels were made by case (HP case ver-
sus non-case) and exposure (exposed versus unexposed) status
using the Wilcoxon two-sample test.Apvalue of .05 was used
to indicate statistical signicance.
Medical Record Review
As of the start of the NIOSH HHE, medical records for 16
workers were available and reviewed by a physician; all work-
ers were from machining departments. Job titles included oper-
ator (8 persons), job setter (4 persons), toolmaker (2 persons),
and machine repair and janitor (1 each). Using the case deni-
tion outlined above,(6) 7 workers met the denition for possible
(2 workers), probable (2 workers), or denite (3 workers) HP.
Among the 7 meeting a case denition for HP were 2 operators,
2 job setters, 2 toolmakers, and 1 janitor. Among the remaining
9 workers for whom records were reviewed, 7 of the 9 appeared,
based on the physicians review, to have work-related respiratory
or upper respiratory symptoms, and 2 of the 9 appeared to have
respiratory symptoms that were not work-related. Among the
14 workers with work-related respiratory conditions (7 with HP
and 7 with other work-related respiratory conditions), 11 (73%)
participated in our laboratory testing, including 6 of the 7 with
HP and 5 of the 7 categorized as having other work-related res-
piratory disorders.
Questionnaire Administration and Selection of
Participants for Laboratory Testing
The 14 workers mentioned above made up the Group 1 par-
ticipants. The primary goal of the questionnaire survey was to
identify participants in Groups 24. Although not used for group
categorization, questionnaires were also completed for all those
available in Group 1. Of the 150 questionnaires distributed to
workers in the departments of interest, 140 (93%) were com-
pleted. Of the 140 persons who completed the questionnaire, 36
(26%) declined to participate any further in the HHE. Among the
remaining 104 workers, 22 from the machining areas were symp-
tomatic (and therefore made up potential members of Group 2),
and 13 were asymptomatic (and therefore made up potential
members of Group 3). Fourteen of the 104 were asymptomatic
workers from the assembly department (and therefore made up
potential members of Group 4). The remainder of the partici-
pants who completed the questionnaire reported some respira-
tory symptoms but did not report work-relatedness and were not
included in any of the four groups. Fifteen of the 22 symptomatic
workers from the machining areas were randomly selected to be
offered the laboratory testing as part of Group 2.
Therefore, 56 workers were initially identied as potential
participants for the laboratory portion of the survey from Groups
14. Seven (13%) of the workers on the original list of 56 were
not available (mostly due to vacation) to participate in the in-
formed consent procedures and were therefore dropped from the
survey. During that time, 9 workers who worked in the depart-
ments of concern, but who had not been at work during the ques-
tionnaire administration, presented requesting to be included in
the HHE.
All completed the questionnaire, were grouped according to
their exposure status and symptoms as reported in the question-
naire, and participated in the laboratory testing. Therefore, 58
workers were scheduled to have blood drawn. Fifty-one (88%)
of the 58 completed the blood sampling. Due to timing of ship-
ment of blood samples, 5 of the 51 blood samples could not be
transported to the University of Michigan laboratory. Three of
the 7 workers unavailable at the time of the initial blood draw
provided blood samples when NIOSH representatives returned
to the facility 2 weeks after the primary testing date; those 3
samples were analyzed only by the NIOSH laboratory because
the samples could not be transported to the University of Michi-
gan laboratory. This yielded the following: 46 samples analyzed
by the Michigan laboratory (Assay 1) and 54 samples analyzed
by the NIOSH laboratory (Assays 2 and 3).
Description of casesAand non-cases participating in health
hazard evaluation
# (%) # (%)
N Mean age Male Current smoker
Non-case 48 40 30 (63) 11 (23)
HP case 6 44 6 (100) 0 (0)
AMeeting case denition for hypersensitivity pneumonitis.(6)
Cytokine/Antibody Analyses
Cytokine and antibody results are presented by case (HP)
and exposure status; descriptive summaries of those groups are
presented in Tables II and III.
Assay 1 (MI-PPD Stimulated Secretion of Cytokines
in Whole Blood Cultures)
The limit of detection (LOD) for each of the three cytokines
after stimulation with M. imunogenum,M. tuberculosis, and
RPMI was 60 picograms per ml (pg/ml); the LOD for the cy-
tokines after stimulation with LPS was 300 mg/ml. IFN-γwas
detected above the LOD (63 and 77 pg/ml) in only 2 of 46
samples after stimulation with M. immunogenumone sample
from a worker with HP, one from a worker in Group 4 (unex-
posed, asymptomatic). TNF-αwas detected above the LOD (89
and 93 pg/ml) in only two samples after stimulation with M.
immunogenumone sample from a worker with HP, one from
a worker who had respiratory symptoms that were not work-
related (Group 1).
Summary data for IL-8 secretion are presented in Tables IV
(HP cases versus non-cases) and V (MRF-exposed versus those
unexposed to MRF). There were no statistically signicant dif-
ferences in the concentrations of IL-8 among the persons with
HP (compared to those without HP), or among the persons ex-
posed to MRF (compared to those unexposed); the median val-
ues were greater among the persons without HP and among the
persons unexposed to MRF. There was no statistically signicant
difference in the index measure for IL-8 secretion (IL-8 secre-
tion expressed as a percent of LPS secretion) between persons
with HP and without. However, this index of IL-8 secretion was
statistically signicantly greater among those exposed to MRF
(median 0.33) compared to the unexposed (median 0).
Description of MRF-exposed and MRF-unexposed
participating in health hazard evaluation
# (%) # (%)
N Age (mean) Male Current smoker
Unexposed 11 41 5 (45) 2 (18)
Exposed 43 40 31 (72) 9 (21)
Cytokine/antibody results by case status
Cytokine/antibody Group Mean Median p valueA
Assay 1 Non-case 4607 2923 0.3
IL-8BHP case 4654 1398
Assay 1 Non-case 2.8 0 0.9
IL-8 %LPSCHP case 3.6 0
Assay 2 Non-case 102.0 0 0.2
IFN-γDHP case 143.3 141
Assay 2 Non-case 6.1 0 0.3
IFN-γ%PHAEHP case 7.1 4.3
Assay 3 Non-case 3.8 1.0 0.4
IFN-γELISPOTFHP case 2.8 1.6
Assay 3 Non-case 1.0 0.5 0.7
IFN-γELISPOT HP case 2.4 1.0
Antibody to Non-case 0.098 0.074 0.001
M. immunogenumHHP case 0.29 0.21
AWilcoxon two-sample test; two-sided.
BInterleukin-8 (pg/ml).
CInterleukin-8 index (% of lipopolysaccharide).
DInterferon gamma (pg/ml).
EInterferon gamma index (% of phytohemagglutinin).
FInterferon gamma ELISPOT-producing cells per 105PBMC.
GELISPOT index is differencebetween antigen-stimulated result and
that from control.
HOptical density at 450 nm.
Assays 2 (MI-PPD Stimulated Secretion of IFN-γ
into Culture Supernatants by PBMC) and 3 (MI-PPD
Stimulated IFN-γELISPOT)
Summary data for IFN-γsecretion are also presented in
Tables IV and V. There were no statistically signicant differ-
ences in the concentrations of IFN-γ(both absolute secretion
and index value [IFN-γ%PHA]) as determined by Assay 2
among the persons with HP (compared to those without HP),
or among the persons exposed to MRF (compared to those un-
exposed). The median values for both measures were greater
among the persons with HP and among the persons exposed to
There were no statistically signicant differences in the ab-
solute and index values for IFN-γsecretion as determined by
Assay 3 among the persons with HP (compared to those without
HP) and among exposed persons (compared to those unexposed
to MRF). The median values for both absolute and index mea-
sures were higher among HP cases (compared to non-cases),
while the median values for the absolute and index measures
among exposed persons (compared to those unexposed to MRF)
did not show the same consistent trend.
Antibody Assays
Antibody levels against the M. immunogenum antigen were
greater among both persons with HP (compared to those without
HP) (Table IV) and persons exposed to MRF compared to those
unexposed (Table V). Because antibody production to inhaled
antigens has been shown to be inhibited by cigarette smoking,(11)
analyses were performed in which current cigarette smokers
were excluded. There was no substantial change in the increased
level of antibody among HP cases and among MRF-exposed
persons with the analyses performed among non-smokers only.
The primary goal of our evaluation was to learn whether mea-
sures of cell-mediated immunity to M. immunogenum, which had
been identied as an MRF contaminant in the areas of the facil-
ity where workers had been diagnosed with HP, were associated
with the presence of HP among these workers. Except for the
IL-8 percent LPS assay, our cytokine studies revealed no statisti-
cally signicant relationships between secretion of the cytokines
tested and either the presence of HP or exposure to MRF. Of the
cytokine assays used in this HHE, Assay 2 (MI-PPD stimulated
IFN-γsecretion into culture supernatants) showed increased
secretion (for both the absolute and index measures of IFN-γ)
among persons with HP, although the differences were not sta-
tistically signicant. A study with a larger number of affected
workers would improve our ability to detect small differences in
IFN-γsecretion from PBMCs of persons with HP. However, due
to overlap between groups with and without HP, it is doubtful
Cytokine/antibody results by exposure status
Cytokine/antibody Group Mean Median p valueA
Assay 1 Unexposed 3185 3131 0.9
IL-8BExposed 4601 2364
Assay 1 Unexposed 0 0 0.02
IL-8 %LPSCExposed 3.3 0.33
Assay 2 Unexposed 34 0 0.2
IFN-γDExposed 124 12
Assay 2 Unexposed 2.0 0 0.2
IFN-γ%PHAEExposed 7.4 0.38
Assay 3 Unexposed 15.7 1.1 0.8
IFN-γELISPOTFExposed 1.4 0.88
Assay 3 Unexposed 0.72 0.25 0.3
IFN-γELISPOT Exposed 1.24 0.5
Antibody to Unexposed 0.057 0.057 0.03
M. immunogenumHExposed 0.16 0.10
AWilcoxon two-sample test; two-sided.
BInterleukin-8 (pg/ml).
CInterleukin-8 index (% of lipopolysaccharide).
DInterferon gamma (pg/ml).
EInterferon gamma index (% of phytohemagglutinin).
FInterferon gamma ELISPOT-producing cells per 105PBMC.
GELISPOT index is difference between antigen-stimulated result and
that from control.
HOptical density at 450 nm.
that this type of testing will be a useful tool in medical screening
for HP at other machining facilities.
The results of our testing for antibodies against Mycobac-
teria were consistent with antibody testing against causative
antigens in the evaluation of other types of HP. For example,
elevated levels of antibody against puried avian antigens have
been associated with increased incidence of pigeon breeders dis-
ease (a form of HP),(15) and increasing exposure to pigeons has
been associated with increase in antibody response to pigeon
However, as with other antibody testing, our antibody testing
was non-specic in that we found elevated levels of antibody in
exposed persons without regard to the presence of HP. MRF as
they are used in production facilities are complex mixtures of
multiple potential antigens, both microbial and chemical. There-
fore, based on antibody testing alone, we cannot rule out that
some other antigen(s) may be present along with the M. immuno-
genum and may be involved in the pathogenesis of HP.
There were several limitations to our evaluation. Although
we based our grouping of workers on the most up-to-date clin-
ical information that was available to us, our records may have
been incomplete so it is possible that participants may have been
misclassied in terms of case/non-case status. Likewise, it is
possible that our departmental-level determination of exposure
status may have led to some misclassication of participants by
exposure at the individual level. Finally, the fact that we were
using PBMCs may be an important limitation of our labora-
tory work. Because the target organ of inhaled antigens thought
responsible for HP is the lung, recent work related to the patho-
genesis of HP has involved the cellular immune response of cells
from the lung (obtained via bronchoalveolar lavage [BAL]).(17)
Cells obtained from BAL may be more accurate indicators of
cell-mediated immune responses occurring in the lung; how-
ever, these types of cells are not readily available from workers
involved in evaluations such as this HHE.
A number of actions were taken by the facility in response to
the illnesses among employees. The primary actions taken in-
volved improvements in maintenance, cleaning, and formulation
of the MRF in the machining areas. Based on review of indus-
trial hygiene records at the facility, the engineering controls in
place appeared to be effectively maintaining the workersex-
posure to MRF aerosol at concentrations substantially less than
the NIOSH-recommended exposure limit. However, there was
concern that some workers at the facility may be exposed to
greater concentrations of MRF aerosol for short periods of time
due to job tasks which entail entering machine tool enclosures.
Four of the seven persons meeting the case denition for HP
were job setters or tool makers, workers whose jobs involve
routine (intermittent) entrance into the machine tool enclosures.
Based on the above, and incomplete knowledge related to the
importance of short-term peakexposures to MRF aerosol rel-
ative to ambient (or average) exposures, reduction in such peak
exposures may be warranted. Primary methods to accomplish
this involve changes in engineering controls and work practices.
Such changes should be part of a comprehensive safety and
health plan recommended by NIOSH to minimize health effects
related to occupational exposure to MRF.(1)
The authors thank Barbara MacKenzie, Debbie Sammons,
Toni A. Bledsoe, Janet Simpson, and Erica Janotka for their
assistance in conducting this evaluation.
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... Lipid pneumonia and legionellosis have been reported rarely in recent decades. However, hypersensitivity pneumonitis, an allergic pneumonia, has been the subject of more recent reports of workers exposed to MWF [CDC 1996;Kreiss, Cox-Ganser 1997;Freeman et al. 1998;Zacharisen et al. 1998;Fox et al. 1999;Shelton et al. 1999;Hodgson et al. 2001;CDC 2002;Bracker et al. 2003;Trout et al. 2003;Beckett et al. 2005;Dawkins et al. 2006;Gupta, Rosenman 2006;Robertson et al. 2007]. In some recent investigations of outbreaks of MWF-associated hypersensitivity pneumonitis, other respiratory illnesses, including asthma and chronic bronchitis, have been found in co-workers [Kreiss, Cox-Ganser 1997;Zacharisen et al. 1998;Hodgson et al. 2001;Robertson et al. 2007]. ...
... The machine 29 sample was negative for mycobacterial DNA. Mycobacterium immunogenum, a rare species of mycobacteria related to Mycobacterium chelonae, has been found in MWF in some investigations of work-related hypersensitivity pneumonitis [Kreiss, Cox-Ganser 1997;Fox et al 1999;Shelton et al. 1999;Wilson et al 2001;Trout et al 2003;Beckett 2005;Gupta, Rosenman 2006]. ...
... Performance was assessed by comparing the proportion of workers correctly identified as definite, possible and definitely not MWF-HP, against the panel opinion. In an attempt to externally validate the MWF-HP Score, it was also applied to 50 previously published US case reports of workers developing HP due to MWF exposure [Bernstein et al., 1995;Trout et al., 1996;Kreiss and Cox-Ganser, 1997;Zacharisen et al., 1998;Fox et al., 1999;Hodgson et al., 2001;Dangman et al., 2002;Weiss et al., 2002;Trout et al., 2003;Gupta and Rosenman, 2006]. These cases had previously been identified during a separate systematic literature review of all previously published MWF ill health outbreaks . ...
... There was a good level of correlation between the derived MWF-HP Score, and the Expert Panel Percentage, with a Pearson correlation coefficient of 0.85 (P < 0.01). By using suitable cut offs for the MWF-HP Scores (definite case > 26, possible case 19-26, and definitely not a case < 19), it was possible to show agreement (shown in bold) with the panel opinion in 30/37 (81%) cases ( Figure 2 shows the MWF-HP Score, or range of possible scores, for 50 previously published cases of MWF-HP from a range of workplaces in the United States [Bernstein et al., 1995;Trout et al., 1996;Kreiss and Cox-Ganser, 1997;Zacharisen et al., 1998;Fox et al., 1999;Hodgson et al., 2001;Dangman et al., 2002;Weiss et al., 2002;Trout et al., 2003;Gupta and Rosenman, 2006]. In 22/50 of these cases, the MWF-HP Score or score range would have identified them as a definite HP case. ...
Full-text available
Background This study used data from a large UK outbreak investigation, to develop and validate a new case definition for hypersensitivity pneumonitis due to metalworking fluid exposure (MWF-HP). Methods The clinical data from all workers with suspected MWF-HP were reviewed by an experienced panel of clinicians. A new MWF-HP Score was then developed to match the “gold standard” clinical opinion as closely as possible, using standard diagnostic criteria that were relatively weighted by their positive predictive value. Results The new case definition was reproducible, and agreed with expert panel opinion in 30/37 cases. This level of agreement was greater than with any of the three previously utilized case definitions (agreement in 16–24 cases). Where it was possible to calculate, the MWF-HP Score also performed well when applied to 50 unrelated MWF-HP cases. Conclusions The MWF-HP Score offers a new case definition for use in future outbreaks. Am. J. Ind. Med. 57:872–880, 2014. © 2014 The Authors. American Journal of Industrial Medicine Published by Wiley Periodicals, Inc.
... The occurrence of mycobacteria in MWF and their potential occupational health significance has been highlighted [3,5,6]. Mycobacterium immunogenum, a nontuberculous Mycobacterium species, has been reported in MWF and implicated as possible causative agent of hypersensitivity pneumonitis in machine workers who have been exposed to it [7][8][9]. Among other MWF microbial communities, pseudomonads often constitute the major fraction of gram-negative organisms responsible for endotoxin release and accumulation in the fluids, resulting in occupational health hazards in metalworkers [10]. ...
... The latter-in most cases the detection of precipitating IgG antibodies to contaminated MWF samples-have however failed, as with other causes of occupational HP, to distinguish reliably between cases and healthy, but exposed, coworkers. 7 Particular attention has been paid to the role of contaminating non-tuberculous mycobacteria (NTM). Notably, molecular techniques identified a single strain of Mycobacterium immunogenum in all but 2 of 107 MWF samples collected from 10 US workplaces with cases of HP, 8 a finding that led some to claim that this species has a particular capacity for colonisation and infection in this setting; a subsequent outbreak in France provided some support for this position. ...
Full-text available
Background Outbreaks of hypersensitivity pneumonitis (HP) are not uncommon in workplaces where metal working fluid (MWF) is used to facilitate metal turning. Inhalation of microbe-contaminated MWF has been assumed to be the cause, but previous investigations have failed to establish a spatial relationship between a contaminated source and an outbreak. Objectives After an outbreak of five cases of HP in a UK factory, we carried out blinded, molecular-based microbiological investigation of MWF samples in order to identify potential links between specific microbial taxa and machines in the outbreak zone. Methods Custom-quantitative PCR assays, microscopy and phylogenetic analyses were performed on blinded MWF samples to quantify microbial burden and identify potential aetiological agents of HP in metal workers. Measurements and main results MWF from machines fed by a central sump, but not those with an isolated supply, was contaminated by mycobacteria. The factory sump and a single linked machine at the centre of the outbreak zone, known to be the workstation of the index cases, had very high levels of detectable organisms. Phylogenetic placement of mycobacterial taxonomic marker genes generated from these samples indicated that the contaminating organisms were closely related to Mycobacterium avium. Conclusions We describe, for the first time, a close spatial relationship between the abundance of a mycobacterium-like organism, most probably M. avium, and a localised outbreak of MWF-associated HP. The further development of sequence-based analytic techniques should assist in the prevention of this important occupational disease.
... Perhaps surprisingly, tests of cell-mediated immunity were not more informative than measurement of serum antibodies. An alternative explanation is that M. immunogenum was not responsible for the disease [33]. This interpretation is supported by finding tests for lymphocyte proliferation using avian antigens to be correlated with specific challenge tests to birds in a wide range of interstitial lung diseases caused by avian antigens [9•]. ...
Full-text available
Purpose of review: This review summarises the clinical knowledge of hypersensitivity pneumonitis in workers exposed to aerosols of metalworking fluid, reviewing published outbreaks and clinical cases. Recent findings: Metalworking fluid exposure has become the commonest recognised cause of occupational hypersensitivity pneumonitis, having been rare before 2000. There are many possible agents in the metalworking fluid which may be the cause of disease including bacteria, mycobacteria, fungae, biocides, emulsifiers, reodorants and dissolved chrome and cobalt. Causes are likely to be different in different outbreaks. Mycobacteria growing in the metalworking fluid have generated immune responses in some workers, but their role in disease causation is not yet established. Many outbreaks have been identified in large workplaces using common sumps. It is not possible to prevent microbial contamination of metalworking fluids in use. Disease prevention should focus on stopping inhalation of aerosols, particularly by re-engineering to remove recirculation.
... BERNSTEIN et al. [15] reported HP associated with the use of semisynthetic MWF, called ''machine operator's lung''. Aerobic Gram-negative rods, such as P. aeruginosa and Acinetobacter [13], fungi, and members of the Mycobacterium chelonae complex, including Mycobacterium abscessus, M. chelonae and M. immunogenum, have been identified in MWF [21][22][23][24]. Mycobacteria and Pseudomonas may also be identified in MWF by real-time PCR assays [25]. ...
... Subsequently, multiple genotypes of M. immunogenum have been isolated from diverse MWF operations in our recent efforts [6,7]. While M. immunogenum is considered as the etiological agent of the MWF-associated HP based on human epidemiological [8,9,10] and animal exposure studies [11,12], the exact mode of interaction of this pathogen with the exposed lung remains unclear. Also, the relative pathogenesis potential of individual genotypes of this pathogen prevalent in the occupational settings is not known. ...
Full-text available
Mycobacterium immunogenum is an emerging pathogen of the immune-mediated lung disease hypersensitivity pneumonitis (HP) reported in machinists occupationally exposed to contaminated metal working fluid (MWF). However, the mechanism of its interaction with the host lung is unclear. Considering that alveolar macrophages play a central role in host defense in the exposed lung, understanding their interaction with the pathogen could provide initial insights into the underlying immunopathogenesis events and mechanisms. In the current study, M. immunogenum 700506, a predominant genotype isolated from HP-linked fluids, was shown to multiply intracellularly, induce proinflammatory mediators (TNF-α, IL-1α, IL-1β, IL-6, GM-CSF, NO) and cause cytotoxicity/cell death in the cultured murine alveolar macrophage cell line MH-S in a dose- and time-dependent manner. The responses were detected as early as 3h post-infection. Comparison of this and four additional genotypes of M. immunogenum (MJY-3, MJY-4, MJY-12, MJY-14) using an effective dose-time combination (100 MOI for 24h) showed these macrophage responses in the following order (albeit with some variations for individual response indicators). Inflammatory: MJY-3 ≥ 700506 > MJY-4 ≥ MJY-14 ≥ MJY-12; Cytotoxic: 700506 ≥ MJY-3 > MJY-4 ≥ MJY-12 ≥ MJY-14. In general, 700506 and MJY-3 showed a more aggressive response than other genotypes. Chemical blocking of either p38 or JNK inhibited the induction of proinflammatory mediators (cytokines, NO) by 700506. However, the cellular responses showed a somewhat opposite effect. This is the first report on M. immunogenum interactions with alveolar macrophages and on the identification of JNK- and p38- mediated signaling and its role in mediating the proinflammatory responses during these interactions.
Background Antigen avoidance (AA) is essential for people with hypersensitivity pneumonitis (HP). An intervention programme to promote continuous AA in people with HP will be the focus of future research. Objectives We systematically analysed the AA behaviour of people with HP, interventions of health-care providers to promote AA behaviour, clinical outcomes after AA, and evaluation methods after AA. Methods We conducted a scoping review using six online databases and manual searches. Papers written in English or Japanese that reported cases on AA were selected. The extracted data were classified qualitatively. Results In total 205 cases included in 109 eligible papers were examined. Conclusions This review clarified the fundamental evidence of AA in people with HP. These people required the continuous support of health-care providers to continue appropriate AA. This review highlighted four aspects that require further research to design interventions for promoting effective and continuous AA in people with HP.
Hypersensitivity pneumonitis (HP), also known as extrinsic allergic alveolitis (EAA), represents a group of lung diseases caused by repeated inhalation of a specific antigen or antigens in susceptible individuals. The lung is the site of a variety of complex inflammatory reactions in response to endogenous or exogenous stimuli. In HP the inflammation represents a tissue reaction against various environmental agents that vary from thermophilic bacteria, mammalian and avian proteins, and fungi, to a large number of organic particles and, rarely, low molecular weight compounds. Diffuse inflammation of the small airways and the peripheral gas-exchanging portion of the lung are characteristic of HP.1 Although the terms hypersensitivity and allergic might suggest an atopic disorder, HP is not associated with eosinophilia or excessive production of immunoglobulin E (IgE). Hypersensitivity pneumonitis is characterized by a bronchioloalveolitis sustained by CD8+ cytotoxic T lymphocytes, interstitial granulomas, and, over time, the development of interstitial fibrosis. Hypersensitivity pneumonitis has a complex and poorly understood pathogenesis, and although it involves predominantly cell-mediated mechanisms, the production of specific antibodies directed against the offending antigens2 and nonimmunologic factors may play a role.
Background: In Korea, there is low awareness of the respiratory health problems caused by the use of humidifiers, leading to a lack of governmental measures. Objectives: The objectives of this study were to review cases of varying degrees of humidifier lung and fever in connection with the use of humidifiers and to summarize the probable environmental agents causing these cases. Methods: We searched all articles reporting on humidifier lung, humidifier fever, and humidifier diseases caused by the use of a humidifier both at home and in the workplace. Results: We summarized a number of cases of varying degrees of respiratory diseases resulting from inhalation of water mist of humidifiers containing various species of bacteria and fungi and their toxins. Type of respiratory disease connected with humidifier lung includes interstitial pneumonitis, hypersensitivity pneumonitis, fever and several respiratory symptoms. Non-tuberculous mycobacteria (NTM), Actinomycetes, endotoxins and contaminated humidifier water were the most commonly suspected probable environmental agents causing humidifier lung. In Korea, the use of humidifier biocide is suspected as a likely cause of fatal lung injury including death and lung transplantation. Conclusion: Governmental policy should be devised and measures including a national surveillance system should be taken to prevent humidifier lung caused by the use of humidifiers.
The diagnosis of extrinsic allergic alveolitis (EAA) is based on history of antigen exposure, symptoms and signs, and clinical findings. Some attempts have been made to present uniform criteria for diagnosis of EAA. These attempts have not been widely accepted. Usually, in the diagnostic criteria, much emphasis has been put on the results of precipitin tests. For reasons discussed previously in other context, a negative precipitin test cannot rule out the presence of EAA. The additional criteria are 1) basal crepitant rales audible on auscultation of the lungs; 2) impairment of the pulmonary diffusing capacity; 3) oxygen tension (or saturation) of the arterial blood either decreased at rest, or normal at rest but decreased during exercise; 4) restrictive ventilation defect in the spirometry; 5) histological changes compatible with EAA [Seal et al, 1968] in a biopsy specimen from the lung; 6) positive provocation test either by work exposure or by controlled inhalation challenge. The diagnosis can be considered confirmed if the patient fulfills all the main criteria and at least two of the additional criteria and if all other diseases with similar symptoms and clinical findings have been ruled out.
Occupational exposure to bacterial or fungal antigens has been associated with hypersensivity pneumonitis (HP), an immunologically-mediated pulmonary disease. Between August 1995 and April 1996, 34 employees working in machining and assembly areas of an engine manufacturing plant were clinically diagnosed with HP. Of these, 20 employees met an epidemiologic case definition. In a case-control study, no exposure variables, including duration and intensity of metal working fluid (MWF) exposure, were statistically associated with an increased risk of disease. Neither cases nor controls demonstrated precipitin reactivity against unused samples of the seven MWF and two biocides used in the plant. HP cases had a significantly higher prevalence of positive precipitin reactions to used oil soluble and synthetic MWFs. Reactivity to used but not unused MWF suggests a biocontaminant, probably bacteria or fungi, is the causative antigen in the development of HP in this setting. Am. J. Ind. Med. 35:58–67, 1999. Published 1999 Wiley-Liss, Inc.
This review describes antigenic and host factors of possible significance in the immunopathogenesis of hypersensitivity pneumonitis (HP). Although certain immunologic studies suggest immune complex mechanisms in HP, recent experimental and clinical data accumulated support a role for cell-mediated immunity. In addition, some data support roles for anaphylactic and cytotoxic antibody-mediated reactivity as well. One type of reactivity alone may not be sufficient for production of HP, and local pulmonary immune responses may be most relevant to the pathogensis. Whether immune damage will be produced in an exposed individual or not may depend on the characteristics of the antigenic exposure as well as inherited and acquired individual differences in immunologic reactivity and possibly target organ sensitivity.
This investigation was carried out to study possible differences in cell-mediated hypersensitivity between symptomatic and asymptomatic pigeon breeders. A variety of pigeon antigens were investigated using the indirect migration inhibition test. Cell-mediated hypersensitivity to pigeon antigens was detected in 6 of 8 symptomatic pigeon breeders, and in only 2 of 12 asymptomatic pigeon breeders. The results suggest that cellular mechanisms may be important in the pathogenesis of pigeon breeder's disease.
Many inhaled agents in the workplace can induce immunologic responses. Given the current state of clinical immunology, this article focuses on how to detect the body's response to those inhaled agents that behave as antigens. First the types of immunologic mechanisms that are activated in occupational lung disease are discussed, and the tests that reflect those states of immunologic activation. The second part of the article reviews major classes of occupational lung diseases and the immunologic tools that are available in making specific diagnoses.
In general, a history of exposure to "moldy" hay, birds, or other incriminated occupational or environmental inhalants in a patient with clinical and radiologic features consistent with HSP should lead to the demonstration of serum precipitins to the suspected antigen and an established diagnosis, confirmed by avoidance of the agent involved. Occasionally, other diagnostic procedures are required. The diagnosis is often difficult in domestic exposures, such as humidification and air conditioning systems. A careful environmental history is essential, and at times the physician must inspect the patient's environment personally. In most cases, the diagnosis is established if (1) the history and physical findings and pulmonary function tests indicate an interstitial lung disease, (2) the x-ray film is consistent, (3) there is exposure to a recognized cause, and (4) there is antibody to that antigen. In other exceptional circumstances, bronchoalveolar lavage may help. Biopsy is rarely needed. Special environmental studies and identification of new antigens require research facilities. Provocation tests are research procedures, not necessary for the diagnosis, and not needed in contested workmen's compensation adjudications.
In a large scale clinical survey of Scottish pigeon fanciers, 277 people completed a detailed questionnaire and provided a venous blood sample. There were 29 (10.4%) who fulfilled the clinical criteria for pigeon breeders' disease used in the study and 84 (30.3%) who showed a significant serum IgG antibody response to pigeon gammaglobulin (4 micrograms/ml) and were considered to have been sensitised. Increasing exposure was associated with a progressive tendency towards sensitisation, but the intensity of the antibody response was related to the presence of symptoms and not the degree of exposure. Thirteen out of 15 subjects in the survey with a serum concentration of antipigeon IgG greater than 60 micrograms/ml fulfilled the clinical criteria for pigeon breeders' disease, and those affected within 10 years of starting the hobby had the highest mean antibody response. Accurate quantitation of antibody response is therefore helpful in the investigation of pigeon breeders' disease. The radioimmunoassay provides a quantitative method for determining antibody response that can be used in the routine screening of pigeon fanciers and in serial monitoring of their response.
Serum IgG antibody against purified avian antigens was quantified by radioimmunoassay in 507 active pigeon fanciers, of whom 110 had documented Pigeon Breeders Disease (PBD). The incidence of PBD increased from 3.7% in the antibody-negative group to 78% of those with IgG antibody greater than 30 micrograms/ml. The highest levels of antibody tended to occur in subjects with relatively short avian exposure, and a significant correlation between decreasing antibody levels and increasing years of avian contact was demonstrated, which was unrelated to age or smoking history. The antibody -positive subjects had lower daily hours of avian contact and this group which contained most of the PBD subjects demonstrated some self-regulation of antigen exposure.