immunological study among grain...

Med. J. Cairo Univ., Vol. 62, No. 2, June : 365-374, 1994

Immunological Study Among Grain Workers

ANAN M. ELMISHAD, M.D.; WASSEF G. NICOLA, M.D.;

ZAKARIA M. ELKHIAT, Ph.D.; AHMED G. HIGAZI, Ph.D. and

ASHRAF A. SALEH, Ph.D.

The Depannents of Occupational Health & Industrkal Medicine,

Basic Medical Sciences (Endocrine & Physiology L&its),

Parm’tology andAnimal Diseare, National Research Center.

Abstract

‘Ihe JgE. IgG. specific precipitin and skin prick test (SFT). were done in

80 male wheat workers and 20 controls, to evaluate the iromuoological

effect of wheat dust. Wheat extract l/500 concentration was used for the

counter-immunoelectrophoresis (CIE) precipitation method, and wheat

extract of l/1000 concentration for the SPT. All subjects were clinically

examined and pulmonary function (FEV/FVC%) was done. The exposed

group was divided into four clinical subgroups. Asthmatics (18-22.5%),

chronic bronchitics (22-27.5%), other allergic manifestations (12-16.25%),

and asymptomatic group (28-33.75%). A significant elevation of both IgE

& IgG among the exposed group was found. IgE level among asthmatics

was significantly higher than the asymptomatic group and IgE level among

the atopic group (24-30%), was significantly higher than non-atopic group

(56-70%). There was no correlation hetweeo the degree of obstruction and

the level of IgE. 31 subjects of the exposed showed +ve precipitio with

varying degrees to wheat extract. No correlation between the degree of

precipitation and the level of IgE. 9 subjects out of the 31 +ve precipitio

cases has +ve SF’T. The aotigenic effect of grain dust was strongly suggest&

Results were discused. Recommendations for further studies to define the

exact allergen and to evaluate the relationship between the immune sod the

endocrine systems were suggested.

366 Anan M. El-Mishad, et al.

Introduction

OCCUPATIONAL allergic diseases

in the form of bronchial asthma and rhin-

itis due to hypersensitivity to flour, are

relatively common among bakers. In cross

sectional studies from recent years, preva-

lences of bakers asthma range from 6% to

1 l%, while work-related allergic rhinitis

was prevalent in approximately 20% [l,

2,3]. However, it was found that not all

workers’will asthmatic complaints reacts

positively to wheat extract in prick test or

in Radioallergos&bent test (RAST) [4].

The current studies indicate that many

subjects will develop an increase in bron-

chial responsiveness when exposed to

grain dust which is independent of pre-

exposure characteristics including atopy

[5]. It has been reported that farmers are

exposed to many classical inhalant aller-

gens and &E-mediated allergy is a poten-

tial disorder that should not be neglected.

In addition to pollens and animal danders

mites are potent allergens [6]. Allergy to

storage mites in the form of bronchial

asthma and allergic rhinitis has been

shown among farmers and grain workers

as well as bakers [4, 6, 7J. In a previous

study we demonstrated the effect of wheat

dust on the prevalence of respiratory

symptoms and signs and the effect on

lung functions [81. These effects were

suggested to be of an antigenic nature

which was potentiated by smoking. Some

investigators suggested a relationship be-

tween the immune system and the neuro-

endocrine axis among these patients, as

both systems are subjected to mutual regu-

lations [9].

The objective of the present study is to

evaluate the immunological manifestations

in a group of grain workers, specifically

wheat grinders. They were exposed to all

types of the grain dust starting from whole

grain dust to flour.

Materials and Methods

Subjects:

This study was conducted on 80 male

wheat workers. Their ages ranged between

22-59 years with mean 44.6 years and du-

ration of exposure between 5-33 years with

mean 12 years. A control group of 20

male subjects not exposed to any type of

dust. Their ages ranged between 20-56

years with mean 43.2 years. All examined

subjects were from the same socio-

economic stratum.

Methoak

All examined subjects were interviewed

using a simplified form of the Medical Re-

search Council questionnaire on respirato-

ry symptoms [lo]. Clinical examination

with special reference to the chest, skin

and upper respiratory tract allergic manifes-

tations was done.

Investigations:

Blood sample was drawn from each

subject for immunological study.

Total IgE evaluation by the Enzyme

L.inked Immunosorbent Assay (ELISA),

Immunological Study Among Grain Workers 367

using Enzygnost IgE micro kitts of Beh-

ring. The value of IgE was measured fol-

lowing the recomm&dation of the manu-

facturer and the results were expressed in

IU/ml. This method is an enzyme immu-

noassay for the determination of human

IgE & is based on the one step sandwich

principle [l 11.

Total IgG evaluation by the turbidi-

metric measurements of the maximum reac-

tion velocity (peak-rate method), using

turbiquant reagents kitts of Behring. The

value of IgG was measured following the

recommendation of the manufacturer and

the results were expressed in mg/dI [It].

Specific precipitin against wheat ex-

tract was measured using the Agar Gel

Diffusion (AGDD) method according to

Pepys 8c Lougbottom [13]. l/100 dilu-

tion of the wheat antigen was used. This

method gave weak results. The counter-

immunoelectrophoresis (CIE) was done

according to Glaussio et al. [14] using

wheat extract antigen in dilution of l/500

(this gave better results).

Skin Prick Test (SPT) was performed

on the anterior side of the forearm for each

subject using a separate needle. Wheat ex-

tract prepared from washed whole wheat

grains in a concentration of l/1000 was

used. Histamine solution was used as a

positive control. The test was read after

15-20 min. for all workers. A wheal

equals to or greater than (= >) 3mm in di-

ameter was considered positive. Subjects

were classified as atopic if the SPT is pos-

itive.

Pulmonary functions using the vicatest

4 respirometer for measuring FEV, Br

FVC. The FEV1/FVC % was calculated to

evaluate the presence of obstruction. A

value of FEV1fi7C% < 75% was consid-

ered obstruction.

Results

In this study-exposed and control

groups were found comparable as regard

age and smoking habits with no signifi-

cant differences. C)n the basis of the infor-

mations from the questionnaire and clinical

examination the exposed group was divid-

ed into four clinical sub-groups (Table I).

Asthmatic subgroup of 18 (22.5%) sub-

jects, diagnosed according to James et al.

151. Chronic bronchitic subgroup of 22

(27.5%) subjects, diagnosed according io

Dosman et al. [El, subgroup with allergic

manifestations other than chest, namely

rhinitis &/or eye irritation &/or skin irrita-

tion. This consists of 12 (16.25%) sub-

jects and an asymptomatic group of 28

(33.75%) subjects. Furthermore, the ex-

posed group was divided into atopic (24-

30%) subjects and non-atopic (56-70%)

subjects according to the SPT results.

According to pulmonary function namely

FEVl/FVC %, obstruction was present in

44 (55%) subjects.

On the other hand, the control group

consisted of 2 (10%) chronic bronchitics,

1 (5%) with allergic rhinitis, 2 (10%)


atopic with +ve SPT, and 17 (85%)

asymptomatic subjects.

Table (II) shows the immunological

values in the different groups. Statistical

evaluation was done between the exposed

group, as a whole, and the control group

separately, and between the asymptomatic

exposed group and the other three clinical

subgroups, also comparison between the

atopic and non-atopic groups was done.

There was a significant elevation of

serum IgE 8c IgG values of the exposed as

a whole compared to control (p < 0.001).

In the clinical subgroups the asthmatic

group only had a significantly higher val-

ues of IgE compared to the asymptomatic

group @ ~0.005). Again the IgE level for

the atopic group was significantly high

compared to non-atopic @<O.OOl), but the

difference between atopic and non-atopic

subjects of the same subgroup was not

significant. The correlation coefficient be-

tween the level of IgE and degree of ob-

struction among exposed group showed no

correlation (r = - 0.078). Table (II) also

shows the number of exposed subjects

with tve precipitin specific to wheat ex-

tract (according to the CIE method). 31

subjects (38.75%) showed tve precipitin

with different degree of positivety (l-4).

This method is a semiquantitative one. On

the other hand the AGDD method gave

only 5 tve cases to wheat antigen. The

principle disadvantages of double diffusion

without electromotive force are the time re-

quired for precipitation (24 hours) and the

relative lack of sensitivity. Counter-

immunoelectrophoresis can produce visible

precipitin lines within 30 minutes and is

approximately 10 times more sensitive than

standard double diffusion techniques. All

controls were -ve. Table (III) shows the re-

lation between the total serum IgE & the

CIE degree of precipitation of the 31 cases.

No correlation between the level of IgE

Table (1): Distribution of Examined Subjects According to Clinical Groups.

Clini. Groups Exposed (n = 80) Control (n = 20)

No. % No. %

Asthmatic 18 22.5 _

Chro. Bronchitis 22 27.5 2 10

Other Allerg. Manifes. 12 16.25 1 5

Asymptomatic 28 33.75 17 85

Atopic 24 30 2 10

Obstruction 44 55 _

Immunological Study Among Grain Worker:; 369

Table (2): Immunological Data Among Different Groups.

Group IgE Iu /ml IgG mg / dl

Precipitins +ve (CIE)

tve SPT

mean- SD mean-SD No % No %

Control

(n = 20)

106.5

t/-41

All Exposed *530.1

‘(n = 80) t / - 435.8

Asthmatics

(n = 18)

Chro. Bronchitis

(n = 22)

Other allergy

(n =m 32)

#730.5

t/-480.2

N.S.

428.2

t/-356

N.S.

407.1

+ / - 315.4

Asymptomatic

(n = 28)

415.7

Atopic

(n = 24)

X930.6

t / - 428.4

Non-atopic

(n = 56)

358.4

+ / - 307.3

947

t / - 118.7

*2845

t/-1114

N.S.

2620.6

t / - 909.6

N.S.

2742.3

t / - 907.6

N.S.

2840

t / - 1187.6

3172.5

2936.7

t I - 1080.3

2838

+I- 1066.4

31

8

7

4

12

9

22

. . 2 10

38.75 24 30

44.4 50

31.8

9

5

3

7

24

22.7

33.3 25

42.85 25

37.5

39.3

30

* Student t between exposed & control is significant @ < 0.001)

# Student t between asthmatic Bt asymptomatic significant @ c 0.05)

XStudent t between atopic & non-atopic is significant @ c 0.001)


Table (3): Relation Between Total IgE & the Degree of Precipitation in the Precipitin +ve EX- posed Workers

No. Clinical Group IgEIu/ml C:IE / Degree tve Skin

of tve lest

1

2.

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30

31

Asthmatic-s

Chro. Bronchitis

Other allergy

Asymptomatic

290 t

1100 ++ 250 -I-

450 t

190 ++++

365 ++++

240 +tt+

1150 t

210 tttt

210 t

120 t

290 t

290 ++t+

200 +

400 +++

1250 t

280 t

240 ++++

240 t

1600 t

220 t

330 tt

290 t

1050 ++++

290 t

610 +

220 tt

240 t

1180 t

1120 +++

210 +++

__

+ __

t __

+ t t __

mm

_-

__

t

__

__

__

__

__

_-

t

__

_-

__

+

_-

t

_-

No correlation between the level of IgE and degree of precipitation (r = -0.1126).


and degree of precipitins (I = -0.1126). 9

subjects only of those with tve precipitin

to wheat antigen showed tve SPT (29%).

Discussion

The role of allergic factors in grain

dust induced pulmonary disease remains

unclear [16, 17. A wide variety of mate-

rials are found in grain dust such as fun-

gal spores, mites, insects or their remnants,

rodent hairs & pesticides, many of which

have the potential to induce disease IS,

16). The high level of IgE & IgG @‘T’

O.OOl), and the high prevalence of atopy

(30%) among grain workers in this study

suggests very strongly that the increase in

bronchial responsiveness may be induced

by allergic mechanism [5& This was also

reported by others. Moselhi et. al. [18]

observed higher levels of gamma globulin

in the exposed workers. He found signifi-

cant increase in the IgE & IgG @ < 0.01)

among grain workers as compared to the

controls. VanHage-Hamster et al. [q re-

ported a high prevalence of allergy with a

positive RAST to storage mites among

farmers. In’ his study he also found that

the atopic farmers had significantly elevat-

ed to non-atopic farmers with the same

symptoms. However,we found high sig-

nificant level of IgE among atopic com-

pared to non-atopic subjects but this was

not found between atopic and non-atopic

of the same clinical group. Again we

found significant high levels of IgE in the

asthmatic group (p eO.05) when compared

to asymptomatic grain workers. Also the

per-cent of atopy among asthmatics was

significantly higher (50%) than the other

exposed subgroups. All these results sug-

gest an immunological reaction. A prelim-

inary data from an uncontrolled study

among grain handlers in Britain have

shown that positive SPT were significant-

ly correlated to work related symptoms,

and that 62% of the asthmatics had posi-

tive SPT [?I. This goes with our find-

ings as well. 55% of our grain workers

have obstruction but no correlation found

between the degree cf obstruction and lev-

el of I@ (r=- 0.078) which may lead to

the assumption that there is an inflammato-

ry effect contributing as well in bronchial

hyper-responsivenessc.. Also it was found

that grain dust contams many agents that

can induce inflammation and increase bron-

chial responsiveness [19].

The significant increase of IgE in our

subjects exposed to wheat dust is a good

proof that there is an antigenic effect of

wheat dust. This was documented in 31

(38.75%) of exposed by positive precipi-

tin to wheat antigen and about 30% had

tve SPT to wheaat extract. However, there

was no correlation between the degree of

precipitation and the level of I@ in the

positive cases and only 9 (29%) of the

positive subjects had +ve SPT. This goes

with what was found by many authors [I,

2, 41. Not all bakers wifh workrelated

asthmatic complaints in their studies react

to extracts of wheat flour in prick test or

in RAST. The results of SPT will depend

372 Anan M. El-Iv&had, et al.

on several factors other than individual

sensitivity such as the quality and poten-

cy of the allergen extract used, as well as

the age and sex of subjects studied.

It is well known that IgE has no role

in antimicrobial immunity and is common-

ly produced against antigens to which the

individual is allergic and participates in

the production of allergic symptoms. Also

the significant increase of IgG might indi-

cate secondary antibody response which

could be due to the increased prevalence

of respiratory infection among exposed

workers as compared with the controls

WJI.

The term storage mites is the ordinary

name for a variety of nonpyroglyphid

mites which are often encountered in the

ecological studies in hay, straw, grain and

other vegelabies [4]. This however was

condemned by many authors to be the

cause of allergic manifestations reported

among farmers and grain workers as well

13, 6, 7, 211. Most of them reported al-

lergy to storage mites in the form of asth-

ma, hay fever, and allergic rhinitis among

their grain workers. At the same time it

was stated by Tee et al. [22], that stor-

age mites are widely distributed in the en-

vironment and are not of special sign&

cance in allergic responses in bakery

workers exposed to flour.

Very recently, it has been postulated

that evidences are gathering pionting out

the relation between the immune system &

the neuroendocrine system t231. This

may be the subject of a future study.

We suggest that further studies are re-

quired to determine which allergens in

grain dust are responsible for allergic dis-

eases.

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