Cytogenetic monitoring of farm animals under conditions of environmental pollution

Mutation Research, 283 (1992) 199-210

199

© 1992 Elsevier Science Publishers B.V. All rights reserved 0165-7992/92/$05.00

MUTLET 0723

Cytogenetic monitoring of farm animals under conditions of environmental pollution J. Rubeg, L. Borkovec, Z. Ho[inovfi, J. Urbanovfi, I. Prorokovfi and L. Kullkovfi Veterinary Research Institute, 62132 Brno, Czechoslol:akia

(Received 16 January 1992) (Revision received 24 June 1992) (Accepted 23 July 1992)

Keywords: Cattle; Pigs; Horses; Chromosomal aberrations; Peripheral blood lymphocytes;Cytogeneticmonitoring; Polluted habitat

Summary Cytogenetic examinations were carried out in 13 cattle farms, two herds of horses, one stag farm and 13 pig farms in areas with different levels of environmental contamination. The frequency of aberrant cells per 100 mitoses was 3.67 _+ 1.89 in pigs (n = 260) and 4.16 _+ 2.4 in herbivores (n = 497). This is a significant difference ( p < 0.01). Ten times higher frequencies of chromatid exchanges were found in pigs. The examined herds were classified into three groups by the level of environmental contamination (satisfactory, impaired and severely impaired environment). Significant differences in aberrant cell counts were recorded between the groups of herbivorous animals. Significant differences in pigs were recorded only between herds in satisfactory and severely impaired environments. Significantly higher frequencies of aberrant cells were found in farms of herbivorous animals in the industrial area of Pardubice compared with findings in the South Moravian agricultural area (4.7% and 3.1% respectively). The effect of local contamination sources on farm environment was also investigated. A cattle farm located in the vicinity of a large chemical plant was examined five times at 6-month intervals. An autumn examination rewealed significantly higher frequencies of aberrant cells compared with the spring examination.

Farm animals can be affected by environmental contamination caused by genotoxic agents, resulting from the excessive use of chemicals in agriculture, and by emissions and waste products of industrial plants. Animals reared in contaminated environments are directly exposed to these contaminants and, in addition, herbivores may be exposed via contaminated feed. The effect of

Correspondence: Dr. J. Rube~, Veterinary Research Institute, 621 32 Brno, Czechoslovakia.

locality is even stronger in cattle due to the feeding of roughage. The level of environmental contamination to which these animals are exposed can be correlated with the levels of chemical residues in tissues and products of animal origin. Serious exposure to genotoxic agents may result in mutations, metabolic disorders, reduced fertility, immunosuppression, etc. Our study focused on the level of chromosomal aberrations in farm animals in various parts of the Czech Republic. Special attention was paid to places where the impact of chemicals used in

200 agriculture is augmented by industrial production. Material and methods

Thirteen cattle farms, two herds of horses, one stag farm and 13 pig farms were investigated in the period 1983-1991. The location of the farms is given in Fig. 1. The ecological situation of the areas where the farms are located is roughly characterized by classifying the level of environmental contamination into the following five categories: (A) environment with minimum contamination, (B) satisfactory environment, (C) impaired environment, (D) severely impaired environment, (E) extremely impaired environment (see Fig. 2). The categories were defined by their pollution burden (sulfur dioxide, airborne dust and other harmful substances) and the degree of landscape damage, surface devastation, immission-damaged forests, etc. (Moldan, 1990). Further, each locality was characterized by the possible presence of a source

of contamination in the immediate vicinity (Tables 1, 2). Two groups of cattle and horse farms were compared. Farms of the first group were in the district of Pardubice (Table 1, herds 9, 10, 11, 12 and 15) which has a high level of emissions from two solid-fuel power stations and a chemical plant. The second group of farms (Table 1, herds 2, 3, 4, 6 and 7) was in the South Moravian region, which is an agricultural area with a relatively low level of industrial pollution. On the other hand, the character of agricultural production and the structure of plants in this region let us presume that there was a higher environmental contamination with pesticides. The difference is not relevant, however. Fig. 3, based on official data published in Balance of Emissions of Pollutants for 1990, shows the production of significant emissions (solid particles, SO 2, NOx, CO, CxHy) in both areas under study in 1990. Cattle of the farm at (~ernfi, located in the immediate vicinity (about 2 kin) of a large chemical plant (production of dye stuffs, semi-products

lt.

O <~ /X []

CATTLE PIGS HORSE DEER Fig. 1. Locationof the farms investigated.

201

for plastic materials and drugs, industrial fertilizers and inorganic acids), in which the highest frequencies of aberrant cells were found, were inw'.stigated 5 times at 6-month intervals in spring and autumn. Cattle from the farm, at Svin(:any, situated in a relatively less contaminated part of the region, about 10 km from a chemical plant, were investigated at the same time. Findings from both farms were compared.

Cytogenetic im,estigation Blood samples were taken from clinically healthy animals, who had been riving in the herd for at least 1 year, and who had had no treatment during the last 3 month~. 1 ml of whole heparinized blood was added to 8 ml of RPMI 1640 medium (Usol, Prague), con-

taining 20% fetal calf serum, phytohemagglutinin H 15 (Wellcome) and glutamine. All cultures were incubated at 38°C for 48 h. Colchicine, 100 /~g/ml, was added to the medium 45 rain before the end of incubation. Slides for microscopy were prepared by the air-drying method and stained using Giemsa stain. In each animal, 100 metaphases were examined for chromosomal aberrations. Gaps were not counted as aberrations. The classification of aberration types by Savage (1978) and Carrano and Natarajan (1988) was adopted. Major types are defined as follows. Chromatid break, break in a chromatid greater than the width of the arm or displaced. Chromosome break, break involving both sister chromatids at the same position and acentric fragments.

"--rU ~"

w.~,,

s t,..

•

~-

¢

[ A ! Environment with - - J minimum contamination

E Impaired environment

B

D Satisfactory environment

'

E

~

Extremely impaired environment

Severely impaired environment

Fig. 2. Map of the environmental quality in the Czech Republic (according to Moldan, 1990).

Malhostovice Zaje61 Uh~ice

Grygov Sokolnice Nov~ Dvory

Val. Mezi~i(:i Svin~:any Brozany Kladruby

Slatifmny

Dobranov

Holany

Cernfi

Byst~any

2 3 4

5 6 7

8 9 10 11

12

13

14

15

16

cows

cows

cows

cows

horses

cows cows cows horses

bulls cows cows

cows cows cows

deer

Animal

24

111

23

7

18

40 108 24 27

10 30 11

15 12 17

20

2400

11077

2300

700

1 800

4000 10850 2400 2 650

1000 3000 1 200

1 500 1 200 1 680

2000

Number of cells

Number of animals

4.25_+2.56

5.65 + 3.07

3.46+ 1.96

4.57_+1.99

2.78+ 1.70

4.23+ 1.84 4.32+2.00 3.13_+1.42 5.11 + 1.96

2.20+0.25 2.67_+1.30 3.72+0.96

3.00_+1.51 1.92+ 1.08 4.44+2.19

2.25+ 1.45

% (mean+SD)

Aberrant cells,

Number of:

76 (3.17)

517 (4.67)

61 (2.65)

16 (2.29)

38 (2.11)

156 (3.90) 417 (3.84) 60 (2.50) 127 (4.79)

19 (1.90) 67 (2.23) 41 (3.42)

41 (2.73) 18 (1.50) 64 (3.81)

42 (2.10)

Chromatid breaks ~

35 (1.46)

135 (1.22)

19 (0.83)

16 (2.29)

12 (0.67)

20 (0.50) 83 (0.77) 16 (0.67) 46 (1.74)

3 (0.30) 23 (0.77) 3 (0.25)

5 (0.33) 5 (0.42) 12 (0.71)

1 (0.04)

3 (0.03)

0

0

0

0 2 (0.465) 0 0

0 1 (0.03) 0

0 0 0

0

exchange ~'

breaks ~ 3 (0.15)

Chromatid

Chromosome

Data in parentheses indicate the number of aberrations per 100 cells. b SM, South Moravia; CB, Central Bohemia; NM, North Moravia; NB, North Bohemia; EB, East Bohemia.

Vimperk

1

Farm

0.0470

0.0594

0.0347

0.0457

0.0277

0.0439 0.0465 0.0316 0.0652

0.0220 0.0307 0.0366

0.0306 0.0192 0.0452

0.0225

per cell

Breaks

E

D

C

B

A

Area

RESULTS OF CYTOGENETIC E X A M I N A T I O N OF CATTLE, HORSES AND D E E R IN THE YEARS 1987-1990

TABLE 1

NB

EB

NB

NB

EB

NM EB EB EB

NM SM SM

SM SM SM

SB

Region b

Teplice

Pardubice

(;eskfi Lipa

(~esk~ L[pa

Chrudim

Vsetln Pardubice Pardubice Pardubice

Olomouc Brno Brno

Brno B~eclav Blansko

Prachatice

District

Chemical factory

Uranium ore and processing

Thermal power station

Coal tar factory

Fireclay factory

Plants close to the farm

Dubfiany Dubfiany D. Mogt~nice Nov4 Dvory Rajhrad Val. Mezi~[c[

Res. Inst. Brno

Skrgfn Machnin

Dobranov

20 21 22 23 24 25

26

27 28

29

boars

minipigs boars boars

fatt. pigs boars sows boars boars sows

sows boars boars

Animal

10

19 18

19

30 10 26 31 10 13

31 10 33

Number of animals

1000

1900 1800

1 900

3000 1000 2600 3100 1000 1350

3100 1000 3 300

Number of cells

3.80+ 1.69

3.63 + 1.57 3.83_+ 1.62

3.59 ± 2.28

5.03_+2.40 3.50+2.12 2.75 + 1.38 3.65+2.09 3.90_+ 1.73 2.62+ 1.33

3.58+ 1.59 2.20± 1.40 3.28 + 1.17

Aberrant cells, % (mean ± SD)

(5.10) (3.10) (4.19) (2.90) (3.40) (2.37)

30 (3.00)

63 (3.32) 51 (2.83)

54 (2.84)

153 31 109 90 34 32

84 (2.70) 20 (2.00) 99 (3.00)

8 (0.80)

8 (0.42) 20 (1.11)

12 (0.63)

28 (0.28) 2(0.20) 29 (1.12) 21 (0.68) 7 (0.70) 5 (0.37)

25 (0.81) 2 (0.20) 11 (0.30)

N u m b e r of: ChromaChromotid breaks '~ some breaks a

a Data in parentheses indicate the number of aberrations per 100 cells. b SM, South Moravia; CB, Central Bohemia; NM, North Moravia; NB, North Bohemia.

Tavikovice Oblekovice Klim~tice

17 18 19

Farm

RESULTS OF C Y T O G E N E T I C E X A M I N A T I O N OF PIGS IN T H E Y E A R S 1983-1990

T A B L E 2-

C

0.0730 0.0390 0.0576 0.0370 0.0410 0.0274

19 (0.63) 3 (0.30) 6 (0.23) 2 (O.O6) 0 0

NB NB NB

0.0347 0.0373 0.0394 0.0420

0 0 2 (0.20)

(~eskA Lfpa

Most Liberec

Brno city

Hodon[n Hodonfn Hodonfn P~fbram Brno Vsetfn SM SM SM CB SM NM SM

Znojmo P~fbram

Znojmo

District

SM SM CB

Region b

0

D

B

0.0390 0.0220 0.0333

Area

6 (0.19) 0 0

Chromatid exchange a

Breaks per cell

Coal tar factory

Thermal power station Thermal power station Thermal power station

Plants close to the farm

204 NOx

CO

SO 2

Statistical methods

b

1:3.

o

b

CxHij

~

a

a

b

b

Data were processed on a PC using a modular statistical and graphical set of programs. The compared sets of data were represented by values (% AB.B.) obtained from individual animals. Basic statistical characteristics were determined for each set. Analysis of variance was followed by a simple sorting procedure and comparisons by Schaffe's method of contrast. The t-test was used after the determination of variance homogeneity by the F-test, when two sets were compared.

solid particles

Results and discussion

o_

a

b

a

o

Fig. 3. Amount of emJssions in the Pardubice district (a) and South Moravia region (b) in 1990.

Chromatid exchange, exchange between chromatids of different chromosomes.

Karyotypic characteristics of examined species Species

2n

Pig (Sus scrofa) Cattle (Bos taurus) Horse (Equus cabal&s) Red deer (Ceruus elaphus)

38 60 64 68

Chromosome arms male

female

64 62 91 71

64 62 92 70

The results of the cytogenetic examinations are summarized in Tables 1-3. The frequency of aberrant cells in 260 pigs was 3.67 +_ 1.89% and in 497 herbivores 4.16_+2.4%. The difference in frequencies of aberrant cells between herbivores and pigs is significant ( p < 0.01). Fig. 4 shows the distribution of frequencies of aberrant cells in both groups. In areas in which both cattle and swine were examined parallelly, higher percentages of aberrant cells were found in cattle located in the districts Vsedn and Ceskfi L~pa, the difference between cattle and swine being highly significant in the former and non-significant in the latter district. Frequencies of different types of aberrations are shown in Table 3 and Fig. 5. No other types of aberrations were found except for breaks and chromatid exchanges. A 10 times higher frequency of chromatid exchanges (0.146%) was recorded in pigs as compared with cattle (0.014%). It is necessary to point out that in pigs the frequency of reciprocal translocations is much higher than in other species and that

TABLE 3 F R E Q U E N C I E S OF A B E R R A T I O N S IN H E R B I V O R E S AND PIGS Cattle

Horses

Deer

Herbivores

Pigs

Number of animals Number of cells Aberrant cells per 100 Aberrations per 100 cells Chromatid breaks Chromosome breaks Chromosome exchanges

432 43 307 4.25 +_44

45 4 450 4.14 _+ 16

20 2 000 2.25 + 1.45

497 49 757 4.16 _+2.41)

260 26 050 3.67 + 1.89

3.59 +_21 0.87 + 1.12 0.016 ± 13

3.71 _+57 1.30 + 1.16 0

2.10 + 145 0.15 ± 0.49 0

3.54 _+2.24 0.88 _+ 1.12 0.014 ± 12

3,38 + 2.37 0,68 ± 0.97 0.146 ± 0.43

Total

4.48

5.01

2.25

4,43

4.21

205

they are mostly of de novo origin (Gustavsson et al., 1983; Gustavsson, 1988). Long (1991) suggests as one cause a higher content of genotoxic agents in the environment of pigs. Our results indicate that pigs are less exposed to mutagens than herbivorous animals; however, chromatid exchanges are more frequent in the former species. A higher susceptibility to the formation of exchanges observed in somatic cells might also occur in meiotic cells, resulting in higher frequencies of gametes with reciprocal translocations. It is generally accepted that a part of cells affected with chromosomal aberrations is eliminated in each cell generation. Therefore, between-species differences in cell cycle kinetics must be taken into account when differences

100= 90= 80g 70~ 60~ 50~ 40g 30% 20g 10~ O=

1

2

3

4

5

6

tm"l A

7 8 g farm i I B m

10 11 12

13 14 15

16

C

c]

36

33 % 30

27

24~

.........

17

18

19

20

21

IT~ A

6 0

,~.

22 23 24 farm r"-1'8 m C

25

26

27

28

29

Fig. 5. Frequencies of single types of chromosome aberrations in herbivores (a), pigs (b). A, chromatid breaks; B, chromosome breaks; C, chromatid exchanges. The numbers of farms correspond with those given in Tables 1 and 2.

0.00 2.00 4.00 6.00 8.0010.00120014.0016.0018.00 Interval b 40. 36. % 32. 28. 2420161 2-I

if

000

200

400

600

8 0 0 1000 12'00 1A'00

Interval Fig. 4. Histogram of distribution of aberrant cells (mean %),

for individual animals. (a) herbivores, (b) pigs.

between cattle and swine in aberrant cell frequencies are evaluated. In our laboratory, approximately 80% of first metaphases are found in pigs after 48 h of incubation. This value corresponds with that published by Lezana et al. (1978) but is higher than that reported by Bianchi et al. (1981). Approximately 90% of first metaphases are found in our laboratory in cattle after 48 h of cultivation when differential staining of sister chromatids by the FPG method is used. Our findings contradict those of Modave et al. (1982) who reported 34 and 84% of first metaphases after 48 and 32 h of cultivation respectively. Only sporadic mitoses were observed in our experiments after 32 h of cultivation and the mitotic index decreased rapidly when the cultivation period was shortened to below 48 h. Also in horses

206 TABLE 4 RESULTS OF CYTOGENETIC EXAMINATIONS OF BOARS AND SOWS Area

Boars Aberrant cells, % (mean + SD)

Number of animals

Aberrant cells, % (mean _+SD)

43 51 47

3.03 + 1.30 3.67 + 1.99 3.74 ± 1.58

31 24 19

3.58 _+1.59 3.25 +_1.42 3.58 +_1.74

141

3.50 _+1.69

74

3.74+ 1.56

Number of animals B C D Total

Sows

a p p r o x i m a t e l y 90% of first m i t o s e s are f o u n d in o u r l a b o r a t o r y a f t e r 48 h of cultivation. A comp a r i s o n of the lengths of l y m p h o c y t e cycles in various a n i m a l species was p u b l i s h e d by L e o n a r d et al. (1982a). T h e d i f f e r e n c e s in species could also be d u e to d i f f e r e n c e s in t h e ability o f the d i f f e r e n t species to m e t a b o l i z e genotoxic a g e n t s into reactive species a n d / o r to r e p a i r D N A d a m a g e . T j c t t a a n d A u n e (1991) e x a m i n e d d i f f e r e n c e s in the activation o f c o o k e d f o o d p r o m u t a g e n s by rat, porcine, a n d ox liver a n d lung enzymes. T h e highest a n d the lowest activities w e r e f o u n d for ox and p o r c i n e liver enzymes, respectively. T h e ox e n z y m e s s h o w e d the highest individual variability, however. L e o n a r d et al. (1982b), who s t u d i e d the yield o f r a d i a t i o n - i n d u c e d c h r o m o s o mal a b e r r a t i o n s , f o u n d a h i g h e r radiosensitivity o f p e r i p h e r a l lymphocytes in c a t t l e t h a n in swine. N o significant d i f f e r e n c e in AB.B. counts was f o u n d b e t w e e n m a l e a n d f e m a l e swine. T h e set of a n i m a l s i n c l u d e d b r e e d i n g b o a r s a n d sows but no f a t t e n i n g pigs a n d c a s t r a t e s (see T a b l e 4). N o similar c o m p a r i s o n was possible in cattle, b e c a u s e the set i n c l u d e d 422 cows, b u t only 10 bulls. T h u s the results reflect r a t h e r the situation in females. N o significant b e t w e e n - g e n d e r d i f f e r e n c e was d e m o n s t r a b l e in horses but, again, 38 m a r e s a n d only seven stallions w e r e e x a m i n e d . A l l e x a m i n e d d e e r w e r e males. A m o n g - b r e e d d i f f e r e n c e s in AB.B. counts could be a n a l y z e d in swine only. R e s u l t s o f examinations of various p u r e b r e d b r e e d s are shown in T a b l e 5. N o significant a m o n g - b r e e d d i f f e r e n c e was found. T h e rest of the pigs w e r e crosses of the b r e e d s given in the table. T h e B o h e m i a n S p o t t e d b r e e d d o m i n a t e d with 60% in the set of

cattle. T h e rest w e r e crosses of this b r e e d with 1 0 - 5 0 % o f R e d H o l s t e i n , Black P i e d L o w l a n d or A y r s h i r e blood. N o r e l e v a n t d i f f e r e n c e s in b r e e d s t r u c t u r e existed b e t w e e n the a r e a s u n d e r study. In a g r o u p of cows in D o b r a n o v ( T a b l e 1, Fig. 5, h e r d 13) significantly h i g h e r f r e q u e n c i e s of chromosomal breaks were detected, compared with cows from o t h e r farms. This might be the result o f m i n i n g a n d p r o c e s s i n g of u r a n i u m o r e in the a r e a o f D o b r a n o v . T h e lack of dicentrics n e e d s to be e x p a n d e d to indicate that, thus, radia t i o n is p r o b a b l y not the cause of the increase. T a b l e s 1 a n d 2 a n d Fig. 6 show the p e r c e n t a g e of a b e r r a n t cells in a r e a s classified into c a t e g o r i e s A - E a c c o r d i n g to t h e level of e n v i r o n m e n t a l cont a m i n a t i o n . T h r e e , six a n d f o u r pigs farms in c a t e g o r i e s B, C a n d D, respectively, w e r e investig a t e d . T h e pigs in c a t e g o r y D had statistically h i g h e r f r e q u e n c i e s of a b e r r a n t cells t h a n those in c a t e g o r y B. N o statistical d i f f e r e n c e was r e c o r d e d b e t w e e n B a n d C, a n d C a n d D. T h r e e , eight a n d t h r e e h e r d s of h e r b i v o r e s in c a t e g o r i e s B, C a n d D, respectively, were comp a r e d . Significant d i f f e r e n c e s were f o u n d beTABLE 5 RESULTS OF CYTOGENETIC EXAMINATIONS OF VARIOUS BREEDS OF SWINE Breeds

Number of animals

Aberrant cells, % (mean + SD)

Large White Duroc Hampshire Landrace Czech meat pig

16 11 26 47 69

3.69 + 1.35 3.82 _+2.60 3.65 _+1.38 3.79 + 1.76 3.22 + 1.58

Sampling time

IV 89 IV 89

X 89 X 89

IV 90 IV 90

X 90 X 90

IV 91 IV 91

Farm

(~ern~ Svin~:any

Cern~ Svin~any

(~ern~ Svin6any

(~ern~ Svin6any

(~ern~ Svin~:any

25 25

23 22

23 23

20 20

20 18

Number animals

2500 2500

2300 2250

2277 2300

2000 2000

2000 1 800

Number of cells

3.92 ± 2.12 5.24 _+2.70

8.30 _+2.91 5.18_+2.22

4.61 + 1.62 3.96 _+ 1.69

8.45 + 2.46 4.05 _+ 1.43

3.15 _+ 1.04 4.33 +_ 1.94

Aberrant cells, % (mean _+SD)

84 (3.36) 116 (4.64)

141 (6.13) 93 (4.13)

87 (3.82) 77 (3.35)

149 (7.45) 68 (3.40)

56 (2.80) 67 (3.72)

Number of: Chromatid breaks

17 (0.68) 15 (0.60)

57 (2.48) 26 (1.16)

19 (0.83) 17 (0.74)

33 (1.65) 13 (0.65)

7 (0.35) 12 (0.67)

Chromosome breaks

2 (0.080) 1 (0.040)

0 0

1 (0.044) 1 (0.043)

0 0

0 0

Chromatid exchanges

0.0420 0.0532

0.0860 0.0529

0.0474 0.0395

0.0910 0.0405

0.0315 0.0439

Breaks per cell

RESULTS OF CYTOGENETIC EXAMINATION OF COWS B R E E D I N G CLOSE TO A CHEMICAL FACTORY (Cernfi) AND OF COWS F R O M A CONTROL FARM

TABLE 6

208

tween herds B and C, C and D, and B and D, the significance levels being p < 0.05, p < 0.01 and p <0.01 respectively. This could be due to a closer contact of the cows with the locality via the feeding of roughage. Feeding mixtures for pigs in the Czech Republic might contain heavy metals, moulds, PCB and pesticides (Rube~, 1987). Further we compared findings from cattle and horse farms (Table 1, Figs. 1, 7 herds 15, 9, 10, 11, 12) in the industrial area of Pardubice with those from farms located in the South Moravian agricultural area (Table 1, Figs. 1, 7, herds 6, 7, 2, 3, 4). A highly significant difference was found ( p < 0.01) for the frequency of aberrant cells in animals of the two regions, the values being 4.7 _+ 2.6% in the industrial and 3.1_+ 1.7% in the agricultural areas. No significant difference in aberrant cell counts was found between horses and cattle in the Pardubice district (4.14_+ 2.16 and 4.80 _+ 2.65 AB.B. per 100 mitoses, respec-

oJ

15

10 ~n~n

9

98 7'

I 6

s ¸

2 1 2

3

4

5

6

Deer

m

10 11 12 form cottle [~] hor~e~

13 14 15

16

g

-

17

18

19

20

21

22

23 2 4 farm

25

12

,0U I

2 form

I

3

4

Fig. 7. Aberrant cells (%) in two different areas, (a) district of Pardubice, (b) South Moravia.

n

O

I 7

11

26

27

28

Fig. 6. Aberrant cells (%) in herbivores (a) and pigs (b).

29

tively). The results indicate that industry in the Pardubice region (chemical industry, thermal power stations, oil processing) results in a high exposure of farm animals to genotoxicants and a risk of food chain contamination. Svandovfi and R6ssner (1989) found 1.75% aberrant cells when evaluating 1201 occupationally non-exposed men from various regions of the Czech Republic. Groups of persons were also examined in the two areas under study. Though aberrant cell frequency was higher in the Pardubice district than in the South Moravian region, the difference was not significant. We performed an extensive study on animals of the farm, at (~ernfi, near the largest source of pollutants in the area, i.e., a complex of chemical plants in a suburb of Pardubice. A farm located in the vicinity of the chemical plant was examined at 6-month intervals, together with a control. The

209

gl 8 7 6 5 4 3 2 1 0

X89

IVB9

Ivgo X90 Samplrn~_ time m~ Svin~any

1

~

--

aveFage lev6[

IV91

. . . . oveFage leve[

Fig. 8. Aberrant cells (%) in farms at Cernfi (close to a chemical factory) and Svin6any(control). A winter-dependent decrease in chromosomal damage in Cernfi is visible.

results shown in T a b l e 6 a n d Fig. 8 indicate seasonal variations in the frequencies of a b e r r a n t cells in the herd. A n a u t u m n e x a m i n a t i o n revealed a highly significant increase in the freq u e n c i e s of a b e r r a n t cells, which d r o p p e d again in winter. T h e variations were observed also in cows which we had the o p p o r t u n i t y to e x a m i n e r e p e a t e d l y in the course of 2.5 years (Fig. 9). No seasonal difference was observed in the control. This might be explained by the feeding of r o u g h a g e c o n t a m i n a t e d with emissions or waste waters from the chemical p l a n t d u r i n g the summer. In winter, w h e n silage is fed, a m a r k e d decrease of genotoxicants in feeds can be expected, caused by flow-off of silage juices or b i o d e g r a d a t i o n . No seasonal variations were ob-

11 10 9 8

g ~

7 6 5

~

3 2 1 0

47 208

47 032 r ~ IV/89 1 X/S0

6g 874 47 586 47 392 a n ~ l number I X/89 Ez~ IV/90 rm IV/S1

38 407

Fig. 9. Aberrant cell counts in cows examined repeatedly in the farm at (~ernfi.

served in any animal species in any other area. Cows of the farm at (~ernfi had constantly lower c o n c e p t i o n rates, indicating hazardous effects of chemical plants on agricultural production. Because of the greater n u m b e r of data detailed analyses of this case will be published elsewhere. U n f o r t u n a t e l y , c o m p a r a b l e results from other countries are not available. Occasional blood samples t a k e n in other countries and e x a m i n e d in our laboratory have revealed 2.05 a n d 3.36% of a b e r r a n t cells in cows in Sweden, 2.84% in F r a n c e a n d 3.3% in P o l a n d ( u n p u b l i s h e d data). W e propose that the m o n i t o r i n g of chromosomal a b e r r a t i o n s in farm animals is a suitable m e t h o d for the assessment of the hygienic level of herds with possible exposure to genotoxicants. T h e resulting data could be used as a basis for necessary m e a s u r e s to prevent food chain contamination.

References Anonymous (1991) Balance of Emissions of Pollutants for 1990, Czech Air-Protection Agency, Prague. Bianchi, M.S., N. Bianchi, M. Larramendy and J. Garcia-Heras (1981) Chromosomal radiosensitivity of pig leucocytes in relation to sampling time, Mutation Res., 80, 313-320. Carrano, A.V., and A.T. Natarajan (1988) Considerations for population monitoring using cytogenetic techniques, Mutation Res., 204, 379-406. Gustavsson, 1. (1988) Reciprocal translocations in four boars producing decreased litter size, Hereditas, 109, 159 168. Gustavsson, I., I. Senergren and W.A. King (1983) Occurrence of two different reciprocal translocations in the same litter of domestic pigs, Hereditas, 99, 257-267. Leonard, A., G. Decat and L. Fabry (1982a) The lymphocytes of small mammals. A model for research in cytogenetics?, Mutation Res., 95, 31-44. Leonard, A., L. Fabry, G. Deknudt and G. Decat (1982b) Chromosome aberrations as a measure of mutagenesis: cytogenetic extrapolation from animal to man, Cytogenet. Cell Genet., 33, 107-113. Lezana, E.A., M.S. Bianchi and N.O. Bianchi (1977) Kinetics of division in PHA-stimulated pig lymphocytes, Experientia, 34, 30-31. Long, S.E. (1991) Reciprocal translocations in the pig (Sus scrofa), Vet. Rec., 128, 275-278. Modave, C., L. Fabry and A. Leonard (1982) Cin6tique cellulaire et radiosensibilitfi des lymphocytes de vache en culture, C.R. Seances Soc. Biol., 176, 90-94. Moldan, B. (Ed.) (1990) Environment of the Czech Republic, Academia, Prague, 281 pp. Rubeg, J. (1987) Chromosomal aberrations and sister-chromatid exchanges in swine, Mutation Res., 191, t05-109.

210 Savage, J.R.K. (1976) Classification and relationships of induced chromosomal structural changes, J. Med. Genet., 13, 103-122. Svandov~, E. and P. R6ssner (1989) Contribution for an establishing of the spontaneous level of chromosomal aberrations in Czech Republic population, Pracov. LAk., 41, 216-219.

Tj0tta, K. and T. Aune (1991) Metabolic activation of food mutagens by liver and lung enzymes from rats, pigs and oxen, Mutation Res., 251, 7-11.

Communicated by F.H. Sobels