Ozone

Toxicity to birds

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

long-term toxicity to birds: reproduction test

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Justification for type of information:

In addition to numerous studies being available for mammals, the amounts of ozone released to the atmosphere are negligible when compared to the level of ozone already present in ambient air. During normal use of ozone in the envisaged applications, no additional exposure of birds to ozone is therefore expected.

Reference 2

Endpoint:

toxicity to birds, other

Remarks:

2 weeks

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline followed

Principles of method if other than guideline:

Chickens were exposed continuously to 0.7 ppm ozone immediately for 1, 3 or 5 wks starting immediately or for 1.5 to 2 weeks starting one week after hatching. The effects of ozone on the development of nutritional encephalopathy was studied as function of the composition of the diet.

GLP compliance:

no

Dose method:

other: inhalation; whole body; continuous

Analytical monitoring:

yes

Vehicle:

no

Details on preparation and analysis of diet:

The diets differed in lipid, glycerol, cerelose, soybean meal, crude protein and cellulose contents, while levels of DL-methionine, mineral mix, vitamin mix showed minimal or no diferences.

Test organisms (species):

other: Gallus gallus domesticus

Details on test organisms:

Crossbred New Hampshire x White Leghorn
1, 3 or 5 wks starting immediately after hatching or for 1.5 to 2 weeks starting one week after hatching.

Limit test:

no

Post exposure observation period:

no

No. of animals per sex per dose and/or stage:

25 males / treatment group

Control animals:

yes, concurrent no treatment

Nominal and measured doses / concentrations:

0.7 ppm

Details on test conditions:

The animals were housed either in an electrically heated battery brooder equipped with raised wire floors or in an ozone chamber. The latter was 58 cm high, 60 cm wide, and 120 cm long. The wall and bottom were made of steel sheeting coated with epoxy paint. The cover was of 4-mm-thick glass to admit light. Air was moved through the length of the chamber via an inlet housing a blower, an ozone generator, and a thermostatically controlled set of heating elements. An air outlet was placed at the opposite side of the chamber, and a smaller outlet for removing air samples was located at the center of the long wall opposite the hinged door. The rate of flow of air was kept at 180±30 liter/min. It was found in preliminary tests that the rate of growth of chicks kept in the exposure chamber under normal air from hatching to two weeks of age was the same as that observed in the battery cages.
Feed and water were provided at libitum. Feed consisted of artifical diets which differed in lipid, glycerol, cerelose, soybean meal, crude protein and cellulose contents, while levels of DL-methionine, mineral mix, vitmain mix showed minimal of no diferences.

Usually 25 chicks were allotted to each treatment and their number was reduced to 20 at the age of 7 days by removing the lightest chicks.

Details on examinations and observations:

The chicks were inspected twice daily and weighed weekly.

Chicks were diagnosed for Encephalopathy and the number of affected chicks were counted. At the end of the treament period the chicks were killed by decapitation and cerebella and lungs were preserved for pathological examination.

Alpha-tocopherol in the livers and fatty acid composition of tissue lipids was determined after saponification of the tissues.

Details on reproductive parameters:

not applicable

Reference substance (positive control):

no

Repellency factors (if applicable):

not applicable

Mortality and sub-lethal effects:

Ozone treatment did not affect the development of encephalopathy or mortality figures.
Ozone improved the growth of young chicks, provided that exposure was started on the day of hatching. When exposure was begun after one week or later, ozone either inhibited growth or had no effect. In one of the experiments of this series, the air and ozone groups were subdivided further into subgroups with or without vitamin E. The vitamin E-deficient subgroups exhibited poor growth compared with the supplemented subgroups, but neither of the two subgroups showed any significant response to ozone exposure.

Liver tocopherol concentrations were determined at the end of the ozone exposure period in several experiments. In two of the trials in which the dietary vitamin E content was of the order of 10 mg/kg, ozone decreased liver tocopheral content, significantly in one case. However, in a third experiment involving 100 mg tocopherol/kg diet, the trend was reversed, although the effect was not statistically significant.

Cerebellar fatty acid composition. Ozone had no effect on the fatty acid composition of the cerebellum. However, chicks raised on a commercial diet had a lower omega 6 fatty acid content, possibly as a result of the lower linoleic acid content of the commercial feed (about 1 to 1.5% in the latter diet vs. close to 6% for the diet containing 10% oxidized safflower oil).

Total lung fatty acids were analyzed in chicks that had been exposed to ozone for different periods of time, at different ages, and with different dietary regimens. Ozone brought about a remarkable increase in the percentage of palmitic acid in comparison with the controls kept in air. Stearic, linoleic, and arachidonic
acids were all decreased in ozone-exposed chicks, while the trend toward a decrease also existed in the case of oleic and docosatetraenoic acids. It appears that ozone specifically increased the content of the palmitic acid and that the decrease in the other fatty acids was a consequence of this specific effect. In another experiment (not detailed here), ozone exposure of older chicks (0.7 ppm during the 54th to 75th days of age) did not affect lung fatty acid composition.

On macroscopic examination, all ozone treated chicks showed congested lungs of a darkened color. Microscopic examination revealed recent hemorrhages within the bronchi and alveoli. These changes were diffuse in both lungs. In addition, abundant leukocytes, mainly peribronchial, were seen.

Effects on reproduction:

not applicable

Results with reference substance (positive control):

not applicable

Reported statistics and error estimates:

not specified

Validity criteria fulfilled:

not applicable

Conclusions:

Chicks exposed to 0.7 ppm ozone show local effects in the lungs but no systemic effect on the anti-oxidant vitamin E which has an important role in the development of nutritional encephalopathy.

Executive summary:

The effect of ozone exposure on the development of nutritional encephalopathy was evaluated in young chicks receiving diets low in vitamin E and containing a source of linoleic acid. Experiments with 0.3 (initial experiments, data not given) and 0.7 ppm ozone at different ages over different periods of exposure failed to establish any consistent effect on the severity of the disease. Ozone had no clear effect on the liver a-tocopherol content of chicks fed vitamin E-supplemented diets, nor did it affect the fatty acid composition of the cerebellum. However, in all cases ozone increased the concentration of palmititic acid in total lung lipids. Ozone-treated chicks had heavily congested lungs. Chicks exposed to ozone from hatching until 23 days of age grew at a faster rate than controls kept in air. This growth stimulation was not evident when exposure to ozone was begun between 8 and 15 days of age.

Reference 3

Endpoint:

toxicity to birds, other

Remarks:

7 days

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline followed

Principles of method if other than guideline:

The effect of ozone inhalation on birds was investigated. Japanese quail were exposed continuously to 0, 0.3, 1.0, and 3.0 mg/cubic meter ozone (0, 0.15, 0.50, and 1.50 ppm, respectively) for 7 days. Pulmonary effects were determined by light and electron microscopy as well as by biochemistry.

GLP compliance:

no

Dose method:

other: inhalation - whole body

Analytical monitoring:

yes

Vehicle:

no

Details on preparation and analysis of diet:

Not applicable

Test organisms (species):

Coturnix coturnix japonica

Details on test organisms:

8-week-old male Japanese quail (Coturnix coturnixjaponica) (110 +/- 10 g) were obtained from the CIVO-TNO Institute, Zeist, The Netherlands. The first three groups comprised eight animals, and the fourth group comprised six animals. Each group was housed in one chamber.

Limit test:

no

Total exposure duration (if not single dose):

7 d

Remarks:

Continuous expoure

No. of animals per sex per dose and/or stage:

The quail were obtained from the CIVO-TNO Institute, Zeist, The Netherlands. The first three groups (0, 0.15 and 0.50 ppm) comprised eight animals, and the fourth group (1.5 ppm) comprised six animals.

Control animals:

yes, concurrent no treatment

Nominal and measured doses / concentrations:

The ozone concentration in each chamber was measured for 2 min every 8 min with a chemiluminescence ozone-analyzer (Model 8002, Bendix Corp., Lewisburg, WV). Ozone concentrations were 0, 0.3, 1.0, or 3.0 mg/cubic meter (0, 0.15, 0.50, or 1.5 ppm, respectively) with a maximal deviation of 10%.

Details on test conditions:

After an acclimatization period of 1 week, four groups of 8-week-old male Japanese quail (Coturnix coturnixjaponica) (110 +/- 10 g) were continuously exposed for 7 days in rectangular stainless steel and glass inhalation chambers (0.90 x 0.60 x 0.45 m). The light cycle was 12-hr light/12-hr dark. Food (Turkey breeding food, Hope Farms, Woerden, The Netherlands) and water were available ad libitum. Ambient air was filtered through an active carbon filter and a high efficiency particle filter. A flow of 6 cubic meter/hr was maintained through the chambers. The temperature and relative humidity were conditioned at 22 +/- I °C and 55 +/- 5%, respectively. Permanganate- and Neomycin-impregnated animal cage boards (Upjohn, Kalamazoo, MI) prevented ammonia buildup produced from excreta. Food was renewed and the exposure chambers were cleaned daily. During this feeding and cleaning period (30 min), the ozone exposures were interrupted. A negative pressure of +/- 100 Pa was maintained in the chambers.

Details on examinations and observations:

MORTALITY / CLINICAL SIGNS: no data provided, however it was observed that animals of the higest exposure group developed laboured breathing with ongoing exposure and that one of these animals had to be autopsied after 5 days of exposure.

BODY WEIGHT: body weights after 7 day exposure are provided.

PATHOLOGY: histopathology of the was performed in 3 animals of the 3 lowest exposure groups and in 2 animals of the 3 mg/cubic meter group. Trachea, lung, heart, liver, kidney, spleen and bursa of Fabricius were examined by light microscopy. In addition, lungs were examined by scanning electron microscopy and lungs of 2 animals of the 0 and 1 mg/cubic meter groups were examined by transmission electron microscopy.

ORGAN WEIGHTS: lung, heart, liver, kidney, spleen and bursa of Fabricius in all groups.

OTHER: the lungs of 5 animals per group (four in the 3.0 mg/cubic meter group) were excised, trimmed free, blotted, weighed and homogenized. The homogenate was centrifuged and the supernatant was used immediately for biochemical analyses. The protein content and the activities of glucose-6-phosphate dehydrogenase (G6PDH), glutathione peroxidase (GSH-Px), glutathione reductase (GSH-Red) and lactate dehydrogenase (LDH) were measured.

Details on reproductive parameters:

Not applicable

Reference substance (positive control):

no

Duration (if not single dose):

7 d

Dose descriptor:

NOEL

Effect level:

0.3 other: mg/cubic meter

Conc. / dose based on:

test mat.

Basis for effect:

pathology

Repellency factors (if applicable):

Not applicable

Mortality and sub-lethal effects:

MORTALITY/CLINICAL SIGNS: animals of the higest exposure group developed laboured breathing with ongoing exposure and one of these animals had to be autopsied after 5 days of exposure.

Effects on reproduction:

Not applicable

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

The Student's t test (two-sided) was used for statistical analyses of the biochemistry data

Validity criteria fulfilled:

not applicable

Remarks:

no guideline study

Conclusions:

Continuous exposure of Japanese quail for 7 days results in local effect in the lungs. The effects are attributable to the corrosive action of ozone. No effects were observed at 0.3 mg ozone/cubic meter. No signs of systemic toxicity were observed in all exposed groups.

Executive summary:

The effect of ozone inhalation on birds was investigated. Japanese quail were exposed continuously to 0, 0.3, 1.0, and 3.0 mg/cubic meter ozone (0, 0.15, 0.50, and 1.50 ppm, respectively) for 7 days. Pulmonary effects were determined by light and electron microscopy as well as by biochemistry. Focal areas of hemorrhages were noticed in the birds exposed to 1.0 mg /cubic meter ozone. Additional effects after exposure to 1 mg/cubic meter included loss of cilia in trachea and bronchi, an inflammatory response, and necrosis of air capillary epithelial cells. Following exposure to 3  mg/cubic meter many atria of tertiary bronchi were completely obstructed by extensive hemorrhages, metaplasia of atrial wall cells, and hypertrophy of smooth muscle cells. Lung biochemistry data revealed that in the 3  mg/cubic metergroup lactate dehydrogenase, glucose- 6-phosphate dehydrogenase, and glutathione reductase activities were significantly increased. In the 0.3 and 1.0  mg/cubic meter exposure groups no effects on lung antioxidant enzymes were observed. In conclusion, Japanese quail appear to respond to ozone exposure in a different way than mammals. Since no signs of repair in air capillary epithelium after 7 days of continuous exposure were observed, the quail seems to lack the morphological and biochemical repair ability as is observed in mammals.

Reference 4

Endpoint:

toxicity to birds, other

Remarks:

40 d

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline followed

Principles of method if other than guideline:

The objective of this experiment was to determine the effect of adding atmospheric ozone at a target level of 0.05 ppm (level achieved, 0.03 ± 0.017 ppm) to rooms housing broiler chickens. Female broiler chickens were grown in either an untreated environment or an environment with added ozone (3 rooms of 5 replicate pens, each containing 110 birds/treatment).

GLP compliance:

no

Dose method:

other: inhalation - whole body

Analytical monitoring:

yes

Vehicle:

no

Details on preparation and analysis of diet:

Not applicable

Test organisms (species):

other: Gallus gallus domesticus

Details on test organisms:

Hubbard × Peterson female broiler chicks obtained from Lilydale Inc., Wynyard, Saskatchewan, Canada.

Limit test:

no

Total exposure duration (if not single dose):

40 d

Remarks:

continuous exposure

Post exposure observation period:

no

No. of animals per sex per dose and/or stage:

3300 broiler chicks were randomly distributed to 5 pens in each of 6 rooms (110 chicks/pen).

Control animals:

yes, concurrent no treatment

Nominal and measured doses / concentrations:

Mean measured and SD values for ozone at bird and human heights in the treated rooms averaged for the entire trial were 0.030 ± 0.017 and 0.031 ± 0.017 ppm, respectively.

Details on test conditions:

All management techniques used were representative of commercial practice. Temperature was decreased from 35°C on d 0 to 22°C by d 35, and was maintained at this level for the remainder of the experiment. Actual room temperatures, monitored at 2-min intervals, were equivalent in all rooms. All birds were fed corn- and soybean based commercial broiler feeds (starter, grower, and finisher) on an ad libitum basis. Each bird was allotted 0.72 kg of starter feed and 1.25 kg of grower and finisher feed as required for the remainder of the trial. Water was also provided ad libitum. Equal amounts of wheat straw were added to pens, and because determining the effect of ozone on atmospheric ammonia was part of the experimental objective, ventilation was adjusted as required, but no other litter management techniques (removal of wet or caked litter) were imposed.

Details on examinations and observations:

The chicks and feed were weighed on a pen basis at trial initiation (day of hatch), 21 d of age, and trial completion (40 d of age). All mortalities were collected daily, weighed, recorded, and necropsied for cause of death. The number and cause of condemnations for each of the 2 treatments were recorded. Atmospheric ammonia was measured at 15, 20, 28, 32, and 38 d of age. Total aerobic bacterial counts were measured at 11, 19, and 34 d of age.

Details on reproductive parameters:

Not applicable

Reference substance (positive control):

no

Mortality and sub-lethal effects:

MORTALITY (%)
0-21d 21-40d 0-40d
Control 2.67 4.66 7.33
Ozone 3.52 7.94 11.46
Pooled SEM 0.225 0.477 0.600
differences significant between groups in 21-40d and 0-40d interval.

BODY WEIGHT GAIN (kg)
0-21d 21-40d 0-40d
Control 0.700 1.353 2.053
Ozone 0.683 1.255 1.938
Pooled SEM 0.003 0.012 0.014
differences significant between groups in all intervals.

FOOD CONSUMPTION (kg/bird)
0-21d 21-40d 0-40d
Control 1.023 2.879 3.953
Ozone 0.996 2.608 3.695
Pooled SEM 0.004 0.030 0.032
differences significant between groups in all intervals.

FOOD INTAKE / BODY WEIGHT GAIN
0-21d 21-40d 0-40d
Control 1.465 2.189 1.929
Ozone 1.463 2.193 1.912
Pooled SEM 0.003 0.013 0.007
differences not significant.

FOOD INTAKE / BODY WEIGHT GAIN (corrected for mortality)
0-21d 21-40d 0-40d
Control 1.451 2.096 1.870
Ozone 1.443 2.022 1.808
Pooled SEM 0.002 0.011 0.008
differences significant between groups in 21-40d and 0-40d interval.

PATHOLOGY
The causes of mortality in the control and treatment group were classified into acute death syndrome, arthritis, ascites or right heart failure, dehydration, pendulous crop, rotated tibia, spondylolisthesis, valgus deformity, varus deformity, yolk sac infection, no visible lesions seen in birds culled during the experiment and other. No significant differences were noted between the groups except for ascites or right heart failure. Ascites or right heart failure was significantly increased in the 21-40d (0.79 vs 5.27% of birds placed) and 0-40d interval (0.97 vs 5.39% of birds placed).

OTHER FINDINGS (not statistically analysed):
Higher condemnation levels (10.36 v.s 3.39% of birds placed) were found in the ozone treated group on arrival at the processing plant. Condemnation causes were ascites, cyanosis and cellulitis. The percentage of birds dead on arrival was higher in the ozone tretament group (0.79 v.s. 0.30% of birds placed).

No significant effects of ozone on atmospheric bacterial count and ammonia were found .

Effects on reproduction:

Not applicable

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

The experimental design was a 1-way factorial analysis, with rooms nested within atmospheric treatment. Data were analyzed using the GLM procedure of SAS. Any data expressed as a percentage was first checked for normal distribution using the UNIVARIATE procedure of SAS and log + 1 transformation was applied for the ANOVA analysis if data were not normally distributed. Differences were considered significant when P ≤ 0.05, unless otherwise noted.

Validity criteria fulfilled:

not applicable

Remarks:

not to guideline

Conclusions:

Continuous exposure of broiler chickens to ozone at a level of 0.03 ppm resulted in increased mortality and reduced weight gain in the surviving birds.

Executive summary:

The objective of this experiment was to determine the effect of adding atmospheric ozone at a target level of 0.05 ppm (level achieved, 0.03 ± 0.017 ppm) to rooms housing broiler chickens. Female broiler chickens were grown in either an untreated environment or an environment with added ozone (3 rooms of 5 replicate pens, each containing 110 birds/treatment). Bird weight and feed consumption were recorded on a pen basis at 21 and 40 d of age; mortalities were collected daily, and necropsies were performed. Air samples were collected and plated on appropriate media to count total aerobic bacteria at 11, 19, and 34 d of age and enterobacteria at 34 d. Atmospheric ammonia levels were measured on d 15, 20, 28, 32, and 38. A significant (P ≤ 0.05) improvement in feed conversion (feed:gain ratio) when corrected for mortality was noted in the ozone-treated birds (1.808 vs. 1.870). Birds exposed to ozone grew significantly slower (1.938 vs. 2.053 kg of BW gain), consumed less feed (3.695 vs. 3.953 kg), and had a higher mortality (11.46 vs. 7.33%) and condemnation percentages (10.36 vs. 3.39%) than nontreated broilers. The addition of ozone caused no significant decrease in ammonia level or total bacterial count.

Description of key information

Numerous studies for mammals are present (see section 7 of the present dossier), hence, further texting is not necessary (REACH Annex X, column 2, 9.6.1). Therefore, it is not considered ethical to perform additional animal experiments for derivation of a no-effect level for environmental risk assessment since the contribution of ozone as disinfectant does not affect the background concentration of ozone already present in the atmosphere. During normal use of ozone in the envisaged applications, no exposure of birds is expected.

Nevertheless, existing studies on birds and ozone from scientific literature provide some weight of evidence, including:

- A treatment of chicks with 0.7 ppm ozone for two weeks leads to heavily congested lungs (Bartov et al. 1981).

- Treatment of japanese quail over 7 days indicated a NOEL of 0.3 mg/cubic meter (Rombout et al. 1991).

- Exposure to 0.03 ppm in a study by Schwaen-Lardner (2009) demonstrated that birds exposed to ozone grew significantly slower (1.938 vs. 2.053 kg of BW gain), consumed less feed (3.695 vs. 3.953 kg), and had a higher mortality (11.46 vs. 7.33%) and condemnation percentages (10.36 vs. 3.39%) than nontreated broilers.