Cytologia 44: 921-926, 1979 Cytological Effects of Paper Mills Effluents on Somatic Cells of Allium cepa K. B. Shanthamurthy and V. Rangaswamy Received April 6, 1978 Department of Post-Graduate Studies and Research in Botany, U niversity of Mysore, Manasa Gangotri, Mysore-6, India Industries and industrial pollution have tremendous impact on living organi sms. The physiological and ultra-structural effects of air pollutants on vegetation have been assessed by various investigators (Thomas 1951, Thomson et al. 1966, Dugger and Ting 1970a, Swanson et al. 1973, Mudd and Kozlowski 1975, Stern 1975, Warren 1971). Types of occupational hazards of industrial workers were studies by several researchers (Bouhuys et al. 1969, Gamble et al. 1976, Mc Michael et al. 1974, Infante Peter et al. 1977). However, the genetic and cytological effects of industrial effluents on living organisms have not been assessed properly. The present report deals with the studies on cytological effects of paper mill effluents on somatic cells of Ailium cepa. Materials and methods The paper mills effluents were brought from Mandya National Paper Mills, Belagola, South India Paper Mills, Nanjanagud and Mysore Paper Mills, Bhadrava thi, of Karnataka State (South India). Various concentrations were made from the effluents collected from the factory. Germinating onion bulbs with roots 2-2.5cm long were immersed in 100%, 75%, 50% and 25% effluents and also in tap water as control for 6, 12, 24 and 48 hours. The roots from the treated and control were clipped and fixed in 1:3 acetic alcohol for 24 hours at room tempera ture and then stored in 70% alcohol until the aceto-orcein squashes were made for cytological observations. Results and discussion The cytological effects of the paper mill effluents on somatic cells have been estimated on the basis of change in mitotic index and other abnormalities induced by them. The effluent inhibited cell division in the treated roots as shown in the table (Table 1). A strong dosage effect is obvious from a decline in the mitotic index values with the increase in concentration and exhibiting a marked decrease at highest concentration. The mitotic index values for different concentrations are shown in the graph. The graph indicates that they follow the same rate of multiplication of cells during the period of treatment as with the control and at the same time retaining their individual differences between concentrations. Various types of abnormalities have been observed, of which disturbed
922 K. B. Shanthamurthy and V. Rangaswamy Cytologia 44 metaphase, anaphase and telophase, binucleate and tetranucleate conditions, occurrence of micronuclei comprised the most dominant types of anomalies. In the case of disturbed metaphase, anaphase and telophase the chromosome plates were found irregularly in the cell (Figs. 1-4). These abnormalities probably Table 1. Mitotic index* following treatment with paper factory effluent * Estimated from 5000 cells. were due to the effect of effluent on spindle apparatus. The c-mitosis is predomina ted in the roots treated with 75% and 100% effluents. At the end of the mitosis the chromatids aggregate themselves in the centre and later they form restitution nuclei (Fig. 5). Binucleate and tetranucleate cells (Figs. 6-9) are frequently found at higher concentrations. This may be attributed to the failure of cell plate forma tion at telophase. The oblique metaphase, anaphase and telphases were more in treated roots (Figs. 1, 4, 10). Chromosome clumping (Figs. 11-12), micro nuclei formation (Fig. 13), cell vacuolation (Figs. 14-15) were less frequently observed. The nucleus in the treated roots were found on one side, while the accumulation of grannules of chemicals on the other region of the cell (Fig. 16).
1979 Effects of Paper Mills Effluents on S omatic Cells of Allium cepa 923 Figs. 1-12. 1-4, irregular movements of the chromosome plates. 1, 2400 ~. 2, 1400 ~. 3, 1200 ~. 4, 1000 ~. 5, restitution nucleus. 1100 ~. 6-8, binucleate cells. 6, 900 ~. 7, 900 ~. 8, 1400 ~. 9, tetranucleate cell. 1000 ~. 10-12, chromosome clumping. 10, 900 ~. 11, 900 ~. 12, 1100 ~. Large pleiomorphic nuclei are also found in the treated roots (Figs. 17-18). The frequency of abnormalities decreased by lowering the concentration of the effluents. The Table 2 indicates that irrespective of the duration of treatment, the abnormalities, more or less remain the same. Very rare abnormalities like contracted chromosomes (Fig. 19) and differential condensation of sister nuclei (Fig. 20) were also seen. This shows the mitoclasic action of the effluents. Numerical increase of chromosomes were also recorded in a few cells (Fig. 21). In general the various kinds of abnormalities were noted more in the treated
924 K. B. Shanthamurthy and V. Rangaswamy Cytologia 44 Figs. 13-21. 13, cell showing micronucleus. 1150 ~. 14-15, vacuolation in the cells. 14, 1300 ~. 15, 1000 ~. 16, cells with grannules of chemicals. 900 ~. 17-18, pleiomorphic nuclei. 17, 700 ~. 18, 700 ~. 19, contracted chromosomes. 1400 ~. 20, differential condensation of sister nuclei. 1200 ~. 21, cells with increased number of chromosomes. 1400 ~.
Table 2.
926 K. B. Shanthamurthy and V. Rangaswamy Cytologia 44 roots for longer periods and at higher concentrations. The results and data presented show that in onion roots treated with paper mills effluent a large number of abnormalities are present. All these abnormali ties observed may be due to the action of alkalies, acids and bleaching agents present in the paper mill effluents. It could be synergistic or individual or both. Summary The cytological effects of paper mills effluents on somatic cells of Allium cepa were presented. The cytological effect decreased by decreasing the concen tration of the effluents. Disturbed metaphase, anaphase and telophase, occurrence of binucleate and tetranucleate cells comprised the most dominant types of abnor malities. Clumping of chromosomes, contracted chromosomes, pleiomorphic and micronuclei formation, cell vacuolation and polyploid cells are found less frequently. The c-mitotic effect and other abnormalities may be due to the action of alkalies, acids and bleaching agents present in the paper mill effluents. Acknowledgement The authors wish to thank Dr. D. A. Govindappa, Professor and Head of the Department of Botany, University of Mysore, Mysore, for encouragement shown during the course of the investigation and critical reading of the manuscript. One of us (K. B. S.) remains grateful to U. G. C. New Delhi, for the award of a Junior Research Fellowship. References Borger, M. and De Brabander, M. 1975. Microtubules and Microtubule Inhibitors. North Holand publishing company. Page: 427-451. Bouhuys, A. et al. 1969. Chronic respiratory disease in Hemp workers. Am. J. Med. 46: 526. Fishbein, L., Flamm, W. G. and Falk, H. L. 1970. Chemical Mutagens. Academic Press. Dugger, W. M. Jr. and Ting, I. P. 1970a. Air pollution oxidants. Their effects in metabolic process in plants. Ann. Rev. Plant Physiol. 21: 215-34. Gamble, John F. 1976. Respiratory function and symptoms an environmental epidemiological study of rubber workers exposed to a phenol-formaldehyde type of resin. Am. Ind. Hygiene Association Journal: 499-512. McMichael et al, 1974. An epidemiologic study of mortality within a cohort of rubber workers 1964-72. J. Occupm Med. 16: 58-64. Mudd, J. B and Kozlowski, T. T. 1975. Responses of Plants to Air Pollution. Academic Press Inc. Infante, Peter F. et al. 1977. Leukaemia in benzene workers. The Lancet 77: 76-79. Stern, 1975. Air Pollution. Academic Press. Amer, Soheir M. and Odette, R. Farah 1971. Cytological effect of pesticides VI. Effect of the insecticide Rogor on mitosis of Vicia faba and Gossypium barbadense. Cytologia 39: 507-514. Swanson, E. S. et al. 1973. The effect of ozone on leaf cell membranes. Can. J. Botany 51: 1213-1219. Thomas, M. D. 1951. Gas damage to plants. Annu. Rev. Plant Physiol. 2: 293-1219. Thomson, W. W. et al. 1966. Effect of ozone on the fine structure of the palisade parenchyma cells of bean leaves. Can. J. Bot. 49: 1677-1682. Warren, 1971. Biology and Water Pollution Control. Philadelphia W. B. Saunders company.