Effect of Sodium Dlethyldithiocarbamate on

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Jan 24, 1980 - dithiocarbamate was tested fer its ... DITHIOCARBAMATES, the salts of .... mination by pesticides and other synthetic organic. 1. ---'-.-"'---~.'...:. ~.
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References

I.

MIZUSHlMA,

S., Blochetn, Biopliys. Res. Commun .. 61 (974),

1221. 2.

MORAN,

3.

MIDT-CLERMONT, M. & TISSIERES, A., Soc. B., 283 (1978), 391. . FRASCH, C. E. & MOCCA, L. F., J.

L.,

MlRAULT,

M.

E.,

ARRIGO,

A.

r.,

GOLDSCH-

Phil. Trans. Roy. Bact., 136 (1978),

1127. 4. 5. 6. 7.

8. 9. 10. 11. 12.

E., DANEO-MoORE, L., & BRAVERMAN, S., J. Bact., 136 (1978), 491. TABOR, C. W., J. Bact., 83 (1962), 1101. HAROLD, F. M., J. Bact., 88 (\964), 1416. BROCK, T. D., SYII/p. Soc. gen. Microbiol., 19 (1969), 15. RAY, P. H. & BROCK, T. D., J. gen. Microbiol., 66 (1971), 133. GRESS HOFF, P. M., Aust . J. Pl. Physiol., 3 (1976), 457. RAMAIAH, K. V. A. & MOOKERJEE, A., Indian J. exd. BioI., 16 (1978),857. GARY-BoBO, C. M., Nature, Lond., 228 (1970), 1101. LURQUlN, P. F. & BEHKI, R. M., Mutation Res., 29 (1975). 35. TERNOVE-HINKS,

Effect of Sodium Dlethyldithiocarbamate Embryos of the Frog Microhyla H.

Department

on Developing ornata"

V. GHATE & LEELA MULHERKAR

of Zoology, University of Poona, Pune 411 007 Manuscript

received 24 January 1980

In view of the fact that various dithiocarbamases

. are

used in

fungicide preparations and commonly handled, sodium diethyldithiocarbamate was tested fer its toxicity and teratogenicity on frog embryos. Developing embryos of the frog, M. ornata, were treated with 0.1-5.0 mg/! of this chemical. The cone. greater than 3 mg/l proved to be lethal in about 24 hr exposure. Between 1 and 3 mgj! cone., severely malformed embryos were formed. Retardation of growth, curvature of the body axis, general oedemic condition, delay or inhibition of pigment development and abnormal notochord were observed in the embryos. In 0.5 mg,' 1 the embryos developed only abnormal notochord which was completely wavy. It appears that the chemical affects mainly the development of the notochord although lit higher conceutrations other abnormalities are also produced.

DITHIOCARBAMATES, the salts of dithiocarbamic acid and related compounds, are potent fungicides and various formulations are widely used to control fungal diseases of crops-'>, Antibacterial and antifungal activities of some dithiocarbarnates have been studied 3,'1. The ernbryotoxic and teratogenic potential of some fungicide preparations and pure compounds have been investigated in amphibians», birds" and mammals". Dithiocarbamates have a strong affinity for copper and sodim dicthyldithiocarbamate is often used in colorimetric estimation of copper", Thus it is commonly handled by human beings. This fact coupled with the knowledge that related compounds are used to control fUl1~OJ diseases or fruits and vegetables, has prompted tl~is study. Naturnlly fertilized eggs of the frog, Ai icrohyla ornat a v.c=e: collected from a locvl pond .. k!ly C02.ts *This p.lpc:r was prseentc.l at Il l All India Development .I 81OIY6Y held at Gwalior during

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on

1930

were removed manually using forceps. Embryos with intact vitelline membrane were then used in the experiments. Sodium die thy ldithiocarbamate (NaDEDC) utilized in this study was a B.D.H. laboratory reagent of 97% purity. The stock solution of NaDEDC was prepared in distilled water. Different dilutions of the 0.1 % stock solution were made with aged tap water to prepare various cone. of the chemical. The term age d tap water refers to tap water that was stored in a large cement tank outdoors for about 2 weeks. The aged tap water was always filtered and aerated before use. The treatment of the embryos was carried out in glass petri dishes of 10 ern diam, Each such petridish contained 10 embryos in gastrulation and 100 ml of medium. Aged tap water without NaDEDC served as a control medium. The embryos were periodically observed under dissecting binocular for mortality and other effects. A few normal and abnormal looking embryos were fixed in Bouin's fluid, embedded in paraffin, sectioned and stained with haematoxylin and eosin. Development of the embryos in aged tap water (controls) was observed to be very rapid. In about 24 hr the embryo, still inside the intact vitelline membrane, showed twitching muscular activity. By the end of 48 hr all the embryos hatched as tadpoles and were swimming in the medium although most of the time they remained attached to the sides of the dish. At this stage all the tadpoles possessed well developed eyes, head and tail. Typical pigment patches were observed on the dorsal and dorsolateral parts of the head. By the end of the experiment (96 hr) all the tadpoles had grown further and were moving actively in the medium. The straight notochord was clearly visible through the transparent tail of the tadpole. In the NaDEDC treated groups there were several abnormalities in the developing embryos. The severity of the defects was dose dependent. At 5 mg/l cone. the chemical proved to be highly embryo toxic. There was a severe retardation of growth and development, followed by death in about 24 hr. In 3 mgjl cone. all survived 24 hr exposure, but there was 50~~ mortality in the subsequent period. All the survivors were highly retarded and malformed. There was abnormal bending of the body axis, general oedema and total lack of pigmentation at 72 hr treatment (Fig. 1). The notochord in these tadpoles appeared competely abnormal and kinky. The tail was usually shortened and the tadpoles lay on one side at the bottom of the petridish with only occasional, spasmodic twitching of the body. J 11 the lower conc., namely 1 and 2 rng.l, same effects were observed with less severity, Pigmentation appeared in some cases after 72 hr,yet it was not complete as in case of the controls. At 0.5 mg!l cone. the tadpoles appeared more: or less norma'! except for being slight.y shortened and possessing a broadened tail. 1he pigment development was also quite normal. However the notochord was again completely wavy in all the tadpoles. Consequently the tadpoles showed abnormal locomotion and most of them remained lying on one side at the bottom of the pctridish. Sagittal sections of the control (Fig. 2) and ex perimental (Fig. 3) tadpoles confirmed the presence of

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. he abnormal a nd wavy notochord in the experimental tadpole. The notochordal cells of the treated zr oup appeared swollen and more in number than control. set. Transverse section of the tadpoles passing through the tail region also indicated the excessive i!rowth of the notochord (Figs. 4, 5). It was also observed that the wavyness of the notochord was in dorsoventral as well as in lateral directions. The abnormal notochord frequently displaced the muscullure and the neural tube in the tail region. The other noticeable effect was thickening of the lining of the intestine in NaDEDC treated tadpoles. The results presented here clearly indicate that the compound NaDEDC is a potent teratogen. It is quite active even at a low concentration and specifically induces abnormal development of the notochord. At higher cone. several other abnormalities are produced alongwith a severely malformed notochord. Si milar results were reported by Bancroft and Prahlad" in the case of Xenopus laevis using a related dithiocarbamate compound. These authors explained the a bnorrnal development of the notochord

to be due to change of the dorsal-ventral gradient of the developing embryo by a decomposition product of dithiocar hamate. namely ixot hiocya nat e. Teratogenicity of dithiocarbamate fungicides was tested on mammals and it was suggested that the teratogenic action was due to alkylthiourea, again a degradation produce. Ethylenethiourea (ETU), which is a decomposition product of dithiocarbarnate, has in fact been proved to be teratogenic to mammalsv-v. The mechanism of teratogenic action of ETU is not yet well known. NaDEDC, as mentioned earlier, has a strong affinity for copper and therefore may affect copper metallo-enzymes. In fact in vitro as well as in vivo inhibition of a copper requiring enzyme 'tyrosinase' by dithiocarbarnate is reported-J'!". This may be the reason for the observed arnelanogenesis or delayed melanogenesis in the exposed embryos. Phenylthiourea (PTU), which is similar to ETU, also affects melanin development-s. With the present situation of environmental contamination by pesticides and other synthetic organic

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Fig .. J - Control tadpole (above) and experimental tadpole (below) at about 72 hr showing highly abnormal development of embryo. x 20. Fig. 2 - Sagittal section of a 96 hr old control tadpole. Note the straight notochord (1· ), x 45. Fig. 3 - Sagittal section of a 96 hr old experimental tadpole showing abnormal notochord ( 1· ). x 45. Fig. 4 - T. S. passing through the tail of a control tadpole, x I ~5. Fig. 5 - T. S. passing through the tail of an experimental tadpole showing abnormal development of the notochord, x 125.

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compounds. many of which are teratogenic. carcinogenic or mutaae nic'". it would be desirable to understand the mechanism of action of these chemicals to safeguard public health. The authors are grateful for the financial support for this research under 'Khosla Research Project". References 1. SHEP,~RD, H. H .. Tire chemistry and action of insecticides (\kGra\\-Hill I300k Co .. Inc. :\cw York), 1951. '1 :VIITcHELL. L. E., in Organic pesticides ill the environment, edited bv E. F. Gould (American Chemica! Society, Washington, USA). 1966. 3. SRIV,-\ST\\',-\. T. ~ .. R.-\STOGI. R. B. & SR[VAST.-WA, O. P., Indian 1. expo Biol., 17 (1979), 710. -l. LUD\\'IG. R. A .• THoR'.:. G. D. & \I!LLER, D. \1., Can, 1. Bot .. 32 (I 95-l). 48. 5, BA'.:cROFT. R. D. & PRAHLAD, K. V., Teratology, 7 (1973), 143, 6. GEBc!~RT. D. O. E. & V,"S LOGTE'.:, M. 1., Toxic. appl . Pharmacol., 13 (1968), 316. 7. PETROV,-\-VERGIEVA, T., Teratology, ~ (1971), 497. 8. \LKKERETH. F. J. H" HERO'.:, J. & TALu'.:G, J. F., Water analvsis (Fresh Water Biological Association U.K., Scientific Publication No. 36). 1978. 9. KHERA, K. S., Teratology, 7(1973), 243. 10. RUDDICK, 277.

1, A. & KHERA, K. S., Teratology,

12 (1975),

11. LU,:-'L H. & STAPLES. R. E., Teratology, 17 (1978). 171. 12. CHE'.:, Y. M. & CHW['.:, W., Experientia, 34 (t 978), 21. 13. BUGOEVA, P. & STOICHEV, I., Experientia, 35 (1979), 1389. l-l. EpPIG. J. J., 1. Embryol. expo Morphol., 24 (1970), 447. 15. FISHREI-':, L.. in Insecticide biochemistry and plivsiolog y, edited by C. F. Wilkinson (Plenum Press, New York), 1976.

J uvenile Hormone janata

L.

Activity of (Lepidoptera

Altosid on Aclioea Noctuidae)

S. SHAHEE'.: & Z. 05'-'1A-.:I Enromologj

Manuscript

Division, Regional Research Hyderabad 500 009

Laboratory

received 26 November 1979: revised received 19 January 1980

manuscript

Topical application of a synthetic juvenile hormone analogue, Altosid (isopropyl ll-methoxy-3, 7-11-trimethyl dodeca-2,4 dienoate ), to larval, and pupal, abdomen of A. janata L. within 2 .• hr of the intervening molts produced larval-pupal. pupal-adult intermediates and also abnormal adults with deformed appendages. A t higher doses (2-10 {J.g,pupae or pre-pupae) there "as cent per cent inhibition in adult emergence, Early larval ~tages "ere not much sensitive; only pre-pupae and pupae were the most sensitive stages showing .JH activity. Acetone has no effect on metamorphosis of .-I. jancua,

IVI AN:

rer,0rlS, are a~ailablc on the juvenile activitv ot Altosid aaumsr economica lly important pests specially the Diptcra in both laboratorYll1d field te~t::,I.~. However 11.0 data ;\1'