Romanova E., Shapovalova K., Ryabinina E. The leukocyte blood composition and micronuclei in the erythrocytes of amphibians in the polluted reservoirs of Nizhni Novgorod region // Principy èkologii. 2018. № 2. P. 125‒139. DOI: 10.15393/

Issue № 2

Original research


The leukocyte blood composition and micronuclei in the erythrocytes of amphibians in the polluted reservoirs of Nizhni Novgorod region

   Elena Borisovna
D.Sc., Nizhny Novgorod’s Lobachevsky State University,
   Kristina Vadimovna
Nizhny Novgorod’s Lobachevsky State University,
   Elena Sergeevna
Nizhny Novgorod’s Lobachevsky State University,
WBC (White blood cells)
micronucleus test
immune status
Summary: Autecological researches are necessary to obtain populations characteristics providing steady existence of animals in the conditions of an anthropogenic stress; they have practical significance connected with the assessment of the environment quality by bioindication. The purpose of this work was the autecological investigation of leukocyte composition and the micronucleation in red blood cells of amphibians. The object of the research was marsh frogs (70 specimens) and pool ones (35 specimens) in the water bodies of Nizhni Novgorod region. The duration of the observation was two years. The main methods were qualitative and quantitative analysis of leukocytes and red blood cells, hydrochemical analysis by spectrophotometry. Statistical processing was carried out by the methods of nonparametric statistics, correlation method, dispersive analysis (R-studio, Statictica). The main pollutants of water bodies were defined and the specific combinatory index of water impurity was calculated. It was established that the number of leukocytes and erythrocytes in the blood of amphibians was decreased. Interspecies distinctions were revealed in the content of myelocytes, basophiles, eosinophils and lymphocytes. The parameters of leukocytal blood system of pool frogs in specific conditions of their habitat were mostly caused by the dynamics of lymphocytes. WBCs of marsh frogs were characterized by the increase in the share of immature forms of neutrophilic granulocytes and the change of the ratio between the share of big and small lymphocytes. Indicators of blood correlated with the content of iron (r = 0.85, р = 0.029) and chlorides (r = -0.87, р = 0.021) in a reservoir. It was found that at higher levels of water pollution for two years, the shares of the erythrocytes containing the micronuclei of attached and disintegrated types increased. It occurred on the background of the decrease in the share of rounded micronuclei. For the first time the size of micronuclei of different types occurring in erythrocytes of amphibians was measured. It was shown that the area of the disintegrated micronuclei surpassed that of rounded and attached micronuclei. The results revealed the features of leukocytal blood composition and cytogenetic homeostasis in the populations of marsh and pool frogs in the specific biotopical conditions.

© Petrozavodsk State University

Reviewer: A. Bakiev
Reviewer: V. A. Ilyukha
Received on: 21 March 2018
Published on: 30 June 2018


Bannikov A. G. Darevskiy I. S. Ischenko V. G. Rustamov A. K. Scherbak N. N. Identification guide of amphibian and reptile fauna of the USSR. M.: Prosveschenie, 1977. 414 p.

Chernyshova E. V. Starostin V. I. Peripheral blood of frogs of the genus Rana as a test system for evaluation of environmental pollution, Izvestiya RAN. 1994. Ser. biol. No. 4. P. 656–660.

Davis A. K., Maney D. L., Maerz J. C. The use of leukocyte profiles to measure stress in vertebrates: a review for ecologists, Functional Ecology. 2008. Vol. 22. P. 760–772.

Gelashvili D. B. Bezel' V. S. Romanova E. B. Bezrukov M. E. Silkin A. A. Nizhegorodcev A. A. Principles and methods of ecological toxicology, Pod red. prof. D. B. Gelashvili. Nizhniy Novgorod: Izd-vo NNGU, 2016. 702 p.

Gelashvili D. B. Koposov E. V. Laptev L. A. Ecology of Nizhni Novgorod. N. Novgorod: NNGASU, 2012. 530 p.

Gelashvili D. B. Ohapkin A. G. Doronina A. I. Kolkutin V. I. Ivanov E. F. The ecological status of water bodies of Nizhni Novgorod. N. Novgorod: Izd-vo NNGU, 2005. 414 p.

Kasuba V., Rozgaj R., Trusic I. Genotoxic effects of cadmium chloride in V79 cell culture, 41 Congress of the European Societies of Toxicology "Eurotox 2003", Florence, Sept. 28 – Oct.1, 2003, Toxicol. Lett. 2003. Vol. 144. P. 136.

Koh-ichi U., Atsushi O., Noriaki S. Generation of Micronuclei during Interphase by coupling between cytoplasmic membrane blebbing and nuclear budding, PLoS ONE. 2011. Vol. 6. P. 11 URL: (data obrascheniya: 29.05.2018).

Kour P., Tripathi N., Poonam. Micronuclei introduction in different tissues of Euphlyctis cyanophlyctis (common skittering frog) exposed to sublethal concentration of mercuric chloride, International Journal of Current Research. 2016. Vol. 8. P. 35680–3568.

Kovaleva O. A. Cytogenetic anomalies in mammalian somatic cells, Citologiya i genetika. 2008. No. 1. P. 58–72.

Kuzina T. V. Changes in the structure of the erythrocytes nucleus in peripheral blood of fish species of Volga-Caspian channel, Vestnik MGOU. Ser. Estestvennye nauki. 2010. No. 2. P. 50–57. URL: (data obrascheniya: 29.05.2018).

Kvasov I. D. Parfenov V. N. Cvetkov A. G. Intranuclear structures containing RNA maturation factors in the early vitellogenic oocytes of the grass frog, Citologiya. 2000. T. 42. No. 6. P. 536–549.

Luzhna L., Kathiria P., Kovalchuk O. Mironuclei in genotoxicity assessment: from genetics to epigenetics and beyond, Frontiers in Genetic. 2013. Vol. 4. P. 17. URL: (data obrascheniya: 29.05.2018).

Mansi A., Sunitha J. D., Geetanshu D., Neelakshi S. R. Micronuclei assay of exfoliated oral mucosal cells: review, Annals of Dental Specialty. 2014. Vol. 2. P. 47–50. URL: (data obrascheniya: 29.05.2018).

Manskih V. N. On the problem of the mechanisms of micronuclei formation in somatic cells of tailless amphibians normally and under the effect of N-nitroso-N-methylcarbamide, Byulleten' ekperimental'noy biologii i mediciny. 2006. T. 141. No. 2. P. 217–220.

Men'shikov V. V. Delektorskaya L. N. Zolotnickaya R. P. Laboratory research methods in clinic. M.: Medicina, 1987. 368 p.

Mineeva O. V. Mineev A. K. Disorders of blood leukocyte formula in the lake frog of the Saratov reservoir, Vestnik Nizhegorodskogo universiteta im. N. I. Lobachevskogo. 2011. No. 2 (2). P. 94–97.

Murav'eva L. V. Tihomirov O. A. Markov M. V. The formation of aquatic-terrestrial complexes of peat pits and their classification, Uchenye zapiski Kazanskogo universiteta. Ser. Estestvennye nauki. 2010. T. 152. Kn. 4. P. 102–115.

Nepomuceno J. C., Ferrari I., Spano M. A., Centeno A. J. Detection of micronuclei in peripheral erytrocytes of Cyprinus carpio exposed to metallic mercury, Environ and Mol. Mutagenes. 1997. Vol. 30. No. 3. P. 293–297.

Peskova T. Yu. Adaptive variability of amphibia in an anthropogenically polluted environment. Tol'yatti, 2004. 36 p.

Proshin S. N. Violations of the morphology of interphase nuclei in cell populations of animals in the evaluation of the genome under destabilizing factors. SPb.; Pushkin: VNIIGRZh, 2007. 33 p.

Romanova E. B. Shapovalova K. V. Mar'in I. A. Myelogram marsh (Pelophylax ridibundus) and pool frogs (Pelophylax lessonae) (Amphibia: Ranidae) of conventionally "background" and anthropogenously transformed territories of the Nizhny Novgorod region, Povolzhskiy ekologicheskiy zhurnal. 2017. No. 3. P. 298–307. URL: (data obrascheniya: 29.05.2018). DOI: 10.18500/1684–7318–2017–3–298–307.

Romanova E. B. Shapovalova K. V. Ryabinina E. S. Gelashvili D. B. Leukocytic indices and micronuclei in erythrocytes as population markers of the immune status of Pelophylax ridibundus (Pallas, 1771) (Amphibia: Ranidae) living in various biotopic conditions, Povolzhskiy ekologicheskiy zhurnal. 2018. No. 1. P. 60–75. URL: (data obrascheniya: 29.05. 2018). DOI: 10.18500/1684-7318-2018-1-60-75.

Shlyahtin G. V. Golikova V. L. Technique of field researches of ecology of amphibians and reptiles. Saratov: Izd-vo Saratovskogo un-ta, 1986. 78 p.

Sils E. A. Comparative analysis of hematological indicators of moor (Rana arvalis, Nilsson, 1842) and marsh frogs (Rana ridibunda, Pallas, 1771) of city populations, Vestnik Orenburgskogo universiteta. 2008. No. 10 (92). P. 230–235.

Smirnova V. M. Makeev I. S. Blagodatkin A. V. Method of complex assessment of contamination of surface water by hydrochemical indicators. Determination of the specific combinatorial index of water pollution (WCISW) and the class of water quality. N. Novgorod, 2011. 19 p.

Vershinin V. L. Hematopoiesis of Anurans – specific features of adaptiogenesis in species in resent ecosystems, Zoologicheskiy zhurnal. 2004. No. 11. P. 1367–1374.

Woznicki P., Lewandowska R., Brzuzan P. The level of DNA damage and the frequency of micronuclei in haemolymph of freshwater mussels Anodonta woodiana exposed to benzo(a)pyrene, Acta toxicol. 2004. Vol. 12. No. 1. P.41–45.

Zhu Yi., Zhang Y. Formation of micronuclei in carp polychromatocytes under the influence of cadmium, chromium and copper: synergistic effect, J. Nanjing Norm. Univ. Nat. Sci. 1999. Vol. 22. No. 3. P. 60–63.

Zhuleva L. Yu. Dubinin N. P. Using the micronucleus test to assess the environmental conditions in the areas of Astrakhan region, Genetika. 1994. T. 30. No. 7. P. 999–1004.

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