Leishman staining for the identification of haemocytes in beetles
Published 2026-06-12
Keywords
- haemocytes,
- Pheropsophus sobrinus,
- Oryctes rhinoceros,
- plasmatocytes,
- adipohaemocytes
- oenocytes ...More
Copyright (c) 2026

This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.
Abstract
Haemocytes in insects play a vital role in immune responses, and their general appearance varies in different groups. Leishman stain is widely used for the staining and identification of blood cells. The present study was conducted to identify the haemocytes of two adult beetles, Pheropsophus sobrinus and Oryctes rhinoceros. Haemocytes were stained with Leishman stain, examined, counted under a microscope and photographed. The cells were identified based on their general appearance, size, shape and position of the nucleus. Prohaemocytes, plasmatocytes, granulocytes, oenocytes, spherulocytes and adipohaemocytes are the major cells identified in the haemolymph. Differential haemocytes count showed a higher percentage of prohaemocytes in both the beetles. Plasmatocytes and adipohaemocytes had the second highest percentages in O. rhinoceros and oenocytes in P. sobrinus. The study enabled us to understand how the haemocytes of different insect species differ in terms of their appearance and number in concomitant with their mode of living.
References
- P. M. Choate, "Manual for the identification of Ground beetles (Coleoptera: Carabidae)(including tiger beetles) of Florida", Department Entomology and Nematology, University of Florida, 2001.
- N. R. Moyetta, F. O. Ramos, J. Leyria, L. E. Canavoso, and L. L. Fruttero, "Morphological and ultrastructural characterization of hemocytes in an insect model, the hematophagous Dipetalogaster maxima (Hemiptera: Reduviidae)", Insects, vol. 12, no. 7, p. 640, 2021.
- J. A. Hoffmann, "The immune response of Drosophila", Nature, vol. 426, no. 6962, pp. 33–38, 2003.
- M. R. Kanost, H. Jiang, and X.-Q. Yu, "Innate immune responses of a lepidopteran insect, Manduca sexta", Immunological reviews, vol. 198, no. 1, pp. 97–105, 2004.
- M. D. Lavine and M. R. Strand, "Insect hemocytes and their role in immunity", Insect biochemistry and molecular biology, vol. 32, no. 10, pp. 1295–1309, 2002.
- S. C. P. Costa, C. Ribeiro, P.-A. Girard, R. Zumbihl, and M. Brehélin, "Modes of phagocytosis of Gram-positive and Gram-negative bacteria by Spodoptera littoralis granular haemocytes", Journal of Insect Physiology, vol. 51, no. 1, pp. 39–46, 2005.
- A. P. Gupta, "Cellular elements in the hemolymph", Comprehensive insect physiology, biochemistry and pharmacology, vol. 3, pp. 402–451, 1985.
- I. M. R. Amaral et al., "Circulating hemocytes from larvae of Melipona scutellaris (Hymenoptera, Apidae, Meliponini): Cell types and their role in phagocytosis", Micron, vol. 41, no. 2, pp. 123–129, 2010.
- A. M. Nayar, G. S. Devi, and D. A. Evans, "Effect of Bacillus thuringiensis israelensis infection on the biochemical and hematological profile of the hemolymph of Oryctes rhinoceros [L.](Coleoptera: Scarabaeidae) larva", Journal Entomon, vol. 35, no. 4, pp. 1–5, 2010.
- J. F. Hillyer, "Insect immunology and hematopoiesis", Developmental & Comparative Immunology, vol. 58, pp. 102–118, 2016.
- C. DaCruzLandim, "Ultrastructure of haemocytes present during flight muscle differentiation of Melipona quadrifasciata anthidioides, Lepidoptera (Hymenoptera, Apidae)", 1996.
- R. Sareen, M. Kapil, and G. N. Gupta, "Incubation and its effect on Leishman stain", Journal of Laboratory Physicians, vol. 10, no. 03, pp. 357–361, 2018.
- A. S. Venugopal and S. K. Thomas, "Bombardier beetles of the genus Pheropsophus Solier 1833 (Carabidae: Brachininae: Brachinini) from Indian subcontinent", Zootaxa, vol. 4608, no. 1, pp. 65–89, 2019.
- G. Manjeri, R. Muhamad, Q. Z. Faridah, and S. G. Tan, "Morphometric analysis of Oryctes rhinoceros (L.)(Coleoptera: Scarabaeidae) from oil palm plantations", The Coleopterists Bulletin, vol. 67, no. 2, pp. 194–200, 2013.
- R. Mathur, A Text Book of Entomology, 1st edn. New Delhi, India: Campus Books International, 2002.
- V. B. Wigglesworth, The principles of insect physiology. Springer Science & Business Media, 2012.
- J. C. Jones, "The hemocytes of Rhodnius prolixus Stal", The Biological Bulletin, vol. 129, no. 2, pp. 282–294, 1965.
- I. Eleftherianos, C. Heryanto, T. Bassal, W. Zhang, G. Tettamanti, and A. Mohamed, "Haemocyte-mediated immunity in insects: Cells, processes and associated components in the fight against pathogens and parasites", Immunology, vol. 164, no. 3, pp. 401–432, 2021.
- R. E. Wheeler, "Studies on the total haemocyte count and haemolymph volume in Periplaneta americana (L.) with special reference to the last moulting cycle", Journal of Insect Physiology, vol. 9, no. 2, pp. 223–235, 1963.
- N. Perveen and M. Ahmad, "Toxicity of some insecticides to the haemocytes of giant honeybee, Apis dorsata F. under laboratory conditions", Saudi journal of biological sciences, vol. 24, no. 5, pp. 1016–1022, 2017.
- T. Li, D. Yan, X. Wang, L. Zhang, and P. Chen, "Hemocyte changes during immune melanization in Bombyx mori infected with Escherichia coli", Insects, vol. 10, no. 9, p. 301, 2019.
- D. G. Perez and C. S. Fontanetti, "Hemocitical responses to environmental stress in invertebrates: a review", Environmental monitoring and assessment, vol. 177, no. 1, pp. 437–447, 2011.
- J. F. Hillyer and M. R. Strand, "Mosquito hemocyte-mediated immune responses", Current opinion in insect science, vol. 3, pp. 14–21, 2014.
- M. Meister and M. Lagueux, "Drosophila blood cells", Cellular microbiology, vol. 5, no. 9, pp. 573–580, 2003.
- B. M. Christensen, B. M. Huff, G. S. Miranpuri, K. L. Harris, and L. A. Christensen, "Hemocyte population changes during the immune response of Aedes aegypti to inoculated microfilariae of Dirofilaria immitis", The Journal of parasitology, pp. 119–123, 1989.
- S. V. Akhil and S. K. Thomas, "Bombardier beetles (Coleoptera: Carabidae: Brachininae) of India-notes on habit, taxonomy, and use as natural bio-control agents", in Frontiers in Biological Research: Proceedings of the National Seminar, Department of Zoology, University of Calicut, India, 2018.
- F. Impellitteri, A.-S. Curpăn, G. Plăvan, A. Ciobica, and C. Faggio, "Hemocytes: a useful tool for assessing the toxicity of microplastics, heavy metals, and pesticides on aquatic invertebrates", International journal of environmental research and public health, vol. 19, no. 24, p. 16830, 2022.
- R. R. James and J. Xu, "Mechanisms by which pesticides affect insect immunity", Journal of invertebrate pathology, vol. 109, no. 2, pp. 175–182, 2012.