In insects, external chemical defences, such as reflex bleeding, have been proved to be an efficient strategy against various predators. At the same time, significant costs of reflex bleeding can be expected because bled haemolymph is lost and all valuable components included have to be renewed. Interestingly, this issue has rarely been investigated for adult insects. In this study, we examined the immune and fitness costs of repeated reflex bleeding in adults of the invasive ladybird Harmonia axyridis, investigating several haemolymph parameters. Reflex bleeding induced twice a week for three weeks resulted in a significant reduction in haemocyte concentration, total protein content, and antimicrobial activity against Micrococcus luteus, and a marginally non-significant decrease in antimicrobial activity against Escherichia coli. Repeated reflex bleeding did not result in significant body mass reduction. Interestingly, resource limitation in the form of complete food absence did not significantly interact with reflex bleeding, even though starvation itself had a strong negative effect on all haemolymph parameters investigated and individual body mass. Daily reflex bleeding did not result in decreased fecundity of young ladybirds during the first 30 days of their adult life, but the start of ladybird reproduction was delayed by about two days. Moreover, ladybirds bleeding larger amounts of haemolymph started their reproduction significantly later. Overall, our results indicate that repeated reflex bleeding weakens a ladybird's immune system and can increase their susceptibility to pathogens, but a ladybird's reproductive potential remains almost unaffected, even by very intensive reflex bleeding.

Department of Ecology Faculty of Environmental Sciences Czech University of Life Sciences Prague Kamýcká 129 Prague Suchdol 165 00 Czech Republic

Zobrazit více v PubMed

Cuénot L. Sur la saignée réflexe et les moyens défense de quelques insectes. Arch. Zool. Exp. Gen. 1896;4:655–680.

Hollande CH. L’autohémorrhée ou le rejet du sang chez les insectes (toxicologie du sang) Archs. Anal. Microsc. 1911;13:171–318.

Bateman PW, Fleming PA. There will be blood: autohaemorrhage behaviour as part of the defence repertoire of an insect. Journal of Zoology. 2009;278:342–348. doi: 10.1111/j.1469-7998.2009.00582.x. DOI

Bugila AAA, Franco JC, da Silva EB, Branco M. Defense Response of Native and Alien Mealybugs (Hemiptera: Pseudococcidae) Against the Solitary Parasitoid Anagyrus sp nr. pseudococci (Girault) (Hymenoptera: Encyrtidae) Journal of Insect Behavior. 2014;27:439–453. doi: 10.1007/s10905-014-9440-x. DOI

Moore KA, Williams DD. Novel Strategies in the Complex Defense Repertoire of a Stonefly (Pteronarcys Dorsata) Nymph. Oikos. 1990;57:49–56. doi: 10.2307/3565735. DOI

Drilling K, Dettner K. First insights into the chemical defensive system of the erotylid beetle, Tritoma bipustulata. Chemoecology. 2010;20:243–253. doi: 10.1007/s00049-010-0054-2. DOI

Fu XH, Nobuyoshi O, Meyer-Rochow VB, Wang YY, Lei CL. Reflex-bleeding in the firefly Pyrocoelia pectoralis (Coleoptera: Lampyridae): Morphological basis and possible function. Coleopterists Bulletin. 2006;60:207–215. doi: 10.1649/892.1. DOI

Nicolson SW. Water Replenishment Following Reflex Bleeding in the Blister Beetle Decapotoma-Lunata Pallas (Coleoptera, Meloidae) African Entomology. 1994;2:21–23.

Sato S, Kushibuchi K, Yasuda H. Effect of reflex bleeding of a predatory ladybird beetle, Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae), as a means of avoiding intraguild predation and its cost. Applied Entomology and Zoology. 2009;44:203–206. doi: 10.1303/aez.2009.203. DOI

Boevé J-L, Schaffner U. Why does the larval integument of some sawfly species disrupt so easily? The harmful hemolymph hypothesis. Oecologia. 2003;134:104–111. doi: 10.1007/s00442-002-1092-4. PubMed DOI

Higginson AD, Delf J, Ruxton GD, Speed MP. Growth and reproductive costs of larval defence in the aposematic lepidopteran Pieris brassicae. Journal of Animal Ecology. 2011;80:384–392. doi: 10.1111/j.1365-2656.2010.01786.x. PubMed DOI

Sword GA. Tasty on the outside, but toxic in the middle: grasshopper regurgitation and host plant-mediated toxicity to a vertebrate predator. Oecologia. 2001;128:416–421. doi: 10.1007/s004420100666. PubMed DOI

Novgorodova TA. Role of social and individual experience in interaction of the meadow ant Formica pratensis (Hymenoptera: Formicidae) with ladybird imagines and hoverfly larvae. Insect Science. 2015;22:440–450. doi: 10.1111/1744-7917.12127. PubMed DOI

Zvereva EL, Kozlov MV. The costs and effectiveness of chemical defenses in herbivorous insects: a meta-analysis. Ecological Monographs. 2016;86:107–124. doi: 10.1890/15-0911.1. DOI

Eisner, T., Eisner, M. & Siegler, M. V. S. Secret weapons: defenses of insects, spiders, scorpions, and other many-legged creatures. (Harvard University Press, 2005).

Holloway GJ, de Jong PW, Brakefield PM, de Vos H. Chemical defence in ladybird beetles (Coccinellidae). 1. Distribution of coccinelline and individual variation in defence in 7-spot ladybirds (Coccinella septempunctata) Chemoecology. 1991;2:7–14. doi: 10.1007/BF01240660. DOI

Karystinou A, Thomas APM, Roy HE. Presence of haemocyte-like cells in coccinellid reflex blood. Physiological Entomology. 2004;29:94–96. doi: 10.1111/j.0307-6962.2004.0358.x. DOI

Knapp M, Dobes P, Rericha M, Hyrsl P. Puncture vs. reflex bleeding: Haemolymph composition reveals significant differences among ladybird species (Coleoptera: Coccinellidae), but not between sampling methods. European Journal of Entomology. 2018;115:1–6. doi: 10.14411/eje.2018.001. DOI

Grill CP, Moore AJ. Effects of a larval antipredator response and larval diet on adult phenotype in an aposematic ladybird beetle. Oecologia. 1998;114:274–282. doi: 10.1007/s004420050446. PubMed DOI

Rowell-Rahier M, Pasteels JM. Economics of Chemical Defense in Chrysomelinae. Journal of Chemical Ecology. 1986;12:1189–1203. doi: 10.1007/bf01639004. PubMed DOI

Holloway GJ, Dejong PW, Ottenheim M. The Genetics and Cost of Chemical Defense in the 2-Spot Ladybird (Adalia-Bipunctata L) Evolution. 1993;47:1229–1239. doi: 10.2307/2409988. PubMed DOI

Lee BW, Ugine TA, Losey JE. An Assessment of the Physiological Costs of Autogenous Defenses in Native and Introduced Lady Beetles. Environmental Entomology. 2018;47:1030–1038. doi: 10.1093/ee/nvy068. PubMed DOI

Rericha M, Dobes P, Hyrsl P, Knapp M. Ontogeny of protein concentration, haemocyte concentration and antimicrobial activity against Escherichia coli in haemolymph of the invasive harlequin ladybird Harmonia axyridis (Coleoptera: Coccinellidae) Physiological Entomology. 2018;43:51–59. doi: 10.1111/phen.12224. DOI

de Jong PW, Holloway GJ, Brakefield PM, de Vos H. Chemical defence in ladybird beetles (Coccinellidae). II. Amount of reflex fluid, the alkaloid adaline and individual variation in defence in 2-spot ladybirds (Adalia bipunctata) Chemoecology. 1991;2:15–19. doi: 10.1007/BF01240661. DOI

Schmidtberg H, Roehrich C, Vogel H, Vilcinskas A. A switch from constitutive chemical defence to inducible innate immune responses in the invasive ladybird Harmonia axyridis. Biology Letters. 2013;9:20130006. doi: 10.1098/rsbl.2013.0006. PubMed DOI PMC

Firlej A, Girard P-A, Brehelin M, Coderre D, Boivin G. Immune Response of Harmonia axyridis (Coleoptera: Coccinellidae) Supports the Enemy Release Hypothesis in North America. Annals of the Entomological Society of America. 2012;105:328–338. doi: 10.1603/an11026. DOI

Vilcinskas A, Mukherjee K, Vogel H. Expansion of the antimicrobial peptide repertoire in the invasive ladybird Harmonia axyridis. Proceedings of the Royal Society B-Biological Sciences. 2013;280:20122113. doi: 10.1098/rspb.2012.2113. PubMed DOI PMC

Yang SY, Ruuhola T, Rantala MJ. Impact of starvation on immune defense and other life-history traits of an outbreaking geometrid, Epirrita autumnata: a possible causal trigger for the crash phase of population cycle. Annales Zoologici Fennici. 2007;44:89–96.

Boggs CL. Understanding insect life histories and senescence through a resource allocation lens. Functional Ecology. 2009;23:27–37. doi: 10.1111/j.1365-2435.2009.01527.x. DOI

Povey S, Cotter SC, Simpson SJ, Lee KP, Wilson K. Can the protein costs of bacterial resistance be offset by altered feeding behaviour? Journal of Animal Ecology. 2009;78:437–446. doi: 10.1111/j.1365-2656.2008.01499.x. PubMed DOI

Simpson SJ, Raubenheimer D. The Central Role of the Hemolymph in the Regulation of Nutrient Intake in Insects. Physiological Entomology. 1993;18:395–403. doi: 10.1111/j.1365-3032.1993.tb00613.x. DOI

Hodek, I., Honěk, A. & van Emden, H. F. Ecology and Behaviour of the Ladybird Beetles (Coccinellidae). 600 (Willey-Blackwell, 2012).

Knapp M. Relative Importance of Sex, Pre-Starvation Body Mass and Structural Body Size in the Determination of Exceptional Starvation Resistance of Anchomenus dorsalis (Coleoptera: Carabidae) Plos One. 2016;11:e0151459. doi: 10.1371/journal.pone.0151459. PubMed DOI PMC

Chown SL, Le Lagadec MD, Scholtz CH. Partitioning variance in a physiological trait: desiccation resistance in keratin beetles (Coleoptera, Trogidae) Functional Ecology. 1999;13:838–844. doi: 10.1046/j.1365-2435.1999.00373.x. DOI

Bayoumy, M. H., Osawa, N. & Hatt, S. Fitness costs of reflex bleeding in the ladybird beetle Harmonia axyridis: the role of parental effects. Insect Science in press, 10.1111/1744-7917.12737 (2020). PubMed

Brown PMJ, et al. Harmonia axyridis in Europe: spread and distribution of a non-native coccinellid. Biocontrol. 2008;53:5–21. doi: 10.1007/s10526-007-9132-y. DOI

Brown PMJ, et al. The global spread of Harmonia axyridis (Coleoptera: Coccinellidae): distribution, dispersal and routes of invasion. Biocontrol. 2011;56:623–641. doi: 10.1007/s10526-011-9379-1. DOI

Lombaert E, et al. Bridgehead Effect in the Worldwide Invasion of the Biocontrol Harlequin Ladybird. Plos One. 2010;5:e9743. doi: 10.1371/journal.pone.0009743. PubMed DOI PMC

Tayeh A, et al. Biological invasion and biological control select for different life histories. Nature Communications. 2015;6:7268. doi: 10.1038/ncomms8268. PubMed DOI PMC

R Development Core Team. R: A language and environment for statistical computing., (R Foundation for Statistical Computing, Vienna, Austria, http://www.R-project.org, 2018).

Fungal ectoparasites increase winter mortality of ladybird hosts despite limited effects on their immune system