Chemical defense in the firefly, Rhagophthalmus ohbai (Coleoptera: Rhagophthalmidae)

Rhagophthalmus ohbai Wittmer (Iriomote botaru, in Japanese) can be found in the soil environment on several islands in Okinawa Prefecture, Japan. The females of this species have been reported to remain in very close proximity to their eggs for about 2 months while producing a pungent odor. We hypothesized that females produced this odor in order to protect their eggs and/or themselves from pathogens and/or predators. Analysis of this volatile substance by solid-phase microextraction and gas chromatography mass spectrometry (GC–MS) revealed that the odor was attributed to 1,4-naphthoquinone (NQ). We also examined resistance to NQ in four fungi isolated from dead fireflies [Aspergillus tubingensis (Schober) Mosseray 21KI501, A. fumigatus Fresenius 21KI502, Fusarium proliferatum (Matsushima) Nirenberg 21KI503, and Trichoderma asperellum Samuels et al. 21KI504], as well as four common soil bacteria [Escherichia coli (Migula) Castellani and Chalmers type B, Staphylococcus aureus subsp. aureus Rosenbach H209P, Bacillus subtilis (Ehrenberg) Cohn ATCC6633, and Rhodococcus jostii Takeuchi et al. RHA1]. The findings revealed that NQ exhibited antimicrobial activity in all of the test organisms, suggesting that female fireflies release NQ to protect their eggs and newly hatched larvae from pathogens in the soil. We also analyzed the volatiles produced by male fireflies using GC–MS and found that some male fireflies emitted NQ. It is possible that male fireflies produce NQ to protect themselves from pathogenic microorganisms in the soil environments that they inhabit during the day. Although several defensive chemicals produced by fireflies have been identified to date, the production of 1,4-naphthoquinone for chemical defense by R. ohbai has not yet been reported.