Comparative transcriptomic analysis provides insights into the response to the benzo(a)pyrene stress in aquatic firefly (Luciola leii).

Many studies have reported that behavior and bioluminescence of fireflies could be affected by changes in environment conditions. However, little is known about how the deterioration of the aquatic environment affects aquatic fireflies, particularly with respect to molecular responses following exposure to water pollutants, such as benzo(a)pyrene (BaP), which is a key indicator in environmental risk assessment because of the hazards it poses. Here, whole transcriptome sequencing and gene expression analysis were performed on freshwater fireflies (Luciola leii) exposed to BaP (concentration of 0.01 mg/L). Four transcriptomic libraries were constructed for the control and treatment groups, including two biological replicates. From the mixed pools (each pool contains 60 individuals from three time points), a total of 54,282 unigenes were assembled. Furthermore, 329,337 of Single-nucleotide Polymorphisms (SNPs) and 1324 of Simple Sequence Repeats (SSRs) were predicted using bioinformatics, which is useful for the future development of molecular markers. Subsequently, 2414 differently expressed genes (DEGs) were identified in response to BaP stress in comparison to the control, including 1350 up-regulated and 1064 down-regulated DEGs. Functional enrichment showed that these DEGs are primarily related to innate immunity; xenobiotic biodegradation and response, biomacromolecule metabolism, biosynthesis, and absorption. Eight key BaP-responsive DEGs were screened to survey the dynamic changes of expression in response to BaP stress at different time points, and to validate the RNA sequencing data using quantitative real-time PCR. The results indicate that the expression of genes encoding UGT, CYP3A, CYP9, CYP6AS5 and ADHP were induced, while those encoding UGT2B10L, PTGDS, and ALDH were reduced, to participate in response to the BaP exposure and potentially help counteract the adverse effects of BaP. This investigation provides insight into the toxicological response of fireflies to the occurrence of water deterioration.

Neurotoxicity and mode of action of N, N-diethyl-meta-toluamide (DEET).

Recent studies suggest that N, N-diethyl-meta-toluamide (DEET) is an acetylcholinesterase inhibitor and that this action may result in neurotoxicity and pose a risk to humans from its use as an insect repellent. We investigated the mode of action of DEET neurotoxicity in order to define the specific neuronal targets related to its acute toxicity in insects and mammals. Although toxic to mosquitoes (LD50 ca. 1.5 µg/mg), DEET was a poor acetylcholinesterase inhibitor (<10% inhibition), even at a concentration of 10 mM. IC50 values for DEET against Drosophila melanogaster, Musca domestica, and human acetylcholinesterases were 6-12 mM. Neurophysiological recordings showed that DEET had excitatory effects on the housefly larval central nervous system (EC50: 120 µM), but was over 300-fold less potent than propoxur, a standard anticholinesterase insecticide. Phentolamine, an octopamine receptor antagonist, completely blocked the central neuroexcitation by DEET and octopamine, but was essentially ineffective against hyperexcitation by propoxur and 4-aminopyridine, a potassium channel blocker. DEET was found to illuminate the firefly light organ, a tissue utilizing octopamine as the principal neurotransmitter. Additionally, DEET was shown to increase internal free calcium via the octopamine receptors of Sf21 cells, an effect blocked by phentolamine. DEET also blocked Na(+) and K(+) channels in patch clamped rat cortical neurons, with IC50 values in the micromolar range. These findings suggest DEET is likely targeting octopaminergic synapses to induce neuroexcitation and toxicity in insects, while acetylcholinesterase in both insects and mammals has low (mM) sensitivity to DEET. The ion channel blocking action of DEET in neurons may contribute to the numbness experienced after inadvertent application to the lips or mouth of humans.

Modulations in the light of the firefly.

Continuous light could be produced from the firefly by making it inhale vapours of ethyl acetate. Here we perform such a control experiment on the Indian species of the firefly Luciola praeusta Kiesenwetter 1874 (Coleoptera : Lampyridae : Luciolinae), and analyse the light in the microsecond time scale. The amplitude of the continuous train of triangular pulses is apparently altered in accordance with the instantaneous values of a hypothetical signal, which exhibits pulse amplitude modulation (PAM). In addition to sampling in amplitude, this scheme apparently provides sampling in time, representing pulse width modulation (PWM). A Fourier transform spectrum of this waveform shows the 'carrier' frequency and the accompanying 'side bands'.

Effect of halogenated fluorescent compounds on bioluminescent reactions.

The paper investigates an application of luminescent bioassays to monitor the toxicity of organic halides. Effects of xanthene dyes (fluorescein, eosin Y, and erythrosin B), used as model compounds, on bioluminescent reactions of firefly Luciola mingrelica, marine bacteria Photobacterium leiognathi, and hydroid polyp Obelia longissima were studied. Dependence of bioluminescence quenching constants on the atomic weight of halogen substituents in dye molecules was demonstrated. Bacterial bioluminescence was shown to be most sensitive to heavy halogen atoms involved in molecular structure; hence, it is suitable for construction of sensors to monitor toxicity of halogenated compounds. Mechanisms of bioluminescence quenching--energy transfer processes, collisional interactions, and enzyme-dye binding--were considered. Changes of bioluminescence (BL) spectra in the presence of the dyes were analyzed. Interactions of the dyes with enzymes were studied using fluorescence characteristics of the dyes in steady-state and time-resolved experiments. The dependences of fluorescence anisotropy of enzyme-bound dyes, the average fluorescence lifetime, and the number of exponential components in fluorescence decay on the atomic weight of halogen substituents were demonstrated. The results are discussed in terms of "dark effect of heavy halogen atom" in the process of enzyme-dye binding; hydrophobic interactions were assumed to be responsible for the effect.

The light of the firefly under the influence of ethyl acetate.

When a firefly is made to inhale ethyl acetate vapour, a constant glow appears after a few minutes from its abdominal lantern. This control experiment has been performed by a few workers to record the emission spectrum of the firefly. However, a time-resolved experiment performed by us on this continuous light emitted by the species Luciola praeusta Kiesenwetter 1874 (Coleoptera : Lampyridae : Luciolinae) reveals that it is composed of a continuous train of tiny pulses! The nature of the pulses suggests that an oscillatory chemical reaction continues in the microsecond time scale in the lantern of the anaesthetized firefly.

The effects of ethylene glycol and ethanol on the body mass and elemental composition of insects collected with pitfall traps.

Insects often used as accumulation indicators of hazardous elements. Pitfall traps with ethylene glycol as trapping fluid are frequently used to collect insects. We studied the effect of glycol and preservation with ethanol on the elemental composition of hand collected firebugs. Control samples were stored in a freezer and the following treatments were used: insect kept in trapping fluid for 2 weeks, and for 1 month, trapping fluid for 2 weeks plus 2 weeks in ethanol, and trapping fluid for 1 month plus ethanol for 1 month. Insects kept in trapping fluid gained mass with respect to control: 26% for the short trapping and 37% for the long trapping. Preservation in ethanol reversed this effect in each case. Trapping fluid did not alter the dry mass. A significant loss in dry mass only occurred in the long trapping plus long preservation treatment. We analysed the concentration of eight elements: Ca, Cu, K, Mg, Mn, Na, Sr and Zn. We found significant difference in the concentrations of elements among the four treatments in the case of all elements, except magnesium and zinc. Our results indicate the potential of both certain trapping fluids as well as preservation in ethanol influencing the concentration of certain elements in insects. Live trapping for collection and storage in under freezing conditions for preservation could be a more reliable method if quantitative analytical studies are to be performed, when invertebrates are used as indicators of the presence and concentrations of hazardous substances in the environment.