Ceramidases (CDases) are vital enzymes involved in the biosynthesis of sphingolipids, which are essential components of eukaryotic membranes. The function of these enzymes in insects, however, is poorly understood. We identified a neutral ceramidase (NlnCDase) from the brown planthopper, Nilaparvata lugens, one of the most destructive hemipteran pests of rice. The C12-ceramide was the most preferred substrate for the NlnCDase enzyme. The activity of the NlnCDase enzyme was highest in the neutral-pH range (pH 6.0). It was inhibited by EGTA, Cs+ and Fe2+, while stimulated by EDTA and Ca2+. Moreover, the NlnCDase has higher transcript level and activity in adults than in eggs and nymphs, and in the reproductive organs (ovaries and spermaries) than in other tissues (i.e. heads, thorax, legs, midguts), which suggested that the NlnCDase might be elevated to mediate developmental process. In addition, transcripts and activity of the NlnCDase were up-regulated under abiotic stresses including starvation, abnormal temperature, and insecticides, and biotic stress of resistant rice varieties. Knocking down NlnCDase by RNA interference increased female survival under starvation and temperature stresses, suggesting that NlnCDase might be involved in the stress response in N. lugens.

Cyrtorhinus lividipennis Reuter (Hemiptera: Miridae) is an important predatory natural enemy of planthopper and leafhopper eggs in Asian rice paddy fields. Cyrtorhinus lividipennis is known to rely largely on herbivore-induced plant volatiles to identify eggs embedded in rice stem tissues for feeding and on pheromones for mating. However, exactly how C. lividipennis decode these chemical information is unclear. In most insects, the odorant-binding proteins (OBPs) and the chemosensory proteins (CSPs) are essential for seeking out food resources and mates. In this study, we identified 10 OBP and 5 CSP genes in C. lividipennis and investigated their expression patterns in various tissues of adult males and females by quantitative real-time PCR (qRT-PCR). Six OBP genes (ClivOBP1, 2, 4, 6, 9, and 10) were mainly expressed in the male antennae, whereas three genes (ClivOBP3, ClivOBP7, and ClivOBP8) had high expression in the female antennae. ClivCSP1 was predominantly expressed in the male antennae. These findings suggest that most ClivOBPs and ClivCSPs are likely involved in food-searching behavior. The recognition of the pheromone molecules provides the basis for further functional studies on the chemoreception system of C. lividipennis.

Although sphingomyelins known to be are lipid constituents of the plasma membrane in vertebrates, much remains obscure about the metabolism of sphingomyelins in insects. With ultra performance liquid chromatography‐time‐of‐flight‐tandem mass spectrometry analysis, we revealed for the first time that sphingomyelins are abundant in Nilaparvata lugens (Stål), the brown planthopper (BPH), and their biosynthesis is carried out by sphingomyelin synthase‐like protein 2 (SMSL2), which is homologous to sphingomyelin synthase‐related protein (SMSr). Unlike other insect species, high concentrations of sphingomyelins rather than ceramide phosphoethanolamines exist in the BPH. Two putative genes, which are homologous to SMSr, are named Nilaparvata lugens SMS‐like 1 (NlSMSL1) and 2 (NlSMSL2). Knockdowns of both NlSMSL2 and NlSMSL1 were conducted but only the first decreased concentrations of sphingomyelins in the BPH, indicating that NlSMSL2 plays a role in the biosynthesis of sphingomyelins. Real‐time quantitative PCR analysis revealed both NlSMSL1 and NlSMSL2 are highly expressed in BPH adults, with NlSMSL1 specifically highly expressed in reproductive organs (ovaries and testes) whereas NlSMSL2 was highly expressed in the malpighian tubules. The knockdown of NlSMSL1 or NlSMSL2 increased BPH female body weight but not that of males, suggesting sex‐specific roles for SMSLs in influencing BPH body weight. The results suggest that NlSMSL2 catalyses the synthesis of sphingomyelins and maintains female BPH body weight through alteration of sphingolipid content.

Ceramides are bioactive sphingolipids that have been implicated in insect development; however, their role in insect reproduction remains poorly understood. Here, we report the pivotal role of neutral ceramidase (NCER) in the female reproduction of the brown planthopper (BPH), Nilaparvata lugens (Stål), a significant pest in rice cultivation in Asia. LC-MS/MS demonstrated that, among different developmental stages of BPH, the levels of ceramides were highest in 1st instar nymphs and lowest in adults. The transcription of NCER was negatively correlated with the levels of ceramides at different developmental stages of BPH, in that the transcript levels of NCER were the highest, whereas ceramides levels were the lowest in BPH adults. Knocking down NCER through RNA interference (RNAi) increased the levels of ceramides in BPH females and ovaries, which resulted in a delay in oocyte maturation, a reduction in oviposition and egg hatching rate, as well as the production of vulnerable offspring. Transmission electron microscopy (TEM) analysis and TdT-mediated dUTP Nick-End Labeling (TUNEL) assays showed mitochondrial deficiency and apoptosis in NCER-deficient oocytes. Taken together, these results suggest that NCER plays a crucial role in female reproduction in BPH, likely by regulating the levels of ceramides.

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