Litsea cubeba (Lour.) Pers. is a promising industrial crop with fruits rich in essential oils. The chemical composition of essential oils of L. cubeba (EOLC) were determined for fruits harvested from eight regions in China. The overall essential oil content, obtained by hydrodistillation and analyzed by gas chromatography–mass spectrometry (GC-MS), ranged from 3.04% to 4.56%. In total, 59 compounds were identified, the dominant components being monoterpenes (94.4–98.4%), represented mainly by neral and geranial (78.7–87.4%). D-Limonene was unexpectedly a lesser constituent (0.7–5.3%) in fruits, which differed from previous reports (6.0–14.6%). Several components were only detected in certain regions and compounds such as o-cymene and eremophilene have never before been reported in EOLC. These results demonstrate significant regional variation in the chemical composition of EOLC. This investigation provides important information with regard to the bioactivity, breeding work and industrial applications of L. cubeba.

Quantitative real-time PCR (RT-qPCR) has become an accurate and widely used technique to analyze expression levels of selected genes. It is very necessary to select appropriate reference genes for gene expression normalization. In the present study, we assessed the expression stability of 11 reference genes including eight traditional housekeeping genes and three novel genes in different tissues/organs and developing seeds from four cultivars of tung tree. All 11 reference genes showed a wide range of Ct values in all samples, indicating that they differently expressed. Three softwares – geNorm, NormFinder and BestKeeper – were used to determine the stability of these references except for ALB (2S albumin), which presented a little divergence. The results from the three softwares showed that ACT7 (Actin7a), UBQ (Ubiquitin), GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and EF1α (elongation factor 1-α) were the most stable reference genes across all of the tested tung samples and tung developing seeds, while ALB (2S albumin) was unsuitable as internal controls. ACT7, EF1β (elongation factor1-beta), GAPDH and TEF1 (transcription elongation factor 1) were the top four choices for different tissues/organs whereas LCR69 did not favor normalization of RT-qPCR in these tissues/organs. Meanwhile, the expression profiles of FAD2 and FADX were realized using stable reference genes. The relative quantification of the FAD2 and FADX genes varied according to the internal controls and the number of internal controls. The results further proved the importance of the choice of reference genes in the tung tree. These stable reference genes will be employed in normalization and quantification of transcript levels in future expression studies of tung genes.

The camellia weevil (CW [Curculio chinensis]) is a notorious host-specific predator of the seeds of Camellia species in China, causing seed losses of up to 60%. The weevil is capable of overcoming host tree chemical defenses, while the mechanisms of how these beetles contend with the toxic compounds are still unknown. Here, we examined the interaction between the gut microbes of CW and camellia seed chemistry and found that beetle-associated bacterial symbionts mediate tea saponin degradation. We demonstrate that the gut microbial community profile of CW was significantly plant associated, and the gut bacterial community associated with CW feeding on Camellia oleifera seeds is enriched with genes involved in tea saponin degradation compared with those feeding on Camellia sinensis and Camellia reticulata seeds. Twenty-seven bacteria from the genera Enterobacter, Serratia, Acinetobacter, and Micrococcus subsisted on tea saponin as a sole source of carbon and nitrogen, and Acinetobacter species are identified as being involved in the degradation of tea saponin. Our results provide the first metagenome of gut bacterial communities associated with a specialist insect pest of Camellia trees, and the results are consistent with a potential microbial contribution to the detoxification of tree-defensive chemicals. IMPORTANCE The gut microbiome may play an important role in insect-plant interactions mediated by plant secondary metabolites, but the microbial communities and functions of toxic plant feeders are still poorly characterized. In the present study, we provide the first metagenome of gut bacterial communities associated with a specialist weevil feeding on saponin-rich and saponin-low camellia seeds, and the results reveal the correlation between bacterial diversity and plant allelochemicals. We also used cultured microbes to establish their saponin-degradative capacity outside the insect. Our results provide new experimental context to better understand how gut microbial communities are influenced by plant secondary metabolites and how the resistance mechanisms involving microbes have evolved to deal with the chemical defenses of plants.

Aromatic essential oils extracted from fresh fruits of Litsea cubeba (Lour.) Pers., have diverse medical and economic values. The dominant components in these essential oils are monoterpenes and sesquiterpenes. Understanding the molecular mechanisms of terpenoid biosynthesis is essential for improving the yield and quality of terpenes. However, the 40 available L. cubeba nucleotide sequences in the public databases are insufficient for studying the molecular mechanisms. Thus, high-throughput transcriptome sequencing of L. cubeba is necessary to generate large quantities of transcript sequences for the purpose of gene discovery, especially terpenoid biosynthesis related genes. Using Illumina paired-end sequencing, approximately 23.5 million high-quality reads were generated. De novo assembly yielded 68,648 unigenes with an average length of 834 bp. A total of 38,439 (56%) unigenes were annotated for their functions, and 35,732 and 25,806 unigenes could be aligned to the GO and COG database, respectively. By searching against the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG), 16,130 unigenes were assigned to 297 KEGG pathways, and 61 unigenes, which contained the mevalonate and 2-C-methyl-D-erythritol 4-phosphate pathways, could be related to terpenoid backbone biosynthesis. Of the 12,963 unigenes, 285 were annotated to the terpenoid pathways using the PlantCyc database. Additionally, 14 terpene synthase genes were identified from the transcriptome. The expression patterns of the 16 genes related to terpenoid biosynthesis were analyzed by RT-qPCR to explore their putative functions. RNA sequencing was effective in identifying a large quantity of sequence information. To our knowledge, this study is the first exploration of the L. cubeba transcriptome, and the substantial amount of transcripts obtained will accelerate the understanding of the molecular mechanisms of essential oils biosynthesis. The results may help improve future genetic and genomics studies on the molecular mechanisms behind the chemical composition of essential oils in L. cubeba fruits.

Quercus fabri Hance, Quercus serrata Thunb, Quercus acutissima Carruth, and Quercus variabilis BL are four Chinese oak species commonly used for forestation. To ensure the survival of seedlings, we first need to understand the differences in drought resistance of the four oak species at the seedling stage, and comprehensively evaluate their drought resistance capabilities. The four oak seedlings were divided into drought-rewatering treatment group and well watered samples (control group). For the seedlings of the drought-rewatering treatment group, drought stress lasting 31 days was used, and then re-watering for 5 days. The water parameters, osmotic solutes content, antioxidant enzyme activity and photosynthesis parameters of the seedlings in the two groups were measured every 5 days. Compared with the control group, the relative water content, water potential, net photosynthetic rate, transpiration rate, and stomatal conductance levels of the four oaks all showed a downward trend under continuous drought stress, and showed an upward trend after rehydration. The soluble protein, soluble sugar, proline, peroxidase, superoxide dismutase and catalase content of the four oaks increased first and then decreased under drought stress, and then increased after rehydration. The content of glycine betaine and malondialdehyde continued to increase, and gradually decreased after rehydration. The weight of each index was calculated by principal component analysis, and then the comprehensive evaluation of each index was carried out through the membership function method. The drought resistance levels of the four oak species were as follows: Q. serrata > Q. fabri > Q. variabilis > Q. acutissima.