Fuzheng Huayu recipe inhibits bleomycin-induced pulmonary fibrosis in rats by inhibiting M2 polarization of macrophages via the oxidative phosphorylation pathway.

Fuzheng Huayu recipe (FZHYR) is a Chinese patent medicine for the treatment of fibrosis. The effects of FZHYR on pulmonary fibrosis and macrophage polarization were investigated in vitro. FZHYR inhibited pulmonary inflammation and fibrosis and M2 polarization of macrophages in bleomycin-induced pulmonary fibrosis (BPF) of rat model. Differentially expressed genes were screened by high-throughput mRNA sequencing and GSEA showed that oxidative phosphorylation (OXPHOS) was correlated with BPF. FZHYR inhibited expressions of Ndufa2 and Ndufa6 in lung tissues of BPF rats. These findings suggest that OXPHOS pathway serves as a possible target for pulmonary fibrosis therapy by FZHYR.

Stereospecific synthesis of S-(-)-trans-verbenol and its antipode by inversion of sterically hindered alcohols.

S-(-)-trans-Verbenol (1) and its antipode, R-(+)-trans-verbenol (1') have been confirmed as the critical pheromone components of bark beetles. Synthesis of these two active secondary alcohols (1 and 1') from commercially available starting materials S-α-pinene and R-α-pinene was reported. The key steps were mainly depended on the effective SN2 stereo-inversion of the hydroxy group of sterically hindered alcohols (3 and 3'), using Mitsunobu reaction or hydrolysis of mesylate ester, alternatively. Our results provide a new and stereo-selectivity way to obtain optically active insect pheromones.

Monoterpenoid signals and their transcriptional responses to feeding and juvenile hormone regulation in bark beetle Ips hauseri.

Hauser's engraver beetle, Ips hauseri, is a serious pest in spruce forest ecosystems in Central Asia. Its monoterpenoid signal production, transcriptome responses and potential regulatory mechanisms remain poorly understood. The quality and quantity of volatile metabolites in hindgut extracts of I. hauseri were found to differ between males and females and among three groups: beetles that were newly emerged, those with a topical application of juvenile hormone III (JHIII) and those that had been feeding for 24 h. Feeding males definitively dominated monoterpenoid signal production in I. hauseri, which uses (4S)-(-)-ipsenol and (S)-(-)-cis-verbenol to implement reproductive segregation from Ipstypographus and Ipsshangrila. Feeding stimulation induced higher expression of most genes related to the biosynthesis of (4S)-(-)-ipsenol than JHIII induction, and showed a male-specific mode in I. hauseri. JHIII stimulated males to produce large amounts of (-)-verbenone and also upregulated the expression of several CYP6 genes, to a greater extent in males than in females. The expression of genes involved in the metabolism of JHIII in females and males was also found to be upregulated. Our results indicate that a species-specific aggregation pheromone system for I. hauseri, consisting of (4S)-(-)-ipsenol and S-(-)-cis-verbenol, can be used to monitor population dynamics or mass trap killing. Our results also enable a better understanding of the bottom-up role of feeding behaviors in mediating population reproduction/aggregation and interspecific interactions.

Effect of nicotine on L-dopa-induced dyskinesia in animal models of Parkinson's disease: a systematic review and meta-analysis.

L-dopa-induced dyskinesias (LIDs) are abnormal involuntary movements (AIM) that develop with long-term L-dopa therapy for Parkinson's disease (PD). In this study, we used these tools to describe the efficacy of nicotine reduced LID in animal models of PD. Studies were identified by electronic searching of six online databases up to September of 2013 to identify preclinical trials involving nicotine for LID in animal model. Data were extracted for AIM compared with LID animals. Pre-specified subgroup analysis was carried out according to method of model, gender, anesthetic used, and species. Combined standardized mean difference (SMD) estimates and 95 % confidence intervals (CIs) were calculated using a random-effects model. Eleven studies involving 181 animals which described the effect of nicotine on LID were included in the meta-analysis. Nicotine was efficacious in reducing total AIM compared with control group (SMD -3.77, 95 % CI -5.30 to -2.23, P < 0.00001). Meanwhile, four studies showed certain effects of nicotine for improving the axial AIM (SMD -2.21, 95 % CI -4.17 to -0.24, P = 0.03); oral AIM and forelimb AIM were obvious improved in six studies in the nicotine group (SMD -3.00, 95 % CI -4.55 to -1.44, P = 0.0002; SMD -2.52, 95 % CI -3.52 to -1.53, P < 0.00001, respectively). We conclude that nicotine appears to have efficacy in animal models of LID. Large randomized clinical trials testing the effect of nicotine in PD patients with LID are warranted.

The association between the LRRK2 G2385R variant and the risk of Parkinson's disease: a meta-analysis based on 23 case-control studies.

Clinical diagnosis of Parkinson's disease (PD) is essential but misdiagnosis of PD-like diseases is quite common. LRRK2 G2385R variants have been extensively examined for the association to the risk of Parkinson's disease. However, results from different studies are inconsistent. The purpose of this meta-analysis was to assess the association between the LRRK2 G2385R variants and the risk of PD. A systematic literature search was performed for 6 databases up to January of 2014 to identify case-control studies involving LRRK2 G2385R variants and the risk of PD. A total of 12,915 cases and 12,451 controls in 23 case-control studies were included in this meta-analysis. The results indicated that the variant A allele carriers (GA + AA) increased risk of PD when compared with the homozygote GG (GA + AA vs. GG: OR = 2.4, 95 % CI = 1.97 to 2.92, P < 0.00001). In the subgroup analysis by ethnicity, increased risks were identified among Chinese (OR = 2.69, 95 % CI = 2.1-3.45, P < 0.00001) as well as in non-Chinese (OR = 2.17, 95 % CI 1.75-2.69, P < 0.00001). In the subgroup analysis by age of onset, significant associations were found in both later-onset PD (LOPD) and early-onset PD (EOPD) cases. And there was no significant difference of the allele frequency between patients with LOPD and EOPD (OR = 1.18, 95 % CI = 0.77-1.80, P = 0.45). Our results suggest that the LRRK2 G2385R variants contribute to the susceptibility of PD especially in Chinese PD. Meanwhile, it is possible that age is not the risk factor to facilitate G2385R gene mutation.

Effects of Sisheng Decoction on the immunity and anti-stress function in mice with spleen deficiency syndrome.

OBJECTIVE: To study the possible mechanism of Sisheng Decoction on spleen deficiency syndrome via the observation of general conditions, immunity and anti-stress function in Dahuang (Radix et Rhizoma Rhei Palmati)-induced mice model. METHODS: Mice were randomized and grouped based on the body weight. The establishment of model and the treatment were done simultaneously. Except the mice in normal group, the decoction and the Dahuang powder were separately given in the morning and the afternoon for 14 d. The general condition of the mice, the rectal temperature, the time of burden swimming, the indexes of thymus and spleen and the interleukin-2 (IL-2) concentration in the serum were observed. RESULTS: The group treated with Sisheng Decoction showed better performance than that of the model group, including less stool, strong appetite and fast growth; the medium- and high-dose Sisheng Decoction increased the rectal temperature of mice. There was no statistical difference in the thymus and spleen indexes between the groups treated by Sisheng Decoction and the normal group (P>0.05). The thymus index in groups treated by sisheng Decoction was significantly elevated as compared with the model group (P<0.05). The medium- and high-dose Sisheng Decoction significantly improved the concentration of IL-2 and prolonged the time of burden swimming, as compared to the model group (P<0.05). CONCLUSION: The medium- and high-dose Sisheng Decoction is good at invigorating spleen and replenishing qi. One of the possible mechanisms may be related with the improvement of the immunity and anti-stress function of the body.

Regulation of autophagy and lipid accumulation under phosphate limitation in Rhodotorula toruloides.

Background: It is known that autophagy is essential for cell survival under stress conditions. Inorganic phosphate (Pi) is an essential nutrient for cell growth and Pi-limitation can trigger autophagy and lipid accumulation in oleaginous yeasts, yet protein (de)-phosphorylation and related signaling events in response to Pi limitation and the molecular basis linking Pi-limitation to autophagy and lipid accumulation remain elusive. Results: Here, we compared the proteome and phosphoproteome of Rhodotorula toruloides CGMCC 2.1389 under Pi-limitation and Pi-repletion. In total, proteome analysis identified 3,556 proteins and the phosphoproteome analysis identified 1,649 phosphoproteins contained 5,659 phosphosites including 4,499 pSer, 978 pThr, and 182 pTyr. We found Pi-starvation-induced autophagy was regulated by autophagy-related proteins, but not the PHO pathway. When ATG9 was knocked down, the engineered strains produced significantly less lipids under Pi-limitation, suggesting that autophagy required Atg9 in R. toruloides and that was conducive to lipid accumulation. Conclusion: Our results provide new insights into autophagy regulation under Pi-limitation and lipid accumulation in oleaginous yeast, which should be valuable to guide further mechanistic study of oleaginicity and genetic engineering for advanced lipid producing cell factory.

Sensilla on antennae and mouthparts of adult spruce bark beetle Ips typographus (Coleoptera: Curculionidae).

The spruce bark beetle, Ips typographus, is an invasive forest pest that occurs across Eurasia. To further understand the mechanism of insect-host chemical communication and the feeding choices of adult I. typographus, we examined the sensilla on the antennae and mouthparts of I. typographus using scanning electron microscopy. No sexual dimorphism in sensilla type or distribution was observed. Ten subtypes of antennal sensilla were identified: sensilla trichodea Types I-III (TR1 , TR2 , and TR3 ); sensilla chaetica Types I and II (CH1 and CH2 ); sensilla basiconica Types I-III (BA1 , BA2 , and BA3 ); sensilla coeloconica (CO); and Böhm sensilla (BS). BA2 were the most abundant among the antennal sensilla in both sexes. Thirteen mouthpart sensilla subtypes were observed: sensilla trichodea Types I-IV (TR1 , TR2 , TR3 , and TR4 ); sensilla chaetica Types I-III (CH1 , CH2 , and CH3 ); sensilla digitiformia (DI); sensilla basiconica Types I-II (BA1 and BA2 ); and sensilla styloconica Types I-III (ST1 , ST2 , and ST3 ). Among these, TR4 were the most abundant in both sexes. The apex of each maxillary and labial palp carried the same sensilla (BA2 , ST1 , ST2 , and ST3 ), although the apex of the maxillary palp contained more total sensilla. The functional roles of each sensilla type are discussed based on their external structure and distribution. The results provide a theoretical basis for further studies on the olfactory and feeding behaviors and electrophysiology of adult I. typographus.

SEM analysis of sensilla on the mouthparts and antennae of Asian larch bark beetle Ips subelongatus.

The Asian larch bark beetle, Ips subelongatus, is a severe pest of larches in Northeastern China. The gustatory and olfactory systems of I. subelongatus play important roles in host location, mating, and feeding. In this study, we examined the types, distributions, and abundances of various sensilla associated with the mouthparts and antennae of I. subelongatus using scanning electron microscopy (SEM). On the mouthparts, five types of sensilla are present: sensilla trichodea (S.t.1-3), sensilla chaetica (S.c.1-3), sensilla basiconica (S.b.1-2), sensilla twig basiconica (S.tb.1-3), and sensilla placodea (S.p). S.t.3 are the most abundant sensilla subtype on the mouthparts in both sexes, while S.b.1 are the least abundant. Most sensilla on the mouthparts are located on the maxillae and labium, and the apex of each maxillary and labial palp carry the same sensilla subtypes (S.b.2 and S.tb.1-3). However, the total number of sensilla on the apex of each maxillary palp is higher than that on the labial palp. On the antennae, five types of sensilla are present: sensilla trichodea (S.t.1-3), sensilla chaetica (S.c.1-2), sensilla basiconica (S.b.1-3), Böhm bristles (B.b), and sensilla coeloconica (S.co). Antennal sensilla are mostly situated on the anterior surface of the antennal club, particularly on the two dense sensory bands. S.b.1 are the most abundant sensilla subtype on the antennae in both sexes, while S.t.1 are the least abundant. No sexual dimorphism in sensilla type or distribution on the mouthparts or antennae is observed between the sexes of I. subelongatus. However, S.t.3 (on mouthparts) and S.c.1 (on antennae) were significantly more abundant in males than in females, while more S.t.1 (on mouthparts) were observed in females than in males. Finally, the putative functions of each kind of sensilla with respect to their fine structures, distributions, and abundances on the mouthparts and antennae are discussed.

The Efficacy of Long-Term Chinese Herbal Medicine Use on Lung Cancer Survival Time: A Retrospective Two-Center Cohort Study with Propensity Score Matching.

OBJECTIVE: To explore the efficacy of long-term use of Chinese herbal medicine (CHM) on survival time of lung cancer. METHODS: We conducted a retrospective cohort study on lung cancer patients. A propensity score matching (PSM) was performed to balance the covariates. Progression-free survival (PFS) was the primary endpoint and overall survival (OS) was the secondary endpoint. Patients who received CHM therapy from the initial date of diagnosis of lung cancer were included in the CHM group. Patients who were not treated with CHM during the same interval were categorized in the control group. A Cox regression model was used to explore the prognostic factors related to lung cancer. Hazard ratios of different subgroups were also analyzed. RESULTS: A total of 1134 patients were included in our study: 761 patients were in the CHM group and 373 patients were in the control group. After PSM, the mPFS and mOS in the CHM group were 70.4 months and 129.1 months, respectively, while the mPFS and mOS in the control group were 23.8 months and 99.7 months, respectively. The results of survival analysis on each stage demonstrated that patients may benefit from the long-term CHM treatment especially for patients with early stage. One-year to ten-year progression-free survival rates in the CHM group were higher than those in the control group (p < 0.001). COX multivariate regression analysis indicated that CHM treatment, female, low age at diagnosis, early tumor stage, and surgery were independent protective factors against recurrence and metastasis of lung cancer. Subgroup analysis showed that CHM treatment could reduce the risk of recurrence and metastasis in each subgroup (p < 0.01). CONCLUSION: Long-term CHM treatment with the Fuzheng Quxie Formula, which can be flexibly applied in the course of lung cancer treatment, not only has a positive influence on the progression-free survival time of lung cancer patients, but also reduces the risk of recurrence and metastasis of lung cancer.

Differences in Gut Bacterial Communities of Ips typographus (Coleoptera: Curculionidae) Induced by Enantiomer-Specific α-Pinene.

The spruce bark beetle (Ips typographus L.) is a destructive pest of Eurasian spruce forests. Although the gut bacteria of this insect are considered to play important roles in its lifecycle, the relationship between I. typographus and its gut bacterial community is poorly characterized. In this study, 16S rRNA gene sequencing was used to determine gut bacterial community composition across successive I. typographus life stages. Responses of the gut bacteria to α-pinene enantiomers were also explored. Ips typographus gut bacterial populations were dominated by the phyla Proteobacteria, Firmicutes, Bacteroidetes, and Actinobacteria, and the relative abundance of these phyla varied across different developmental stages of the beetle. Bacterial species diversity and richness indices increased with developmental stage progression. Relative abundances of the dominant genera, Erwinia (Enterobacteriales: Enterobacteriaceae), Pseudoxanthomonas (Xanthomonadales: Xanthomonadaceae), Serratia (Enterobacteriales: Enterobacteriaceae), and Romboutsia (Clostridiales: Peptostreptococcaceae), also varied across successive I. typographus life stages. Large disparities in the gut bacterial community of male adults were observed when the beetles were treated with S-(-)-α-pinene and R-(+)-α-pinene. The relative abundances of Lactococcus (Lactobacillales: Streptococcaceae) and Lelliottia (Enterobacteriales: Enterobacteriaceae) increased drastically with R-(+)-α-pinene and S-(-)-α-pinene treatment, respectively. This indicated a distinct enantiomer-specific effect of α-pinene on the I. typographus gut bacteria. This study demonstrated the plasticity of gut bacteria during I. typographus development, when α-pinene host monoterpenes are encountered. This study provides new insights into the relationship between 'I. typographus-gut bacteria' symbionts and host trees.

Epibiotic Fungal Communities of Three Tomicus spp. Infesting Pines in Southwestern China.

The association between insects and fungi has evolved over millions of years and is ubiquitous in nature. This symbiotic relationship holds critical implications for both partners, the insects and the associated microbes. Numerous fungi are externally allied with bark beetles and form a close symbiosis, but the community structures of these fungi are largely unknown. In Yunnan Province in southwestern China, the beetles Tomicus yunnanensis, T. minor, and T. brevipilosus are major forest pests that cause large losses of two indigenous pines, Pinus yunnanensis and P. kesiya. In this study, we used the Illumina MiSeq PE300 platform to process 48 samples of epibiotic fungal communities pooled from 1348 beetles; the beetles were collected during both the branch- and trunk-infection sections from five locations across Yunnan Province. Considerably greater species richness was detected using high-throughput sequencing of amplified internal transcribed spacer 1 (ITS1) ribosomal DNA (rDNA) libraries than previously documented by using culture-dependent methods. In total, 1,413,600 reads were generated, and a 97% sequence-similarity cutoff produced eight phyla, 31 classes, 83 orders, 181 families, 331 genera, 471 species, and 1157 operational taxonomic units (OTUs), with 659, 621, and 609 OTUs being confined to T. yunnanensis, T. minor, and T. brevipilosus, respectively. Tomicus yunnanensis, T. minor, and T. brevipilosus had the similar OTUs richness and evenness of fungal communities in Yunnan Province; nevertheless, the two fungal community compositions associated with T. yunnanensis and T. minor were structurally similar to each other but distinct from that associated with T. brevipilosus. Lastly, the results of principal co-ordinates analysis suggested that epibiotic fungal community structures of the three Tomicus spp. were conditioned strongly by the locations and pine hosts but weakly by beetle species and infection sections. Our findings provide baseline knowledge regarding the epibiotic fungal communities of three major Tomicus spp. in southwestern China.

[Effects of Sisheng Decoction on spontaneous activity and serum concentration of malondialdehyde in mice with yin deficiency syndrome].

OBJECTIVE: To assess the therapeutic effects of Sisheng Decoction, a compound traditional Chinese herbal medicine, on a mouse model of yin deficiency syndrome induced by thyroid hormone, and to make the preliminary study on its mechanisms. METHODS: Simultaneous modeling and treatment were carried out. Sixty mice were randomly divided into six groups: normal group, yin deficiency model group, low-, medium- and high-dose Sisheng Decoction group and Shengmai oral liquid group. Normal group and yin deficiency model group were administered with double distilled water. Spontaneous activity and serum concentration of malondialdehyde in different groups were detected. RESULTS: The symptoms of yin deficiency syndrome such as xerostomia, dysphoria and fervescence were improved in the Sisheng Decoction groups. Compared with the yin deficiency model group, the spontaneous activity was increased and the serum concentration of malondialdehyde was decreased in the Sisheng Decoction groups (P<0.05). There was no significant difference between the Sisheng Decoction groups and the Shengmai oral liquid group (P<0.05). CONCLUSION: Medium- or high-dose Sisheng Decoction is effective for nourishing yin, clearing heat, engendering liquid and allaying thirst. The above effects of Sisheng Decoction may be realised by improving the spontaneous activity and resisting oxidative damage.

Lipoic acid alleviates L­DOPA­induced dyskinesia in 6­OHDA parkinsonian rats via anti­oxidative stress.

Levodopa (L­DOPA) is the gold standard for symptomatic treatment of Parkinson's disease (PD); however, long­term therapy is associated with the emergence of L­DOPA­induced dyskinesia (LID). Nigral dopaminergic cell loss determines the degree of drug exposure and time required for the initial onset of LID. Accumulating evidence indicates that α­lipoic acid (ALA) decreases this nigral dopaminergic cell loss. However, until now, the precise mechanisms of ALA have only been partially understood in LID. Chronic L­DOPA treatment was demonstrated to develop intense AIM scores to assess dyskinetic symptoms. Rats in the LID group were administrated twice daily with L­DOPA + benserazide for 3 weeks to induce a rat model of dyskinesia. Moreover, other 6­OHDA­lesioned rats were treatment with ALA (31.5 mg/kg or 63 mg/kg) in combination with L­DOPA treatment. Furthermore, the authors investigated the level of malondialdehyde (MDA) and glutathione (GSH) activity, as well as IBa­1, caspase­3 and poly (ADP-ribose) polymerase (PARP) in substantia nigra by the way of western blotting and immunofluorescence. ALA reduced LID in a dose­dependent manner without compromising the anti­PD effect of L­DOPA. Moreover, ALA reduced the level of MDA and upregulated the GSH activity, as well as ameliorated IBa­1 positive neurons in the substantia nigra. Finally, it was identified that ALA could reduce L­DOPA­induced cleaved­caspase­3 and PARP overexpression in the substantia nigra. Based on the present findings, ALA could be recommended as a promising disease­modifying therapy when administered with L­DOPA early in the course of PD. The exact mechanism for this action, although incompletely understood, appears to relate to anti­oxidative stress and anti­apoptosis.

Melittin exerts an antitumor effect on non­small cell lung cancer cells.

Lung cancer accounts for a significant percentage of all cancer­associated mortalities in men and women, with non­small cell lung cancer being the most frequently occurring type of lung cancer. Melittin is the principal active component of apitoxin (bee venom) that has been reported to exert anti­chronic inflammatory and anti­cancer effects. In the present study, the antitumor effect of melittin was evaluated using in vivo and in vitro analyses. The results demonstrated that melittin significantly inhibited the epidermal growth factor­induced invasion and migration of non­small cell lung cancer cells. Subcutaneous injection of melittin at doses of 1 and 10 mg/kg significantly suppressed non­small cell lung cancer tumor growth by 27 and 61%, respectively. In addition, melittin significantly inhibited the secretion of vascular endothelial growth factor (VEGF) in non­small cell lung cancer cells. Furthermore, melittin decreased the protein expression of VEGF and hypoxia­inducible factor 1­α. Therefore, the antitumor activity of melittin may be associated with the anti­angiogenic actions of inhibiting the VEGF and hypoxia­inducible factor signaling pathways.

Identification, expression pattern, and feature analysis of cuticular protein genes in the pine moth Dendrolimus punctatus (Lepidoptera: Lasiocampidae).

Cuticular proteins (CPs) are vital components of the insects' cuticle that support movement and protect insect from adverse environmental conditions. The CPs exist in a large number and diversiform structures, thus, the accurate annotation is the first step to interpreting their roles in insect growth. The rapid development of sequencing technology has simplified the access to the information on protein sequences, especially for non-model species. Dendrolimus punctatus is a Lepidopteran defoliator, and its periodic outbreaks cause severe damage to the coniferous forests. The transcriptome of D. punctatus integrating the whole developmental periods are available for the potential investigation of CPs. In this study, we identified 216 CPs from D. punctatus, including 147 from CPR family, 4 from TWDL family, 3 from CPF/CPFL families, 22 from CPAP families, 8 low complexity proteins, 1 CPCPC and 31 from other CP families. The putative CPs were compared with homologs in other species such as Bombyx mori, Manduca sexta and Drosophila melanogaster. We further identified five co-orthologous groups have highly similar sequences of CRPs in nine lepidopteran species, which exclusively presented in RR-2 subfamily rather than RR-1. We inferred that in Lepidoptera the difference in RR-2 numbers was maintained by homologs in co-orthologous groups, coincided with observation in Drosophila and Anopheles that gene cluster was the model and source for the expansion of RR-2 genes. In combination with the variation of members in each CP family among different species, these results indicated the evolution of CPs was highly correlated to the adaptation of insect to environment. Furthermore, we compared the amino acid composition of the different types CPRs, and examined the expression patterns of CP genes in various developmental stages. The comprehensive overview of CPs from our study provides an insight into their evolution and the association between them and insect development.

Current Experimental Studies of Gene Therapy in Parkinson's Disease.

Parkinson's disease (PD) was characterized by late-onset, progressive dopamine neuron loss and movement disorders. The progresses of PD affected the neural function and integrity. To date, most researches had largely addressed the dopamine replacement therapies, but the appearance of L-dopa-induced dyskinesia hampered the use of the drug. And the mechanism of PD is so complicated that it's hard to solve the problem by just add drugs. Researchers began to focus on the genetic underpinnings of Parkinson's disease, searching for new method that may affect the neurodegeneration processes in it. In this paper, we reviewed current delivery methods used in gene therapies for PD, we also summarized the primary target of the gene therapy in the treatment of PD, such like neurotrophic factor (for regeneration), the synthesis of neurotransmitter (for prolong the duration of L-dopa), and the potential proteins that might be a target to modulate via gene therapy. Finally, we discussed RNA interference therapies used in Parkinson's disease, it might act as a new class of drug. We mainly focus on the efficiency and tooling features of different gene therapies in the treatment of PD.

Dynamic Changes in Chemosensory Gene Expression during the Dendrolimus punctatus Mating Process.

The insect chemosensory system is pivotal for interactions with their environments, and moths have especially sensitive olfaction. Exploration of the connection between the plasticity of olfactory-guided and molecular level pathways in insects is important for understanding the olfactory recognition mechanisms of insects. The pine caterpillar moth, Dendrolimus punctatus Walker, is a dominant conifer defoliator in China, and mating is the priority for adults of this species, during which sex pheromone recognition and oviposition site location are the main activities; these activities are all closely related to chemosensory genes. Thus, we aimed to identify chemosensory related genes and monitor the spectrum of their dynamic expression during the entire mating process in D. punctatus. In this study, we generated transcriptome data from male and female adult D. punctatus specimens at four mating stages: eclosion, calling, copulation, and post-coitum. These data were analyzed using bioinformatics tools to identify the major olfactory-related gene families and determine their expression patterns during mating. Levels of odorant binding proteins (OBPs), chemosensory proteins (CSPs), and odorant receptors (ORs) were closely correlated with mating behavior. Comparison with ORs from other Dendrolimus and Lepidoptera species led to the discovery of a group of ORs specific to Dendrolimus. Furthermore, we identified several genes encoding OBPs and ORs that were upregulated after mating in females; these genes may mediate the location of host plants for oviposition via plant-emitted volatiles. This work will facilitate functional research into D. punctatus chemosensory genes, provide information about the relationship between chemosensory genes and important physiological activities, and promote research into the mechanisms underlying insect olfactory recognition.

Identification and Expression Profiling of Chemosensory Genes in Dendrolimus punctatus Walker.

Dendrolimus punctatus Walker is a serious pest affecting conifers in southern China. As extensive pesticide spraying is currently required to control D. punctatus, new control strategies are urgently needed. Chemosensory genes represent potential molecular targets for development of alternative pest control strategies, and the expression characteristics of these genes provide an indication of their function. To date, little information is available regarding chemosensory genes in D. punctatus or their expression profiles at different development stages and in various tissues. Here, we assembled and analyzed the transcriptomes of D. punctatus collected at different developmental stages and in a range of organs, using next-generation sequencing. A total of 171 putative chemosensory genes were identified, encoding 53 odorant binding proteins, 26 chemosensory proteins, 60 odorant receptors (OR), 12 gustatory receptors (GR), 18 ionotropic receptors (IR), and 2 sensory neuron membrane proteins (SNMPs). Expression analysis indicated that the antennae possess the largest number of highly expressed olfactory genes and that olfactory gene expression patterns in the eggs, larvae, and head were similar to one another, with each having moderate numbers of highly expressed olfactory genes. Fat body, ovary, midgut, and testis tissues also had similar olfactory gene expression patterns, including few highly expressed olfactory genes. Of particular note, we identified only two pheromone binding proteins and no pheromone receptors in D. punctatus, similar to our previous findings in Dendrolimus houi and Dendrolimus kikuchii, suggesting that insects of the Dendrolimus genus have different pheromone recognition characteristics to other Lepidopteran insects. Overall, this extensive expression profile analysis provides a clear map of D. punctatus chemosensory genes, and will facilitate functional studies and the development of new pest control methods in the future.

Recovery and purification of limonin from pummelo [Citrus grandis] peel using water extraction, ammonium sulfate precipitation and resin adsorption.

Limonin is a bioactive compound that is traditionally extracted from citrus seeds using organic solvents or alkaline/metal ion solutions. In the present study, pummelo [Citrus grandis] peel was investigated for limonin preparation using a novel process consisting of water extraction, ammonium sulfate precipitation and resin adsorption. The pummelo peel was determined to have 4.7mg/g limonin, which could be extracted by water and further recovered by ammonium sulfate precipitation with a yield of 2.4mg/g, which was similar to that of traditional process using ethanol extraction and vacuumed evaporation. The precipitated limonin was purified by resin adsorption and crystallization with a purity of 96.4%. In addition, the limonin was identified via the analyses of retention time, infrared spectrum and nuclear magnetic resonance. This study indicates a novel and eco-friendly process for recovering limonin, providing a new candidate for limonin preparation.