Cloning and functional analysis of the juvenile hormone receptor gene CsMet in Coccinella septempunctata.

The potential role of the juvenile hormone receptor gene (methoprene-tolerant, Met) in reproduction of Coccinella septempunctata L. (Coleoptera: Coccinellidae)(Coleoptera: Coccinellidae), was investigated by cloning, analyzing expression profiles by quantitative real-time PCR, and via RNA interference (RNAi). CsMet encoded a 1518-bp open reading frames with a predicted protein product of 505 amino acids; the latter contained 2 Per-Arnt-Sim repeat profile at amino acid residues 30-83 and 102-175. CsMet was expressed in different C. septempunctata larvae developmental stages and was most highly expressed in third instar. CsMet expression in female adults gradually increased from 20 to 30 d, and expression levels at 25 and 30 d were significantly higher than levels at 1-15 d. CsMet expression in 20-d-old male adults was significantly higher than in males aged 1-15 d. CsMet expression levels in fat body tissues of male and female adults were significantly higher than expression in the head, thorax, and reproductive system. At 5 and 10 d after CsMet-dsRNA injection, CsMet expression was significantly lower than the controls by 75.05% and 58.38%, respectively. Ovary development and vitellogenesis in C. septempunctata injected with CsMet-dsRNA were significantly delayed and fewer mature eggs were produced. This study provides valuable information for the large-scale rearing of C. septempunctata.

Changes in Transcriptome and Gene Expression in Sogatella furcifera (Hemiptera: Delphacidae) in Response to Cycloxaprid.

The white-backed planthopper, Sogatella furcifera (Horváth), causes substantial damage to crops by direct feeding or virus transmission, especially southern rice black-streaked dwarf virus, which poses a serious threat to rice production. Cycloxaprid, a novel cis-nitromethylene neonicotinoid insecticide, has high efficacy against rice planthoppers, including imidacloprid-resistant populations. However, information about the influence of cycloxaprid on S. furcifera (Hemiptera: Delphacidae) at the molecular level is limited. Here, by de novo transcriptome sequencing and assembly, we constructed two transcriptomes of S. furcifera and profiled the changes in gene expression in response to cycloxaprid at the transcription level. We identified 157,906,456 nucleotides and 131,601 unigenes using the Illumina technology from cycloxaprid-treated and untreated S. furcifera. In total, 38,534 unigenes matched known proteins in at least one database, accounting for 29.28% of the total unigenes. The number of coding DNA sequences was 28,546 and that of amino acid sequences in the coding region was 22,299. In total, 15,868 simple sequence repeats (SSRs) were identified. The trinucleotide repeats accounted for 45.1% (7,157) of the total SSRs and (AAG/CTT)n were the most frequent motif. There were 359 differentially expressed genes that might have been induced by cycloxaprid. There were 131 upregulated and 228 downregulated genes. Twenty-two unigenes might be involved in resistance against cycloxaprid, such as cytochrome P450, glutathione S-transferase (GST), acid phosphatase (ACP), and cadherin. Our study provides vital information on cycloxaprid-induced resistance mechanisms, which will be useful to analyze the molecular mechanisms of cycloxaprid resistance and may lead to the development of novel strategies to manage S. furcifera.

Transcriptome sequencing of Coccinella septempunctata adults (Coleoptera: Coccinellidae) feeding on artificial diet and Aphis craccivora.

BACKGROUND: The insect predator Coccinella septempunctata can effectively control many types of pests, such as aphids, whiteflies, and small lepidopteran larvae. We previously found that C. septempunctata fed an artificial diet showed diminished biological properties(e.g. fecundity, egg hatching rate, survival rate, etc.) compared with those fed natural prey (Aphis craccivora), likely due to different nutritional characteristics of the diet. In this study, we used transcriptome sequencing analysis to identify nutrition- and metabolism-related genes of C. septempunctata that were differentially expressed depending on diet. METHODOLOGY/PRINCIPAL FINDINGS: The Illumina HiSeq2000 was used to sequence 691,942,058 total clean reads from artificial diet-fed and A. craccivora-fed C. septempunctata libraries, and the clean reads were assembled using Trinity de novo software (Tabel 2). Comparison of transcriptome sequences revealed that expression of 38,315 genes was affected by the artificial diet, and 1,182 of these genes showed a significant change in expression levels (FDR ≤ 0.05,|log2FC|≥1, "FC" stands for "fold change"). These differentially expressed genes (DEGs) were likely associated with the decreased egg laying capacity, hatching rate, longevity, and increased sex ratio (♀:♂) of adult C. septempunctata observed in the group fed the artificial diet. Furthermore, in the most DEGs metabolic pathways for C. septempunctata feeding on the artificial diet accumulated amino acid metabolic pathways, lipid metabolic pathways, and starch and glucose metabolism were down-regulated. CONCLUSIONS/SIGNIFICANCE: We found some differentially expressed genes and metabolic pathways are related to nutrition, from which a more informative feedback for diet formulation was obtained and the artificial diet could be more efficiently optimized.

Expression Differences of Resistance-Related Genes Induced by Cycloxaprid Using qRT-PCR in the Female Adult of Sogatella furcifera (Hemiptera: Delphacidae).

As a newer cis-nitromethylene neonicotinoid pesticide at present, cycloxaprid has good industrialization prospects, including the management of imidacloprid-resistant populations, because this chemical have an excellent efficiency against rice planthoppers. Sogatella furcifera (Horváth) is the most economically important pest of rice worldwide and has developed resistance to many insecticides. This study focused on the expression change of these resistance genes, induced by cycloxaprid, involved in metabolic detoxification and receptor protein. Twenty-two differentially expressed genes (DEGs) that may be related with the insecticide resistance were found in the transcriptome of S. furcifera, including 2 cytochrome P450 genes, 2 glutathione S-transferase (GST) genes, 1 acid phosphatase (ACP) gene, 12 decarboxylase genes, 2 glycolipid genes, 1 cadherin gene, and 2 glycosyltransferase genes, which were up- or downregulated in response to an exposure of cycloxaprid. Furthermore, two P450 genes (CYP4 and CYP6 family, respectively), two decarboxylase genes, and one glycosyltransferase gene were validated by qRT-PCR. Expression differences of these genes verified successfully by qRT-PCR in response to different concentrations and times treated with cycloxaprid could explain the insecticide resistance mechanism under cycloxaprid stress in S. furcifera.

Monitoring Trends in Insecticide Resistance of Field Populations of Sogatella furcifera (Hemiptera: Delphacidae) in Guizhou Province, China, 2012-2015.

Sogatella furcifera (Horváth) is a migratory insect that is one of the most important pest species on rice in many Asian countries. Control of S. furcifera (Hemiptera: Delphacidae) primarily depends on the use of chemical insecticides, and with this extensive reliance on pesticides, determining the degree of resistance of S. furcifera populations to the chemicals used for its control is essential. In this study, the resistance level to six conventional insecticides in five populations of S. furcifera from Guizhou Province was monitored yearly using the rice-stem dipping method in 2012-2015 to precisely understand current resistance levels and to estimate trends in the development of insecticide resistance in S. furcifera in Guizhou. Overall, S. furcifera from five regions in Guizhou showed a trend toward decreased susceptibility to isoprocarb (resistance ratio [RR] 0.82-3.59), susceptibility to low resistance against thiamethoxam (RR 0.27-9.69), susceptibility to moderate resistance to imidacloprid (RR 0.71-26.06), and decreased susceptibility to moderate resistance to chlorpyrifos (RR 4.63-19.58). The resistance to pymetrozine (RR 10.48-84.65) was moderate to high, and that to buprofezin (RR 6.36-412.43) was low to very high. In conclusion, the use of buprofezin and pymetrozine to control S. furcifera should be reduced in Guizhou Province, whereas prudent use at a reasonable frequency of chlorpyrifos and imidacloprid can continue. Isoprocarb and thiamethoxam are the best choices for effective management of S. furcifera. Rotations using alternative insecticides with different modes of action are recommended for regions in which resistance is at a moderate level.

A comparative dosimetric study of left sided breast cancer after breast-conserving surgery treated with VMAT and IMRT.

BACKGROUND AND PURPOSES: This study compared VMAT and IMRT plans for intact breast radiotherapy for left sided breast cancer and evaluated the irradiated dose of planning target volume and OARs, especially focusing on heart and coronary artery. MATERIALS AND METHODS: Eleven patients with left sided breast cancer whose breast was relatively smaller (the mean volumes is 296 cc) treated with breast-conserving surgery were prescribed radiotherapy of 50 Gy in 25 fractions using two or four-field step and shoot IMRT (2 or 4-F IMRT), and one or two-arc VMAT (1 or 2-arc VMAT). The 10 Gy electron boost to the tumor bed after delivery of 50 Gy was not included in the analysis. Multiple planning parameters for the PTV and the PRV-OARs were measured and analyzed. RESULTS: Treatment plans generated using VMAT had better PTV homogeneity than the IMRT plans. For the PRV-OARs, the 1-arc VMAT had significantly higher Dmean and V5 for left lung and heart, and showed worse Dmean for liver, esophagus, spinal cord, contralateral lung and breast. In contrast, the 2-arc VMAT and the 2-F or 4-F IMRT plans showed better results for the PRV-OARs than the 1-arc VMAT. However, for the heart and coronary artery, the 1-arc VMAT showed better V20 and V40 compared with the other plans. Moreover, the 2 F-IMRT had specially advantage on V5 and V20 for heart and V5 for coronary arteries, the 2-F IMRT also showed a greater MU and treatment times. Using the table of quality score to evaluate the plans, we found that 2-F IMRT had the highest scores of 13, followed by the 2-arc VMAT plan (10 points) and 1-arc VMAT plan (8 points), and finally the 4-F IMRT plan (6 points). Moreover, when a dose comparison for heart minus coronary artery was calculated, the V20 and V40 for the rest of heart in all plans were very small and closed, indicating the dose to the coronary artery contributed dramatically to the high dose volumes for the entire heart. CONCLUSIONS: Compared to other plans, the 2-F IMRT plan with fewer monitor units and shorter delivery time is an appropriate technique for left sided breast cancer, which achieved good PTV coverage and sparing of organs at risk besides for the heart and coronary artery.

Frameless stereotactic body radiation therapy for multiple lung metastases.

Two patients with multiple lung metastases (≥ 5) were treated using frameless stereotactic body radiation therapy (SBRT) on an Elekta Axesse linear accelerator equipped with an interdigitation-capable multileaf collimator and four-dimensional cone-beam CT (4D CBCT). The technique and the early clinical outcomes were evaluated. Patient A with five lung metastases and Patient B with seven lung metastases underwent SBRT (48 Gy/8 fractions for Patient A, 42 Gy/7 fractions for Patient B). The treatments were administered using a 6 MV photon beam. The nominal dose rate was 660 MUs/min. Patients were positioned and immobilized using thermoplastic masks and image guidance was done using 4D CBCT. The targets were delineated on the images of the 4D CT, and the positron emission tomography-computed tomography (PET-CT) images were taken as references. A two-step, volumetric-modulated arc therapy (VMAT) plan was designed for each patient. Step 1: the lesions in one lung were irradiated by a 210° arc field; Step 2: the rest of the lesions in the other lung were irradiated by a 120° arc field. Plans were evaluated using conformity index (CI) and homogeneity index (HI). Patients were followed up and adverse events were graded according to the Common Terminology Criteria for Adverse Events v4.0 (CTCAE v4.0). The beam-on time of each treatment was less than 10 min. The CI and HI for the two plans were 0.562, 0.0709 and 0.513, 0.0794, respectively. Pulmonary function deteriorated slightly in both patients, and the patient with seven lung lesions was confirmed to have Grade 1 radiation pneumonitis. The technique was fast, accurate, and well tolerated by patients, and the two-step plan is a helpful design in reducing the dose to the lungs.

Single arc volumetric-modulated arc therapy is sufficient for nasopharyngeal carcinoma: a dosimetric comparison with dual arc VMAT and dynamic MLC and step-and-shoot intensity-modulated radiotherapy.

BACKGROUND: The performance of single arc VMAT (VMAT1) for nasopharyngeal carcinoma (NPC) on the Axesse linac has not been well described in previous studies. The purpose of this study is to assess the feasibility of VMAT1 for NPC by comparing the dosimetry, delivery efficiency, and accuracy with dual arc VMAT (VMAT2), dynamic MLC intensity-modulated radiotherapy (dIMRT), and step-and-shoot intensity-modulated radiotherapy (ssIMRT). METHODS: Twenty consecutive patients with non-metastatic NPC were selected to be planned with VMAT1, VMAT2, dIMRT and ssIMRT using Monaco 3.2 TPS on the Axesse™ linear accelerator. Three planning target volumes (PTVs), contoured as high risk, moderate risk and low risk regions, were set to receive median absorbed-dose (D50%) of 72.6 Gy, 63.6 Gy and 54 Gy, respectively. The Homogeneity Index (HI), Conformity Index (CI), Dose Volume Histograms (DVHs), delivery efficiency and accuracy were all evaluated. RESULTS: Mean HI of PTV72.6 is better with VMAT1(0.07) and VMAT2(0.07) than dIMRT(0.09) and ssIMRT(0.09). Mean HI of PTV63.6 is better with VMAT1(0.21) and VMAT2(0.21) than dIMRT and ssIMRT. Mean CI of PTV72.6 is also better with VMAT1(0.57) and VMAT2(0.57) than dIMRT(0.49) and ssIMRT(0.5). Mean CI of PTV63.6 is better with VMAT1(0.76) and VMAT2(0.76) than dIMRT(0.73) and ssIMRT(0.73). VMAT had significantly improved homogeneity and conformity compared with IMRT. There was no significant difference between VMAT1 and VMAT2 in PTV coverage. Dose to normal tissues was acceptable for all four plan groups. VMAT1 and VMAT2 showed no significant difference in normal tissue sparring, whereas the mean dose of the parotid gland of dIMRT was significantly reduced compared to VMAT1 and VMAT2. The mean delivery time for VMAT1, VMAT2, dIMRT and ssIMRT was 2.7 min, 3.9 min, 5.7 min and 14.1 min, respectively. VMAT1 reduced the average delivery time by 29.8%, 51.1% and 80.8% compared with VMAT2, dIMRT and ssIMRT, respectively. VMAT and IMRT could all be delivered accurately based on our quality assurance standards. CONCLUSIONS: In the treatment of NPC using the Axesse™ linear accelerator, single arc VMAT has shown superiority to double arc VMAT, dIMRT and ssIMRT in delivery efficiency, without compromise to the PTV coverage. However, there is still room for improvement in terms of OAR sparing.

[The effects of nitric oxide on the survival of a random pattern skin flap].

OBJECTIVE: To observe the effect of nitric oxide (NO) on the survival of a random pattern skin flap. METHODS: Caudal based random skin flaps (9 cm x 3 cm) were raised on the back of Wistar rats. Six methods were used in the experiment to observe the effect of NO synthase inhibitor L-NAME and NO synthase substrate L-arginine on flaps: image analysis technology; light and electron microscopic studies; enzyme histochemistry of NOS in flaps; concentration of NO2-/NO3- in plasma and wet/dry ratio of the flap tissue. RESULTS: Survival area of flap in the L-arginine-treated group significantly increased (67.06 +/- 5.65)% (p < 0.01) whereas the area in the L-NAME-treated group significantly decreased (35.17 +/- 1.87)% (p < 0.01) compared with the control group (53.25 +/- 3.24)% at seven days after the operation. General and microscopic observations showed that pathological changes in the L-arginine-treated group were fewer. Abundant capillaries and fewer inflammatory cells were noticed in the L-arginine-treated group. Transmission Electron Microscopy (TEM) studies find endothelial swelling, thrombosis-formation and endothelial loss of contact with the basement membrane in the L-NAME treated group. Before operation, the serum NO concentrations were not significantly different in three groups (p > 0.05). After operation, NO concentration of the control group began to increase and reached to the top at the third day. L-Arg kept serum NO concentration in a higher level than the control. Enzyme histochemistry of NOS in flaps: microvessel intima in dermis, hair follicles, sweat glands and inflammatory cells showed oxford blue, more positive in flaps of the L-Arg treated group than the control group at the third day after operation. The flaps of L-NAME-treated group demonstrated negative or weak positive. Wet/dry ratio: twenty-four hours after flap elevation wet/dry weight ratios increased significantly in all regions of the flap of the L-arginine-treated rats compared with saline-treated rats. The ratios of the flaps of L-NAME-treated rats were reduced compared with saline-treated rats. CONCLUSION: NO could improve microcirculation of the flap and increase its survival rates. The mechanism might be that NO could accelerate flap vascularization and protect flaps from ischemia-reperfusion injury.