Peripheral monocyte count is an assessable parameter. Recently, evidence suggested an elevated preoperative monocyte counts predicting poor prognosis in malignancies. The aim of this study was to determine the prognostic effect of early postoperative blood monocyte count in patients with lung adenocarcinoma or squamous cell carcinoma following lobectomy.We retrospectively reviewed patients with operated lung adenocarcinoma or squamous cell carcinoma from 2006 to 2011 in Western China Lung Cancer database. Univariate analysis on disease-free survival (DFS) and overall survival (OS) was performed using the Kaplan-Meier and log-rank tests, and multivariate analysis was conducted using the Cox proportional hazards regression model.P < 0.001), positive smoking history (P = 0.005), and higher N stage (P = 0.002) and higher tumor stage (P = 0.026). Two-tailed log-rank test indicated patients with an early postoperative elevated monocyte count predicted a poor DFS and OS overall  as well as in subgroup analysis, and further presented as a promising independent prognostic factor for both DFS and OS  on multivariate analysis. However, no significance was detected for preoperative monocyte in multivariate analysis.There were 433 patients enrolled in our analysis. High postoperative elevated monocyte was associated with male gender (Elevated early postoperative peripheral monocyte count was an independent prognostic factor of poor prognosis and inferior clinicopathological features for patients with operable lung adenocarcinoma or squamous cell carcinoma by lobectomy. Lung cancer is the leading cause of cancer-related death worldwide, with 5-year survival rates of less than 17% . If operThe interplay between cancer and inflammation was postulated back in the late nineteenth century, and even today continues to be an active area of research. . Such reWe focused on the early postoperative period because it is the most turbulent stage of the host immune system which may be caused by the surgical trauma and/or tumor removal effect, and we hypothesized that this change in immune environment is related with tumor progression. The aim of this study was to evaluate whether an elevated early postoperative monocyte count predicted a poor prognosis in patients with operable lung adenocarcinoma or squamous cell carcinoma after lobectomy.The data were retrieved from Western China Lung Cancer database (WCLC). We enrolled patients with operable lung adenocarcinoma or squamous cell carcinoma treated with lobectomy at West China Hospital, Sichuan University between 2006 and 2011. All patients were > 18 years of age, with complete clinicopathological data, and proven to be lung adenocarcinoma or squamous cell carcinoma after surgery. Preoperative evaluation included physical examination, blood routine examination, tumor markers test, chest X-ray and computed tomography, brain magnetic resonance imaging (MRI), bone scintigraphy, and bronchoscopy and integrated positron emission tomography scan and CT (PET/CT) scan when necessary. The eligibility criteria were: 1) lobectomy with no microscopic residual tumor; 2) no preoperative chemotherapy and/or radiotherapy; 3) no previous history of other malignancies; 4) no evidence of infections such as pneumonia; 5) availability of laboratory data and follow-up information. All peripheral venous blood samples were collected from patients within 1 week before and 4 days after surgery. If multiple post-operative blood samples were drawn, only the first sample after surgery was used for analysis. Absolute peripheral blood count and the percentage were analyzed for each blood sample. Histological classification was made with reference to the latest WHO guideline . Study aBaseline characteristics included age, sex, underlying diseases, smoking history, pathological stage, pathological tumor status, pathological lymph node status, and peripheral blood counts and the percentages. Blood samples analysis were performed within the clinical laboratory of our hospital with a Cell-Dyn 3700 . Other data included the surgical date and procedures.Lobectomy was performed on all patients with intent to cure, and systematic nodal dissection was carried out. The resection was done both macroscopically and histologically with a negative tumor margin and no evidence of distant metastasis. Patients were regularly followed up at outpatient department 1 month after surgery, every 3 months for the first year, every 6 months for the next 4 years, and once annually thereafter. Patients received a physical examination, blood routine examination, chest and brain and upper abdomen CT scan at each follow-up. Bone scintigraphy was performed every 12 months. In particular, all patients treated in our department received phone call follow-up regularly, during which we recorded their living status, tumor recurrence/metastasis condition, and any adjuvant therapy such as chemotherapy and/or radiotherapy, etc. The patients were followed until August 31, 2017 or until they died.P values were two-tailed with less than 0.05 considered to be statistically significant. Statistical analysis was performed using SPSS  and STATA 14.0 . Survival curves were drawn by GraphPad Prism 5.0 .Receiver operating characteristic (ROC) curve was performed to obtain the best cut-off value for monocyte count to stratify patients at a high risk of tumor recurrence, distant metastasis, or death. In the ROC curve, the point with the maximum sensitivity and specificity was selected as the best cut-off value. Disease-free survival (DFS) was calculated from the date of surgery to the date of recurrence/metastasis or death with any cause, and overall survival (OS) was presented from the date of surgery to the date of death with any cause. Fisher’s exact test or χ2-test for categorical variables and t-test for continuous variables were used to analyze the clinicopathological features for the two groups divided by the cut-off value of monocyte. A life table was made to calculate 1, 3 and 5-year survival rate. Survival curves were plotted using the Kaplan–Meier method and compared using the log-rank test. The prognostic factors of OS and DFS were analyzed by Cox proportional hazard model with univariate and multivariate analysis. Factors significant in the univariate analysis were included in the multivariate analysis. Subgroup analysis was used to further discriminate tumor prognosis between monocyte count and other prognostic factors: pathological stage, histological type, etc. All The selection process of patients was shown in Fig. 9/L with the area under the curve (AUC) of 0.566; while for postoperative monocyte count, the cut-off value was 0.845*109/L with AUC of 0.692  of both preoperative and early postoperative group.P = 0.003), and the 1-year DFS of low and high subgroups were 82% and 85% (P = 0.106). The 3-year OS of low and high subgroups were 82% and 74% (P = 0.003), and the 3-year DFS of low and high subgroups were 67% and 60% (P = 0.106), respectively. Additionally, the 5-year OS of low and high subgroups were 72% and 58% (P = 0.003), and the 5-year DFS of low and high subgroups were 62% and 51% (P = 0.106). However, when stratified by gender, age, smoking history, stage, T stage, N stage and surgical procedure, no statistical significance were seen in subgroup study of the preoperative group.The 1-year OS of low and high subgroups were 95% and 93% (P = 0.004), and the 1-year DFS of low and high subgroups were 90% and 71% (P < 0.001). The 3-year OS of low and high subgroups were 89% and 62% (P < 0.001), and the 3-year DFS of low subgroup was 78% compared to 41% in high group (P < 0.001). In addition, the 5-year OS of low and high subgroups were 79% and 46% (P < 0.001), and the 5-year DFS of low and high subgroups were 72% and 33% (P < 0.001). Overall, an early postoperative elevated monocyte count was significantly associated with poor OS (P < 0.001) and DFS (P < 0.001) , tumor stage (P < 0.001), histological type (P = 0.014), preoperative (P = 0.023) and postoperative monocyte count level (P < 0.001) were significantly associated with DFS, while age (P = 0.017), surgery procedure (P = 0.002), T stage (P = 0.003), N stage (P < 0.001), tumor stage (P < 0.001), preoperative(P = 0.024) and postoperative monocyte count level (P < 0.001) showed significant relationships with OS.Univariate prognostic analysis was performed to detect the prognostic significance of clinicopathological factors, and preoperative and early postoperative blood monocyte count Table . N stageP = 0.008), positive N status , Adenocarcinoma  and high postoperative monocyte count  were risk factors with statistical significance for DFS. Correspondingly, age , thoracotomy , advanced tumor stage , and high postoperative monocyte count  were observed as risk factors for OS. However, preoperative monocyte was no more associated with either DFS or OS within multivariate analysis.The clinicopathological factors proved to be prognostic predictors in univariate analysis were included as covariates in further multivariate analysis Table . FinallyImmune response is an essential component of tumor progression. Studying the immune factors expressed in the tumor microenvironment can further stratify the prognosis of cancer . Back inMonocytes belonged to circulating peripheral blood cells that played the crucial role in immune response with the capability of differentiating into macrophages and antigen-presenting cells (APCs). Thus, they formed the first line of innate immune defense . In addiThe mechanism of elevated monocyte count and poor prognosis of several kinds of tumors is still unclear, despite several proposed postulations. First, it was hypothesized that monocytes are attracted by several cytokines or chemokines to the tumor site and then differentiated into tumor-associated macrophages (TAMs), which further promoted those invading leukocytes to bring out potentials of angiogenesis, motility, and invasion , 24. AngIt is known that classical monocytes are recruited to tumor sites, contributing to tumor macrophage content and promoting tumor growth and metastasis . MeanwhiIn the present study, an elevated early postoperative monocyte count was shown to predict a poor DFS and OS both in the univariate and multivariate analysis. In addition, as shown in the subgroup analysis, the monocyte count was found to be significantly associated with poor prognosis when stratified by gender, age, smoking history, TNM stage, surgical procedure and histological type. All of the above indicated that an elevated early postoperative blood monocyte count to be a very strong prognostic factor in tumor progression.Limitations of the current study are inherent to its design, including the retrospective data collection and several confounding factors when comparing postoperatively. For example, a t-test performed from Table Although there is no evidence currently to change the management of patients following a lobectomy, we are reporting some prognostic significance of the postoperative monocyte count on survival. The increased mortality for patients with elevated early postoperative monocyte, particularly in patients with squamous cell carcinoma, warrants a cautious approach to monitor disease progression.The present study supported the prognostic significance of early postoperative peripheral blood monocyte count in patients with operable lung adenocarcinoma or squamous cell carcinoma after lobectomy in both OS and DFS. This easily measured blood parameter may provide useful information for the clinicians to stratify patients. Further investigations will be needed to figure out the oncological significance of monocyte and its subsets, and the association with the host inflammatory microenvironment.


The recommendation of non-indicated caesarean section (CS) after 39 gestational weeks has been announced based on evidence of maternal and infant physiological effects. The potential psychological risks have not been acknowledged. This study aims to investigate emotional and behavioral problems in pre-school children born with elective CS (ECS) on maternal request prior to 39 weeks.Pregnant women within 12 gestational weeks between November 2008 and October 2010 were invited to participate in the China-Anhui Birth Cohort Study (C-ABCS). They were asked to complete a self-administered questionnaire respectively in 1st and 3rd trimester of pregnancy to collect basic maternal characteristics. Pregnant complications and delivery modes were abstracted from medical notes. Their singleton live births were followed up at preschool age. Strengths and Difficulties Questionnaires (SDQ) were completed by parents to assess children’s emotional and behavioral problems. A total of 3319 mother-child pairs were put into the final analysis. Descriptive analysis and binary logistic regression analysis were used to assess the impact of delivery modes on abnormalities in SDQ dimensions at various gestational ages.The prevalence of ECS on maternal request prior to 39 weeks, at 39–40 weeks, and after 41 weeks was 16.6, 23.7 and 15.9%, respectively. Compared with those born vaginally, children born with ECS on maternal request were more likely to have total difficult problems . ECS on maternal request was the independent predictor of emotional problems  and total difficult problems  in children born prior to 39 gestational weeks.Children delivered by ECS on maternal request have an increased risk to have emotional and behavioral problems prior to 39 gestational weeks at preschool age. The potential psychological implication prior to 39 weeks has been added to the roster of impacts of ECS on maternal request. Further research is needed to probe the potential biological mechanisms.The online version of this article (10.1186/s12888-019-2012-z) contains supplementary material, which is available to authorized users. Caesarean section (CS) rate is rising dramatically around the world . WHO GloChildhood psychopathology related to mode of delivery has drawn wide attention recently. Emotional and behavioral problems are areas that require more concern because they are potentially modifiable and are related with later-life social development . ResearcSimultaneously, mean gestational age has decreased globally . This trGiven the potential risks in CS on maternal request, educational and policy initiatives have been led to eliminate the non-indicated CS prior to 39 gestational weeks by American College of Obstetricians and Gynecologists and National Institute of Clinical Excellence , 24. NevParticipants of this study came from the China-Anhui Birth Cohort (C-ABCS). This cohort was set up between November 2008 and October 2010 in six cities in Anhui province, China. Anhui province is located in the southeast of China. In 2017, it has a resident population of 62.5 million, 52.0% of which live in urban areas. GDP per capita is 44, 206 yuan (us $6547). The cohort profile was published elsewhere .Mother-child pairs in this article were from one of the six cities, Ma’anshan city. The city was selected because of a stable population, having a high-quality child health care system and a strong research team in children’s development. The enrolling criteria of participants included: 1) within 12 gestational weeks who came to maternal and child health center for the first antenatal visit; 2) lived in Ma’anshan city for over 6 months; 3) willing to have childbirth in the center; 4) could understand the questionnaire and correctly answer the questions; 5) without mental diseases. A total of 5084 women were recruited in original cohort and 3319 mother-child pairs with full data were put into the final analysis. Figure Women were asked to complete a self-administered questionnaire respectively in the 1st and 3rd trimester of pregnancy. Children were followed up when they were at the age of 3–5 years. Their parents were invited to kindergartens and fill in questionnaires , we defined the level of BMI < 18.5 for underweight,24 to 27.9 for overweight and ≥ 28 for obesity for Chinese adults [During the 1st visit of pregnancy, maternal demographic information was collected. It included maternal age, maternal educational years , residence and family income. Previous adverse pregnant outcomes were reported by women, including spontaneous abortion, induced abortion, preterm, difficult vaginal delivery, fetal death or stillbirth, newborn death, ectopic pregnancy, cystic mole, malformation. Women who had any one of above-mentioned outcomes were defined to have previous adverse pregnant outcome. Women who had no previous experience of childbirth, regardless of gestational age, delivery mode or newborn’s health status, were defined as primipara. Usage of assisted reproductive technology for the index pregnancy was reported by answering yes or no. Maternal weight and height were measured by research team members. The weight was regarded as the maternal pre-pregnancy weight. Pre-pregnancy body mass index (BMI) was then calculated dividing pre-pregnancy weight (kg) by body height (m)e adults .Pregnancy-related anxiety was assessed in 1st and 3rd trimester of pregnancy by pregnancy-related anxiety questionnaire developed by research team. The coefficient of retest reliability and Cronbach α coefficient were 0.786 and 0.811, respectively . The finPregnant complications and delivery modes were abstracted from medical notes. Main complications covered gestational diabetes, hypertensive disorders in pregnancy and intrahepatic cholestasis of pregnancy. Women who had any one of these diseases were considered as having pregnant complications. As to the delivery mode, due to the very small sample size, instrumental delivery and assisted breech delivery were excluded from data analysis. Final delivery mode was classified as vaginal delivery, elective caesarean section (ECS) and emergency CS. ECS was further grouped into ECS with medical indications and ECS on maternal request in the absence of any medical indications.Children’s gender, birth weight, Apgar scores, gestational ages at birth were from medical notes. Infants born less than 36 6/7 weeks were defined as preterm, 37 0/7 weeks through 38 6/7 weeks were early term. These two groups were merged as prior to 39 weeks. 39 0/7 weeks through 40 6/7 weeks were full term and 41 0/7 weeks and beyond were late/post term . FeedingStrengths and Difficulties Questionnaire (SDQ) was completed by parents to identify children’s emotional and behavioral problems. It consisted of 25 items rated on a 3-point Likert scale: 1 “not true”, 2 “somewhat true” and 3 “certainly true”. These 25 items were divided into five subscales named as emotional symptoms, conduct problems, hyperactivity, peer problems and pro-social behaviors. Higher scores in the former 4 subscales indicated greater symptom severity, and they were summed up to generate a total difficulties score. The subscale of pro-social behaviors was scored positively and high scores indicated the presence of more adaptive behaviors. Normative data for Chinese urban population with bandings and cut-offs for borderline and abnormal scores were described by Shanghai Mental Health Centre, China . The folData analysis was performed using the Statistical Package of Social Sciences and Problem Solutions .2 tests were adopted to compare the differences among continuous variables and categorical indicators, respectively. In each gestational group, prevalence of abnormalities in SDQ dimensions was compared by χ2 tests by delivery modes. When multiple comparison tests were carried out for categorical indicators, number of analyses was counted and threshold for significance was adjusted based on number of tests completed. Binary logistic regression analysis was used to assess the impact of delivery modes on abnormalities in SDQ dimensions at various gestational ages. Maternal age, maternal educational level, residence, family income, previous adverse pregnant outcomes, pre-pregnancy BMI, parity, pregnant complications, pregnancy-related anxiety, children’s gender, birth weight and 5 min Apgar scores were adjusted in the models. In addition, sensitive analysis was conducted in this study. Firstly, we compared the basic maternal characteristics of those who were recruited in final data analysis with those who dropped out due to incomplete data to assess potential selection bias by independent t tests and χ2 tests. Secondly, we had run the model restricting to boys. In addition, pregnancy-related anxiety [Maternal and children’s characteristics among four types of delivery modes were described as means, standard deviations for continuous variables, and as frequencies for categorical indicators. Analysis of variance  and χ anxiety –37, preg anxiety  were keyAmong 3319 children, 920 (27.7%) were born prior to 39 gestational weeks, including 116 (3.5%) preterm and 804 (24.2%) early term children. As to the delivery mode, 2171 (65.4%) children were born with CS, and 693 (20.9%) were ECS on maternal request. As shown in Table P = 0.006). The score of pregnancy-related anxiety in 3rd trimester of pregnancy was significantly higher in women taking CS on request than women with vaginal delivery (P = 0.001) and women having ECS with medical indications (P = 0.001). Women who chose non-indicated ECS were more likely to have previous adverse pregnant outcomes (P = 0.001) and pregnancy complications (P = 0.005) than those who chose vaginal delivery.Women with non-indicated ECS were younger than those with vaginal delivery (P<0.001) and older than those with emergency CS (P = 0.004) and with ECS on maternal request (P = 0.045), and had significantly higher pre-pregnancy BMI than those with vaginal delivery (P<0.001), with non-indicated ECS (P<0.001) and with emergency CS (P = 0.013). Babies born by vaginal deliveries had lowest birth weights (P<0.001) and those born by emergency CS were the heaviest (P<0.05)  Table .Table 2BP<0.001), children with indicated ECS (P<0.001) and those with emergency CS (P = 0.004). As to conduct problems, the prevalence was significantly higher in children with ECS on maternal request than those with indicated ECS (P = 0.005)  (data not shown). There were no significant differences in SDQ dimensions, however, among various delivery modes in children born at 39–40 weeks and beyond 41 weeks.When the group prior to 39 weeks was divided into preterm and early term, we found a more possibility of conduct problems (42.9%) and total difficult problems (42.9%) in preterm children born with ECS on maternal request compared with other three delivery modes. The rate of emotional symptoms in early-term children born with vaginal delivery, with indicated ECS with ECS on maternal request and with emergency CS was 3.9, 5.0, 13.7 and 1.6%, respectively. The difference was statistically significant (P = 0.007)  irrespective of gestational weeks. The difference was significant between children with ECS on maternal request and children with indicated ECS (7) Table .In children prior to 39 weeks, compared with vaginal delivery, ECS on maternal request was significantly associated with emotional problems  after adjusting for potential maternal and infant characteristics. ECS with medical indications decreased the risk of peer problems . Non-indicated ECS was also independent predictive factor of total difficult problems . No significant effect was observed of ECS on maternal request on SDQ dimensions among children born at 39–40 weeks and beyond 41 weeks. Overall, despite of gestational ages, children born with ECS on maternal request had more possibilities to have total difficult problems compared with those born with vaginal delivery  in the recruited group. There were no differences in previous adverse pregnant outcomes, pre-pregnancy BMI, parity or pregnancy-related anxiety between the two groups  and total difficult problems . ECS with medical indications, however, decreased the possibility of children’s peer problems  , 20.6% women had reported antenatal anxiety in late pregnancy . Yu Y etMeanwhile, some weakness in the study must be acknowledged. Firstly, as to the participants, most of them were from urban areas. Maternal health service usage, breastfeeding situation and parenting style could be quite different between urban and rural families. Therefore, it should be prudent to explain our findings in rural areas. The overall prevalence of emotional and behavioral problems in pre-school children was reported to be 34.1 and 11.9% in rural areas respectively in Shandong and Hunan province, China , 61, beiECS on maternal request is found to be the independent predictor of emotional and behavioral problems in pre-school children born prior to 39 gestational weeks. The potential psychological implication has been added to the roster of impacts of ECS on maternal request before 39 weeks. It’s vital to communicate with both maternal health providers and users about the negative psychological consequences of early ECS in the absence of medical indications. Further research is needed to probe the potential biological mechanisms.Additional file 1:Questionnaire of maternal and children’s health in China-Anhui Birth Cohort Study (DOCX 25 kb)Additional file 2:Table S1. Comparison of basic maternal characteristics between women who were recruited in data analysis and those who were dropped out. Table S2. Binary logistic regression analysis on effect of delivery mode on SDQ in children born with various gestational ages (restricted to boys). Table S3. Binary logistic regression analysis on effect of delivery mode on SDQ in children born with various gestational ages (restricted to women without pregnancy-related anxiety). Table S4. Binary logistic regression analysis on effect of delivery mode on SDQ in children born with various gestational ages (restricted to women without pregnant complications). Table S5. Binary logistic regression analysis on effect of delivery mode on SDQ in children born with various gestational ages . Table S6. Binary logistic regression analysis on effect of delivery mode on SDQ in children born with various gestational ages (emergency caesarean section as the reference) (DOC 192 kb)


Euschistus heros (Fabricius) (Hemiptera: Pentatomidae) primarily attack the pods and seeds of soybean plants, causing severe economic losses in Neotropical Region, and chemical control is essential to avoid these losses. Thus, insecticides more effective against this pest and less toxic to Telenomus podisi Ashmead (Hymenoptera: Platygastridae) - the main biological control agent of E. heros - should be used. In this report, we studied the differential acute impacts of pesticides used in Brazilian soybean against E. heros and T. podisi and evaluated their sublethal effects on the parasitoid to identify effective pesticides towards the pest with less harmful effect to the natural enemy. The LC50 of the insecticides to E. heros ranged from 1.20 to 533.74 ng a.i./cm2; the order of toxicity was thiamethoxam + lambda-cyhalothrin > acetamiprid + fenpropathrin > zeta-cypermethrin > acephate > imidacloprid. All pesticides were classified as slightly to moderately toxic to T. podisi based on the risk quotient. The exposure of T. podisi females to imidacloprid and the insecticide pre-formulated mixtures reduced the emergence of the offspring parasitoids by up to 40% whereas zeta-cypermethrin and the insecticides pre-formulated mixtures reduced offspring survival. The preferred order of choice of insecticides for the management of E. heros according to agronomic, toxicological, and environmental feasibility was the following: thiamethoxam + lambda-cyhalothrin > zeta-cypermethrin > acetamiprid + fenpropathrin > acephate > imidacloprid. Our study provides important and pioneer information to select insecticides for effective control of E. heros with lower impacts on T. podisi. Glycine max L.) (Fabaceae: Phaseoleae) is one of the most economically important leguminous crops worldwide. Brazil is the world’s second-largest producer of soybeans with an estimated production of 115 million tons in the harvest season 2017/18, with USD 32.4 billion in soybean exports1. However, the potential productivity of soybeans is usually limited by the occurrence of pest insects during the crop season.Soybean  (Hemiptera: Pentatomidae), is the most abundant and prevalent stink bug in soybean of Neotropical Region3. Insect populations are managed by frequent spraying of insecticides, and in many situations, the chemical control is the only method capable of effectively avoid economic losses4.Among the several pests infesting soybeans, the stink bugs are of high relevance due to their high population levels and direct feeding on grains and pods, which can transmit diseases, reducing seed qualityE. heros is based on broad-spectrum insecticides, including organophosphates, pyrethroids, and neonicotinoids5, without considering the economic threshold and/or using pesticides tank mixtures3. As a result, intensive spraying of insecticides causes several problems, including increased residues in food products, intoxication of users, occurrence of resistant insect populations, resurgence and imbalance of beneficial insects that serve as natural enemies9. Therefore, the sustainability of soybean crops depends on the development of less hazardous pest management strategies, including biological control and the use of selective agrochemicals11.The management of 16. Parasitoids of stink bugs eggs have been used in 0.03 million hectares of soybean crops in South America in augmentative biological control programs17. Telenomus podisi Ashmead (Hymenoptera: Platygastridae) is the most efficient parasitoid of E. heros and Piezodorus guildinii (Westwood) (Hemiptera: Pentatomidae) eggs18, insects which cause the highest economic losses to soybean crops in Brazil19. In Brazil, T. podisi is found from the Midwest20 to the extreme South Regions21.The egg parasitoids of the Platygastridae family are considered the main natural enemies of stink bugs pests (Hemiptera: Pentatomidae) in different crops22 present simultaneously in soybean crops. Therefore, the choice of chemicals for pest control in IPM programs should not be based only on the agronomic efficiency (e.g. efficiency in pest control) of the products but also on the lowest impact over the pest natural enemies (e.g. selective pesticide)23. To date, information on pesticide selectivity has been disregarded when choosing chemicals to pests control in Brazil, because this information is not easily available to farmers24, such as on package leaflet or product labels or even on online pesticides database of the Brazilian Ministry of Agriculture and Food Supply25. This situation has become even more worrying since Brazil is one of the world leaders in agrochemicals use26.Although biological control is essential for Integrated Pest Management (IPM) by maintaining pest populations below economic threshold levels, chemical control in many circumstances is necessary for effective management of stink bugs and other harmful organisms27. Previous studies evaluated the differential acute toxicity of pesticides against the target pests and their natural enemies in different crops with the aim of choosing a pesticide with a high degree of lethal toxicity on pests and minimal non-target lethal toxicity30. In addition to direct pesticide-induced mortality, the sublethal effects must be considered for a complete impact analysis, helping pesticide choice for IPM32. However, to the best of our knowledge, no studies to date compared the acute toxicity of pesticides on E. heros and its main biocontrol agent T. podisi and the sublethal effects on this egg parasitoid. Thus, the aim of this research was to know the differential impacts of pesticides frequently used in soybean crops in Brazil to the brown stink bug E. heros and its main parasitoid T. podisi, and determine the sublethal effects on the parasitoid. Once this data were available, we could select those insecticides that were most effective in controlling the pest and with lowest toxicity to the natural enemy.Assessment of the acute toxicity of pesticides towards beneficial arthropods traditionally has relied on the determination of an acute median lethal dose or lethal concentration50) values for acephate, imidacloprid, zeta-cypermethrin, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin after exposure in glass vials were significantly different between E. heros and T. podisi  was thiamethoxam + lambda-cyhalothrin > acetamiprid + fenpropathrin > zeta-cypermethrin > acephate > imidacloprid   was thiamethoxam + lambda-cyhalothrin > imidacloprid > acetamiprid + fenpropathrin > zeta-cypermethrin > acephate  (Table The LCy) Table .T. podisi (50 < RQ ≤ 2500), with values ranging from 79.55 and 1646.67.The risk quotient-based classification (RQ) is shown in Table T. podisi females exposed to the LC50 of acephate, imidacloprid, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin showed significantly decreased in the percentage of parasitized eggs, by up to 13.42%  (Table T. podisi (Table 1) of T. podisi was significantly affected by the insecticides  (Table P = 0.06) , being observed a decrease of up to 35% compared to the control  was 23.45, 24.01, 21.80, 20.12, and 21.30 days for acephate, imidacloprid, zeta-cypermethrin, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin, respectively. The mean survival in the control group was 30.96 and did not differ significantly from that of acephate and imidacloprid , thiamethoxam + lambda-cyhalothrin was considered the most suitable pesticide to control the brown stink bug Table . The DA 6 and therefore demands high insecticide applications in attempt to regulate population level on the field. These insecticides contain several active ingredients, formulated commercially in isolation or mixtures, presenting different control efficiencies. In this study, E. heros adults were more susceptible to the residual toxic effects of neonicotinoids and pyrethroids formulated in mixtures, including thiamethoxam + lambda-cyhalothrin and acetamiprid + fenpropathrin, with LC50 values of approximately 72, 318, and 445, 2.5, 11, and 15-fold lower than those of zeta-cypermethrin (pyrethroid), acephate (organophosphate), and imidacloprid (neonicotinoid), respectively.The brown stink bug is one the predominant insect pests in Brazilian soybean crops33 or mixtures of two active ingredients34. Spray mixture formulations are registered and currently used to control the brown stink bug in Brazilian soybean crops35. These pre-mixtures allow a broader spectrum of action, targeting different toxicological sites in the pest. This is due the characteristic of the active ingredients in these mixtures that usually contain a neonicotinoid - systemic in the plant - which acts in the insect mainly by plant tissues/sap ingestion, plus a pyrethroid, which mainly acts by contact. Recent efficiency assays in E. heros demonstrated a stronger acute effect of commercially formulated neonicotinoids and pyrethroids in mixtures within 14 days after application compared to isolated pesticides such as acephate, zeta-cypermethrin, and imidacloprid36.Farmers and phytosanitary managers often prefer to control pest insects in different crops with the same application in tank mixturesE. heros in the field (ng a.i./cm2) are approximately 20, 2, 8, 26, and 409 times higher than the LC50 (ng a.i./cm2) of acephate, imidacloprid, zeta-cypermethrin, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin, respectively  and Telenomus remus Nixon (Hymenoptera: Platygastridae)42.The results of this study indicated that acephate, imidacloprid, zeta-cypermethrin, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin were toxic to 43, and are more sensitive to the effects of pesticides than the immature stages because the embryo is protected by the egg chorion during insect development44. Therefore, the RQ was used for the first time to evaluate the ecological risk of insecticide used to control E. heros over the its natural enemy T. podisi. The RQ is an important measure of risk to natural enemies under field conditions because it also considers the recommended field rate for target pest control45. RQ has been used to evaluate the safety of predators and parasitoids in different agroecosystems49.Adult parasitoids may be directly exposed to insecticide droplets during spraying or indirectly by toxic residues on the plant canopy, water droplets, nectar, or honeydewT. podisi, although the RQ for zeta-cypermethrin approached 50 and was approximately 4, 19, 5, and 20 times lower than that of acephate, imidacloprid, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin, respectively. Acephate and other organophosphates presented a high risk of toxicity to Trichogramma spp.51. Similarly, neonicotinoids, including imidacloprid and thiamethoxam, were toxic to egg parasitoids46. Therefore, the use of these insecticides in IPM programs should be carefully evaluated52.The analysis of the RQ in this study indicated that none of the evaluated pesticides was considered harmless (RQ < 50) to T. podisi to lethal or sublethal doses of pesticides allows the determination of the chemical and biological compatibility and the effect of insecticides on the natural enemies31. Beneficial arthropods surviving insecticide exposure may be mildly or severely affected, manifested in individual biological changes and offspring survival , and behavioral characteristics53.The exposure of 50 decreased, albeit to a small extent, the percentage of host eggs parasitized by T. podisi. Previous studies reported significant impairment of parasitism by Telenomus spp. and Trichogramma spp. exposed to toxic residues at the recommended concentrations of acephate, imidacloprid, zeta-cypermethrin, thiamethoxam + lambda-cyhalothrin, and other organophosphorus pesticides, pyrethroids, and neonicotinoids commercially formulated in isolation or mixtures55.The exposure to LCT. podisi females from the maternal generation to insecticides may impair their offspring53. In our study, imidacloprid, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin significantly reduced the emergence of offspring of exposed females  of the pyrethroids deltamethrin and cyfluthrin to the progeny of females of Telenomus busseolae Gahan (Hymenoptera: Platygastridae) and found no detrimental effects on insect emergence. However, studies with parasitoids of the Platygastridae family indicated that insects emergence decreased when immature stages were exposed to pesticides44. It is of note that our results do not allow determining whether the reduction in insect emergence is due to the direct effects of pesticides or the occurrence of other dysfunctions such as organ malformation31.The exposure of et al.56 pointed out that it is vital to consider the fitness of emerging parasitoids. In our study, the survival of the offspring of females exposed to the LC50 of zeta-cypermethrin, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin was significantly reduced. The effect of insecticides on the parasitoid longevity will depend on the type of insecticide, parasitoid species, and the mode of insecticide application40. For instance, these reductions in longevity are commonly observed in parasitoids emerged from eggs exposed to pesticides while developing inside the host57. However, Beserra and Parra53 found no significant changes in the longevity of F1 females of Trichogramma pretiosum Riley (Hymenoptera: Trichogrammatidae) developing in eggs of Anagasta kuehniella (Zeller)  treated with lambda-cyhalothrin in the larval, pre-pupal, and pupal stages of the parasitoid. Until now, to the best of our knowledge, there are no available studies on changes in the biological characteristics of Platygastridae egg parasitoids, including offspring longevity, as a consequence of the exposure of maternal females to sublethal pesticides concentrations.Sohrabi 31. For instance, the organophosphorus insecticide chlorpyrifos modified the sex ratio of the offspring of several Hymenoptera parasitoid59, whereas imidacloprid significantly changed the sex ratio of the progeny of Encarsia inaron Walker (Hymenoptera: Aphelinidae) by increasing the number of male offspring56. However, these authors did not determine the mechanisms underlying the change in the sex ratio of beneficial arthropods caused by insecticides. In the present study, the proportion of F1 females remained high in all treatments, and these results are similar to those obtained in a study that determined the rate of parasitism of E. heros eggs by T. podisi in vitro60.Agrochemicals products can also cause changes in the sex ratio of beneficial insectset al.23 recently performed a research review about the challenges, limitations and field recommendations on the selectivity of pesticides to natural enemies, and reported that considering the populations dynamics of insects and other pests and the frequent introduction of new chemical compounds for pest management, farmers need complete information on pesticides that could effectively control target pests with minimal impact on the agroecosystem and related agents to continually adjust their IPM routines. In this research, DA was studied for acephate, imidacloprid, zeta-cypermethrin, acetamiprid + fenpropathrin, and thiamethoxam + lambda-cyhalothrin in order to support the choice of the most appropriate insecticide in an integrated management program for E. heros. This method was proposed by Martins et al.24 for pesticides registered in rice, corn, and soybean crops in Southern Brazil. The strong lethal effect of thiamethoxam + lambda-cyhalothrin against E. heros, with a moderate classification by RQ and sublethal effects on T. podisi, together with the toxicological class of the commercial product  in the year 2015, was established by mass rearing in the laboratory 61. The T. podisi colony was obtained from “BUG Brasil Agentes Biológicos©” and reared in the laboratory 62.The E. heros in soybean crop25 and widely used for managing pentatomids in crop36  (Hemiptera: Miridae) and E. heros65 to contact insecticides, with slight modifications, was used in insecticide toxicity bioassays for E. heros and T. podisi in the laboratory .The method using glass vial, initially developed to assess the susceptibility of 2) to obtain the range of doses causing mortality of 0 to 100%. In the second phase, seven to ten concentrations of each insecticide were prepared by sequential dilution in distilled water to obtain the concentration-response curves and the estimated median lethal concentration (LC50). Distilled water was used in the control treatment.The concentrations of each insecticide used in the assays were based on the level of active ingredient indicated in the package label of the formulations and were prepared in two phases. The first phase consisted of serial dilutions (1:10) of the insecticide stock concentration (1000 ng a.i./cm2) was impregnated with 600 μL of each insecticide concentration (treatment). The following minimum and maximum concentrations (in ng a.i./cm2) were used: acephate (Orthene® SP) 46.27 to 9253.55; imidacloprid (Imidacloprid Nortox® SC) 0.46 to 9253.55; zeta-cypermethrin (Mustang® EC) 0.46 to 9253.55; acetamiprid + fenpropathrin (Bold® EW) 0.09 to 4626.77; and thiamethoxam + lambda-cyhalothrin (Engeo™ Pleno SC) 0.09 to 9253.55. The vials treated were dried on a rotating equipment to ensure the uniformity of mix in the vials. Each treatment included five replicates, each with five pairs  of stink bugs adults aged ≤72 h.The surface of each glass vial  containing beans, soybeans and peanuts as food and water 2) were performed as described for the E. heros lethal toxicity bioassay. The following minimum and maximum insecticide concentrations (in ng a.i./cm2) were used: acephate (Orthene® SP) 2.32 to 1157.85; imidacloprid (Imidacloprid Nortox® SC) 0.23 to 23.16; zeta-cypermethrin (Mustang® EC) 0.23 to 231.57; acetamiprid + fenpropathrin (Bold® EW) 1.16 to 115.79; and thiamethoxam + lambda-cyhalothrin (Engeo™ Pleno SC) 0.23 to 23.16. Each treatment included five repetitions, each with five pairs  of parasitoid adults aged ≤48 h.The application and drying of the insecticides on the surface of the glass vials  containing pure honey as food. Mortality was assessed at 24 and 48 h after insecticide exposure. The parasitoids that did not move when stimulated with a fine-tipped brush were considered dead.T. podisi were established and maintained for 24–36 h in glass vials (diameter of 2.4 cm and height of 8.0 cm) containing pure honey as food for mating. Subsequently, females  were transferred to glass vials  impregnated with insecticide (LC50) or distilled water (control treatment). Five repetitions with 20 females each were used.The pairs of 50), the parasitoids were removed from the glass vials and transferred to another vials (diameter of 2.4 cm and height of 8.0 cm) containing pure honey as food. The mortality ratio was determined 24 and 48 h after contact with the pesticides. Twenty surviving females, randomly selected from each treatment, were transferred to glass vial of the same size containing pure honey as food and an egg mass (aged < 12 h) of E. heros (cards with approximately 25 eggs) to parasitize for 24 h. The egg cards were removed and individualized to measure the rate of eggs parasitized by the females (generation F0) exposed to insecticides (LC50), emergence rate, sex ratio and survival of adult parasitoids (F1 generation).After 4 h of treatment with the insecticides , and χ2 values were calculated by Probit analysis using the POLO Plus software . The LC50 values were compared for each species (E. heros and T. podisi) using the LC50 confidence intervals, being considered significantly different when these intervals did not overlap.The LC50 for T. podisi and the registered dose for the control of E. heros in soybean crop  of insecticides were calculated from the values of LCop Table 46. QR va50) on the rate of parasitized eggs, emergence rate and adult sex ratio (F1) were determined by the Kruskal-Wallis test with Dunn post hoc test (P < 0.05) using R® software66. The Kaplan-Meier estimators (Log-Rank method) were used to evaluate survival (days) of adult parasitoids (F1) and the survival curves were compared by the Holm-Sidak test (P < 0.05) using the software SigmaPlot version 12.3 . In addition, descriptive analysis established by the International Organization for Biological and Integrated Control of Noxious Animals and Plants (IOBC) was conducted using equation  toxicity to E. heros [Wa = 4], based on the differences in LC50 (in ng of a.i. per cm2) for E. heros; b) toxicity to T. podisi based on RQ values36 [Wb1 = 1.5] and reduction of parasitism and emergence using IOBC criteria38 [Wb2 = 1.5]; c) toxicological class (package leaflet and insecticide label) [Wc = 1]; d) environmental class (package leaflet and insecticide label) [Wd = 1]; and e) safety interval (package leaflet and insecticide label) [We = 1]. For each item, the scores 1 (lower adequacy), 3, 5, or 7 (higher adequacy) were assigned descriptively. Furthermore, the weighted average indicative of the degree of adequacy (DA) of the commercial insecticide for the control of E. heros was calculated using equation (E. heros) multiplied by weight 4; Sb1 is the score attributed to the toxicity of the pesticide to the natural enemy (T. podisi) according to the RQ multiplied by weight 1.5; Sb2 is the score attributed to the toxicity of the pesticide to the natural enemy (T. podisi) according to the highest IOBC classification to reduce parasitism or emergence multiplied by weight 1.5; Sc is the score assigned to the toxicological class of the pesticide multiplied by weight 1; Sd is the score attributed to the environmental toxicological class of the pesticide multiplied by weight 1; Se is the score assigned to the safety interval of the pesticide multiplied by weight 1.We elaborated an indication of the most adequate insecticides for the control of This article does not contain any studies with human participants or vertebrate performed by any of the authors.Supplementary Information


C3 gene account for a significant risk of advanced AMD.The purpose of this study is to discuss whether genetic variants  in the We performed a meta-analysis using electronic databases to search relevant articles. A total of 40 case-control studies from 38 available articles  were included in our study.P < 0.001; OR = 1.45, 95% CI = 1.37–1.54, P < 0.001). Moreover, in the subgroup analysis based on ethnicity, rs2230199 and rs1047286 polymorphisms were more likely to be a predictor of response for Caucasian region . Besides, pooled results suggested that the G allele of rs2230199 could confer susceptibility to advanced AMD in Middle East .In our meta-analysis, the pooled results showed that the carriage of G allele for rs2230199 and the T allele for rs1047286 had a tendency to the risk of advanced AMD  contains supplementary material, which is available to authorized users. Age-related macular degeneration (AMD) is a complex and progressive retinal disorder influenced by family history, aging, race, smoking and diet, which caused irreversible visual impairment in a growing number of elderly persons , 2. The CFH) gene by candidate region and whole genome association analyses [Advanced AMD has been implicated with important risk factors listed above, it is a multifactorial disease which influenced by a combination of environmental and genetic susceptibility , 3, 7, 8analyses , 13.C3 gene have been investigated with advanced AMD in different ethnic groups, the pooled results are incompatible and ambiguous. According to the International HapMap Project database, the human C3 gene is located on chromosome 19 and exhibits nine common genetic SNPs . Used for screening the electronic database and manual searching, the most widely condidate polymorphisms of the C3 gene which at least has been surveyed in three pertinent studies are rs2230199, rs1047286, rs2230205 and rs2250656. In order to better understand the genetic risk of C3 gene in the relationship with exudative AMD, we performed a meta-analysis to illuminate this association and determine whether the genetic variants of C3 gene conferred susceptibility to advanced AMD.Apart from CFH, the central element of the complement cascade, complement component C3a has been interconnected with the vascular endothelial growth factor expression, geographic atrophy, retinal pigment epithelium deterioration, and progression to choroidal neovascularization , 14, 15.C3” or “C3” or “complement component 3”) and (“variant” or “mutation” or “genetic” or “SNP” or “polymorphism” or “genetic polymorphism” or “genetic variant” or “single nucleotide polymorphism”). Each database was thoroughly scanned and was up to date as of September 1 2018. Our meta-analysis was mainly focused on case-control studies, without any language limitation imposed in the literature searching.A systematic search of electronic database such as PubMed, Embase, CNKI, Cochrane library and Web of Science was conducted with the following keywords: (“AMD” or “maculopathy” or “macular degeneration” or “age-related maculopathy” or “age-related macular degeneration”) and  detailed genetyping data for each site could be acquired to estimate the odds ratio (OR) and 95% confidence interval (CI) based on genetic model contrast; (3) individual for all selected samples met the modified version of the age-related eye disease study (AREDS) grading system as described elsewhere. Major exclusion criteria were limited to several items as follow: (1) overlapping subjects in several articles for the same research group; (2) only focused on families’ individuals rather than sporadic advanced AMD patients; (3) abstract from conferences, letters, review articles and case reports. When several articles included some of the same samples, the one with largest individuals and thorough genotype information would be winnowed for our meta-analysis.Retrieved articles were considered eligible for our meta-analysis when they met the following inclusion criteria: (1) investigating the disease risk of Data from the retrieved studies were extracted independently by two reviewers (J.Z. and S.L.). The following items obtained from each eligible articles included: the first author, the year of publication, country and ethnicity of subjects, information on study design, sample size, genotyping methods and distribution in case and control groups. Two authors carefully inspected the raw statistics and reached a consensus in all aspects. If any disagreement still existed, the third author (S.H.) would be invited to chew over current controversy and resolve the dispute.Quality assessment of the screened studies was also independently conducted by two reviewers (J.Z. and S.L.) in the basis of the HuGENet Handbook . A totalC3 polymorphic site were counted between cases and healthy controls. The genetic strength association including pooled ORs and 95% CIs was assessed using different genetic models, including allele model (A vs. a), homozygote model (AA vs. aa), heterozygote model (Aa vs. aa), dominant (AA+Aa vs. aa), recessive (AA vs. Aa+aa). The heterogeneity assumption between studies was estimated and evaluated by Cochran’s Q statistic as well as the I2 statistic. The result that our P value of Q statistic was less than 0.05 or the I2 value was greater than 50% suggested apparent heterogeneity, thus a random-effect model was utilized in our model analysis. Otherwise, the fixed-effect model was performed [P values was executed by STATA 12.0 software . A significant difference was estimated under the level of 0.05. The final results needed to be tested and verified by two authors (J.Z. and S.L.) respectively.Allele and genotype frequency of each erformed . Sensitierformed  and Eggeerformed  were furC3 genetic polymorphism. The other 46 full-text articles were left in our meta-analysis. Seven of them did not have detailed genotype data after cautiously reading the included literatures.Besides, two papers were investigated by the same author and the same batch of patients from Iran [C3 gene with advanced AMD from 38 available publications were generally contained in our current meta-analysis [The flow diagram for literature searching is summarized in Fig. rom Iran , 21. We analysis , 22–57. Overall results in Table C3 polymorphisms including rs2230199, rs1047286, rs2230205, rs2250656 were used in our meta-analysis and the combined results are presented in Table Several genetic models for P < 0.001; homozygote model: OR = 2.33, 95% CI = 1.98–2.74, P < 0.001; heterozygote model: OR = 1.53, 95% CI = 1.41–1.64, P < 0.001; dominant model: OR = 1.62, 95% CI = 1.51–1.74, P < 0.001; recessive model: OR = 1.99, 95% CI = 1.70–2.34, P < 0.001). Moreover, the subgroup analysis straitified by ethnicity indicated that rs2230199 conferred obvious susceptibility to advanced AMD in the group of Caucasian in allelic  , heterozygote , dominant , recessive  models (Table P < 0.001). However, this relationship was not significant in East Asian group for any genetic models . In subgroup analysis stratified by ethnicity, our meta-analysis indicated significant correlation of rs1047286 with advanced AMD in the five genetic models . Subgroup analysis of Caucasian and East Asian group showed that there was a lack of relationship in any of the genetic models . But a weakly protective risk between this SNP and advanced AMD was observed in heterozygote model and dominant model . In the stratified analysis by ethnicity, there was no association in any of the genetic models.   . Furthermore, as emerged in Table P values for both tests are more than 0.05.Publication bias is a potential problem, thus Begg’s funnel plots and Egger’s regression tests were applied to investigate the publication bias for C3 has been a plausible candidate gene since its cleavage product C3a was confirmed in drusen. In our current meta-analysis, 20,673 patients and 20,025 controls from 38 articles were combined to detect the association of C3 genetic polymorphisms with advanced AMD. We came to the conclusion that two nonsynonymous SNPs rs2230199 and rs1047286 were demonstrated an increased pathogenic effect on advanced AMD . Moreover, our meta-analysis discovered that SNP rs2250656 decreased the risk of advanced AMD susceptibility, which a protective association was acquired in heterozygote model and dominant model. Obviously, the results of SNP rs2250656 with advanced AMD needed to be validated with larger samples and studies in different ethnicity.AMD is a multifactorial disease, in which complement system mediated inflammation plays a pivotal role. Several pathways including the alternative complement component have been described to be implicated in the development of AMD . As the Being consistent with previous studies, the G allele of rs2230199 conferred susceptibility to advanced AMD in Caucasian group. In our meta-analysis, we first confirmed that the G allele of rs2230199 could be linked with AMD in Middle East but not East Asian region, though rather larger population needed to be validated in the future. Besides, our meta-analysis found a novel association between the T allele of rs1047286 and advanced AMD in Caucasian but not East Asian group.C3 gene have been identified as genetic risk factors for advanced AMD in Caucasian populations. However, the allele frequencies of rs2230199 vary widely among different ethnicities. Frequencies of the risk G allele at rs2230199 were 25% to 31% in AMD cases and 19% to 21% in controls in Caucasians [The common polymorphisms rs2230199 and rs1047286 in the ucasians . Besidesucasians , 57. Whiucasians . For rs1ucasians  also fouC3 is located on the short arm of chromosome 19 and consists of 41 exons, which forms 13 functional domains. C3 is the most abundant complement component and significant C3 messenger RNA is detected in the neural retina, choroid, RPE, and cultured RPE cells [C3 undergoes important structural rearrangements which causes conformational changes exposing binding sites for complement components and drusen including C3 and its activation products was confirmed in the finding that local inflammation and activation of the complement cascade can contribute to the pathogenesis of AMD. Notably, animal studies conducted by Bora et al. [C3 deficiency in C3−/− mice prevented the formation of choroidal neovascularization in advanced AMD (wet AMD), indicating that C3 is a pivotal element of this activation process.The gene for PE cells . CleavagPE cells  argued ta et al. , 61 haveC3F and C3S), the two alleles showed a differential capacity to bind monocyte- complement receptor. Helgason et al. [C3F) was associated with the reduction of C3 gene binding to CFH, which leads to an increase in complement activation. Additionally, rs2230199 variant may alter the net charge of the molecule and influence the position of the thioester-containing domain. Except for advanced AMD, the risk variant of rs2230199 has been previously considered as associated with other immune-mediated conditions, such as IgA nephropathy, systemic vasculitis.In our meta-analysis, four SNPs including rs2230199, rs1047286, rs223205 and rs2250656 were analyzed in the pooled data. Among them, rs2230199 and rs1047286 are located in the first ring of macroglobulin domains, which conduct a prominent function for correct orientation of the thioester-containing domain. The amino acid changes induced by the genetic mutations may alter the configuration of the macroglobulin ring . With evn et al.  noted th2 = 0.80), which haplotype analyses suggested that the effect of the C3 alleles was independent from the established genetic and environmental risk factors. Furthermore, our pooled analysis of neighboring SNPs of rs2230199 indicated that the allele frequency of the variant rs2230205 and rs2250656 was not significantly different between the advanced AMD cases and controls. Pei et al. [In the current meta-analysis, rs1047286 variant showed significant association with advanced AMD in Caucasian populations. Despriet et al.  argued ti et al.  confirmeIn a previous meta-analysis where a total of 15 independent studies with 5593 cases and 5181 controls were included, Zhang et al.  indicateC3 genetic polymorphisms and advanced AMD, some limitations for our present meta-analysis need to be declared. First, heterogeneity among the ethnic groups was discovered when investigating the association of C3 genetic variants with advanced AMD. However, based on the results of the sensitivity analysis, it is clear that the overall effect was not affected by heterogeneity. Additionally, there was no obvious publication bias detected in the contrast of C3 gene with advanced AMD. Second, the number of patients and controls was relatively small in each included study; therefore, a great number of samples from different ethnic regions are required for further analysis. Third, the effects of common confounding factors, including sex, age, body mass index, smoking, and diet were not evaluated in the present study because of insufficient data. Fourth limitation is that only three ethnic backgrounds with relatively few studies were taken into consideration, thus further efforts to reduce the incidence of ethnic bias will be needed once raw data become available. Finally, the electronic databases from which we selected eligible studies were listed in English and Chinese; therefore, a language bias may be existed in our meta-analysis.Considerable efforts have been paid to discuss the potential relationship between The present meta-analysis provided a series of evidence-based pooled data for a significant association between rs2230199, rs1047286 and susceptibility to advanced AMD, especially in Caucasians. Additional well-designed work with a larger number of studies in which incorporate different ethnicities together with gene-gene and gene-environment is recommended to better confirm the functional role of the two nonsynonymous polymorphisms.Additional file 1:The full details of databases searching terms. (DOC 38 kb)Additional file 2:Figures S1-S3. The sensitivity analysis of C3 genetic polymorphisms . (DOC 106 kb)


Live-attenuated viral vaccines are highly safe and efficacious but represent complex and often multigenic attenuation mechanisms. Most of these vaccines have been generated empirically by serial passaging of a wild-type (WT) virus in cell culture. One of the safest and most effective live-attenuated vaccines is yellow fever (YF) virus strain 17D, which has been used for over 80 years to control YF disease. The availability of the WT parental strain of 17D, Asibi virus, and large quantities of clinical data showing the effectiveness of the 17D vaccine make this WT parent/vaccine pair an excellent model for investigating RNA virus attenuation. Here, we investigate a mechanism of 17D attenuation and show that the vaccine virus is resistant to the antiviral compound ribavirin. The findings suggest that attenuation is in part due to a low probability of reversion or mutation of the vaccine virus genome to WT, thus maintaining a stable genotype despite external pressures. The molecular basis of attenuation for live-attenuated vaccines is poorly understood. The yellow fever (YF) 17D vaccine virus was derived from the wild-type, parental strain Asibi virus by serial passage in chicken tissue and has proven to be a very safe and efficacious vaccine. We have previously shown that wild-type Asibi is a typical RNA virus with high genetic diversity, while the 17D vaccine virus has very little genetic diversity. To investigate this further, we treated Asibi and 17D viruses with ribavirin, a GTP analog with strong antiviral activity that increases levels of mutations in the viral genome. As expected, ribavirin treatment introduced mutations into the Asibi virus genome at a very high frequency and decreased viral infectivity while, in contrast, the 17D vaccine virus was resistant to ribavirin, as treatment with the antiviral introduced very few mutations into the genome, and viral infectivity was not lost. The results were confirmed for another YF wild-type parental and vaccine pair, a wild-type French viscerotropic virus and French neurotropic vaccine. Using recombinant Asibi and 17D viruses, ribavirin sensitivity was located to viral nonstructural genes. Thus, two live-attenuated YF vaccine viruses are genetically stable even under intense mutagenic pressure, suggesting that attenuation of live-attenuated YF vaccines is due, at least in part, to fidelity of the replication complex resulting in high genetic stability. Flavivirus contains approximately 70 viruses, most of which are arthropod-borne (arboviruses) and major public health problems, including dengue, Japanese encephalitis, yellow fever (YF), and Zika viruses. These enveloped, positive-sense RNA viruses contain the following 10 genes: three structural proteins  and seven nonstructural (NS) proteins . The functions of the NS proteins, which constitute the host cell membrane-bound replication complex, are incompletely understood. The viral protease and helicase are contained within NS3, and methyltransferase and polymerase reside in NS5  strain Asibi virus in various cell types, including chicken embryos lacking nervous tissue, wherein it lost viscerotropic properties as well as the ability to be transmitted by mosquitoes . AlthougConcurrent to the development of the 17D vaccine virus, another live-attenuated YF vaccine, French neurotropic vaccine (FNV) virus, was developed by 128 passages of the WT strain French viscerotropic virus (FVV) in the mouse brain. Asibi virus and FVV were isolated in the same YF outbreak in West Africa in 1927. As such, these viruses are genetically very similar, with only two amino acid mutations differentiating the WT strains (E-200 and NS3-280)  and a high mutation rate with considerable nucleotide mismatching . Though 10–15–RNA virus diversity in attenuated mutants has been investigated. Studies with poliovirus have shown that increasing the fidelity of the RdRp attenuates the virus by reducing the ability of the virus to enter the brain and cause disease , 18. Whe20–Lethal mutagenesis is an antiviral principle that utilizes the concept of error threshold, a theoretical maximum rate of genetic diversity. Antivirals utilizing this mechanism of action are typically nucleoside analogs which, when incorporated into the genome, increase the viral mutation rate beyond the error threshold. At this point, the benefits of a robust mutant spectrum are outweighed by the accumulation of deleterious mutations and the generation of defective genomes that inhibit viral replication and reduce infectivity . Ribavir26–in vitro, supporting the hypothesis that the incorporation of a GTP analog is the primary mechanism of action of ribavirin against WT YFV. We propose that YFV vaccine strains are resistant to ribavirin due to an acquired high-fidelity replication mechanism and that this contributes to the attenuated phenotype.Due to the differences in genetic diversity between WT Asibi virus and its 17D vaccine virus derivative, we investigated the changes in genetic diversity of 17D and Asibi viruses following treatment with ribavirin. It was found that 17D virus was resistant to antiviral effects, whereas the drug was active against WT Asibi. Furthermore, ribavirin resistance was observed in a second live-attenuated YFV vaccine strain (FNV) and susceptibility in a second WT YFV strain (FVV). Experiments with recombinant-derived mutant viruses indicated that resistance was associated with the NS genes of the vaccine virus, suggesting a role for the replication complex in susceptibility. The contribution of anti-IMPDH activity was tested using mycophenolic acid (MPA), a nonnucleoside triphosphate antiviral that inhibits IMPDH and has been shown to be effective against 17D , 32. WT P < 0.0001). The 50% inhibitory concentration (IC50) values of Asibi  and 17D  viruses differed by 57-fold, showing that ribavirin is much more active against Asibi virus than against 17D virus (P < 0.037). The IC50 values of FVV  and FNV  viruses differed by 14-fold showing that, again, WT YFV was more susceptible to ribavirin than was a live-attenuated vaccine virus. It was noted that YFV vaccine strains 17D and FNV were not statistically different from each other (P > 0.999). The same was also true in a comparison of the two WT YFV strains, where no difference can be detected between the curves (P > 0.999).The effect of ribavirin treatment on the viral infectivity of low-passage-number WT Asibi virus and commercial 17D vaccine virus strain was investigated in Vero cells. The viral infectivity reduction curves for WT Asibi virus and vaccine YFV were significantly different (7D virus . The mulP = 0.68), and the IC50 values of Asibi  and 17D  viruses were statistically indistinguishable and within the range considered safe and effective for MPA in vitro drug screens (R2 = 0.94) and FNV  viruses, with reduction curves and IC50 values which appeared to be statistically similar (P > 0.999)  and vaccine strains (17D and FNV viruses) was investigated. MPA is an anti-IMPDH antiviral with no nucleoside triphosphate (NTP) analog activity. Viral infectivity reduction curves of MPA-treated Asibi and 17D viruses were not significantly different P = 0.68)(i) WT Asibi versus 17D vaccine virus. Four ribavirin treatment concentrations  were incubated with Asibi and 17D viruses, sampled at 48 h postinfection, and sequenced by massively parallel sequencing. Shannon’s entropy  Effect of ribavirin on single nucleotide variant populations. Genetic diversity was also evaluated in terms of single nucleotide variants (SNVs), defined as a nucleotide that differs from the consensus sequence by at least 1% of the population at that position.P = 0.033) (Treatment of Asibi virus with ribavirin resulted in a dose-dependent increase in both the number of SNVs and the number of nonsynonymous variants compared to synonymous variants. The 0.5 μM concentration caused the largest increase in genome variation . Twenty = 0.033) . With 1.= 0.033) . Interes= 0.033) . The E-410.1128/mBio.02294-19.1TABLE S1Table S1, DOCX file, 0.1 MB.17D-204 treated with ribavirin generates less SNV than does ribavirin-treated Asibi. Variants were detected using V-Phaser2, and variants below 1% frequency and those that did not pass the strand bias test were discarded. Variants highlighted in green are common throughout the samples. Variants in red text are residues which distinguish 17D-204 and Asibi. Download Copyright © 2019 Davis et al.2019Davis et al.Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the P > 0.999).Although the number of SNVs in the 17D virus samples increased with increasing concentration of ribavirin, the frequency of variants detected remained low. The number of nonsynonymous SNVs was consistently lower than synonymous SNVs, suggesting that the virus was not responding to the selection pressure posed by ribavirin. In total, only seven nonsynonymous SNVs were detected across all ribavirin treatments sequenced, with each SNV being present in less than 5% of the population . Structu(iii) NGS analysis of WT FVV versus FNV virus. In order to confirm the resistance of vaccine strains to ribavirin treatment, untreated and 0.5 μM ribavirin FVV- and FNV virus-treated samples were sequenced by next-generation methods. As with Asibi virus, WT FVV experienced large increases in viral diversity by entropy (P < 0.0001). Consistent with the 17D findings, the number of SNVs detected in ribavirin-treated  and untreated  FNV virus samples were consistent in both number and identity  Sensitivity to ribavirin is controlled by YFV nonstructural proteins. To exclude virus passage history in the antiviral activity of ribavirin and determine if the YFV replication complex was involved in the resistance of 17D to ribavirin, infectious clone-derived viruses and structural chimeras  were generated using an infectious clone system (P > 0.999). Thus, the results of infectious-clone-derived viruses show that the effects of ribavirin were not due to the passage history of the viruses. The IC50 of the ribavirin-treated Asibi/17D prME virus  was similar to that of the Asibi virus infectious clone  (R2 = 0.68) was resistant to ribavirin, similar to the 17D virus   . The 17D = 0.88) , indicatLive-attenuated virus vaccines have been successfully used to combat infectious disease for decades, but due to their empirical derivation, little is known about their mechanisms of attenuation. This lack of understanding makes the development of next-generation live-attenuated vaccines difficult. The live-attenuated YF 17D vaccine virus has been used for 80 years to successfully reduce YF disease, and over 850 million doses of the vaccine have been distributed with exceptionally low levels of vaccine-associated adverse events . The suc50 values, 0.37 μM and 0.51 μM for Asibi and FVV, respectively) is consistent with findings from studies of other RNA viruses encoding a low-fidelity RdRp, such as Lassa and Crimean-Congo hemorrhagic fever viruses   , 40, 41. strains . It has  strains . In orde strains , 32. The and FNV . Togethe and FNV .50 similar to that of Asibi virus and had increases in diversity and SNVs, of which 66% were detected in the replication complex. Our studies indicate that ribavirin is being incorporated into the genome of WT YFV and should be tested as a treatment for YFV infection. This warrants further investigation, as most studies to date have utilized 17D vaccine virus as a model system, which is not representative of WT YFV strains  of hepatitis C virus, dengue virus NS5 methyltransferase, poliovirus RdRp, and chikungunya virus nsP4, which contains the RdRp , 44, 45. strains , 49.Little variation was detected in the genotype of ribavirin-treated YFV vaccine strains, and only a negligible decrease in infectivity titer was observed, suggesting a resistance to the lethal mutagenesis of ribavirin by YFV vaccine strains. This confirms previous reports examining the activity of ribavirin against 17D virus . In compIt is possible that the extensive serial passaging of WT Asibi virus and FVV  caused the WT virus to adapt to an unchanging environment where an unwavering consensus sequence was the most fit. This adaptation caused the virus to be less fit in a biological environment where the vaccines could no longer respond to selection pressure  . ResistaIn this study, the effects of ribavirin on two WT YFV strains and the two LAV strains derived from them were compared under a hypothesis that an acquired fidelity mechanism contributes to the attenuation of an empirical flavivirus vaccine. It had been previously shown that 17D was resistant to the drug; however, the WT Asibi strain was not tested in a similar context, which was performed herein . The WT Ribavirin  was reconstituted in high-performance liquid chromatography (HPLC)-grade water and sterilized using a 0.2-μm filter. A 10 μM working solution was then obtained by diluting the stock in minimal essential medium (MEM) supplemented with 2% fetal bovine serum (FBS). This working stock was used to make ribavirin concentrations of 6 μM, 2 μM, 1 μM, 0.7 μM, 0.5 μM, 0.2 μM, 0.15 μM, 0.1 μM, 0.05 μM, and 0.02 μM for the antiviral assays.MPA  was reconstituted at 25 mM in HPLC-grade water. A 1 mM working solution was obtained by diluting the stock in MEM supplemented with 2% FBS. This working stock was used to make a dose-response curve of 2-fold dilutions from MPA concentrations of 1 mM to 0.24 μM.KF769016) and is the lowest passage number available, namely, six passages in rhesus macaques and 3 passages in C6/36 cells. FVV (GenBank accession no. U21056) and FNV-Yale (GenBank accession no. U21055) viruses used were received from the WRCEVA and passaged once in Vero cells . The Asibi virus used was obtained from the World Reference Center for Emerging Viruses and Arboviruses . Virus was harvested at 80% cytopathic effect (CPE) and sequenced by Illumina methods to confirm sequence identity .Chimeric 17D-204 and Asibi viruses with prM and E (prME) genes swapped were constructed using infectious clone systems , 55. BriAll viral infectivity assays were undertaken in biosafety level 3 (BSL3) facilities at the University of Texas Medical Branch.As described by Leyssen et al. , viral iAY640589) using Bowtie2 with paired-end, very sensitive local parameters. PCR duplicates were removed with MarkDuplicates . All samples were downsampled to the lowest mean coverage of the data set, 9,984 reads per base. Shannon’s entropy was calculated using custom R scripts over possible nucleotide variables  using the method described by Nishijima et al  and sequenced on an Illumina HiSeq 1500 platform. Reads were trimmed of adapter sequences and quality controlled for quality scores greater than 30 (Trimmomatic). The reads were aligned to a reference genome  to determine antiviral 50% inhibitory concentrations (IC50) values and significant differences between four-parameter nonlinear fits of sensitivity curves. A one-way ANOVA was performed with Friedman’s post hoc multiple-comparison test (P < 0.05) to determine significant differences in average entropy across viruses and treatments. A one-way ANOVA was performed with Kruskal-Wallis post hoc multiple-comparison test (P < 0.05) to determine significant differences in the frequencies of variants.All statistical analyses were completed in the GraphPad Prism (v7.2.1). A one-way analysis of variance (ANOVA) was performed with Friedman’s


Meningiomas present as intracranial extra-axial lesions with dural attachment, which are primarily managed surgically. The extent of resection (EOR) may vary depending on patient- and tumor-related factors. The aim of this study is to identify preoperative predictive factors of EOR and to propose an estimation of the predicted gross total resection (GTR) based of patient- and tumor-characteristics. 1469 patients from a retrospectively (1990 to 2002) and prospectively managed (2003 to 2010) databank of Oslo University Hospital, Norway, totalling 11,414 patient-years of follow-up were included. Patients had a mean age at surgery of 64 ± 20.1 years with a female-to-male ratio was 2.4:1 and a mean KPS of 81.2 ± 12.1. Skull-base meningiomas represented 47% of all cases. WHO grades were I in 92.3%, II in 5.2%, and III in 2.2%. Bone infiltration was described in 18.7% of cases. 39.3% of patients had Simpson I resection, 34.3% had Simpson II, 5.4% had Simpson III, 20.6% had Simpson IV, and 0.5% had Simpson V. The risk factors for incomplete resection were: symptomatic presentation (OR 0.56 [0.43–0.72]), skull-base location (OR 0.79 [0.70–0.88]), and bone invasion (OR 0.85 [0.73–0.99]). Using a recursive partitioning analysis, we propose a classification-tree for the prediction of GTR rate based on preoperatively determinable patient- and tumor characteristics. The identification of preoperative predictors of poor GTR rate may aid clinicians managing meningioma patients. In selected cases were the predicted GTR rate is low, staged treatment with surgical debulking followed by adjuvant therapy may be favored in order to minimize postoperative morbidity and mortality. Owned to an aging population and the increasing availability of imaging diagnostics, more incidental meningioma are detected, thus leading to a higher overall incidence5. Depending on the size, location, and anatomical relationship of the tumor with the surrounding structures, achieving a complete resection can be challenging. The extent of resection (EOR) is quantified using the Simpson scale6. Several studies assessed EOR as a prognostic factor of overall and progression-free survival (OS and PFS)11, but none addressed specifically preoperative factors determining EOR. However, identification of predictive factors of surgical resection might be helpful for “personalized surgery”, i.e. in tailoring surgical resection on a case-by-case basis.Besides surveillance, the therapeutic management of meningiomas is primarily surgical and aims at maximal tumor removal as the complete removal of tumor and its dural tail, which is important for later disease control, and to obtain a pathological diagnosisThe aim of this study is to identify predictive factors of EOR and to propose an estimation of the predicted EOR based on patient’s and tumor’s characteristics.1469 patients  surgically treated for a meningioma were identified. The mean age at surgery was 64 ± 20.1 years. The female-to-male ratio was 2.4:1. The mean Karnofsky performance status (KPS) was 81.2 ± 12.1. Neurologic deficit was the most common presenting symptom (60.2%), followed by signs of intracranial hypertension (ICH) (31.7%) and seizures (29.6%). 5.4% of patients were asymptomatic. The mean follow-up was 7.8 ± 5.5 years  grades were I for n = 1352 (92.3%), II in n = 77 (5.2%), and III for n = 32 patients (2.2%). Bone infiltration was described in n = 274 (18.7%) of the cases  of surgeries  had Simpson I resection, n = 503 (34.3%) Simpson II, n = 79 (5.4%) Simpson III, n = 302 (20.6%) Simpson IV, and n = 8 (0.5%) Simpson V. GTR defined as a Simpson grade I, II or III resectionSkull base meningiomas were associated with higher Simpson grades than non skull-base meningiomas. This held true especially for anatomically difficult locations such as the orbit, the petroclival region or the cavernous sinus. As expected, GTR was more often achieved in convexity (96.7%) and lateral sphenoid wing (87.2%) meningiomas.Three independent risk factors for incomplete resection were identified: symptoms at presentation  (OR 0.56 0.43–0.72]), a skull base meningioma location (OR 0.79 [0.70–0.88]), and associated bone invasion (OR 0.85 [0.73–0.99]) (Table ), a skulThe identified preoperative predictors of meningioma EOR were put in hierarchical order using the recursive partitioning analysis with: the existence of symptoms at presentation, a tumor bone invasion, a skull-base located tumor, the patient’s sex and the preoperative KPS Fig. . The GTR13. The preoperative KPS was not a significant predictive factor of EOR despite being predictive for postoperative neurological outcome, OS and PFS15. Age, preoperative KPS, and female-to-male ratio in our cohort were comparable to meningioma patients characteristics in the literature18. Similar to other series19, a Simpson grade I resection was achieved in 39.4% and a GTR in 78.9% of the cases.The presence of symptoms at presentation, a skull-base location and/or a bone invasion were all negative predictive factors for GTR. To the best of our knowledge, this is the largest study analyzing predictive factors of the surgical EOR in meningiomas21.Besides the presence of symptoms at presentation, the patients’ baseline characteristics were not predictive for EOR. The clinical presentation  was the sole patient-dependent factor identified as a significant factor of low EOR. This never previously described association may be explained by the surgeon’s apprehensiveness to aim for GTR in an already neurologically impaired patient. Increased GTR rates in patients with preoperative seizures may be linked to the meningioma’s location. Convexity and anterior skull base meningiomas are notoriously more epileptogenic, while also being more surgically accessible22. Patients’ gender was not a significant independent prognostic factor in multivariate analysis, despite the increased incidence skull base meningiomas in women 16. In our study, the preoperative KPS was not a significant predictive factor of surgical resection’s quality despite being an important predictor of postoperative neurological outcome in several publications15.Patient’s age was not a significant prognostic factor of EOR. Simpson I resection was generally attempted even in older patients, which is in line with previous studies24. This difference was particularly stark between convexity and cavernous or petroclival meningiomas, where a Simpson grade I or even grade II resection was rarely possible.Meningiomas with skull-base location and bone invasion were less often completely resected. This is unsurprising as these locations can be technically more challenging due to their restricted surgical access and vicinity to vascular and/or neurological structures25.Bone invasion was another significant independent factor of poor resection quality. This infiltration requires additional drilling of the bone close to the dural insertion, often on the skull flap. In certain cases, bone invasion management represents a major part of the surgical procedure, for instance in spheno-orbital meningiomas where the extent of bone invasion and cavernous sinus involvement may not allow complete removalThis study is the first to propose a classification-tree of the predictors of EOR Fig. . As can 11. This holds especially true in view of other available adjuvant therapeutic options such as radiotherapy28, second surgery31, or a watchful wait-and-scan follow-up.Considering these factors may be helpful in the decision-making process and the planning process of the surgical resection. The proposed classification tree that allows for a rapid estimation of the estimated EOR in consideration of the patient and tumor preoperative characteristics. Although complete surgical resection while preserving the patient’s neurological status is the goal, a Simpson I resection is neither always attainable nor does it guarantee increased OS or PFS32. Hence, a staged treatment with debulking followed by adjuvant treatment might be preferable in order to minimize postoperative morbidity and mortality, especially in skull-base meningiomas. The predictive factors of low EOR identified in this study may help neurosurgeons to identify those patients who may fare better with staged treatment33.Recently, there has been a trend towards more conservative treatment for meningiomas, mainly because it has been shown that the Simpson grade is not universally applicable to all meningiomasThe main strengths of this clinical study are the clinical setting, the number of patients as well as the length of follow-up up to 21 years. Loss of follow-up was minimal since all patients with a postoperative complication or recurrence were systematically re-referred to our tertiary center. Only one patient was lost to follow-up, moving abroad. The data stem from one neurosurgical center with mostly homogenous surgical practices. This can make the generalization of our finding to all patients difficult. All patients with a histologically proven meningioma were included, which reduces selection bias. The retrospective data collection before 2003 is a limitation. Also, due to the long period of data collecting starting in 1990, radiological data, especially MRI, tumor size and molecular biomarkers such as Mib-1 or Ki67 were not available for all patients and were included in the statistical analysis.The choice to regroup all meningiomas together for the statistical analysis and the data interpretation may also be subject to question as meningioma is not a homogenous pathology, with different pattern of evolution and therapeutic management for specific subgroups, especially depending of the location and WHO grade. The global results may not be representative of these specific categories of meningioma.Clinical symptoms at presentation, skull-base location and bone invasion were significant predictors of a poor EOR for meningiomas. The identification of these factors may aid clinicians managing patients with meningiomas.Data were acquired from a retrospectively (1990 to 2002) and prospectively managed (2003 to 2010) databank of Oslo University Hospital (OUH). OUH is the main Norwegian tertiary referral center and has two neurosurgical units , which covers an area of approximately three million inhabitants, i.e. 56% of the Norwegian population.et al.34. All patients were operated on by the neurosurgical teams of OUH. The EOR was assessed using the Simpson grade scale, based on the surgical report in conjunction with post-operative imaging. Gross total resection (GTR) was defined as a Simpson grade I, II or III resection12. The histopathological diagnosis of meningioma was confirmed by experienced neuropathologists.All patient operated for a meningioma during the study period at OUH were included. Preoperative imaging studies were reviewed to confirm tumor location and size, contrast enhancement, and presence of calcification. The WHO grading system was used to classify the histology of meningiomas. The WHO criteria changed during the study period. From 1990 to 2001, the tumors were classified as benign, atypical or anaplastic. The present WHO-grading system for meningioma was implemented in 2001, which divides the tumors into grade I, II and III. For this study, we reclassified the tumors operated before 2001 to the present WHO classification; benign = WHO grade I, atypical = WHO grade II and anaplastic = WHO grade III. The definition of skull base meningioma was based on Al-Mefty The study was regulated by the Personal Data Act/Personal Health Data Filing System Act and approved by the Data Protection Official at OUH (2017/5204). Informed consent was not required according to the Personal Data Act/Personal Health Data Filing System Act.https://www.r-project.org). The significant p-value threshold was defined at 0.05. Multivariate analysis was performed using a linear generalized model approach. The variable considered for the multivariate analysis were patient’s age, sex, preoperative Karnofsky prognostic scale (KPS) and clinical status, as well as the tumor location, WHO histopathological grade and the presence of a bone invasion.Statistical analysis was performed using R v3.5.1 (The above-mentioned factors were used to build a classification of predictors of EOR using the classification and regression tree (CART) recursive partitioning analysis. The variables considered were patient’s age, sex, preoperative KPS and preoperative status, as well as tumor location and bone invasion. The generated CART tree was pruned by adjusting the complexity variable to minimize the estimated error in order to avoid data overfitting.