This study included 6,971 students in grades 9 and 10 (ages 13 to 16 years) from 158schools who participated in the 2009/2010 Health Behaviour in School-aged Children Study. Students provided information on where they typically ate lunch. The number of food retailers was obtained for six road network buffer sizes  surrounding schools. Associations between the presence of food retailers near schools and students’ lunchtime eating behaviours were examined using multilevel logistic regression. Comparisons of model fit statistics indicated that the 1,000 m buffer provided the best fit. At this distance, students with ≥3 food retailers near their schools had a 3.42 times greater relative odds (95% CI: 2.12–5.52) of eating their lunchtime meal at a food retailer compared to students with no nearby food retailers. Students who had ≥2 food retailers within 750 m of their schools had a 2.74 times greater relative odds (95% CI: 1.75–4.29), while those who had ≥1 food retailer within 500 m of their schools had 2.27 times greater relative odds of eating at food retailer (95% CI: 1.46–3.52) compared to those with no nearby food retailers. For distances greater than 1,000 m, no consistent relationships were found. Over the past three decades, the prevalence of obesity among youth has increased dramatically worldwide . There hThe food retail environment has been measured using both subjective ,18 and oet al., [Recent literature reviews have identified the selection of the appropriate buffer size as a key methodological issue for studies of the food retail environment ,23. Someet al.,  found thet al., ,24,25, wet al., ,20, limiThe primary objective of this study was to identify the most appropriate buffer size to use when studying the relationship between the school food retail environment and the eating behaviours of Canadian students. To achieve this, the food retail environment surrounding schools was measured using several buffer sizes. Multilevel logistic regression was used to examine associations between the number of food retailers in each buffer size and students’ lunchtime eating behaviours. Model fit statistics were used to determine the most appropriate buffer size.This was a multilevel cross-sectional analysis of schools participating in the 2009/2010 Canadian Health Behaviour in School-aged Children (HBSC) study. Addresses of food retailers near schools were obtained using an online food retailer database. The number of food retailers surrounding each school was obtained for the following road network-based buffers: 500 m, 750 m, 1,000 m, 1,500 m, 2,000 m and 5,000 m. Associations between the presence of food retailers and students’ reports of eating their lunch at a food retailer were then assessed.The 2009/2010 Canadian HBSC study involved a survey of students from 436 schools and collected information on a variety of health behaviours in students in grades 6 to 10 (approximate ages 11–16 years). Classes were the primary sampling unit, and they were stratified by province, with an oversampling of some provinces and the three northern territories. Two Canadian provinces with small populations (New Brunswick and Prince Edward Island) were unable to participate in the 2009/2010 survey. The HBSC also excludes students in private schools, incarcerated youth, special needs schools and students who are home schooled. Ethics approval was obtained from the Queen’s University Health Sciences Research Ethics Board. Subject consent was obtained at the school board and school levels as well as from parents or guardians .In the current analysis, the sample was restricted to the 169 schools where students were permitted to leave school grounds, making it possible for them to access nearby food retailers. Because only 1.1% of grade 6 to 8 students reported the study outcome, they were excluded from the analysis. Due to missing information on food sources within schools or in the school neighbourhoods, a further 11 schools were excluded. Finally, 940 students were excluded because of missing data on key variables. The final analyses involved 6,971 students from 158 schools.The addresses of the 158 eligible HBSC schools were mapped in ArcGIS  and road network-based buffers were constructed using the following distances: 500 m, 750 m, 1,000 m, 1,500 m, 2,000 m and 5,000 m. These distances were selected based upon existing precedents ,26,27,28www.yellowpages.ca). The Yellow Pages directory was chosen because it provided the most accurate information available on food retailer locations [Convenience stores, fast food restaurants, and coffee/donut shops were used to develop the primary independent variable of interest. These food retailers directly corresponded to the lunchtime eating question used to create the study outcome. Addresses of these food retailers were obtained using an online Yellow Pages directory  was used to confirm the location and obtain latitude and longitude coordinates to map them manually in ArcGIS. Because the number of food retailers within the buffers was positively skewed, they were categorized into groups. All categories were based on thresholds established for the 500 m network buffer size. At this distance, 74% of schools had no nearby food retailers (categorized as “none”), and the remaining 26% were categorized as having “1 or more” food retailers. For the remaining buffer sizes, a “none” category was created, and a threshold of the top ~26% was used to denote the category with the highest number of food retailers. The thresholds for the remaining categories were created to ensure an approximately even distribution of schools across the categories and similar sizes among the categories.Wheredoyouusuallyeatyourlunchormid-daymealonschooldays?” Students who chose the response “snack-bar, fastfoodrestaurant, café” were considered to regularly purchase their lunch from food retailers. Those who chose the remaining responses  were classified as those who did not typically obtain their lunch from food retailers.The outcome of this study was obtained from the response to the following question: “Individual-level variables, including age, sex, and socioeconomic status were considered as potential confounders since fast food consumption varies by these characteristics ,32,33. TAll analyses were conducted using SAS statistical software, version 9.2 . Multilevel logistic regression was carried out to examine the relationship between the presence of food retailers near schools  and the likelihood of students eating their lunch at these food retailers using different sized road network buffers. For each buffer size, the multivariate model building process began with the introduction of the individual-level confounders and proceeded using a backwards elimination approach. Next, the school-level food exposure variables were forced into the model because we were interested in accounting for food sources within school as well as those surrounding schools. The Akaike information criterion (AIC), which is a measure of goodness-of-fit when comparing two or more regression models, was determined for the final multivariate models. A difference in AIC values of between 2 to 7 indicates a moderate difference in fit of the models, while a difference of 7 or more indicates a large difference in model fit . Using tThe median number of student respondents per school was 37 (interquartile range: 23–55). Results of the multilevel logistic regression are shown in The AIC-related values comparing goodness-of-fit between the final regression models  buffer size may be the most appropriate [i.e., 1,600 m) buffer sizes are most appropriate [The association between the number of food retailers within 500 m, 750 m, and 1,000 m road network buffers around schools and students’ eating behaviours was consistent with observations by Davis and Carpenter . They foropriate ,25, and ropriate .Results from this study provide important evidence regarding the appropriate buffer size to use when measuring the food retail environment surrounding schools. Namely, using a buffer that is either too large or too small may obscure relationships with eating behaviours and result in weakened study findings. Furthermore, having a standard buffer size will enhance comparability across studies. Finally, results from studies such as this one are needed to inform the implementation and evaluation of policies aimed at optimizing the built environment surrounding schools. Specifically, our findings suggest that policies that limit the number of food retailers around schools should consider a 1,000 m distance and a maximum of 2 fast food restaurants, coffee/donut shops, and/or convenience stores. More than one quarter (28.4%) of the schools studied here would be affected by such a policy.This study has some key strengths including: the large and geographically diverse study sample, the wide variety of buffer sizes, and the use of goodness-of-fit statistics to assess the relative contribution of each buffer size to model fit. Another key strength of this study is that it considered constraints on mobility and time when measuring accessibility to food retailers. In other words, by limiting our study outcome to a specific time and place to eat , we accounted for these limitations on spatial accessibility. Measures of spatial accessibility that consider limitations imposed by time and space are weakly correlated with those which do not consider them  and thisA key limitation of this study was that the GIS database used to obtain information on the food environment does not provide a completely accurate measure of the food environment. Furthermore, for practical reasons, only the top chain fast food restaurants and donut/coffee retailers were included in the food retail environment measures. In addition, the survey data were obtained by self-report and this may introduce bias due to the social desirability of eating healthy foods . It is aThe results from this study indicated that the most appropriate road network buffer distance when assessing the food retail environment surrounding Canadian schools was 1,000 m. Future studies investigating the school food retail environment should consider using street network buffers of this size. Having a consistent buffer size across studies will not only help inform policies and interventions directed at the modification of the food environment surrounding schools, but will also help facilitate comparisons across studies.


Lymantriadispar), one of the most recognized pests of forests and ornamental trees in the world. We assessed genetic diversity and structure in eight geographic populations of gypsy moths from China using five polymorphic Inter simple sequence repeat markers, which produced reproducible banding patterns. We observed 102 polymorphic loci across the 176 individuals sampled. Overall genetic diversity  was 0.2357, while the mean genetic diversity within geographic populations was 0.1845 ± 0.0150. The observed genetic distance among the eight populations ranged from 0.0432 to 0.1034. Clustering analysis , revealed strong concordance between the strength of genetic relationships among populations and their geographic proximity. Analysis of molecular variance demonstrated that 25.43% of the total variability  was attributable to variation among geographic populations. The results of our analyses investigating the degree of polymorphism, genetic diversity (Nei’s and Shannon) and genetic structure, suggest that individuals from Hebei may be better able to adapt to different environments and to disperse to new habitats. This study provides crucial genetic information needed to assess the distribution and population dynamics of this important pest species of global concern.This study provides the first genetic characterization of the gypsy moth from China ( Lymantriadispar Linnaeus (Lepidoptera: Lymantriidae), is a destructive defoliator with a broad host range, and one of the most recognized pests of forests and ornamental trees in the world. Indeed, it is the only forest insect to be included in the American Quarantine regulations [Lymantriadispardispar and the Asian gypsy moth (AGM), Lymantriadisparasiatica, based on preferred host plant usage and the females’ flight abilities. The EGM was originally found in Europe, but was accidentally introduced into North America from France in 1869 [o and 58o north, covering an area of more than 20 provinces or autonomous regions [The gypsy moth, ulations . The gyp in 1869 , where i in 1869  and to t in 1869 . AccordiGenetic variability is a crucial indicator of the capacity of a species or population capacity to adapt to the environment . SpeciesL. ddispar.  subspecies and the two Asian subspecies asiatica and japonica [Previous research using microsatellite DNA markers to assess genetic variability in four gypsy moth populations from Japan, eastern Russia, China, and North America revealed that genetic variation in the North American population was lower than that in the Asian populations . Amplifijaponica . In all In this study, we assessed the genetic diversity of the gypsy moth within China using Inter Simple Sequence Repeat (ISSR) markers. The ISSR technique entails the amplification of DNA fragments located between two closely and inversely oriented microsatellites . PrimersLdispar.  sourced from eight locations . Compared to traditional insect DNA extraction methods, such as SDS  and CTABAristaeomorpha foliacea ISSR work [Ldispar.  DNA; however, twenty-seven just produced single patterns. Ultimately, five ISSR primers capable of producing reproducible and unambiguous multiple , 0.75 U Taq DNA Polymerase  and ddH2O to the final volume. The amplification reaction was performed in a DNA Engine Thermal Cycler PTC-200 . The conditions were 94oC for 5 min, 35 cycles of 94oC for 45 s, annealing at their respective temperature for 45 s and 72oC for 90 s, and final extension at 72oC for 10 min. For each primer, replicates and negative controls were included to ensure the consistency of results.All PCR amplifications were carried out in a 20 µL volume, containing 10 ng of template DNA, 2 µL 10× PCR buffer , 1 µM of primer , 240 µM of each dNTP , 1.5 mM of MgClTo analyze the PCR product, 5 µL of product was added to 1 µL of 6× loading buffer , and this mixture was loaded in a 2% agarose gel  for electrophoresis with 0.5× TAE (Tris/Acetic/EDTA) electrophoresis buffer. After running at 100 V for 1 h, the gel was stained in ethidium bromide (EB) buffer for 3 min, rinsed with distilled water and photographed in a gel documentation machine . A DL2000 DNA marker  was run as a standard with each primer.The presence or absence of bands was scored as a 1 or a 0, thusour data was consisted by a presence/absence (1/0) matrix of 102 multiple bands from 176 individuals.Ldispar.  populations was obtained by Nei’s original measurements [The program PopGene32  was usedurements . And a dFST, with 95% confidence intervals (CI) obtained by bootstrapping 1000 replicates over all loci.To examine the partitioning of genetic variance within and among samples from different geographical regions, Analysis of molecular variance (AMOVA)  was carrOverall, 102 different bands, varying from 280 to 2,000 base pairs, were obtained by the five primers screened for 176 individuals across the eight geographic populations. All primers screened in this work were polymorphic, and the number of polymorphic bands per primer ranged from 16, for IT2, to 23, for UBC847. The primer which amplified the highest mean number of bands per population was IT1 (16.87 polymorphic bands); whereas that producing the lowest mean value was IT3 (9.25 polymorphic bands). Hebei was the geographic population with the highest number of bands (84 polymorphic bands); whilst Arxan, Inner Mongolia was the geographic population with the lowest mean number of bands  and Shannon’s information index (I = 0.3735) showed relatively high overall genetic diversity for gypsy moth from China. Among the different geographic populations, Hebei showed the highest genetic diversity , while Arxan, Inner Mongolia showed the lowest  .Ldispar.  ranged from 0.0432 to 0.1034, suggesting a large genetic base  was estimated as 1.466 via the formula: Nm = (1- FST)/2FST [AMOVA conducted on ISSR markers confirmed the presence of significant genetic differentiation among the eight geographic populations /2FST . This vaLymantriaxylina Swinhoe and Lymantriamonacha Linnaeus when monitoring with attractant the next years . To determine the genetic composition and structure of the gypsy moth from China, we conducted a large-scale analysis of genetic variability across eight different geographic populations, utilizing novel ISSR molecular markers [Ldispar.  in China.The AGM has received close attention as a potential pest species over the past few decades because, unlike EGM, their eggs could complete diapause under a lower chill requirement , their l markers . This inLdispar.  with AFLP which demands of preliminary pedigree analysis, independent inheritance and alletic assignment of AFLP markers [The efficiency of molecular markers is evaluated by the degree of polymorphism detected among the populations under investigation. All five primers, UBC818, U847, IT1, IT2 and IT3 used for analyses in this study were polymorphic. Unlike another previous genetic study of  markers , the ISSLdispar.  populations from different localities, revealed that the Hebei population (HB) had the highest percentage of polymorphic loci (82.35%), suggesting this population could be the longest-existence of the eight populations or could be the largest population. Individuals from Arxan, Inner Mongolia (MA) showed the least genetic diversity, probably because of the frequent eradication measures undertaken by the local government every year.Polymorphism of a population depends on the existence of genetic variants, represented by the number of alleles per locus and by their frequency distribution in the population . The resH = 0.2410, I = 0.3707), agreeing with the data for percentage of polymorphic loci. This suggests that individuals from Hebei possessing greater information content may be better able to adapt to different environments and, therefore, to disperse to new habitats.Our analyses revealed high genetic diversity in the gypsy moth from China (as measured by both the Nei’s and Shannon’s index), which may be explained by the great dispersal capabilities at adult stage of AGM. Hebei (HB) had the highest genetic diversity of all the geographic populations  lending further support to the finding of strong genetic structuring among the Chinese AGM. What is noteworthy is that there exists large degree of genetic separation among the five northern Chinese populations, which might be driven by the accumulation of genetic differences via adaptation of these populations to their differing environments.Cluster analyses utilizing UPGMA revealed stronger genetic relationships among geographically proximate locations, suggesting that more closely related populations share a common origin, or evolutionary history. Our results also revealed high overall genetic differentiation among populations  and genetic relationships among different populations all indicate that individuals from Hebei pose the greatest threat as potential pests, and should be closely monitored by the phytosanitary department. The genetic information on the Chinese AGM obtained in this study will assist in elucidating crucial information on the distribution and population dynamics of this important pest species of global concern.


While baculovirus genomes are well characterized, the genome of S. frugiperda is not sequenced and the virus-host molecular interplay is sparsely known. Herein, we describe the application of stable isotope labeling by amino acids in cell culture (SILAC) to obtain the first comparative proteome quantitation of S. frugiperda cells during growth and early baculovirus infection. The proteome coverage was maximized by compiling a search database with protein annotations from insect species. Of interest were differentially proteins related to energy metabolism, endoplasmic reticulum and oxidative stress, yet not investigated in the scope of baculovirus infection. Further, the reduced expression of key viral-encoded proteins early in the infection cycle is suggested to be related with decreased viral replication at high cell density culture. These findings have implications for virological research and improvement of baculovirus-based bioprocesses.Baculovirus infection  Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) infects a few insect lepidopteran species. The economic interest of BVs derives from their use as biopesticides and, along the past decades, as versatile protein expression systems AcMNPV propagation is usually a cell line derived from the fall armyworm S. frugiperda, particularly the Sf9 clonal isolate. More than simply a vehicle for viral propagation, insect cell lines are chosen for their robust growth, straightforward culture scale-up and post-translational modifications of expressed proteins Baculoviruses (BVs) are a diverse family of arthropod pathogens containing large DNA genomes reaching up to 180 kbp p10 and polh is relatively insensitive to the presence of 5′-cap-binding eukaryotic initiation factor eIF4E Knowledge on the BV infection cycle has accumulated over many years of research. Briefly, replication and assembly take place in the nucleus, where viral particles travel by reorganizing the actin cytoskeleton upon cell entry AcMNPV susceptible cell lines have not been sequenced. This constitutes a major disadvantage for the use of genome–wide technologies for better understanding the host-pathogen interplay. So far, global expression patterns of Sf9 cells at mRNA level have been recorded at various time-points following infection AcMNPV infection in a different, permissive cell line In spite of the biotechnological interest, the genomes of Among the “omic” technologies available, quantitative proteomics based on stable isotope labeling by amino acids in cell culture (SILAC) has become a preferred choice for large–scale expression profiling, combining quantitative accuracy, with straightforward sample processing and control over culture conditions Sf9 cells associated with culture growth and BV infection. Since most of the host proteome is down–regulated at later times in the AcMNPV life cycle, we focused on the transition from early to late phase (around 6 hpi) to investigate the establishment of infection. The lack of a complete genome sequence was overcome by cross–referencing to a database constructed from S. frugiperda and related insect species. For quantitation, cells were cultured in the presence of heavy arginine and lysine, commonly used SILAC reagents S. frugiperda-based bioprocesses.In the present work, we investigate proteome expression changes in Sf9 proteome is the lack of an annotated genome sequence. This missing knowledge was overcome by compiling a search database with available annotations from related insect species, constructed as a subset of the non–redundant protein database maintained by the National Center for Biotechnology Information (NCBI). The proteome of Sf9 cells was examined and its coverage was further maximized by enriching raw datasets with MS spectra recorded from different experimental conditions, biological replicates and biochemical fractionation of samples  classification system was adopted to group proteins sharing homology (common ancestry) and homeomorphy (full–length sequence and domain architecture similarity) Sf9 proteins (approximately 361 out of 648).The major challenge for a large-scale assessment of the  samples . When an samples . In thisBombyx moriDrosophila melanogaster and Anopheles gambiae. Genomic information on S. frugiperda derives primarily from constructed EST libraries of larval hemocyte, fat body and midgut tissues, as well as the Sf9 cell line BmNPV, which shares an average 90% homology with AcMNPV Spodoptera litura NPV correspond to the same P35 and the large (RR1) and small (RR2b) subunits of ribonucleotide reductase, a virally encoded enzyme involved in synthesizing deoxyribonucleotides AcMNPV genome, and the pronounced homology with their eukaryotic counterparts indicates that they originate from the cellular proteome Among the organisms with the highest number of identifications are the only sequenced lepidopteran, the silkworm S. frugiperda EST libraries contain at least 35% of the potential total gene number S. frugiperda are ribosomal. The complete set of proteins with NCBI identifiers, species, PIRSF and GO assignments is provided as Unfortunately, an enrichment analysis of GO annotations across the different database species is hindered by incomplete sequence information. For example, it is assumed that the Sf9 cells were cultured for at least 6 passages in customized medium where Arg and Lys have been replaced by their respective heavy isotope counterparts (13C)–Arg (“Arg 6”) and (13C15N)-Lys (“Lys 8”). In parallel, cultures in light media were maintained as a control. The feasibility of performing SILAC in this cell line was confirmed by comparable average duplication times (22–24 h), high viability (>90%) and normal morphology. To assess the effect of culture growth and AcMNPV infection on the cellular proteome, a full factorial design of 2×2 comparisons was applied, whereby cells were grown to and infected at low  and high  cell densities  values lower than 1 or 0.5  was calculated for most of the proteins. Naturally, each of these ratios constitutes a source of experimental error and biological variability in the calculation of Rc. The question therefore is whether the previously applied mathematical correction of the SILAC ratios introduces additional data variability by means of error amplification. It could lead one to assume that a statistical outlier has biological significance, or produce a distribution of corrected ratios that is too spread out, hampering the identification of differentially expressed proteins. To investigate this, a sensitivity analysis of Rc with respect to R and Ri was performed . It can ln Rc was different from the average by a t-test with at least 95% confidence. This definition accounts simultaneously for the overall dispersion of the data as well as for the propagated S.D. associated with each corrected protein ratio, thus avoiding arbitrary definitions based only on location n) exists for uncorrected and incorporation ratios; those proteins that pass the test for the higher n value, but not for the lower one, were considered differentially expressed only if they lie outside of the bulk 95% of the data (Average ± 1.96 S.D.). This was also done for those proteins lacking a replicate for either R, Ri or both. Only protein ratios lying outside 68% of the distribution were considered to have biological relevance (Average ± S.D.). The combined set of proteins with statistically significant expression changes from the unfiltered and filtered distributions is considered to be the regulated sub-proteome for each experimental comparison . In the presence of experimental error and biological variability, their interacting effect can significantly obscure the assessment of how growth affects the cell proteome when comparing two infected cultures at different cell densities, for which reason we considered only uninfected cells in this case. Direct comparison of two infected cultures at different cell densities was carried out to discriminate possible changes in the expression levels of early BV proteins. On the other hand, the infection effect was evaluated not just separately for LCD and HCD cultures, but also globally by pooling together as “infection replicates” the six experimental comparisons performed in both conditions. Those proteins identified as differentially expressed from this pool will form a subset that has higher associated confidence.According to the statistical analysis described, a total of 13, 14, 14 and 9 proteins had significantly changed expression along culture growth, infection , infection at LCD and infection at HCD, respectively  and S5. vice versa.We also carried out a comparative analysis of the overall quantification results in the various experimental settings. The subsets of corrected SILAC ratios simultaneously quantified for the “treatment” pairs Growth/Infection  and Infection (LCD)/Infection (HCD) were standardized and plotted against each other . It is cCellular proteins with changed expression levels across the 4 experimental comparisons were grouped according to their potential physiological roles in growth and infection . In confOf the metabolic enzymes suggested to be regulated during culture growth, dihydrolipoyl dehydrogenase  and aldehyde dehydrogenase  are known to fuel energetic metabolism and were overexpressed at high cell densities. In contrast, several enzymes involved in different metabolic pathways were less expressed at HCD , consistent with the typical slowdown of growth along culture time. In particular, decreased anabolic fluxes through ME were previously reported by the authors A total of 11 cellular proteins were differentially expressed at the same time for the global infection assessment and either of the LCD or HCD infection conditions, showing consistent regulation in all cases. This indicates that gross quantitative errors are not present in our study. In terms of metabolic enzymes, PDH-E3 and ALDH were upregulated in at least one infection condition, underlying the importance of the energetic metabolism previously described for BV replication In terms of protein processing pathways, the up-regulation of HSP chaperones  has been associated with several viral infections, including BV Finally, of the proteins suggested to be only regulated by infection at LCD, the physiological roles of HSC70, Mod(Mdg4)–heS00531 and TRXL may imply limitations to BV replication during high cell density culture. In particular, the lower levels of LEF3 detected following infection at HCD, compared to cells infected at LCD, are consistent with the lower baculovirus progeny produced after infection at HCC when compared with LCC Sf9 cell factory was presented. The effect of BV infection and culture growth on the cellular proteome was investigated to gain insights into regulatory modulations associated with a productive infection cycle, thus relevant from both the virological and bioprocess points of view. Despite not having a complete genome sequence for this insect species, homology-based mass spectra interpretation gave the framework for genome–wide data extraction from limited knowledge. Inarguably, reanalyzing the data generated against a complete S. frugiperda database would unravel many more important players in this complex molecular interaction. To this respect, we expect this work to constitute a potential catalyst for further developments in the BV-Sf9 system.In this work, a comprehensive SILAC-based proteome analysis of the popular S. frugiperda cell lines require supplementation with serum or yeastolate. Cell adaptation to a minimal supplementation of these components could be cumbersome and induce alterations to normal culture behavior, affecting the infection cycle. Furthermore, serum dialysis was recently shown to induce marked changes in the proteomes and phosphoproteomes of several mammalian cell lines The analysis of differentially expressed proteins in SILAC experiments is straightforward when an almost complete incorporation (>95%) of heavy amino acids in cellular proteins is possible. However, atypical growth behavior of certain cellular types may complicate or completely prevent this ideal scenario 18O sample labeling that the hepatitis C virus significantly over expresses glycolytic and TCA cycle enzymes for energy generation, while also over expressing non–oxidative enzymes in the pentose phosphate pathway for biosynthesis. Furthermore, the same central catabolic pathways were observed to be activated following human cytomegalovirus infection in a metabolic flux profiling study, as well as the efflux to fatty acid biosynthesis necessary for viral envelope formation Our study was conducted at 6 hpi to assess cellular proteomic modulations associated with the establishment of infection, which will ultimately determine the outcome in terms of viral replication and recombinant protein expression. At this stage in the infection life cycle, Nobiron et al. The induction of energetic metabolism as a response to culture growth was also observed, a phenomenon previously described to be associated with a low proliferative, quiescent physiological state 2+ homeostasis, and the polypeptide transporter SRP57. These findings suggest that, through unknown mechanisms, BVs are able to take advantage of ER stress while avoiding negative effects associated with UPR, probably in different ways than already described for other viruses The early up-regulation of HSC70 and the related sHSP after LCD infection (but not HCD) indicate a disturbance of ER homeostasis known to be associated with viral infections. Interestingly, we observed expression changes in two proteins that together are counterintuitive with an activation of UPR, namely decreased levels of the chaperone ERp57, a central UPR effector also involved in CaHeliothis virescens, although at very late times after AcMNPV infection. More relevant, however, is the discovery that the multifaceted early P35 protein also acts as an antioxidant by quenching ROS Also interesting are the decreased levels of TRXL (only after LCD infection), since human thioredoxin (TRX) related proteins are known to integrate signals and prevent apoptosis from ER, oxidative and mitochondrial stresses. This complexity draws further curiosity into the strategies that different viruses have evolved to manipulate the cell for their advantage. For instance, the endogenous human T-cell leukemia retrovirus type I was shown to promote cell growth through elevation of TRX levels, while the apoptotic HIV infection is associated with decreased TRX expression S. frugiperda cells The role of LEF3 as a key determinant of a productive infection cycle is suggested by its lower expression at HCC compared to LCC infections. Noteworthy, a recent study has demonstrated that silencing of the replicative LEFs (including LEF3) during AcMNPV infection not only blocks viral DNA replication and late gene expression, but also prevents the induction of apoptosis and shutoff of cellular protein synthesis in Finally, among changed expression levels in several translation factors and ribosomal proteins, we found diminished levels of kinesin heavy chain (Khc), a tubulin motor protein, and up-regulation of a calponin-like protein (CLP), an inhibitor of myosin ATPase activity, which in turn is an actin motor protein. These observations suggest BVs reorganize the cytoskeleton not only through actin polymerization In conclusion, as a hypotheses generation tool, this proteomic study offers new insights and potential research targets for a better understanding of the complex BV infection cycle. One exciting point remains to be discriminating active modulations induced by the virus and the natural cellular response to infection. The complete resolution of this puzzle will probably require complementary information from targeted, scale-down experiments and other high-throughput assessments of protein-protein, protein-DNA and protein phosphorylation networks. From a distinct perspective, our data highlights possible regulatory bottlenecks associated with the described cell density effect on viral replication. This is one of few examples where large–scale quantitative proteomics was used with a focus on improved performance of an animal cell factory, in the scope of Systems Biotechnology. In a broader sense, this work provides a case–study of the application of advanced “omic” technologies in poorly characterized organisms.Sf9 insect cell line (ECACC 89070101) was kindly provided by Dr. Otto W. Merten . Culture maintenance was done as previously described 13C)-Arg and (13C15N)–Lys at 1.35 mM each (Cambridge Isotope Laboratories). Cells were cultured in this medium for 5 passages growing as 20 mL flask suspension cultures and then up-scaled to 70 mL for an additional 1–2 passages. In parallel, cells were cultured in light medium containing (12C)–Arg and (12C14N)–Lys at the same concentration. For infection experiments, cells were grown to the desired cell density and inoculated with a recombinant AcMNPV at a MOI of 5 infectious particles per cell. Information on virus composition, handling and quantification are provided in Carinhas et al. The commercially available Cells were lysed in basic RIPA buffer containing 25 mM Tris (pH 7.6), 150 mM NaCl, 1% NP-40, 1% Na deoxycholate, 1 mM EDTA, and supplemented with 1 mM PMSF and 1x complete protease inhibitor cocktail . Before further handling, the protein content was quantified for each extract  and equal protein amounts of labeled and unlabeled material from each culture were pooled. Incorporation of iosotopically labeled amino acids was checked on pooled samples from light and heavy cultures as well as unmixed samples. For this, proteins from the lysate were precipitated by adding 9 volumes of ice–cold 13.3% trichloroacetic acid/0.07% β-mercaptoethanol (β-ME) in acetone and incubated overnight at −20°C. Protein content was collected by centrifugation, washed twice with 90% acetone/0.07% β-ME and dissolved in 50 mM Tris–HCL/8 M urea (pH 8). Samples were reduced with 10 mM dithiothreitol and alkylated with 50 mM iodoacetamide, followed by overnight dialysis against 50 mM Tris–HCL/6 M urea (pH 8). Before trypsin digestion, the dialysate was first digested with 20 µg of Lys-C at 30°C for 2 h and then diluted with 5 volumes of 50 mM Tris-HCl (pH 8). Trypsin digestion was carried out overnight at 37°C using a 50∶1 protein to enzyme ratio. The peptide mixtures were acidified and approximately 1 mg of the initial protein content was desalted in 0.1% trifluoroacetic acid (TFA) using MacroSpin Column, Silica C18, 30–300 ug capacity . Peptides were biochemically stepwise eluted with 0.1% TFA containing 20%, 40%, and 60% acetonitrile. Before LC–MS/MS analysis, peptide fractions were dried in a speed vac and dissolved in 0.1% TFA./z 445.120025 LC–MS/MS analysis was performed on a LTQ-Orbitrap hybrid instrument . 2 µl of peptide digest were injected with an autosampler  onto a C18 trapping column  that was connected to a separation column (0.1 mm×10 cm) packed with Magic 300Å C18 reverse-phase material . A linear 80-min gradient from 2 to 50% solvent B (80% acetonitrile/0.1% acetic acid) in solvent A (2% acetonitrile/0.1% acetic acid) was delivered with an Agilent 1200 nano pump  at a flow rate of 300 nL/min. The eluting peptides were ionized at 1.7 kV. The LTQ-Orbitrap was operated in data-dependent mode. A survey scan between m/z 400–1600 was acquired in profile mode in the Orbitrap at 60,000 resolution and the 10 most abundant ions were then selected for fragmentation in the LTQ part of the instrument at normalized collision energy of 35%. Singly charged ions were omitted from fragmentation and previously selected ions were dynamically excluded for 25 sec. Scan-to-scan calibration was allowed by setting the lock mass to mst, 2010): all protein sequences for the insect species Manduca, Drosophila, Anopheles, Bombyx, Spodoptera, Adisura, Culex, Heliothis, Agrotis, Plubella, Apis and Mamestra were extracted from the non-redundant database and compiled into an insect–specific database. To this, a second database was collated having all protein sequences whose annotations contained the text string “insect” as well as baculovirus protein sequences.The LC-MS/MS data were searched with the SEQUEST search engine (version 3.3.1 SP1) The precursor ion and fragment ion mass tolerances were set to 10 ppm and 0.6 Da, respectively. Two missed cleavages were allowed. Dynamic modification was set for oxidized methionine, while cysteine carbamidomethylation was set as a fixed modification. The results were filtered for Xcorr values of 1.5 for singly, 2.0 for doubly, 2.5 for triply, and 3.0 for quadruply charged peptides. Delta CN was set to 0.1, and the peptide and protein probabilities were set to 0.5 and 0.01, respectively. In addition, all single peptide hit identifications were inspected manually for the correctness of identification.For SILAC ratio quantification, the area of those precursor ions that had been identified by SEQUEST were integrated by the PepQuan option of Bioworks . Precursor ion and fragment ion mass tolerances were set to 10 ppm and 0.6 Da, respectively. Searched files corresponding to different fractions of the same sample were merged together. Relative abundances were calculated as area ratios of heavy and light peptides, and the protein ratios were calculated as averages of all quantified peptides. The mass shift tolerance of “Arg 6” and “Lys 8” was set to 0.02 Da.The corrected unlabeled/labeled culture ratio for each protein was derived from Liao et al. where R represents the heavy/light isotope ratio from a 1∶1 mixed sample and Ri the heavy/light isotope ratio from the corresponding unmixed labeled sample. The error propagation from R and Ri into Rc is based on the unbiased standard deviation from triplicate experiments and was calculated according to the equation:Sensitivity analysis of equation (1) was performed by calculating the relative gradient with respect to R and Ri:ln) has the advantage of preserving data dispersion by only dividing each S.D. (Rc) by the respective Rc to yield S.D. (ln Rc). Maximum S.D. (ln Rc) cutoffs were then applied to each data set resulting in more focused normal distributions. The two-tailed Student's t-test was performed assuming unequal variances and unequal sample sizes.The corrected SILAC ratio distributions were symmetrized around 0 through logarithmic transformation. Using the natural logarithm . A high-titer virus stock was used to ensure minimal culture dilution. The percentage of GFP-positive cells was followed by flow cytometry, monitoring GFP expression under the control of the very late polh promoter. Synchronous infection can be observed by discounting 18–24 h from viral inoculation until full polh activity.(PDF)Click here for additional data file.Figure S2Sf9 cells growth profile in customized SF900 II medium. Cells were grown either without Arg, Lys or both amino acids. Medium supplemented with 1.35 mM Arg and 1.35 mM Lys was used as control. Undefined medium components may contain variable amounts of these amino acids.(PDF)Click here for additional data file.Figure S3Allocation of differentially expressed proteins for each experimental comparison after the application of S.D. (ln Rc) cutoffs to data distributions.(PDF)Click here for additional data file.Table S1Complete set of identified proteins with NCBI identifiers, species, PIRSF and GO assignments.(XLS)Click here for additional data file.Table S2Quantitative and statistical information on proteins with changed expression levels along culture growth.(XLS)Click here for additional data file.Table S3Quantitative and statistical information on proteins with changed expression levels as a response to infection .(XLS)Click here for additional data file.Table S4Quantitative and statistical information on proteins with changed expression levels as a response to infection at low cell density.(XLS)Click here for additional data file.Table S5Quantitative and statistical information on proteins with changed expression levels as a response to infection at high cell density.(XLS)Click here for additional data file.


Sardinella brasiliensis was found in Shelf Water; whereas in the south, Engraulis anchoita inhabited the Plata Plume Water. At the slope, Tropical Water was characterized by the bristlemouth Cyclothone acclinidens. The concurrent analysis showed the importance of both cross-shelf and latitudinal gradients on the large-scale distribution of larval fish species. Our findings reveal that ichthyoplankton composition and large-scale spatial distribution are determined by water mass composition in both latitudinal and cross-shelf gradients.Ichthyoplankton is an essential component of pelagic ecosystems, and environmental factors play an important role in determining its distribution. We have investigated simultaneous latitudinal and cross-shelf gradients in ichthyoplankton abundance to test the hypothesis that the large-scale distribution of fish larvae in the South Brazil Shelf is associated with water mass composition. Vertical plankton tows were collected between 21°27′ and 34°51′S at 107 stations, in austral late spring and early summer seasons. Samples were taken with a conical-cylindrical plankton net from the depth of chlorophyll maxima to the surface in deep stations, or from 10 m from the bottom to the surface in shallow waters. Salinity and temperature were obtained with a CTD/rosette system, which provided seawater for chlorophyll-a and nutrient concentrations. The influence of water mass on larval fish species was studied using Indicator Species Analysis, whereas environmental effects on the distribution of larval fish species were analyzed by Distance-based Redundancy Analysis. Larval fish species were associated with specific water masses: in the north,  Engraulis rigens in the Chilean coast Dicentrarchus labrax in estuaries in Portugal Ichthyoplankton distribution is under the influence of environmental factors that regulate life history traits and determine its geographical range It is known that larval fish assemblages result from adult spawning strategies and environmental influences The South Brazil Shelf (SBS), in the Southwest Atlantic Ocean (SWAO), is a region associated with the largest Brazilian fish stocks Ichthyoplankton has been studied in the region over 30 years The South Brazil Shelf (SBS), located at the Southwest Atlantic Ocean (SWAO) stretches from Cape São Tomé (21°S) to Chuí (33°S). The shelf waters of the SBS can be divided into three major latitudinal regions according to their oceanographic characteristics, from north to south: the Cape São Tomé-Cape Frio region (CSTF), the Southern Brazilian Bight (SBB) and the Southern Subtropical Shelf (SSS) . Along tCoastal upwelling due to wind and topographic effects, occurs in the northern portion of the study area (CSTF) The Southern Brazilian Bight (SBB) is located between Cape Frio (22°52′S) and Cape Santa Marta (28°36′S), and is characterized by the presence of SW in its northern portion, and by Subtropical Shelf Water (STSW) in its southern area. The STSW is the result from a mixture of Plata Plume Waters (PPW) and TW, and spreads throughout the neritic region of the SBB and SSS 3 and the values for each sample are displayed in Cruises were carried out between 21°27′S and 34°51′S during late austral spring (December 2010) and early austral summer (January 2011). Temperature, salinity, fluorescence and oxygen were measured vertically with a conductivity-temperature-depth (CTD) profiler Sea Bird Electronics model 911 at 107 stations distributed at 17 cross-shelf transects . AdditioPlankton samples .Sea surface temperature (SST) images were used to evaluate ichthyoplankton distribution in relation to surface thermal mesoscale features in the Brazilian shelf and slope waters, such as meanders, eddies and upwelling. We used monthly SST composition images for December 2010 and January 2011 with a 4 km spatial resolution from the Moderate Resolution Imaging Spectroradiometer (MODIS)/Aqua sensor. These images were obtained from the Ocean Color web site . Oxygen stratification was used to characterize the relationship between ichthyoplankton abundance and oxygen distribution in the water column. This was calculated using the surface oxygen value and the value for the bottom of the oxycline, and respective depths.Ichthyoplankton abundance was standardized to the number of individuals per 100 ma prioriSamples were also classified as being located within specific water masses  Distance-based Redundancy Analysis (db-RDA) was used to investigate the variability in ichthyoplankton composition constrained by centered environmental explanatory variables , using the Bray-Curtis index for similarity between samples Six water masses were identified in the study area down to the maximum zooplankton sampling depth (130 m): Subtropical Shelf Water (STSW), Shelf Water (SW), Tropical Water (TW), Plata Plume Water (PPW), Subantarctic Shelf Water (SASW) and South Atlantic Central Water (SACW) . Among tOver the shelf, a latitudinal pattern in the distribution of the water masses can be observed . Cold anAuxis sp.). The 19 remaining taxa might be members of two or more species . The most specious family in the samples was Myctophidae (21), followed by Serranidae (5), Paralepididae (4), Scorpaenidae (4), Carangidae (4), Sciaenidae (4) and Scombridae (4)  . Fish egidae (4) .−3), on the coast of Tramandaí (631 ind. 100 m−3) and on Cape Frio  and Cape São Tomé  shelves. The abundance of Engraulidae eggs was higher near Patos Lagoon  and Anguilliformes eggs showed a low abundance and were mostly present in the SBB area. The abundance of Maurolicus muelleri eggs peaked at the slope near to the Feia lagoon area (374 ind. 100 m−3) and occurred along the slope in the southerly direction  and the Patos Lagoon area (119 to 479 ind. 100 m−3), the slope near Feia Lagoon (215 ind. 100 m−3) and the shelf at Cape Frio (131 to 246 ind. 100 m−3). Low abundances were observed mostly at the slope, nevertheless, two peaks of high abundance were registered in the northern area at stations 83 and 91 (−3), there was a mixture of yolk sac larvae (29%), larvae of Scorpaenidae sp. 1 (14%) and larvae of pelagic fishes such as Myctophidae (29%), Phosichthyidae (14%) and Paralepididae (7%). Station 83, located at the slope (215 ind. 100 m−3), the same stations where M. muelleri eggs peaked (−3). The SST image showed the formation of BC meanders at the boundary between TW at the slope and SW over the continental shelf in the northern area  with a 22% frequency of occurrence, followed by the Brazilian sardine Sardinella brasiliensis (4.7±2.9 ind. 100 m−3). Some species had a low abundance but were frequently found in samples, for example, larvae of the pelagic fishes such as the lanternfishes Diaphus mollis (26%) and Ceratoscopelus townsendi (13%); the bristlemouth Cyclothone acclinidens (16%); the codlet Bregmaceros cantori (10%); and others from coastal origins such as the lizardfish Synodus foetens (10%) and the largehead hairtail Trichiurus lepturus (10%) (Engraulidae larvae were the most abundant (35%) followed by those of Myctophidae (24%), Clupeidae (9%), Bregmacerotidae (3%), Scombridae (3%) and Gobiidae (3%). The highest abundance was for the anchovy us (10%) .E. anchoita was the dominant species. Between 25°S and 30°S, anchovy shared dominance with T. lepturus, S. foetens and B. cantori in the coast and inner shelf. Trichiurus lepturus and S. foetens also occurred between Cape São Tomé and Cape Frio, but S. brasiliensis and the goby Ctenogobius boleosoma were the most abundant species on the shelf  and pelagic species (sardines), considering adult habitat  = 0.0001, Residual d.f. = 77), when all mean similarities within groups were higher than similarities between groups , the group formed by SW samples was characterized by larvae of the Brazilian sardine  habitat . In the n groups .Distance-based Redundancy analysis (db-RDA) constrained 24% of ichthyoplankton variance in relation to the explanatory variables . The MonE. anchoita were positively correlated with latitude, oxygen stratification, silicate and phosphate concentration and low temperatures from PPW. At mid-latitudes and in inshore waters, under the influence of STSW, larvae of the codlet B. cantori, the largehead hairtail T. lepturus and the lizardfish S. foetens were also positively correlated with high chl-a concentration. The abundance of Engraulidae eggs and other fish eggs was highest mostly in inshore waters, with a high chl-a concentration. In addition, a high abundance of S. brasiliensis and C. boleosoma larvae and M. muelleri eggs was influenced by the high-temperature SW. Larvae of oceanic species, such as the bristlemouth C. acclinidens and the lanternfishes D. mollis, D. brachycephalus, L. guentheri and C. townsendi were associated with TW, low nutrients and chl-a concentration, and high salinity.According to the species distribution , larvae Sardinella brasiliensis characterized Shelf Water (SW); in the south Engraulis anchoita was found in Plata Plume Water (PPW); and, in Subtropical Shelf Water (STSW) species from both coastal and shelf origins were found. At the slope, Tropical Water (TW) was characterized only by the bristlemouth Cyclothone acclinidens, although a high abundance and occurrence of lanternfishes were also found. Ichthyoplankton composition was influenced by the cross-shelf gradient and by the latitude in continental shelf waters.Our findings provide information on ichthyoplankton distribution and its relationship with the oceanographic environment within a wide latitudinal range in the South Brazil Shelf. Four surface water masses were present in the whole study area, and over the shelf, three were positioned in a latitudinal gradient. Larval fish species were associated with specific water masses: in the north Parona singnata, Gerreidae, Cynoscion sp., Micropogonias furnieri, Mugil sp., Symphurus spp. and Engraulidae eggs, are expected near or in front of large estuaries because of the contribution of estuarine spawners and estuarine-dependent fishes that use these environments for reproduction and/or as nursery areas −3) and larvae (>100 ind. 100 m−3), once the enhancement of the primary and secondary production ensure the availability of food for the future larvae A maximum ichthyoplankton abundance and the presence of coastal- and estuarine-related species, such as −3 in late austral spring was found. Fish egg abundance was similar, and sometimes higher, than that described for the Northern Brazilian coast −3) than in coastal and shelf waters, and fish eggs were absent in some offshore stations. This is probably an effect of the nutrient-poor TW that is driven by the Brazil Current (BC) throughout the entire slope Ichthyoplankton abundance distribution in shelf waters was comparable to that of previous studies in the region between Cape Frio and Chui A high larval abundance occurred between Cape São Tomé and Cape Frio, at the outer shelf and slope. This pattern might be related to the eddy-induced upwelling Auxis sp., Thunnus spp. and Katsuwonus pelamis and release their eggs in areas with a higher primary production than in the open ocean, where waters are more suitable for the early larvae Fish larvae species composition reflected the spawning area of adult fish populations and was similar to that previously registered for the area C. boleosoma and the blenny Parablennius sp., with larvae from pelagic eggs such as from the sardine S. brasiliensis. In a region in southwestern Australia, inshore stations were also dominated by larvae from demersal eggs such as those from Tripterygiidae, Gobiidae and Blenniidae, and larvae of neritic species such as Clupeidae Water mass composition is an important controlling factor that conditions larval fish distribution S. brasiliensis is located on the Southern Brazilian Bight (SBB) between Cape Frio (∼23°S) and Cape Santa Marta (∼28°S) S. brasiliensis in the continental shelf area around Cape Frio.The spawning area of the Brazilian sardine S. foetens and the largehead hairtail T. lepturus, which shared dominance with larvae of oceanic origin such as the codlet B. cantori. Due to the lack of intrusion of SACW in the SBB area, the STSW group did not contain any S. brasiliensis larvae in its composition. This same larval association was registered in the Subtropical Shelf Front (STSF), the transition between the STSW and Subantartic Shelf Water (SASW) located near the 50 m isobaths, around 33°S T. lepturus and other codlet B. atlanticus, respectively The STSW group comprised mainly stations from the SBB area, and was associated with larvae of fish that spawn in inshore waters, such as the lizardfish E. anchoita and its pelagic eggs were dominant. Anchovy has a wide distribution range from Cape Frio (23°S) to the Gulf of San Jorge (47°S) E. anchoita is associated with a lower intensity upwelling, cold water from the south and continental runoff from La Plata and Patos Lagoon E. anchoita larvae an important component of the pelagic ecosystem in the Southwest Atlantic Ocean (SWAO).In the area under the influence of PPW, larvae of the anchovy C. acclinidens, Myctophidae species such as D. mollis, D. brachycephalus and C. townsendi were also overwhelmingly abundant in the slope waters. Larvae from Myctophidae and Gonostomatidae are good indicators of the presence of oceanic waters such as TW Regarding the TW group, characterized by a high abundance and occurrence of the bristlemouth The main role of water mass influence can be attributed to the ordination of larval species distribution in space. Cross-shelf and latitudinal environmental effects were the two main components that accounted for the composition of fish larval assemblages, and reflect the influence of water masses e.g., . The latS. brasiliensis population and its spawning is influenced by SACW intrusion Our results revealed a high unexplained source of variation, and we suggest that other sources of variability should be investigated. Large-scale ichthyoplankton composition and distribution are also influenced by food availability and geostrophic circulation In conclusion, our study supports the hypothesis that in the South Brazil Shelf, the large-scale distribution of the ichthyoplankton is mainly controlled by water mass composition. In addition, larval fish assemblages are influenced by the cross-shelf gradient and by the latitude of continental shelf waters. Species are more abundant and frequent at different areas under the influence of specific thermohaline characteristics in the water column. Further studies should be performed to assess the role of food availability in the control of fish larvae abundance.Table S1Plankton data for the stations visited at the South Brazil Shelf. Depth of plankton sample (m) and water volume (m3) filtered for each of the 89 stations between Cape São Tomé (21°S) and Chuí (33°S) from December 2010 to January 2011.(DOC)Click here for additional data file.Table S2Taxonomic list of the ichthyoplankton from the South Brazil Shelf. Total catch of fish larvae and eggs, abundance (mean ± SE) and frequency (%) for the 89 stations visited between Cape São Tomé (21°S) and Chuí (33°S) from December 2010 to January 2011.(DOC)Click here for additional data file.


PRODH gene may be related to the disorder. Therefore, our study investigates the effects of 12 polymorphisms of PRODH on schizophrenia and its phenotypes. To further evaluate the roles of the associated variants in the disorder, we have conducted magnetic resonance imaging (MRI) scans to assess cortical volumes and thicknesses. A total of 192 patients were evaluated using the Structured Clinical Interview for DSM-IV (SCID), Positive and Negative Syndrome Scale (PANSS), Calgary Depression Scale, Global Assessment of Functioning (GAF) and Clinical Global Impression (CGI) instruments. The study included 179 controls paired by age and gender. The samples were genotyped using the real-time polymerase chain reaction (PCR), restriction fragment length polymorphism (RFLP)-PCR and Sanger sequencing methods. A sample of 138 patients and 34 healthy controls underwent MRI scans. One polymorphism was associated with schizophrenia (rs2904552), with the G-allele more frequent in patients than in controls. This polymorphism is likely functional, as predicted by PolyPhen and SIFT, but it was not associated with brain morphology in our study. In summary, we report a functional PRODH variant associated with schizophrenia that may have a neurochemical impact, altering brain function, but is not responsible for the cortical reductions found in the disorder.Schizophrenia is a neurodevelopmental disorder with high heritability. Several lines of evidence indicate that the  Schizophrenia is currently considered to be a neurodevelopmental disease that arises from a complex phase-specific interaction of genetic and environmental factors One of the strongest known genetic risk factors for schizophrenia is the 22q11 deletion PRODH) gene, which encodes the proline dehydrogenase enzyme that catalyzes the first step in proline catabolism PRODH gene, consisting of 14 coding exons and located in a low copy repeat sequence, is widely expressed in the brain and in other tissues One of the genes commonly deleted in 22q11DS is the proline dehydrogenase  PRODH single nucleotide polymorphisms (SNPs - rs2008720 and rs372055) with respect to brain anatomy in a sample of 51 patients with schizophrenia. While no association was found for rs2008720, significant differences in grey matter (GM) and total brain volume were revealed for rs372055, with C carriers showing increased total GM and total brain volume in comparison with patients carrying the TT genotype Only two previous studies have investigated the role of PRODH SNPs with brain morphology have not been investigated, and neither previous study investigated the different roles of these polymorphisms in both normal and pathological conditions. In the present study, we aimed to associate 12 PRODH polymorphisms, and especially functional variants, with schizophrenia and other clinical variables such as treatment resistance, age of onset, and positive and negative symptoms. To further understand the impacts of the studied polymorphisms, we conducted MRI scans in a subsample of patients.However, the associations of other important functional This study was approved by the Research Ethics Committee of UNIFESP [CEP No. 1737/06]. Written informed consent was obtained from all recruited participants or their caregivers, and the clinical and laboratory investigations were strictly conducted according to the principles expressed in the Declaration of Helsinki. All invited subjects agreed to enroll in this study. If the patients were within a stable period according to their clinical features and could understand the consent forms, they double-signed the documents themselves. The next of kin, caretakers or guardians consented on the behalf of those participants whose capacity to consent was compromised.www.ipap.org], as a failure to respond to 4- to 6-week trials of monotherapy with two different antipsychotics at adequate doses .A total of 192 chronically medicated outpatients were recruited from the Schizophrenia Program (PROESQ) at the Universidade Federal de São Paulo (UNIFESP). The diagnosis of schizophrenia was established according to the criteria of the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV), using the Structured Clinical Interview of the DSM-IV (SCID). Other clinical variables were assessed using the Positive and Negative Syndrome Scale (PANSS); Calgary Depression Scale A 1.5 Mb microdeletion in 22q11.2 was detected in one patient with schizophrenia using multiplex ligation-dependent probe amplification A total of 179 healthy subjects with no family history of severe psychiatric illness were recruited by UNIFESP. These subjects were also investigated using a modified version of SCID screening to exclude any with a previous illness episode with psychotic features or a family history of similar diseases.Considering that the Brazilian population is admixed, the population structure was verified for a sample of our study population .Whole blood was collected into EDTA tubes and genomic DNA isolation was performed using the Gentra Puregene Kit  according to the manufacturer’s protocol.The population genetic ancestry structure was assessed by a panel of 48 ancestry-informative insertion-deletion polymorphisms, previously described and validated in the Brazilian population PRODH SNPs were interrogated (BsaBI enzyme and the following PCR primers: 5′-TGGTGGGGAGGAGGAGGTCA-3′ (forward) and 5′-CAGCCAGGACTGGGAGACGT-3′ (reverse). The other eight SNPs in exon 12  were investigated using Sanger sequencing. The PCR primers were as follow: 5’-TCCCCACTGCCATTGCTCCT-3′ (forward) and 5′-CCTGCCCTGAGAAGACAGAG-3′ (reverse). The PCR conditions are available upon request.A total of 12 rrogated  by real-The characteristics of the study population are described in Statistical analyses were performed using the Statistical Package for the Social Sciences (SPSS version 15.0), and the level of significance was set at 0.05.http://sift.jcvi.org/) and PolyPhen (http://genetics.bwh.harvard.edu/pph/) and analyzed the amino acid conservation using the UCSC genome browser (http://genome.ucsc.edu). Both SIFT and PolyPhen predict whether an amino acid substitution affects protein function. SIFT prediction is based on the degree of conservation of amino acid residues in sequence alignments For the SNPs associated with schizophrenia, we tested the predictive values using SIFT  and 34 healthy controls  using a 1.5T scanner  with an eight-channel head coil. A series of exploratory sagittal images (nine to eleven slices of 5 mm with 1 mm spacing) was performed to evaluate the image quality and the positioning of the head of each subject. T1 images were acquired sequentially using a pulse sequence (SPGR) with the following parameters: TR  = 2000 ms, TE  = 3.42 ms, matrix size  = 256 x 256, FOV  = 245 mm, flip angle  = 15 degrees, NEX  = 1, 1.0 mm slice thickness with no gaps, yielding 192 slices.http://surfer.nmr.mgh.harvard.edu) based on established processing steps . The cortical volumes and thickness information of the brain regions were estimated using the automated parcellation of the Desikan-Killiany Atlas The MRI scans were analyzed using the FreeSurfer software package  was used to assess the association of each cortical and subcortical volume and thickness with schizophrenia. Then, all measures were inserted as dependent variables, and each SNP, schizophrenia diagnosis and the interaction between them were included as fixed factors. The total intracranial volume (TIV), gender and age were included as covariates. Statistical significance was set 2 = 4.38, df  = 1, p  = 0.036; cases: χ2 = 9.70, df  = 1, p  = 0.002) and rs450046 .The genotype and allele frequencies for each variant are represented in Regarding allele comparisons, one polymorphism was associated with schizophrenia  after Bonferroni correction . Comparing the genotypes of rs2904552, we found that the GG-genotype carriers were at greater risk for a schizophrenia diagnosis in comparison to the AG-genotype carriers . Comparing the GG homozygotes to the A-allele carriers (grouping GA and AA genotype carriers), a stronger association was found . In all these analyses, the power was >80% as determined using G*Power 3.1.6.For the haplotype analyses, only polymorphisms in Hardy-Weinberg equilibrium and with a minor allele frequency >5% were included . The results of the linkage disequilibrium test are described in PRODH polymorphism, but no significant associations were detected.We analyzed the association of age at onset, TR, GAF, CGI, Calgary, Negative, Positive, Disorganized, Excited and Anxiety/Depression clusters with each in silico analysis. This SNP seems to have a functional role, as it was classified as “probably damaging,” with a score of 0.996, using PolyPhen and as “damaging,” with a score of 0.04, using SIFT. Data were also available for five orthologous species , and the residues showed evolutionary conservation in all these organisms. Furthermore, this variation was reported in a previous study to decrease the enzyme activity To predict the likely pathogenicity of rs2904552, we conducted an For the cortical and subcortical volumes, 21 regions were associated with schizophrenia after Bonferroni correction and are described in PRODH genotypes with all brain volumes and cortical thicknesses, only one association remained significant after Bonferroni correction: the interaction between rs2238731 (V427M) and schizophrenia was associated with right pars opercularis volume . Controls carrying the GA genotype had an increased volume (6261.50±144.95 mm3) in this region compared to carriers of the GG genotype (4193.37±628.09 mm3). However, in schizophrenia patients, GA genotype carriers had a smaller volume (3796.68±527.12 mm3) than did GG carriers (3993.41±733.26 mm3).In the comparison of all PRODH variant (rs2904552 or R431H) and its haplotypes were associated with schizophrenia, with the GG genotype present at a higher frequency among patients than among healthy controls. To further understand the role of this SNP, we investigated its role in brain volumes and cortical thicknesses.In this study, one Among the 12 investigated SNPs, 10 had been previously associated with schizophrenia , hyperprolinemia  or both ; two SNPs had not been previously investigated .PRODH in schizophrenia: 1) it is located at 22q11.2, the deletion of which is one of the strongest genetic risk factors for schizophrenia; 2) PRODH deletion missense mutations have been described in patients with schizophrenia ; 4) it is widely expressed in the brain; 5) mice with PRODH deficiency show elevated plasma and brain proline levels, locally decreased glutamate and γ-aminobutyric acid (GABA) levels and a deficit in sensorimotor gating Different lines of evidence suggest an important role for PRODH variants and hyperprolinemia in the pathophysiology of schizophrenia Indeed, a recent metabolomics study reported abnormalities related to proline metabolism in schizophrenia Along with its role in proline catabolism, PRODH is also known as a key enzyme in controlling homeostasis, providing energy and shuttling redox potential between cellular compartments and the production of reactive oxygen species PRODH rs2904552 is a nonsynonymous mutation that causes the substitution of an arginine for a histidine in codon 431. This change seems to promote a functional effect, according to our in silico findings, and also results in a moderate decrease (30–70%) of PRODH enzyme activity N  = 2) and patient (N  = 19) groups.Our results support earlier findings of reduced total, left and right hemisphere gray matter volumes, corpora callosa, frontal, parietal and temporal lobes, and hippocampi, all of which are brain regions involved in the pathophysiology of schizophrenia Some limitations of this study should be noted. First, we should have analyzed plasma proline levels. Considering that rs2904552 variant may be affecting PRODH enzyme, its substrate might be dysregulated. For this measurement, gender, acute alcohol use and medication, such as valproic acid, may be taken into consideration PRODH polymorphisms on brain morphology in both patients and healthy controls, testing for the interaction between this SNP and a diagnosis of schizophrenia. Only one SNP (rs2238731) seemed to be significantly associated with right pas opercularis volume, but this finding should be confirmed in larger samples. Therefore, considering the functional effect of rs2904552 and the importance of proline levels in schizophrenia, this SNP may act by altering neurochemistry and is not responsible for the deficits in brain structure found in schizophrenia.This study represents the first report of the effects of PRODH rs2904552 polymorphism may be a genetic risk factor for developing schizophrenia, and further studies should address how disturbance of the proline pathway could affect brain function.In summary, this study indicates that the Figure S1Linkage disequilibrium plot across the proline dehydrogenase (PRODH) gene. Numbers within the diamonds are D' values for the respective SNP pairs.(TIF)Click here for additional data file.Table S1Genotype and allele frequencies of PRODH variants.(DOCX)Click here for additional data file.Table S2Haplotype frequencies of PRODH variants in patient and control group.(DOCX)Click here for additional data file.Table S3Association between cortical and subcortical volumes and schizophrenia.(DOCX)Click here for additional data file.Table S4Association between cortical thickness and schizophrenia.(DOCX)Click here for additional data file.


Exploring the presence and role of human papillomavirus (HPV) in head and neck cancer (HNC) is a necessary step to evaluate the potential impact of HPV prophylactic vaccines.To assess the prevalence and oncogenic role of HPV in HNC in Senegal.25. Tubulin analysis was performed to assess DNA quality. HPV DNA-positive cases were tested for p16INK4a expression.This is a multicenter cross-sectional study. Paraffin-embedded blocks of cases diagnosed with invasive HNC between 2002 and 2010 were collected from 4 pathology laboratories in Senegal. Presence of HPV DNA was determined by PCR and DEIA, and genotyping performed with LiPAINK4a overexpression.A total of 117 cases were included in the analysis: 71% were men, mean age was 52 years old (SD ±18.3), and 96% of cases were squamous cell carcinoma. Analysis was performed on 41 oral cavity tumors, 64 laryngeal tumors, 5 oropharyngeal tumors and 7 pharyngeal tumors. Only four cases  harbored HPV DNA. HPV types detected were HPV16, HPV35 and HPV45. However, among HPV-positive cases, none showed p16Our findings indicate that HPV DNA prevalence in HNC in Senegal is very low, suggesting that HPV is not a strong risk factor for these cancers. Additional larger studies are needed to confirm these findings and explore other potential risk factors specific to the region. Head and neck cancer (HNC) is the eighth most common cancer worldwide with approximately 650,000 new cases and 350,000 deaths reported each year . In addiIn sub-Saharan Africa, knowledge on the role of HPV in HNC is very limited. Actually, 84% of the existing information on HPV and HNC is derived from studies in Europe and North America . The aim25) . The LiPA25 detects 25 high-risk and low-risk HPV types . Additionally, amplification of the human tubulin gene was performed to determine DNA quality of DEIA negative samples. In HPV DNA-positive samples, p16INK4a overexpression was evaluated using CINtec histology kit  in order to verify the potential causal role of HPV in positive cases. Overall HPV detection percentages and 95% confidence intervals (95% CI) were estimated. Data analyses were performed with STATA 11.0. Ethical approval was obtained from the Ministry of Health of Senegal.This is a cross-sectional study designed and coordinated by the Catalan Institute of Oncology in Barcelona and is part of a large international study on HPV in HNC. Four major centers in Dakar, Senegal, have provided consecutive cases diagnosed with HNC from 2002 to 2010: Centre Hospitalier Universitaire A. Le Dantec, Hôpital Principal de Dakar, Hôpital Général de Grand Yoff and the pathology laboratory at Cheikh Anta Diop University. For each specimen, after pathological confirmation of invasive diagnosis, a paraffin tissue section was treated with 250 μl of freshly prepared Proteinase K solution to extract DNA. SPF-10 polymerase chain reaction (PCR) was performed using 10 μl of a 1:10 dilution of the crude DNA extract in a final reaction volume of 50 μl. The amplified PCR products were tested using a probe hybridization step with a cocktail of conservative probes that can recognize around 54 mucosal HPV genotypes using a microtiter plate format for the detection of HPV DNA through a DNA enzyme immunoassay (DEIA) . Optical densities (OD450) were read on a microtiter plate reader and categorized as HPV DNA negative, positive, or borderline. After PCR, 10 μl of the DEIA HPV DNA positive amplimers were used to perform the reverse hybridization line probe assay  and three laryngeal SCC (2 HPV45 and 1 HPV16). However, none of these cases showed p16INK4a overexpression: 3 cases showed no p16INK4a staining and one HPV45 laryngeal cancer case showed focal staining of less than 25% of cells with low-moderate intensity.The overall HPV prevalence was 3.4%; 95% CI = 0.9%-8.5%. Table INK4a expression suggested that HPV was not involved in the oncogenic process of the tumors. Expression of p16INK4a is strongly seen in HPV-associated tumors but nearly absent in HPV-negative carcinomas [To our knowledge, this is the first study to explore the presence of viral DNA and cellular protein expression in head and neck tumors in West Africa. HPV DNA was detected in 3.4% (95% CI = 0.9%-8.5%) of studied cases but further analyses testing for p16INK4a. Both studies agree on significant ethnic and biological disparities in HPV prevalence, after adjusting for clinical and sociodemographic characteristics, and support the validity of our study. Further studies are needed to elucidate the mechanism of HPV clearance in African-American and African populations, which share common ancestral origins.Similar low HPV prevalences have been reported in the African-American community in the USA. In a prospective study conducted by Settle et al. , overallThese findings are very different from what has been reported previously in other regions of the world. In a systematic review by Kreimer et al.  compilinLow HPV prevalence in invasive HNC in Senegal suggest that other established risk factors such as alcohol and tobacco consumption may play a more significant etiological role than HPV infection in HNC. However, the prevalence of tobacco use in Senegal is relatively low with estimates of 19.9% in men and 1.3% in women . Alcohol2 and CO [For example, indoor air pollution causes 6,300 deaths each year in Senegal due to daily exposure to smoke from open burning of wood and charcoal in homes . More th2 and CO . Accordi2 and CO .Another potential risk factor could be the consumption of “ataya”, a strong and bitter hot tea comparable to “yerba mate” in South America, which has been associated with an increased risk of developing cancer of the oral cavity, larynx and esophagus . On averPoor oral hygiene is another potential risk factor that has been documented  and coulOccupational exposure such as jobs in the construction, metal, textile, ceramic, logging, and food industries has been associated with the development of laryngeal cancer . In SeneOur study has some methodological limitations. At sample collection stage, some cases were identified eligible for the study but corresponding blocks were not found as they were sent to laboratories abroad for histopathological evaluation. Additionally, several archival records were lost in one of the main laboratories due to a fire. However, these issues affected all cases regardless of patient characteristics or diagnosis. Another major limitation is the lack of individual data on risk factors: no information on patients besides their gender, age and pathological information were available in the registries. Finally, the small sample size (n = 5) for oropharyngeal tumors may have affected our positivity rate. It is recognized that the highest associations of HPV in HNC have been shown in the oropharynx, particularly in the tonsils, with a positivity of 57% to 82% compared to 0.8% to 9% in other anatomic sites .INK4a expression. Additionally, we evaluated the quality of our specimens by means of cellular tubulin detection and subsequently excluded the HPV DNA negative and tubulin negative cases from the statistical analyses. Lastly, all of the study cases were selected from the main anatomy and pathology laboratories in Senegal. Thus, our cohort is representative of the target population as the vast majority of patients are diagnosed in participant centers.Our study has several strengths comprising the combined use of highly sensitive assays for HPV DNA detection and a marker of HPV related transforming process such as p16In Senegal, HPV prevalence in HNC is very infrequent and HPV infection is not associated with carcinogenesis. Therefore, HPV prophylactic vaccines would not have any impact on HNC incidence. Our findings need to be further validated with supplementary studies that include larger case series and the assessment of region-specific risk factors. Additional data will allow to close the gap of knowledge between Western countries and the sub-Saharan African region and to assist health authorities in implementing public health strategies.HT served as a consultant and on advisory boards and received speaker fees and travel assistance from Merck-Frosst Canada and Glaxo Smith Kline Pharmaceuticals, Belgium. LA occasionally received travel grants to attend conferences granted by Merck and Sanofi Pasteur MSD. XC received travel grants for scientific meetings and honorarium for consultancy occasionally granted by GlaxoSmithKline, Merck, Sanofi Pasteur MSD. XB received travel grants to conferences/symposia/meetings and honorarium occasionally granted by GlaxoSmithKline, Merck, Sanofi Pasteur MSD, Roche or Qiagen. SS received travel grants to conferences/symposia/meetings occasionally granted by GlaxoSmithKline, Sanofi Pasteur MSD or Qiagen. Other co-authors have no potential conflict of interest to declare.CN collected data, performed data analysis and drafted the manuscript. SJ, XC, FXB and LA initiated and designed the study. LA, HT and SJ guided data analysis, interpreted data and edited the paper. XC and FXB made substantial contributions to the manuscript and contributed to data interpretation. NN, YD and MJD interpreted data and edited the paper. JEK performed the laboratory tests and MA the histopathological evaluation. ST managed and cleaned the database. All authors read and approved the final manuscript.


B) should scale isometrically with aboveground biomass (MA) and the scaling exponent should not vary with environmental factors. We tested this hypothesis using a large forest biomass database collected in China. Allometric scaling functions relating MB and MA were developed for the entire database and for different groups based on tree age, diameter at breast height, height, latitude, longitude or elevation. To investigate whether the scaling exponent is independent of these biotic and abiotic factors, we analyzed the relationship between the scaling exponent and these factors. Overall MB was significantly related to MA with a scaling exponent of 0.964. The scaling exponent of the allometric function did not vary with tree age, density, latitude, or longitude, but varied with diameter at breast height, height, and elevation. The mean of the scaling exponent over all groups was 0.986. Among 57 scaling relationships developed, 26 of the scaling exponents were not significantly different from 1. Our results generally support the isometric hypothesis. MB scaled near isometrically with MA and the scaling exponent did not vary with tree age, density, latitude, or longitude, but increased with tree size and elevation. While fitting a single allometric scaling relationship may be adequate, the estimation of MB from MA could be improved with size-specific scaling relationships.Based on the isometric hypothesis, belowground plant biomass (M B) with respect to aboveground biomass (MA) influences both the structure and function of individual plants and of vegetation ecosystems Forest biomass plays an important role in estimating forest productivity, carbon sequestration, and in sustainable forest management B to MA  or by using the allometric scaling function relating MB and MA: MB = a MAb, where a is a normalizing scaling constant and b represents an allometric scaling exponent A scales nearly isometrically with respect to MB  L) scales as the ¾ power of stem biomass (MS), and MS scales as the ¾ power of root biomass (MR) A = ML+MS = a1MS3/4+ a2MR3/4 = (a2/a1)3/4MR+a2MR3/4. If (a2/a1)3/4>>a2, MA scales isometrically with respect to MB (MB = MR). The isometric hypothesis has been tested empirically and validated at individual and forest community levels A scales as the 1.06 (95% CI 1.05∼1.08) power of MB across 257 plant species. Similarly, Yang & Luo Biomass partitioning can be described as a ratio of MA and MB at large spatial scales has not been systematically investigated.The isometric hypothesis has also been disputed in several studies. For example, Li et al. In this study, we analyzed a large forest biomass database to develop large-scale biomass partitioning patterns and identify influencing factors. The database includes branch, stem, leaf and root biomass, and other derivative information such as altitude, longitude, and elevation, covering a wide range of forests in China. Understanding the pattern and variation of biomass partitioning among age groups, tree sizes, locations, and taxonomic groups could have important implication on terrestrial ecosystem modeling. So far, most ecosystem models only consider biomass partitioning as a constant and the effects of these influencing factors on biomass partitioning have not been incorporated into terrestrial ecosystem models B and MA using a power function; (3) investigate whether the relationship varies with biotic factors  and abiotic factors ; 4) evaluate the influences of regression methods on the scaling exponent and constant estimation; and 5) explore whether phylogeny affects root:shoot ratio and scaling relationship.The overall aim of this study was to test whether the root:shoot ratio and scaling exponent of the allometric function vary with tree age, diameter at breast height, height, density, latitude, longitude, elevation and family. We hypothesized that: 1) As tree age and size increase, the root:shoot ratio and scaling exponent should decrease, as more biomass accumulates aboveground than belowground as trees age. Thus, ontogeny and biotic factors (i.e. tree age and size) may influence the allometric scaling relationship; 2) Conditions are drier and colder in western than eastern China, therefore the root:shoot ratio and coefficient of the scaling relationship between root and shoot biomass will decrease from west to east. Since latitude, longitude and elevation may influence precipitation and temperature, we expect that biomass partitioning will vary with these factors. To test these hypotheses, we specifically tended to (1) estimate the mean and variation of the root:shoot ratio in forest ecosystems in China; (2) develop an overall relationship between MThis study was based on a forest biomass database that includes data collected from journal publications and reports. Thus, no specific permissions were required for this study. The field studies did not involve endangered or protected species. Data will be made available upon request.A was calculated as total biomass of leaf, stem and branch biomass. MB was root biomass. This large database provides a unique opportunity to evaluate the relationship of productivity and biomass in Chinese forests The database and the management system were constructed by Dr. Hai Ren and his colleagues at the South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, China. Data were collected from journal publications and the China Forestry Inventory Reports up to 2007 . In brieet al.Biomass measurements were conducted mostly using the “standard tree” method B = aMAb) was applied to develop the relationship between MB and MA. Several methods have been used for model parameter estimation and the pros and cons of the methods have been discussed An allometric scaling model  for each group A, MB, and root-shoot ratio with group variables were developed for each subdivision of the data  were significant at α  = 0.05 level. For example, MB scaled isometrically with MA within all 8 age groups (2) for the power functions was 0.903 with a range from 0.731 to 0.968. The scaling components varied slightly among age groups. The scaling exponent was 0.964±0.0040 . Our results also showed that the root:shoot ratio initially declined with DBH and height, which supports the findings by Mokany et al.et al.A in woody tissue as stands develop Our results supported our hypotheses on DBH, height and latitude influences, but not on age, longitude, and elevation. Whether the root:shoot ratio varies with age and other variables has been controversial. For example, the root:shoot ratio has been reported to decrease with stand age across forests and woodlands B scaled isometrically or near isometrically with MA in Chinese forest ecosystems, consistent with the prediction of the isometric hypothesis. The mean of scaling exponents of all groups was 0.986 with a small range of 0.964 to 1.015, close to 1 predicted by the isometric hypothesis. The 95% confidence intervals for scaling exponents included 1 in 26 out of 57 relationships for the 95% confidence intervals and in 37 out of 57 instances for the 99% confidence intervals, and did not vary with tree age, density, latitude, or longitude. Our results were consistent with many previous studies which considered different forest ecosystems Another finding of this study was that MIn the absence of physiological measurements, explanations of these results may seem to be speculative. But our hypotheses may promote a mechanistic understanding of biomass partitioning and influence the design of future experimental studies. One hypothesis to explain the invariant scaling relationship is that of a developmental constraint B and MA may also be linked to nutrient and water uptake The near isometric relationship between MStatistically significant variation may also exist in the scaling exponent, depending on how the data are sorted into different categories et al.Whereas the scaling exponent of the scaling relationship is considered as the results of universal physical constraints and natural selection, the scaling constant of the relationship has been linked to various taxon-specific or ecological factors B scaled isometrically with MA in 14 of the 15 most abundant families, most phylogeny groups , leaf forms, and forest origins . These results were comparable to Luo et al.Our analyses of the influence of phylogeny and forest type on the scaling relationship confirmed that Met al.B could be underestimated and result in a lower root:shoot ratio. Slightly different sampling methods in individual studies collected in the database might also add biases to biomass estimation. Despite this, it is evident that MB is closely related to MA in Chinese forests. More accurate estimation in MB could improve our understanding of root biomass and biomass partitioning. Nevertheless, the relationship developed in this study could be very helpful for the estimations of root biomass and ecosystem carbon dynamics in forests. The root:shoot ratios and allometric scaling models for different families, phylogeny groups and forest types could be used in ecological modeling. Considering the tree size and other factors could provide more accurate estimations in forest biomass and reduce the uncertainty at regional scales.As more ecological data accumulate, data synthesis becomes an important tool to reveal general patterns and ecological rules B and MA and tested systematically how biotic and abiotic factors would influence their relationship. We demonstrated that the scaling component did not vary with tree age, density, latitude or longitude, but varied with tree size and elevation. The overall small variations of scaling exponent within each group and among different groups indicated that MB scaled isometrically or near isometrically with MA. While fitting a single allometric scaling relationship may be adequate, the estimation of MB from MA could be improved with size-specific scaling relationships.Using a large forest biomass database, we developed the allometric scaling relationships between M