Lerista elegans and Ctenophorus adelaidensis) and specious in banksia sites. Several species  preferred long unburnt melaleuca sites  compared to recently burnt sites (<12 YSLF). Several of the small elapids  were only detected in older-aged banksia sites (>16 YSLF). The terrestrial dragon C. adelaidensis and the skink Morethia obscura displayed a strong response to fire in banksia woodlands only. Highest abundances of the dragon were detected in the recently burnt (<7 YSLF) and long unburnt (>35 YSLF) banksia woodlands, while the skink was more abundant in older sites. Habitats from a range of fire ages are required to support the reptiles we detected, especially the longer unburnt (>16 YSLF) melaleuca habitat. Current burning prescriptions are reducing the availability of these older habitats.Fire is an important management tool for both hazard reduction burning and maintenance of biodiversity. The impact of time since last fire on fauna is an important factor to understand as land managers often aim for prescribed burning regimes with specific fire-free intervals. However, our current understanding of the impact of time since last fire on fauna is largely unknown and likely dependent on vegetation type. We examined the responses of reptiles to fire age in banksia woodlands, and the interspersed melaleuca damplands among them, north of Perth, Western Australia, where the current prescribed burning regime is targeting a fire-free period of 8–12 years. The response of reptiles to fire was dependent on vegetation type. Reptiles were generally more abundant (e.g.  Fire is a management tool for landscape custodians of conservation reserves and remnant vegetation All components of fire regimes  influence the impacts of fire on fauna. However, an important consideration arising from fire and fauna research is that the fire-free period may be instrumental in structuring faunal communities Our study was undertaken on a Mediterranean-type ecosystem located on the Swan Coastal Plain in the south-west Western Australia (SWWA). The SWWA region is internationally recognised because of its high levels of biodiversity and endemicity, and the high degree of threatening processes, such as habitat loss associated with urban and rural development In our study, we examine the responses of reptiles, using reptile species richness, abundance and community structure as variables, to time since last fire in two habitat types, banksia woodlands and melaleuca damplands on the Swan Coastal Plain. The key questions we addressed were: 1) Do reptile communities vary between vegetation types and fire-age categories based on prescribed burning management targets?; 2) Are there detectable seral responses of reptiles to time since last fire and microhabitat variables, such as litter cover; and 3) What are the implications of these relationships for current fire management objectives. Understanding the responses of reptiles to time since last fire in large remnant vegetation patches may provide important information for fire management that is currently in practice in this region.Data collected adhered to the legal requirements of Australia (WA Department of Environment and Conservation Scientific Purposes Permit: SC 000826) and to the ethical guidelines for treatment of animals by the WA Department of Environment and Conservation (AEC 37/2007).Banksia overstorey with sporadic stands of Eucalyptus and Allocasuarina, and an understorey consisting mainly of low shrubs from the Myrtaceae, Fabaceae and Proteaceae families. There are many seasonal damplands, swamps and permanent wetlands, fringed by Banksia littoralis and Melaleuca trees with a variable understorey of species from the Cyperaceae, Juncaceae and Myrtaceae The study area is situated on the Swan Coastal Plain in Western Australia and extends from the Swan River in the south, to the Moore River and Gingin Brook in the north, and from the Ellen Brook in the east to the Indian Ocean in the west . The areThe region experiences a dry Mediterranean-type climate The sampling regime was designed to assess 30 sites in the major areas of continuous remnant bush land in the northern and eastern banksia woodland . Sites wEach site contained one pitfall trap array, with 10 pitfall traps (20 L buckets) located in a Y shape, with three pits placed along each arm radiating out from a central pit and placed at approximately seven m intervals along each arm (∼22 m). The pitfall traps were connected with 30 cm high aluminium fly wire drift fence that extended out one m beyond the last pitfall of each arm.Sites were opened for 12–20 nights in spring 2007, autumn 2008 and spring 2008, with trapping intensity varying among season and site. Logistical constraints limited the number of sites that were opened simultaneously and we ensured that sites opened at the same time were from both vegetation types and a range of time since last fire habitat to reduce sample bias within a single vegetation-fire age category. While open, all traps were checked once per day in the early morning. Captured animals were identified, measured and released onsite. Individuals were marked under the throat with a non-toxic permanent marker pen. Taxonomic nomenclature followed the Western Australian Museum. Sample-based rarefaction (species accumulation curves) were examined for the initial trapping period (spring 2007) using Estimate S Floristic surveys on plant species richness at each site were conducted using a 10 m×10 m quadrat established within 10 m of pit-fall trap arrays. All vascular plant species within each quadrat were recorded in spring 2008. Specimens were collected and dried in plant presses prior to identification by a botanist (D. Mickle), with nomenclature checked using the Western Australian Herbarium's plant collection database MAX. Plant species abundances were not recorded. Estimates of plant species richness at each site were determined using all plant species that were able to be differentiated as different taxa within each site.2 quadrat was estimated as a percentage of vegetation (live and dead), bareground and litter (including leaf and woody debris). Litter depth (cm) was measured using a ruler that was pressed through the litter (where relevant) until it touched a firm soil surface. To provide an index of vertical vegetation density in the understorey (within a 2 m height range), vegetation contact (both live and dead) was recorded  using a graduated pole, placed in the centre of each quadrat (referred to as touch pole counts). At the same point, canopy cover was measured using a densiometer, which calculated an approximate percentage canopy cover.Microhabitat variables were measured using one m×one m quadrats, placed on either side of each pit-fall trap at each site (20 quadrats per site). Attributes assessed included vegetation structure and ground substrate composition. Ground cover within the one mPit-fall trapping intensity varied among sites, and we pooled the capture data from the trapping periods and performed analyses based on the relative abundance for each species and for the number of species captured from a standardised 10 trap nights. Diversity (D) and evenness (E) of the reptile assemblage at each site was calculated from these measures using Simpson's Diversity Index (1-D) The years since last fire varied considerably among sites (3–36 YSLF). To examine differences based on the current prescribed burning targets, we grouped sites into two major categories based on fire age: young, those recently burnt (<12 YSLF); and old, those long unburnt (>16 YSLF). Although the categorical groupings of fire age are broad, they represent sites that are within the prescribed burning fire-free period (<12 YSLF) and sites that may be targeted for future prescribed burning, and hence reflect the different fire ages perceived by the fire management objectives. This orthogonal design included 9 replicates for the old age banksia sites, 7 replicates for the young age banksia sites, 8 replicates for the young age melaleuca sites and 6 replicates in the melaleuca sites with an old fire age.A two-factor ANOVA  was used to examine differences between vegetation type, fire age and any interactions for habitat variables, plant species number, reptile abundance, species richness, diversity, evenness, and the abundance of dominant species of reptiles .2>0.2) are graphically depicted on the ordinations.Community composition, defined as the average abundance (per 10 trap nights) of each species per site, was compared among factors (vegetation type and fire age) using Multi-Response Permutation Procedure (MRPP) 2 values for these relationships.To examine the relationship between microhabitat variables  and time since last fire within each vegetation type, General Additive Models (GAM) were used. We used GAMs rather than assuming linear fits, as univariate plots between the microhabitat variables and time since last fire indicated there may be non linear relationships. GAMs were fitted using the gamm function of the mgcv package in R C. adelaidensis, C. buchanni, H. quadrilineata, L. elegans, M. greyii and M. obscura) was explored within each vegetation type using GAMs (as outlined above) by comparing all possible models of one, two, and three predictors. Akaike information criterion, corrected for small sample size (AICc), the associated AICc weights and adjusted r2 values were used to select the optimal model from this complete set 2 values>0.10.The relative importance of the five microhabitat variables and time since last fire for predicting reptile responses  and Menetia greyii (n = 143), the tree-dwelling skink Cryptoblepharus buchananii (n = 106), and the small terrestrial heath dragon Ctenophorus adelaidensis (n = 126). The small elapid Neelaps calonotos, listed as a priority species requiring further monitoring under the Western Australian Wildlife Conservation Act 1950, was captured twice. Of the 33 reptile species observed, 12 species  were captured ≤2 times and/or were detected from ≤2 sites during surveys, and were removed from community analyses. The three elapid  and one blind snake  species captured ≤2 times were only captured on older sites (>16 years since last fire).Pit-fall trapping at the 30 sites resulted in 1042 captures  from 33 species of lizards and snakes . The most commonly caught reptiles were skinks, including the litter dwelling species C. adelaidensis was observed in higher abundances in young banksia sites, but was also abundant at 35 year old sites , melaleuca = 0.57 (±0.18); H. quadrilineata, banksia = 0.19 (±0.12), melaleuca = 0.40 (±0.17)). Both of the skinks M. greyii and Morethia obscura were observed in higher abundances in the older sites compared to the younger sites , young <12YSLF = 0.16 (±0.09)).Reptile species richness was higher in banksia woodland sites . A signild sites . A numbeP<0.001). Pair-wise comparisons indicated that the melaleuca old sites differed from the banksia old sites (P = 0.017), the banksia young sites (P = 0.001) and the melaleuca young sites (P = 0.047). The banksia old sites also differed to the melaleuca young sites (P = 0.003) and were approaching a different community structure to the banksia young sites (P = 0.055). Banksia young sites differed in community structure to melaleuca young sites (P = 0.007).Twenty-one reptile species were detected in more than two sites and were included in community analyses. MRPP was performed on four groups based on the combination of habitat and fire age, including: melaleuca, old; melaleuca, young; banksia, old; banksia, young. MRPP detected differences in community structure between the four groups , but decreased in melaleuca sites , however no relationship was apparent for bareground cover in melaleuca sites , whereas no relationship was detected between canopy cover and time since last fire in banksia woodlands .The relationship between microhabitat variables and time since last fire was dependent upon vegetation type . In bankoodlands . Litter 2 value>0.10. In contrast, only one top model with moderate support was detected for reptile abundance  and species number  with time since last fire and vegetation cover respectively in the melaleuca damplands  curvilinear response with time since last fire (2 = 0.46) was also included in the top models for C. adelaidensis in banksia woodlands, indicating that both of these variables are potential predictors of the abundance of this small dragon (2<0.05). In melaleuca sites, the abundance of M. obscura was best explained by vegetation cover (adjusted r2 = 0.45), whereby an increase in vegetation cover, was associated with a decrease in the abundance of this skink  and litter depth (adjusted r2 = 0.23) were identified in top models for H. quadrilineata in melaleuca damplands (M. greyii was best explained by litter depth (adjusted r2 = 0.52), vegetation cover (adjusted r2 = 0.59), litter cover (adjusted r2 = 0.49) and a combination of both litter depth and vegetation cover (adjusted r2 = 0.69) (For the skinks amplands . In mela = 0.69) .The responses of microhabitat structure and reptiles to fire age are strongly driven by vegetation types. The differences in microhabitat structure among fire ages were particularly pronounced in melaleuca sites. For example, litter depth was greater in the older sites, but the differences between old and young melaleuca sites was more than double the differences between old and young banksia sites. Similarly, the differences in vertical vegetation density were more pronounced in melaleuca sites. The older aged banksia sites contained higher vegetation density (<80 cm), whereas the younger melaleuca sites contained higher vegetation density (<80 cm). Unsurprisingly, the banksia woodland sites were more floristically diverse compared to the melaleuca damplands. Much of the banksia woodland diversity is contained in the highly variable understorey <60 cm C. adelaidensis, M. greyii, and M. obscura) responded to fire age, and two additional skinks, C. fallens and C. buchananii, were associated with older fire ages within a particular habitat type. Responses of reptiles to fire age may be associated with microhabitat attributes that also vary between fire age categories Reptile communities also varied between vegetation types and fire ages. Young melaleuca habitat tended to contain fewer reptiles, and had few species associated with them. Although some differences were also detected among young and old banksia sites, these were less pronounced. However, our results indicate that three of the six most frequently detected reptile species  C. adelaidensis, C. buchanni and M. obscura) in one of the vegetation types. In addition, various microhabitat variables were identified in top models for at least one vegetation type for reptile species number, C. buchanni, M. greyii, M. obscura and H. quadrilineata. In both vegetation types, time since last fire and microhabitat variables were unable to explain much variability in the abundance of L. elegans (highest r2 = 0.11).Our study has highlighted how time since last fire and microhabitat variables may influence species' abundances. Of the eight reptile response variables examined, time since last fire was identified as an important predictor for four variables , indicating the composition of microhabitat in these vegetation types are likely to driven by additional factors.The responses of microhabitat variables to time since last fire also varied between vegetation types. For example, vegetation cover positively increased with time since last fire in banksia woodlands, but decreased in melaleuca damplands. Such opposite responses are likely a function of plant species composition, with vegetation cover in young melaleuca sites dominated by few species. As time since last fire increases, the understorey vegetation cover may decline in response to an increase in canopy cover. Only a few of the relationships among microhabitat variables and time since last fire were very strong  increases until approximately 20 years since last fire, after which the litter cover and depth declines and plateaus respectively. Rates of ground litter fuel accumulation are different for different components of fuel C. adelaidensis at older sites was possibly in response to this change in litter accumulation. A similar response to combinations of time since fire and microhabitat changes has been observed for the rodent Pseudomys novaehollandiaeChanges in the abundance of reptiles following burning is often linked to fire-induced changes in the resource availability of the postfire environment M. greyii in our study, respond strongly to the removal of litter and are usually observed in high abundance in the least-disturbed sites, with their density often correlated with microhabitat variables, such as litter cover As reptile abundance was lowest in very young (<5 YSLF) melaleuca sites, burning may have modified elements of the microhabitat in a manner undesirable to some species. The young melaleuca sites had reduced litter depth, litter cover and canopy cover, and a limited number of plant species. Typically, litter-associated lizards, such as M. obscura in banksia woodlands is best described by time since last fire, rather than microhabitat variables, with the highest numbers of this skink detected in very old sites. In contrast, in jarrah forests in south-west Western Australia, this species was only recorded in restored mine sites that had been thinned and recently burnt M.obscura may be driven by broader scale ecological process  that fire affects in various ways depending on the ecosystem. Knowledge of the ecology and natural history of small lizards is depauperate and hampers our interpretation of species' responses to disturbances.Similarly, the abundance of Neelaps calonotus) we recommend retaining older-aged banksia sites (>12 years since last fire) that are currently being targeting for prescribed burning. Our work will contribute to the development of ecological burning regimes incorporating both flora and fauna responses to fire. Recommendation for burning rotations that will retain biodiversity values, reduce wildfire risk and be spatially variable can be developed by conservation and fire managers.For prescribed burning to be effective for conservation purposes, land managers require understanding of the impacts of fire on biota, and clear management targets regarding burning practices and ecological objectives


XRCC1 polymorphisms and childhood acute lymphoblastic leukemia (ALL) risk. However, the results of previous reports are inconsistent. Thus, we performed a meta-analysis to clarify the effects of XRCC1 variants on childhood ALL risk.Recently, there have been a number of studies on the association between XRCC1 polymorphisms  and childhood ALL risk. We critically reviewed 7 studies with a total of 880 cases and 1311 controls for Arg399Gln polymorphism, 3 studies with a total of 345 cases and 554 controls for Arg280His polymorphism, and 6 studies with a total of 783 cases and 1180 controls for Arg194Trp polymorphism, respectively. Odds ratio (OR) and its 95% confidence interval (CI) were used.A meta-analysis was performed to examine the association between XRCC1 Arg399Gln polymorphism and childhood ALL risk was observed in total population analyses  and Asian subgroup analyses . No association was detected in Caucasians, Metizo and mixed populations. Ethnicity was considered as a significant source of heterogeneity in the meta-regression model. For the other two XRCC1 polymorphisms, no association with childhood ALL risk was found.Significant association between XRCC1 Arg399Gln polymorphism might be associated with elevated childhood ALL risk among Asian population.The meta-analysis results suggested that  While survival rates for childhood acute lymphoblastic leukemia (ALL) have been improved significantly over the past 50 years, ALL is still the most common pediatric cancer in developed countries with high incidence and mortality As a complex and multifactorial process, leukemogenesis is still not fully understood. It is widely known that dysregulated immune response to infection may be a cause of childhood ALL To date, the candidates for childhood ALL susceptibility genes have been categorized into those coding for carcinogen metabolism enzymes, folate metabolism enzymes, DNA repair proteins, and others XRCC1), located on chromosome 19q13.2–13.3, with 33 kilobases in length, is one of the most important proteins in base excision repair (BER) XRCC1 gene reported in the dbSNP database (http://www.ncbi.nlm.nih.gov/SNP). Nevertheless, only three genetic changes have been extensively studied including Arg194Trp on exon 6 , Arg280His on exon 9 , and Arg399Gln on exon 10 . There have been a number of studies on the association between XRCC1 polymorphisms and childhood ALL risk XRCC1 variants  on childhood ALL risk, we conduct a meta-analysis of all eligible studies to resolve this pivotal issue.X-ray repair cross-complementing group 1 (XRCC1” or “X-ray repair cross-complementation group 1”), covering all papers published before November 30, 2011. All eligible articles were retrieved and their references were searched simultaneously to find other relevant articles. Inclusion criteria was defined as follows: (1) case-control studies evaluating the association between XRCC1 polymorphisms and childhood ALL risk; (2) studies based on unrelated individuals; (3) sufficient published data available to estimate an odds ratio (OR) with 95% confidence interval (CI). We excluded studies that were not full-length publications articles or letters in peer-reviewed English journals. When the same patient population was included in different articles, the one with the largest population of participants or the most recent one was selected.Computer searches of PubMed, EMBASE, Medline, Google Scholar and Cochrane Library were performed by two authors independently using the following key words (“childhood acute lymphoblastic leukemia” or ”childhood ALL”)and with the corresponding 95% CI for each study. The OR and its 95% CI in each comparison were assessed in additive , dominant (aa+Aa versus AA), and recessive (aa versus Aa+AA) genetic models. Heterogeneity among studies was tested by chi-square-based Q test, and I2 statistics was calculated to quantify the proportion of the total variation due to heterogeneity The strength of With our search criterion, a total of eleven full-text articles XRCC1 Arg399Gln polymorphism and childhood ALL risk, and the heterogeneity test were shown in OR = 0.008, P = 0.384 for heterogeneity) . We did not observe any significant association in any genetic model among other subgroups. Moreover, meta-regression analysis revealed that ethnicity was a significant source of between-study heterogeneity (P = 0.007).The results of the associations between ogeneity). We alsogeneity) . No signOR = 0.709; Dominant model: OR = 1.125, 95% CI 0.805–1.574, POR = 0.490; Recessive model: OR = 1.203, 95%CI 0.357–4.055, POR = 0.765) and Arg194Trp polymorphism. Subgroup analysis based on ethnicity also showed no significant association between the two SNPs and childhood ALL risk.There was no statistical difference in all contrasts of genotypes for Arg280His polymorphism ; Arg194Trp polymorphism additive model , dominant model , recessive model   . A randoInfluence analysis was performed to assess the influence of each individual study on the pooled OR by sequential omission of individual studies. The results suggested that no individual study significantly affected the pooled ORs .Publication bias was examined by Funnel plot and Egger's regression test. The shapes of the funnel plot did not indicate any evidence of obvious asymmetry in additive model  and the XRCC1 plays an important role in the DNA damage repair pathway for the processing of small base lesions, which has been thought of as the predominant DNA-damage repair pathway for the processing of small base lesions derived from oxidation and alkylation damage XRCC1 genetic polymorphisms and provide the most comprehensive assessment of its association with childhood ALL risk. By critically reviewing 7 studies on XRCC1 Arg399Gln polymorphism , 3 studies on XRCC1 Arg280His polymorphism , and 6 studies on Arg194Trp polymorphism , we performed a meta-analysis to indicate that the polymorphisms in XRCC1 Arg399Gln was significantly associated with risk of childhood ALL. However, we did not observe associations of XRCC1 Arg280His polymorphism and Arg194Trp polymorphism with childhood ALL risk.XRCC1 variant 399Gln allele was 28.99% . No association was detected in Caucasians, Metizo and mixed populations, while increased risk was found in Asian population carrying variant 399Gln allele homozygote, and it should be further investigated in large scale Asian populations.This study showed that the mean frequency of the s 28.99% . PreviouDespite lots of investigations in many aspects of childhood ALL, little attention has been paid to its pathogenesis, particularly with respect to genetic susceptibility. Several reports have demonstrated the association between some DNA repair gene variants and childhood ALL, so the possible relationship between polymorphisms of DNA repair genes and childhood ALL may be helpful in understanding the pathogenesis of childhood ALL and the prevention of this disease. Our results suggest that the risk of childhood ALL may be associated with DNA repair mechanisms. XRCC1 polymorphisms may be used as an important predictive factor, and ethnic background might have an impact on the results in the studies of its polymorphisms in childhood ALL. Analysis of these polymorphisms, particularly XRCC1 codon 399 Arg/Gln may help in identifying individuals at risk of developing ALL and providing an essential information source for future improvement of ALL treatment.XRCC1 variants and risk by subtype. Second, specific environmental and lifestyle factors may alter those associations between gene polymorphisms and cancer risk. Evidence supports that there is no association between Gln/Gln genotype of Arg399Gln and bladder cancer risk in total population, but it is associated with a decreased risk of bladder cancer among ever smokers XRCC1 gene polymorphism and childhood ALL risk was analyzed without consideration of gene–gene and gene-environment interactions because of the lack of sufficient data, which should be further investigated. Third, the Funnel plot did not reveal any evidence of obvious publication bias, while there is still a possibility that our meta-analysis was biased toward a positive result since negative findings were likely to be unreported.Although our result is suggestive, there are still some limitations in this meta-analysis. First, heterogeneity among the studies, resulting from different sources of controls, matching criteria of age and gender or some other factors, may influence the results of this analysis. The matching criteria of the control group, such as age, gender, and environment exposures, are different between studies. Furthermore, childhood ALL is heterogeneous considering its underlying cellular and molecular biology, so subtypes of B- or T-cell precursor ALL may not be suspected to share a common etiology. To date none of the studies have examined the relationship between XRCC1 399Gln might be a susceptibility allele for childhood ALL. However, we could not observe any association of XRCC1 Arg194Trp and Arg280His with childhood ALL. We have searched as many publications as we could by means of various searching approaches, comprehensively assessed publication biases and pinpointed the potential sources of heterogeneity via subgroup and sensitivity analysis. However, these results may be biased by the relatively small number of subjects, and therefore need to be validated by larger studies and subsequent update of the current meta-analysis. In order to understand the mechanisms underlying childhood ALL better, future research should be considered and investigated.In summary, it is a worthy and meaningful enterprise to search for polymorphic variants influencing the risk of childhood ALL. This meta-analysis suggests that Table S1Frequencies of XRCC1 Arg194Trp, Arg280His and Arg399Gln allele among control population in different studies.(DOC)Click here for additional data file.


Severe unconjugated hyperbilirubinemia may cause deafness. In the Netherlands, 25% lower total serum bilirubin (TSB) treatment thresholds were recently implemented for preterm infants.To determine the rate of hearing loss in jaundiced preterms treated at high or at low TSB thresholds.In this retrospective study conducted at two neonatal intensive care units in the Netherlands, we included preterms  treated for unconjugated hyperbilirubinemia at high or low TSB thresholds. Infants with major congenital malformations, syndromes, chromosomal abnormalities or toxoplasmosis, rubella, cytomegalovirus, herpes, syphilis, and human immunodeficiency infections were excluded. We analyzed clinical characteristics and TSB levels during the first ten postnatal days. After two failed automated Auditory Brainstem Response (ABR) tests we used the results of the diagnostic ABR examination to define normal, unilateral, and bilateral hearing loss (>35 dB).There were 479 patients in the high and 144 in the low threshold group. Both groups had similar gestational ages (29.5 weeks) and birth weights (1300 g). Mean and mean peak TSB levels were significantly lower after the implementation of the novel thresholds: 152±43 µmol/L and 212±52 µmol/L versus 131±37 µmol/L and 188±46 µmol/L for the high versus low thresholds, respectively (P<0.001). The incidence of hearing loss was 2.7% (13/479) in the high and 0.7% (1/144) in the low TSB threshold group .Implementation of lower treatment thresholds resulted in reduced mean and peak TSB levels. The incidence of hearing impairment in preterms with a gestational age <32 weeks treated at low TSB thresholds was substantially lower compared to preterms treated at high TSB thresholds. Further research with larger sample sizes and power is needed to determine if this effect is statistically significant. In line with this is the increased likelihood of an abnormal hearing screening test, i.e. an automated auditory brainstem response (AABR), with increasing UCBfree levels in preterm infants of 34 or more weeks of gestation. In 2007, lower treatment thresholds were introduced for the management of unconjugated hyperbilirubinemia in Dutch preterm infants of less than 32 weeks of gestation. In addition, the effect of a variety of demographic and clinical characteristics on hearing impairment was analyzed.According to the Research involving Human Subjects Act (WMO), because this study only includes retrospective research with data, it is not subject to an ethics review and does not need to include informed consent from the subjects, as the data was used anonymously. This study fulfils all the requirements for test subject anonymity and is in accordance with regulations of the review boards of the participating hospitals.Preterm infants of less than 32 weeks of gestation were included when admitted to one of the two participating NICU centers in two time periods: between January 2005 and December 2006, i.e. before introduction of the novel thresholds, and between March 2007 and March 2008. Exclusion criteria consisted of major congenital malformations, clinical syndromes, chromosomal abnormalities or TORCH . These infections were excluded as these conditions are likely to affect hearing status. In addition, infants who died before the automated auditory brainstem response (AABR) was carried out were excluded.2/nm; center 2∶7.4±3.3 µW/cm2/nm; p = 0.11). Eligible infants admitted in 2005 and 2006 to one of the participating NICU centers were included and used as a control group: the high threshold group. In this group, treatment of unconjugated hyperbilirubinemia was applied according to identical guidelines for four (birth weight) categories: <1000 grams, 1000–1500 grams, 1500–2500 grams, and term newborns. In April 2007, a new “consensus based” guideline for preterm infants was implemented in the Netherlands. Treatment thresholds were ∼25% lower in the novel guideline. Total serum bilirubin was tested in the first 10 postnatal days or when clinically indicated. Bilirubin measurements were performed using routine laboratory techniques. In each infant, the mean and mean peak of TSB measurements were calculated and used for analysis.All Dutch NICUs participate in a two-stage AABR neonatal hearing programme. The instruments’ algorithm of the AABR assumes an infant as passing the test when the acquired data obtained at 35 dB fits with 99.96% likelihood a template composed of auditory brainstem responses from normal hearing newborns. Descriptive analyses were performed to determine the incidence of hearing loss and background characteristics in the high and low threshold group. Sensitivity and specificity of mean and mean peak TSB levels were evaluated using receiver-operating characteristics (ROC) curves.No difference was observed in the proportion of infants who were hospitalized ≥ 12 days at intensive care, which is seen as an independent risk indicator for hearing loss. This retrospective cohort study demonstrates that introduction of lower treatment thresholds in preterm infants results in significantly lower mean and mean peak TSB levels. Treatment according to novel treatment thresholds did not result in a significant reduction of hearing impairment, albeit a ∼75% reduction in the incidence of hearing loss. Finally, and in analogy to previous data in term infants, it appeared that TSB level is also a poor predictor of hearing loss in preterm infants.In 2007, the Dutch Society of Pediatrics approved and recommended a novel, “consensus-based” guideline on unconjugated hyperbilirubinemia in preterm infants. Infants treated according to the ∼25% lower thresholds had lower TSB levels: we found a reduction of ∼15% in mean and mean peak TSB levels, supporting an effective implementation of the new guideline.Yet, auditory impairment in preterm infants may also occur at TSB levels below applied treatment thresholds, and - despite the rather low TSB levels- similar TSB levels have contributed to profound neurological impairment including sensoneurinal hearing loss in extreme or very low birth weight infants.The incidence of hearing loss in the control group treated at higher TSB thresholds was in agreement with previous data: in a representative NICU population (10830 neonates with a mean (SD) gestational age 28.5 (1.6) weeks and birth weight of 1039 (256) g) the prevalence of hearing loss was 3.2% (95%-CI 2.6–4.1). Despite these limitations, this is the first study to report on evaluation of a consensus-based national guideline in preterm infants with unconjugated hyperbilirubinemia regarding TSB levels and - unlike other studies - a formal diagnostic ABR (and not a screening ABR) as outcome parameters.We conclude that successful implementation of lower TSB thresholds results in a reduction of mean TSB and mean peak TSB levels, and that larger studies - which preferably also provide data on Bf levels - are needed to establish a statistically significant effect of hyperbilirubinemia on hearing loss in very preterm infants.


Drosophilae genomes GSSA identifies hundreds of functional modules with significant associations to high and low rates of evolution. Many of the detected functional modules with high evolutionary rates have been previously identified as biological functions under positive selection. Notably, GSSA identifies conserved functional modules with many positively selected genes, which questions whether they are exclusively selected for fitting genomes to environmental changes. Our results agree with previous studies suggesting that adaptation requires positive selection, but not every mutation under positive selection contributes to the adaptive dynamical process of the evolution of species.Classically, the functional consequences of natural selection over genomes have been analyzed as the compound effects of individual genes. The current paradigm for large-scale analysis of adaptation is based on the observed significant deviations of rates of individual genes from neutral evolutionary expectation. This approach, which assumed independence among genes, has not been able to identify biological functions significantly enriched in positively selected genes in individual species. Alternatively, pooling related species has enhanced the search for signatures of selection. However, grouping signatures does not allow testing for adaptive differences between species. Here we introduce the Gene-Set Selection Analysis (GSSA), a new genome-wide approach to test for evidences of natural selection on functional modules. GSSA is able to detect lineage specific evolutionary rate changes in a notable number of functional modules. For example, in nine mammal and  Drosophilae; or an almost equal distribution of positively selected genes in functions evolving at significantly high and low rates in primates. Such findings show the complex correspondence between positive selection and the dynamic process of adaptive evolution in genomes.Characterizing genome adaptation is paramount for understanding evolutionary genomics. Classically, the search for positively selected genes has been used to identify adaptive differences in morphology, physiology and behavior between species. However, this approach assumed gene independence and was unable to identify sets of functions significantly enriched by positively selected genes. To overcome such limitation, we apply an alternative test on the evolutionary rates of genes, called Gene-Set Selection Analysis (GSSA), which is able to detect functional sets of genes evolving at high and low evolutionary rates in genomes. Our analysis illustrates that by focusing on sets of genes instead of individual loci, we are able to describe a richer relationship between positive selected genes and the adaptive evolution of functions in different genomes. For example, GSSA identified many positively selected genes within biological functions under strong evidence purifying selection in mammals and  In a second step, a conventional functional enrichment test Adaptation analysis at a large or genome scale relies on methods and concepts originally conceived for the study of single genes . The current paradigm for large scale analysis of adaptation typically involves conducting a given test individually for all of the genes of a genome in order to find those with statistically significant deviations from neutrality  Drosophila genomes defense response, proteolysis, DNA metabolic process, and odorant binding, among others. In the analysis of 6 mammalian genomes chemosensory perception and defense/immunity related processes were functionally enriched after pooling together all PSGs (400 genes) in primates and rodents respectively. Finally, using the deviations from the expected branch length on gene trees, similar patterns of selection across genomes were found for a group of gamma proteobacteria To circumvent this statistical problem recent works have drawn their conclusions by looking for signatures of selection in related groups of species a prior threshold application have already been noticed in other omics fields such as transcriptomics The limitations of methods based on Drosophila species. By using this gene-set strategy we found a large number of functional modules that have significantly increased or decreased their rates of evolution with respect to the ancestral state. We will show evidences of selection working in groups of functionally related genes, suggesting that they share a common pattern of evolution imprinted by natural selection. In addition, all biological GO processes previously found as significantly enriched by PSGs were distinguished within the set of functions evolving at higher rates than the expected in genomes. Finally, the relationship between GSSA results and the relative influence of PSGs during adaptive evolution is discussed.Here we set forth the Gene-Set Selection Analysis (GSSA), a gene-set based test that searches for significant evidences of the action of natural selection modeling the evolutionary rates of groups of genes in genomes. Two different and widely accepted definitions of functional modules: GO D = ωD−ωA) along the phylogeny . Throughout the manuscript only p-values for partitions with the highest confidence were reported after FDR.Mammals, represented by human, chimpanzee, rat and mouse, and five Drosophila genomes were studied. For each species, genes were ranked into four lists according to the estimation of i- synonymous (dS), ii- nonsynonymous (dN) rates of substitution, iii- selective pressures (ω = dN/dS), and iv- the change of selective pressures between (A) ancestor and (D) descendant species  in mammals. Alternatively, 4.1% and 2.6% of GO terms and KEGG pathways were found with significantly high values of ω (SHω) in Drosophila, respectively.The application of GSSA to lists of genes ranked by dS, dN, ω and the Δω values yielded a large number of functional modules (defined by GO and KEGG annotations) with rates that were significantly skewed toward the extremes of the lists  in mammaDrosophila rather than in mammals.Drosophilas. This observation points out the conservative character of the selective constraints in functional related groups of genes during evolution.Only a minor proportion of functional terms changed significantly at higher or lower rates relative to estimates of the corresponding ancestral lineages. Specifically, increased or decreased ω values on the external branches  were observed for only half of the cases where a significant increase or decrease of ω was identified in mammals and Drosophilas is shown in neurological process and sensory perception clearly contrasted between primates and rodents  in primates than in rodents. Moreover, significant differences in rates can be detected between human and chimpanzee , and between mouse and rat .A summary of the results of the GSSA for mammals and  rodents . While mDrosophilas were unevenly distributed within the two clusters of the phylogeny  and rodents (11.3%), while in primates (6.4%), where population sizes are known to be smaller, the number of significant modules was smaller The fact that most of the functional modules under selection (SHω and SLω) correlate with changes in dN, suggests that selective pressures are mainly driven by nonsynonymous rather than by synonymous substitutions during evolution. Moreover, according to the expectation of the nearly neutral theory, a low but still considerable number of significant associations of functional modules to dS were found in The strategy presented here lead to detect significant patterns of increments and decrements modeled by natural selection in evolutionary rates of functional groups of genes. This pattern is consistent with the hypothesis that natural selection acts on phenotypes by the combined action of many functional related genes. Moreover, this functionally based approach identified with statistical significance, and on individual species, all the functional modules previously found significantly enriched by positively selected genes and therefore the main targets of adaptive biological functions in species   test of positive selection was conducted on terminal branches of phylogenies . We founp≪0.01), mammals (p≪0.01), and Drosophila (p≪0.01) species. For primates however, it was not significant (p = 0.47), indicating that PSGs are distributed almost evenly in functional modules evolving at significantly high and low values of ω in human and chimpanzee.Finally, we ask if PSGs preferentially concentrate in functional modules evolving at faster rates in different genomes. For doing that we computed the mean number of PSGs in functional modules with SHω and SLω results (red and blue dots in p≪0.01) towards higher numbers of PSGs in mammals, rodents, and Drosophila species, but showing no differences in primates (p = 0.73).To contrast these results, PSGs from previous works in mammal and Drosophila species were collected Drosophila species. Alternatively, PSGs were evenly distributed in functional modules changing at the extreme rates of evolution in primates. This observation suggests that a more complex scheme than the cumulative differences of PSGs must rely on the observed adaptive differences in human and chimpanzee genomes. The search for integrative factors taking into account the action of multiple genes other than only those which have been targeted by positive selection In summary, PSGs are frequently observed in functional modules evolving under a wide range of evolutionary scenarios; however, they concentrate more frequently in functional groups of genes changing at elevated rates in rodents and Evolutionary biologists recognize that natural selection works on phenotypes indirectly by changing the frequency of genes in populations The existence of many PSGs in functional modules evolving at significant low (or no-significant) ω ratios does not represent false positive results in the analysis of molecular adaptation. This observation, registered in our data and detected in previous publications, suggests that PSGs are frequently recruited in the genomes for other purposes than the classical increase of rates of functional set of genes compromised in adaptive processes such as evolutionary arm-races. A possible explanation is that many of the PSGs in the genomes are changing in association with the constraints imposed by the architecture of the network Currently, with the possibility of conducting analysis at the genome level, evolutionary biology cannot disregard major aspects of systems biology approaches that consider the modular organization of genomes. With the testing strategy used here, we increased the statistical power for the evolutionary analysis on individual genomes and suggest that PSGs could have additional roles in the genome than the adaptive evolutionary change of phenotypes.H. sapiens was retrieved from the Ensembl-Compara database vs56 D. melanogaster protein-coding genes of D. melanogaster was retrieved from the Ensembl Metazoa-Compara database vs4 D. simulans, D. sechellia, D. yakuba, D. erecta, and D. ananassae.The subset of 23,438 known Ensembl human protein-coding genes of the Ensembl vs56.37a Drosophila respectively.DNA coding sequences (CDS) were aligned using the Muscle vs3.7 Maximum likelihood estimation of dN, dS, and ω was computed using CodeML program from PAMLDrosophila species External branches of Gene-set selection analysis across lists of genes ranked by different evolutionary rate parameters  was computed using the program Babelomics Drosophila species respectively. The global GO directed acyclic graph was processed with Blast2GO Drosophilas. The final set of GO and KEGG terms used in the GSSA corresponds to those containing a minimum number of 15 genes. To test possible biases attributed to the size of the functional category, the magnitude of change in evolutionary rate or the proportion of genes experiencing a rate change we randomized the original assignation of ENSG's to the list of ranked values and functional annotation  using branch-site models. Here we addressed the possibility of a variation in the representation of significant results after GSSA  results for dS, dN, dN/dS & Δω using 731 GO terms.(6.47 MB PDF)Click here for additional data file.Figure S2Complete list of significant results of GSSA for GO terms in Drosophila species. The figures cover from the most general to the most specific biological GO functions. GSSA (5% FDR) results for dS, dN, ω & Δω using 386 GO terms.(8.85 MB PDF)Click here for additional data file.Figure S3Complete list of significant results of GSSA for KEGG pathways in mammals species. GSSA (5% FDR) results (82 KEGG pathways) for dS, dN, ω & Δω in mammals species.(1.54 MB PDF)Click here for additional data file.Figure S4Complete list of significant results of GSSA for KEGG pathways in Drosophila species. GSSA (5% FDR) results (43 KEGG pathways) for dS, dN, ω & Δω in Drosophila species.(0.87 MB PDF)Click here for additional data file.Figure S5Randomisation experiment. (A) The pipeline shows the steps followed to tests possible biases attributed to the size of the functional category, the magnitude of change in evolutionary rate and the proportion of genes experiencing a rate change in the GSSA. The proportion of false positive results never reached 5% (FDR) in mammals (B) and Drosophila (C).(1.34 MB PDF)Click here for additional data file.Figure S6Evolutionary and statistical simulation of GSSA. The pipeline shows the steps taken along three different spaces of analysis, the real data, the simulated data and the testing block. See Supplementary Results for a complete explanation of methods and results.(1.42 MB PDF)Click here for additional data file.Table S1Evolutionary rates of genes computed in Mammals. Complete values of evolutionary rates  for all genes analysed.(5.32 MB ZIP)Click here for additional data file.Table S2Evolutionary rates of genes computed in Drosophila. Complete values of evolutionary rates  for all genes analysed.(9.55 MB XLS)Click here for additional data file.Table S3The complete functional enrichment results using gene-by-gene and gene-set approaches. The table depicts all the biological functions enriched by PSGs as cited in references 1 to 7, and the corresponding significant result observed after GSSA of ω values. References 1 to 7 correspond to cites 6, 7, CSAC, 4, 5, 9 and 8 in the manuscript, respectively. Abbreviations: SHω: statistically significant high ω values; SLω: statistically significant low ω values; H: human; C: chimpanzee; Pr: primates; M: mouse; R: rat; Ro: rodents; Dmel: D. melanogaster; Dsim: D. simulans; Dsec: D sechelia; Dyak: D. yakuba; Dere: D. erecta; Ds: Drosophila species. *: p<0.05; ** p<0.001. CSAC: Chimpanzee Sequencing and Analysis Consortium, Nature. 2005 vol. 437 (7055) pp. 69–87.(0.09 MB PDF)Click here for additional data file.Text S1Supplementary (0.11 MB PDF)Click here for additional data file.


The Echiura, or spoon worms, are a group of marine worms, most of which live in burrows in soft sediments. This annelid-like animal group was once considered as a separate phylum because of the absence of segmentation, although recent molecular analyses have placed it within the annelids. In this study, we elucidate the interfamily relationships of echiuran worms and their evolutionary pattern of feeding mode and sexual dimorphism, by performing molecular phylogenetic analyses using four genes  of representatives of all extant echiuran families. Our results suggest that Echiura is monophyletic and comprises two unexpected groups: [Echiuridae+Urechidae+Thalassematidae] and [Bonelliidae+Ikedidae]. This grouping agrees with the presence/absence of marked sexual dimorphism involving dwarf males and the paired/non-paired configuration of the gonoducts . Furthermore, the data supports the sister group relationship of Echiuridae and Urechidae. These two families share the character of having anal chaetae rings around the posterior trunk as a synapomorphy. The analyses also suggest that deposit feeding is a basal feeding mode in echiurans and that filter feeding originated once in the common ancestor of Urechidae. Overall, our results contradict the currently accepted order-level classification, especially in that Echiuroinea is polyphyletic, and provide novel insights into the evolution of echiuran worms. Echiurans (spoon worms or innkeeper worms) are a group of marine worms with a sausage-shaped non-segmented body and a highly extensible scoop-like proboscis In recent years, much attention has been paid to the systematic position of echiuran worms In contrast to their systematic position within the annelids, the interfamily relationships of echiurans remain poorly understood Ikeda taenioides , which is endemic to Japan I. taenioides was once described as including outer longitudinal and inner circular layers, which is the opposite of that found in all other echiurans I. taenioides specimens revealed that the arrangements of their muscle layers are not different from the other echiurans It has also been suggested that the currently accepted classification should be revised It is also interesting that echiuran worms include species with and without marked sexual dimorphisms c oxidase subunit I, COI). Representatives of all extant echiuran families were included in the analyses. We discussed the classification and character evolution of echiuran worms based on the tree produced.In order to understand the evolutionary pattern of feeding mode and sexual dimorphism in echiuran worms, and to resolve taxonomic confusion, we constructed the molecular phylogenetic tree of echiuran worms using two nuclear ribosomal genes (18S and 28S rRNA), one nuclear protein gene (histone H3) and one mitochondrial protein gene  and [Thalassema+Arhynchite]  (Maximum likelihood (ML) and Bayesian phylogenetic analyses of the combined data set  . Phyloge = 1.00) .Traditionally, echiurans are separated into the three orders: Echiuroinea , Xenopneusta (with Urechidae) and Heteromyota (with Ikedidae) A molecular phylogenetic analysis of echiurans based on three genes  was recently performed in Lehrke's dissertation work Osedax, anglerfishes, and pedunculate barnacles) Our results suggested that echiurans are separated into two groups; [Thalassematidae+Echiuridae+Urechidae] and [Ikedidae+Bonelliidae] . These tMost echiuran worms are deposit feeders that collect detritus by scooping mud or sand sediments using their flattened and elongated proboscis Our analyses strongly supported a sister group relationship of Bonelliidae and Ikedidae  . In addiBonellia viridis Rolando, 1821  . These tArhynchite and Thalassema] and   as an outgroup species  and Urechis sp. 1 , 0.1 µl of ExTaq polymerase , and 15.1 µl of distilled water. Thermal cycling was performed with an initial denaturation for 3 min at 94°C, followed by 30 cycles of 30 s at 94°C, 30 s at a gene-specific annealing temperature , H3 and COI genes. Polymerase chain reactions (PCRs) were used to amplify ∼1700 bp of 18S rRNA and ∼1000 bp of 28S rRNA, ∼350 bp of H3 and ∼700 bp of COI. Amplifications were performed in 20 µl mixtures consisting of 0.4 µl of forward and reverse primers Click here for additional data file.Table S2Information on primers and PCR conditions used in this study.(PDF)Click here for additional data file.Table S3Information on models of sequence evolution for maximum-likelihood (ML) and Bayesian analyses.(PDF)Click here for additional data file.Dataset S1Combined molecular data set. The data provided includes alignments of the four concatenated molecular data partitions .(PDF)Click here for additional data file.


Prostate cancer patients with rectal involvement are rare, and the factors associated with the survival of these patients are yet to be elucidated.We collected data on patients who were admitted to our hospital for prostate cancer in the last thirteen years and of those in studies in the literature. The associations of clinical characteristics with survival were evaluated using Cox regression models.This study included 94 patients  of prostate cancer with rectal involvement. 11 patients in the group of synchronous rectal involvement at first cancer diagnosis (n = 58) and 23 patients in the group of metachronous diagnosis of rectal involvement (n = 29) died at the latest follow up. The estimated overall survival rate (% ± SE) at 1, 3, and 5 years were 68.3 ± 5.3%, 54.4 ± 7.2%, and 38.1 ± 11.1%, respectively. In the Cox univariate analysis, Asian prostate cancer (p = 0.001) was associated with better survival, while rectal bleeding (p = 0.043), metachronous presentation of development of rectal involvement (p = 0.000), prior hormonal therapy (p = 0.000) and extrarectal metastases (p = 0.054) were associated with poor survival. In multivariate analysis, prior hormone therapy  and rectal bleeding  retained independent poor prognostic values. There were 13 patients survived for more than 3 years, the longest survival time was 96 months. Total pelvic extenteration (TPE) combined with hormonal therapy in 12 hormone-untreated prostate cancer give us six of thirteen long-term survivors for more than 3 years in this series.Our findings suggest that rectal involvement does not necessarily predict a worse outcome when presenting as a previously hormone-untreated disease and that the prognosis was worse when presenting as a hormone relapsed disease. Prior hormone therapy and rectal bleeding were associated independently with a significantly poor overall survival in prostate cancer patients with rectal involvement. TPE combined with hormonal therapy appears to confer better overall survival in hormonally untreated patients.http://www.diagnosticpathology.diagnomx.eu/vs/1604504118106105.The virtual slide(s) for this article can be found here:  In ChMetastatic disease to the bones and lymph nodes has long been recognized as the most typical pattern of extraprostatic tumor spread . ProstatIn this study, we collected data on all patients who were admitted to our institution because of this rare tumor over the past 13 years, and thoroughly reviewed the literature for individually documented cases of prostate cancer with rectal involvement to comprehensively analyze patient and tumor characteristics, survival and prognostic factors in these rare tumors.The patients examined in this study were enrolled in the following manner. First, we studied the data on all patients who were diagnosed with rectal involvement of prostate adenocarcinoma at the Tianjin Medical University Cancer Center and Institute between January 1990 and April 2013. These cases were retrospectively reviewed on the basis of histological examination of a surgical specimen or tissue biopsy. Rectal cancer patients with metastasis to the prostate or with synchronous and metachronous adenocarcinoma of the prostate and rectum were excluded. In all cases in our cohort, the diagnosis of prostate cancer with rectal involvement was determined by rectal biopsy and/or prostate biopsy or postoperative histology. Most of them were confirmed with immunohistochemical staining for prostate specific antigen (PSA) or prostatic acid phosphatase (PAP). This study was carried out in accordance with the principles of the Helsinki Declaration and approved by the Institutional Review Board of Tianjin Medical University Cancer Institute and Hospital. Written informed consent was obtained from all of the study participants.Second, a systematic search of the literature was performed to identify all relevant articles dealing with prostate cancer with rectal involvement. First, a computerized search was made of the electronic databases PubMed (1990 to 2013) and google scholar (1990 to 2013) with the following Mesh headings and key words: “prostate/prostate neoplasms/prostate cancer” in combination with “rectum or rectal involvement or invading”. The search was exploded using the “related articles” term in PubMed. The abstracts of all identified articles were studied and selection was based on their relevance for the subject. The reference lists of selected articles were then screened systematically for additional studies of interest. No restrictions for language were applied. Articles containing duplications of case descriptions were excluded. In addition, patients with prostate special pathological type, including sarcoma, squamous cell carcinoma,small cell carcinoma and so on were excluded because of the different tumor biology in comparison with prostate adenocarcinoma. We reviewed the titles to include case reports or review articles on or case series of prostate cancer invading rectum. Altogether, 35 reference articles were suitable for further review. All 35 articles were reviewed to identify patients with prostate cancer invading rectum.Clinical data including patient demographics, clinical characteristics, extent of disease , type of rectal involvement, treatment modalities, and follow-up on these patients were extracted by 2 authors, then crosschecked to ensure accuracy, and finally entered into a database. Some factors had higher missing values because secondary data were collected in the literature review. Clinical data at our institute were determined by reviewing their medical records. Type of rectal involvement was defined according to the criteria described in the literature ,12. OverData on patient demographics, tumor characteristics, extent of disease, treatment modalities were summarized using descriptive statistics. The associations between these factors and survival were evaluated using the univariate Cox regression models. The hazard ratio of death and its 95% confidence interval were calculated for each factor. Multivariate Cox proportional hazards regression was conducted to investigate independent predictors of survival. Variables were entered in the multivariable model when the P value was less than 0.05 in univariate Cox analysis and have complete follow up information. The median survival time was calculated using the Kaplan–Meier method. In order to present survival distribution by significant variables, survival curves were generated using the Kaplan–Meier method. Generally, p < 0.05 (two-sided) was considered statistically significant. Statistical analyses were performed with SPSS version 16.0 .Analysis of the records from the past 13 years in the pathology database of our hospital revealed that only 8 patients with histologically documented prostate cancer with rectal involvement fulfilled the inclusion criteria. After reviewing all medical records, 3 patient was excluded since pathology revealed prostate sarcoma, squamous cell carcinoma, small cell carcinoma respectively. The remaining 5 patients were eligible for the review of the prostate cancer with rectal involvement. After reviewing the 35 articles previously described in “Patient population,” we found 22 suitable articles with clinical description of prostate cancer with rectal involvement. Of these 22 suitable reference articles, 11 case reports -10,13-20The clinical characteristics of the 5 patients enrolled from our institute are summarized in Table At diagnosis, a high proportion of patients (63.77%) showed extrarectal PCa metastases, and PCa with rectal involvement only were found only in 25 (36.23%) cases. 21 patients (22.3%) were misdiagnosed preoperatively as rectal cancer, concomitant rectal and prostate cancer, or bladder cancer in 16, 3 and 2 patients respectively.The appropriate treatment was given on an individual basis according to the severity of the clinical symptoms, preoperative diagnosis, organs involved, hormonal status at diagnosis of rectal involvement, patient's age and patients’ preferences. 19 of these patients (20.2%) have accepted diverting colostomy for rectal occlusion. Treatment of primary prostate cancer for the patients in the hormone relapsed group consisted of hormonal therapy, either with luteinizing hormone-releasing hormone (LHRH) agonists or orchiectomy or combined with antiandrogens. 1 patient underwent TURP, and 7 patients received radical prostatectomy or curative radiation therapy. After diagnosis of rectal involvement at hormonally relapsed disease, 3 patients were given chemotherapy. 4 of these patients received palliative radiotherapy due to rectal bleeding or local pain, 1 patient received total pelvic extenteration (TPE) and 3 patients were given abdomino-perineal excision of the rectum because of misdiagnosis as a carcinoma of the rectum.All of the 63 patients with hormone-untreated prostate cancer were treated with hormonal therapy, either with luteinizing hormone-releasing hormone (LHRH) agonists or orchiectomy or combined with antiandrogens. Among these who underwent hormonal therapy, part of the patients (31.7%) also underwent different types of local therapy, 12 of these patients were given TPE, 9 patients received radiotherapy and rectum resection were given because of misdiagnosis as a carcinoma of the rectum. Among the 12 patients who underwent TPE, 2 died of prostate cancer at 51 and 81 months respectively, 2 died of other disease at 1 and 70 months respectively.5 patients were still alive with disease-free status for 8,10,24,36 and 96 months; 3 patients were still alive with biochemical or clinical progression at 13,13 and 45 months. Interestingly, TPE combined hormonal therapy give us six of thirteen long-term survivors for more than 3 years in this series. Surprisingly, most of these diseases were extrarectal disease, only 1 with rectum only disease. In addition, 6 patients underwent neoadjuvant and adjuvant hormonal therapy, while another 7 patients received adjuvant hormonal therapy.Of the 94 patients included, data was available on the survival status of 87 patients. The data and the hazard ratios with overall survival of demographics, clinical characteristics, tumor characteristics, treatment for all the 87 cases are shown in Table 11 patients in the group of synchronous rectal involvement at first cancer diagnosis (n = 58) and 23 patients in the group of metachronous diagnosis of rectal involvement (n = 29) died at the latest follow up. Since more than half of the patients are still alive it is not possible to calculate survival accurately, but it is estimated by the Kaplan-Meier method that the median overall survival after diagnosis of rectal involvement will be about 48.0 months (95% CI 22.3-73.7). The estimated overall survival rate (% ± SE) at 1, 3, and 5 years were 68.3 ± 5.3%, 54.4 ± 7.2%, and 38.1 ± 11.1%, respectively  were misdiagnosed preoperatively as rectal cancer, concomitant rectal and prostate cancer, or bladder cancer in 16, 3 and 2 patients respectively. The definitive diagnosis was finally established by postoperative histopathological work-up. In these cases, the preoperative misdiagnosis poses the therapeutic decision-making a greatest problem. Although the levels of PSA, radiographic appearance, and clinical presentation may be helpful in distinguishing the two entities, histological examination of the lesion may be the only method of making the correct diagnosis. It may be difficult to differentiate rectal adenocarcinoma from prostatic adenocarcinoma in small rectal biopsy specimens, although it is generally not a problem in operation specimens ,39. The Traditionally, prostate carcinoma presenting with rectal involvement represents an advanced stage of a highly invasive form of the disease. The prognosis is poor. Bowrey et al. retrospectively described the outcome of six prostate cancer patients with rectal infiltration and reviewed the literature for 1966–2002. The median survival for the 86 patients with outcome data reported was 15 months (95% CI 14–16 months). Survival beyond 30 months was rare . While, As expected, hormonal status at presentation had a major impact on overall survival in PCa patients with rectal involvement on both univariate and multivariate analysis. In the present study, patients previously untreated by hormones had a relatively good prognosis with an estimated median survival of 70 months , the estimated 1, 3, 5 year survival rate were 88.9%, 78.6% and 55.0%, respectively, and half of them were still alive at the latest follow-up, which is at least equivalent to patients presenting with minimal or extensive metastatic prostate cancer treated with androgen deprivation in large series . PatientRectal involvement comprises a multitude of differing clinical presentations. Unless one patient present asymptomatic during a routine screen for occult metastases, all other patients present with specific symptoms. In our series, patients presented with rectal bleeding had significantly poor survival on univariate analysis (p = 0.043). Furthermore, the presence of rectal bleeding is an independent prognostic factor predicting for worse prognosis . It is unclear whether the tumors of patients presenting with rectal bleeding have a different biology. Bleeding may indicate tumors of larger luminal surface, erosions, or highly vascular tumors.An interesting finding from this study is that Asian patients appeared to predict better overall survival than non-Asian patients on univariate analysis (p = 0.001). The results support a previous report that many Asian prostate cancer patients had better survival than whites . The surRectal involvement are often a sign of more widespread systemic disease, considering the sixty-four percent of patients had extrarectal metastases at the diagnosis of rectal involvement. Patients with extrarectal metastasis disease were associated with poor overall survival in comparison with patients who had singly rectal involved disease . Multivariate analysis was not performed because of the incomplete follow up information for this covariate, so whether it is an independent prognostic factor deserves further evaluation. However, Stage IV subcategory has been identified as independent predictor of overall survival in stage IV prostate cancer . ProstatThe role of curative surgery in treating prostate cancer patients with rectal involvement has been controversial. Historically, TPE has been generally unsuccessful when used as a curative approach to treat locally advanced prostate cancer in several small-sample case series and the role of TPE was limited to palliate debilitating perineal pain and other local symptoms, such as haematuria, urinary obstruction, and urinary and rectal incontinence . SpauldiThis study had several limitations. Due to the extreme rarity of prostate cancer with rectum involvement, we could obtain the data for only 5 patients who were admitted over the last 13 years to our hospital. Hence, we collected all available cases from the literature and found an additional 89 cases. Because of the retrospective nature of data collection, the limited information provided in the literature, and short follow-up period with high censoring rate, the statistical results should be interpreted cautiously. Therefore, the absence of association of some factors with survival could be attributed to the small case numbers which may result in insufficient statistical power. Nevertheless, the findings of this study are still valuable for better understanding the disease processes of prostate cancer with rectal involvement and the factors affecting the survival of patients with this rare disease.In conclusion, our findings suggest that rectal involvement does not necessarily predict a worse outcome when presenting as a previously hormone-untreated disease and that the prognosis was worse when presenting as a hormonally relapsed disease. Although limited by the short follow-up period with high censoring rate, our study indicates that prior hormone therapy and rectal bleeding were independently associated with a significantly poor overall survival in prostate cancer patients with rectal involvement. TPE combined with hormonal therapy appears to confer better overall survival in hormonally untreated patients. It is also necessary to determine who can benefit from TPE for prostate cancer with rectal involvement patients.The authors declare that they have no competing interests.HTW and BGL: acquired the data and draft the manuscript. HTW and YHY performed the statistical analysis. All authors read and approved the final manuscript.


NDVI, fine particles content and soil water content had positive direct effects on soil organic C and total N stocks. Soil EC affected soil organic C and total N stocks mainly through its indirect negative effect on NDVI, soil texture, and water content. NDVI, soil texture, and moisture also indirectly affected soil organic C and total N stocks via changes in each other. These indirect effects augmented each other, although in some cases indirect effects worked in opposing directions.Grassland salinization causes considerable changes to soil and vegetation, which can lead to changes in soil organic carbon (C) and total nitrogen (N). These changes have complex causal relationships. A significant correlation between soil organic C and total N and any soil or vegetation property does not necessarily imply a significant direct effect of the property on soil organic C and total N. In this study, a field survey was conducted to investigate the changes in soil organic C and total N in grassland along a salinity gradient in Hexi corridor, China, and the direct and indirect effects of soil and vegetation properties on both stocks were quantified using a path analysis approach. Significant decrease in soil organic C and total N contents were observed with increasing salinity. Both had significant positive correlations with the Normalized Difference Vegetation Index (NDVI), soil water, and fine particles (silt+clay) content ( Such variables include climate, vegetation, topography, intrinsic soil properties, land use and management practices, etc. Because of the close relationships between soil and vegetation properties and soil organic C and total N contents, efforts have been made to predict soil organic C and total N contents from these variables using various combinations. It is important to determine soil and vegetation properties that have significant direct influence on soil organic C and total N. This information enables the estimation of soil organic C and total N using appropriate soil and vegetation properties contained in (i) routine soil survey databases ; and (ii) routine vegetation database such as the Normalized Difference Vegetation Index (NDVI) and ecosystem productivity. Therefore, the objectives of this study were: 1) to examine changes in soil and vegetation properties and soil organic C and total N contents in the process of grassland salinization; and (2) to examine the direct and indirect effects of vegetation and soil properties on soil organic C and total N stocks.Linze Grassland Ecological Test Station  is a department of Lanzhou University. This study was approved by the College of Pastoral Agriculture Science and Technology, Lanzhou University.This experiment was conducted in Linze Grassland Ecological Test Station  in the middle reaches of the Hexi Corridor region at an average altitude of 1400 m above sea level in Gansu Province, PR China. The station is located at the southern edge of the alluvial fans of the Heihe River. The Heihe River runs down from the Qilian Mountains; melt water from glaciers and snow cover is the principal source of surface runoff. The ground water table is at a depth of 0–2.5 m, with periodic rises during the year. The climate in this region is characterized as temperate continental arid monsoon, dry and hot in the summer and cold in the winter. The annual mean precipitation is 121.5 mm, of which 61% is received in summer and autumn; the annual mean evaporation is over 2,338 mm. Annual mean temperature is 7.1°C, with the absolute maximum up to 38°C and minimum –28°C. Salt-affected grasslands are estimated to encompass 1.4 million ha, accounting for 79% of the total salt-affected soils in Hexi corridor, China Phragmites communis and Legmus dasystachys. Species unique to specific grassland types are Kalidium gracile to the moderately salinized grassland; Achnetherum splendens to the heavily salinized grassland; and Nitraria tangutorum to the severely salinized grassland For the present work, a space-for-time substitution approach was used 3) at three depths , with five replicates. All samples were collected in August 2011.Five soil samples were collected and combined at each plot from 0–30 cm depth. Soil samples were placed in sealed plastic bags and brought to the laboratory. Soil bulk density was determined at each site using a core sampler (stainless steel cylinder with a volume of 100 cmred) and near-infrared (NIR), a floating point raster was also generated for each scene containing values for NDVI NIR is the reflectance signal in the near-infrared (band 4 in Landsat TM) and red is reflectance in the red band (band 3 in Landsat TM)Landsat TM 30-m resolution imagery for the studied plots was acquired during a cloud-free day: August 1 in 2011. Using bands from the red . The second part was further sieved to 0.25 mm and used for determining average organic C and total N. Soil particle size distribution was determined by the pipette method in a sedimentation cylinder, using Na-hexamethaphosphate as the dispersing agent e(N+1) transformed for normality. All data were analyzed using DPS software.Because data were not normally distributed, the Kruskal-Wallis test was used to determine the differences among the four grassland types. Pearson correlation analysis was used to study the relationships between the corresponding variables. Aside from this, path analysis was performed to investigate the direct and indirect effects of vegetation and soil properties on soil organic C and total N stocks P<0.05). Although bulk density did tend to increase with salinity, it was not as sensitive to the different classes as were NDVI and soil parameters. No changes in bulk density were found among the remaining three grassland types except for the lightly salinized grassland.As shown in Compared to the lightly salinized grassland, soil EC and sand content increased by 449.7% and 136.4% , and NDVP<0.05) with increasing salinity  with the progression of salinization. Compared to the lightly salinized grassland, even in the moderately salinized grassland the loss of organic C and N reached 1.46 kg/m2 and 0.11 kg/m2, respectively, while in the severely salinized grassland losses were 3.39 kg/m2 and 0.31 kg/m2.As shown in Path analysis showed differential direct and indirect effects of vegetation (NDVI) and soil properties on soil organic C and total N stocks . NDVI anNDVI is a commonly used and easily calculated satellite image-based proxy for vegetation productivity Additionally, soil physical properties can be affected. A soil’s ability to percolate water (permeability and infiltration), how much water the soil can store (available water holding capacity), and the soil’s ability to adsorb or desorb chemical ions (exchange capacity) properties are all strongly related to soil texture Soil and vegetation resources are closely correlated, soils influence plants, and plants affect the characteristics of soils inversely Soil organic C and total N contents were closely correlated to soil texture, water content, EC and vegetation productivity , which hThe direct effects of biophysical properties found in path analysis were the positive effects of NDVI, fine particles (%) and soil water content (%) on soil organic C and total N stocks. Plant traits clearly affect soil organic C and total N cycling Soil texture also has a strong effect on the soil’s capacity to accrue C; numerous studies in rangelands Soil moisture, as an abiotic driver of soil organic matter dynamics, is well studied Path analysis also showed that, in the salinized grasslands, the main influence of soil EC on soil organic C and total N stocks was indirect through its negative effect on NDVI, soil texture, and water content. Even though simple correlation analysis indicated that the correlation between soil organic C and total N stocks and soil EC was significant, the direct effect of soil EC was not significant. This further shows that simple correlation analysis is not enough to describe the relationship between soil organic C and total N stocks and soil properties. Meanwhile, the interaction among soil texture, moisture and vegetation showed indirect effects on soil organic C and total N stocks. For example, vegetation property affected soil organic C and total N stocks indirectly through its positive effect on fine particles, while it affected fine particles indirectly through its positive effect on NDVI and soil water content . These iThe observed patterns in soil organic C and total N stocks in salinized grasslands depend on complex relationships among vegetation and soil factors whose influence is difficult to isolate. Although the combination of good statistical technique with previous knowledge of the system and new ecological theories allows us to better understand this ecosystem