Salmo trutta) and the development of periphyton, grazer macroinvertebrates, terrestrial invertebrate inputs, and drift in twelve 20 m long enclosed stream reaches in a five-month-long experiment in a boreal coniferous forest stream. We found that light intensity, which was artificially increased 2.5 times above ambient levels, had an effect on grazer density, but no detectable effect on chlorophyll a biomass. We also found a seasonal effect on the amount of drift and that the reduction of terrestrial prey input, accomplished by covering enclosures with transparent plastic, had a negative impact on the amount of terrestrial invertebrates in the drift. Further, trout growth was strongly seasonal and followed the same pattern as drift biomass, and the reduction of terrestrial prey input had a negative effect on trout growth. Diet analysis was consistent with growth differences, showing that trout in open enclosures consumed relatively more terrestrial prey in summer than trout living in covered enclosures. We also predicted ontogenetic differences in the diet and growth of old and young trout, where we expected old fish to be more affected by the terrestrial prey reduction, but we found little evidence of ontogenetic differences. Overall, our results showed that reduced terrestrial prey inputs, as would be expected from forest harvesting, shaped differences in the growth and diet of the top predator, brown trout.Subsidies of energy and material from the riparian zone have large impacts on recipient stream habitats. Human-induced changes, such as deforestation, may profoundly affect these pathways. However, the strength of individual factors on stream ecosystems is poorly understood since the factors involved often interact in complex ways. We isolated two of these factors, manipulating the flux of terrestrial input and the intensity of light in a 2×2 factorial design, where we followed the growth and diet of two size-classes of brown trout ( Forests adjacent to streams and rivers potentially have a great impact on the aquatic community as forests not only provide a substantial energy base for the lotic communities, i.e. FPOM/CPOM, woody debris and invertebrates, but also influence solar radiation, flow regime, nutrient runoff and temperature. As small streams have a high surface area to volume ratio and a high degree of shading these are particularly affected by surrounding forests Salmo trutta) are light and terrestrial invertebrate input. These factors affect forest-dependent energy pathways to streams, and the strength of them depends on both the extent and structure of riparian tree vegetation Modern forestry practices have changed over the last 40 years, with a growing environmental awareness, where the use of harvesting practices that entail clear-cutting down to the stream edge has decreased Parallel with studies reporting that forested riparian zones generally have higher inputs of falling terrestrial invertebrates than unforested riparian zones The factors influenced by the structure and composition of the riparian zone are rarely permanent in their ability to drive stream ecosystem processes; instead they often show seasonal variation. For example, the flux of terrestrial inputs into streams is seasonally variable Temporal variability in a resource subsidy such as terrestrial invertebrates may have population level consequences, affecting for example fitness, growth or intraspecific competition Salmo trutta), 2) the biomass of benthic algae, 3) the density of invertebrate grazers, and 4) the potential food resources of brown trout, i.e. composition and biomass of drift and terrestrial invertebrate input. This was done in a five-month-long field enclosure experiment. Our overall hypothesis was that trout growth and diet would be affected by both increased light level and reduced terrestrial invertebrate input. More specifically we hypothesized that increased light level would increase biomass of benthic algae or grazer abundance, which in turn would affect the diet and growth of brown trout. We also hypothesized that reduced input of terrestrial invertebrates would 1. lead to a decreased amount of terrestrial prey in the drift, 2. induce decreased trout growth rates and that these reductions would show ontogenetic differences, with larger trout being more influenced than smaller due to size-dependent foraging strategies, and 3. vary over the season, having its largest impact on trout growth during summer when terrestrial invertebrate inputs were expected to peak.Light level and terrestrial subsidies represent two important factors that link streams with their surrounding forests, and both of these factors are affected by forest harvesting. Consequently, we wanted to manipulate these two factors to isolate how two factors associated with forest harvesting affect stream biota. Here, we examine the effects of increased light levels and reduced terrestrial invertebrate input on a brown trout population in a small coniferous forest stream. Specifically, we examined the 1) growth and diet of two size classes of brown trout  by the Gothenburg Ethical Committee.Picea abies and Pinus sylvestris, although there are some deciduous species present such as Betula pendula and Alnus glutinosa. The stream is oligotrophic , with a pH of 6.9–7.8. Brown trout  is the only resident fish in the stream. It should be noted that the year prior to the experiment (2005) a severe drought extirpated most of the fish population. Before starting the experiment the stream was electrofished to ensure that no fish were present in the experimental area.The experiment was conducted in a ∼700 m long section of Sundtjärnsbäcken, a first order lake outlet stream situated in the Glaskogen Forest Reserve in central Sweden . The stream is a typical forest stream for the region, with a stony, gravel bottom and an average width of 1–1.5 m. The riparian forest provides a dense canopy over the stream (>70%) and is dominated by coniferous R: reduced terrestrial prey input and ambient light level), (3) Light enhancement  and finally (4) Reduced terrestrial input and enhanced light level applied together (TRL). The replicates of each treatment were semi-randomly distributed, with the stipulation that replicates of the same treatment could not be placed adjacent to each other. As a result the replicates of the different treatments were well distributed along the entire length of the stream  Unmanipulated  Terrestrial input reduction  were programmed via electronic timers providing a daily light/dark photoperiod. To avoid undesired attraction of terrestrial invertebrates to the lamps the timers were set to switch on the light one hour after dawn and switch off one hour before dusk. To correct for changes in day length over seasons all timers were manually re-programmed on a weekly basis.To increase light levels, we used 8 lamp houses (150×30×10 cm) equipped with two fluorescent tubes  per lamp in each enclosure. The fluorescent tubes imitate photosynthetic active radiation with wavelengths mainly between 400 and 700 nm. The lamp houses were hung, c. 50 cm apart and 60 cm above the stream bottom, from the wooden frames. Originally the lamps were hung about 40 cm from the stream bottom but due to spring floods at the end of May the lamps had to be raised 20 cm. By doing so the amount of PAR (photosynthetic active radiation) reaching the water surface decreased from about 65 to 45–50 µmol mMean daily air temperature inside and outside one of the tents, as measured with two data loggers, was similar . Similar−2, which was well within the range found in similar streams in the surrounding area. Two size groups, hereafter referred to as young and old fish, were used to represent the natural population structure and to examine the size-dependent effects of terrestrial prey on trout growth and diet. Young and old fish had a mean length of 79 (±0.6 SE) and 137 (±3) mm and a mean biomass of 4.7 (±0.1) and 23.9 (±1.41) g, respectively. Altogether 11–13 young and 5–6 old fish were translocated to each enclosure, depending on the area of the enclosure and the size of the fish. Separate one-way analyses of variance (ANOVA) did not reveal differences among the treatments for length, weight and density variables for young or for old fish  and weight (0.01 g) and individually marked with PIT-tags (TROVAN ID-100). They were held for 24 h before stocking them into the enclosures on 17 May 2006 at a pre-calculated density of ∼0.70 trout mSampling occurred on five occasions: 10–11 May, 19–20 June, 1–2 August, 2–3 September and 8–9 October. However, terrestrial invertebrate input was also sampled once between each of these occasions  and light levels (PAR) were measured weekly.2 per trap) were used in each enclosure, spaced to proportionally sample the whole enclosure area. The bottom of the traps was covered with approximately 2–3 cm of water together with a few drops of detergent. After 72 h the content of the traps was sieved with a 500-µm net and preserved in 70% ethanol. All samples were treated separately for later laboratory analyses.Input of terrestrial invertebrates was measured using bucket-traps suspended over the stream. Six traps (51 cm−2 s−1) was measured between 10:00 and 14:00 h under a range of weather conditions using a light meter . At each sampling, nine measurements were taken directly above the water surface in each of the 12 enclosures. These were taken at two meter intervals along the entire length of the enclosure, alternately from the left side, middle and right side of the enclosures.Photosynthetic active radiation .Three weeks before the start of the experiment 350 glazed ceramic tiles (5×5 cm) were incubated in the stream. Although unglazed tiles typically have higher colonization rates than glazed tiles −3) was calculated from measurements of water velocity (Höntzsh μP-flowtherm) at the center of each net, the width of the net and the mean water depth at the sampling spot. Drift samples were immediately preserved in 70% ethanol and treated separately for subsequent laboratory examinations. Because drift density was low on most sampling occasions, day and night drift data were later pooled to give a more robust estimation.Invertebrates drifting into the enclosures were sampled at the upstream end of each enclosure using a drift net . The nets were deployed twice for 60 min, in the morning and right after sunset. Drift density (g mBenthic grazers were collected using a Surber sampler (20×20 cm). Six samples were collected from randomly selected positions in each enclosure on each occasion. The samples were sorted in the field and preserved in 70% ethanol and treated separately for laboratory examination. Benthic grazers were determined at the highest taxonomic resolution possible 1 is the initial weight (g) at time t1, and W2 is the weight measured at time t2.Fish were collected with a single pass, or occasionally, with two-pass electrofishing, resulting in 100, 94, 77, 89 and 66% of the fish being sampled on the different sampling occasions. Captured fish were anaesthetized with MS-222 and measured for standard length (mm) and body mass (0.01 g). Stomach contents were obtained from all captured fish by stomach flushing, which is an effective and non-lethal method to examine the diet of salmonids a content, grazer abundance, drift biomass, proportion of terrestrial drift, and specific growth rate of trout. Because growth rates of trout differ between age groups (i.e. younger fish grow faster), we ran separate repeated measures ANOVAs for the two age groups. All variables had to be log (x+1) transformed before the analyses to satisfy assumptions of normality and reduce heteroscedasticity, except the proportion of terrestrial drift and proportion of terrestrial prey in the diet, which were arcsine square root transformed. Analyses were carried out with either the program STATISTICA or SPSS version 18.0–19.0.We used separate repeated measures two factorial ANOVAs with time as the repeated measure and terrestrial prey (reduction vs. no reduction) and light (enhanced vs. ambient condition) as categorical predictors, to test the effects of these factors on chlorophyll a priori defined groups (reduction and no reduction treatments for the two age classes). The CAP analyses were run based on relative biomass data, using Hellinger distances Due to empty stomachs, sample sizes for old fish were too low to perform a repeated measure ANOVA on diets for the entire data set. Instead we investigated differences in trout diet with canonical analysis of principal coordinates (hereafter CAP). This recently developed constrained ordination method is identical to a non-parametric multivariate analysis of variance −2 d−1 for U and L, and 9.4 (±1.2) and 8.5 (±1.0) mg m−2 d−1 for TR and TRL sections, respectively. Repeated measures ANOVA on the total biomass of these prey indicated significant effects of time , terrestrial input reduction  and a time×terrestrial input reduction interaction , whereas the effects of light and associated interactions were not significant. In terms of taxonomic composition, Diptera, Araneida, Coleoptera and Hymenoptera were the dominant taxa, ranging from 73.3 to 80.6% of the total biomass among the four treatments. Flying aquatic insects were of relatively minor importance, comprising <10% of the biomass.The flux of terrestrial prey input showed large temporal variation in the uncovered enclosures, with a maximum input in July , which cR, L and TRL treatments were 25.6±2.6 (SE), 22.8±1.7, 55.5±2.2 and 59.8±3.0 µmol m−2 s−1, respectively. Repeated measures ANOVA indicated a significant effect of time , light  and a time×light interaction , whereas other effects were not significant.Light intensity varied substantially during the experiment, particularly early on (prior to mid-June) when foliage was developing . Mean PAa biomass varied between 0.26±0.17 (SE) µg cm−2 (U in June) and 0.91±0.03 (TR in August) µg cm−2 during the experiment , light , terrestrial input reduction  or their interactions (P>0.38 for the different interactions).Mean chlorophyll periment . HoweverElmis larvae and adult, Oulimnius larvae and adult, and Nemoura . The number of grazers varied between 103±46 (SE) (U in May) and 326±65 (L in June) and grazer biomass between 0.019±0.009 g/m2 (T in October) and 0.057±0.019 g/m2 (T in June) during the experiment  and light  but no effects of terrestrial input reduction  or any of their interactions (P>0.3). A similar analysis based on biomass showed no significant effects of time, light, terrestrial input reduction or any of their interactions (P>0.2), probably due to the relatively few but heavy molluscs masking any biomass effect of other taxa.The grazers were dominated by Orthocladinae, periment . Repeate4,32 = 4.84; P = 0.004) but no effects of light , terrestrial reduction  or interactions (P>0.5 for the different interactions). However, repeated measures ANOVA on the proportion of terrestrial drift showed an effect of terrestrial reduction , but no effect of time , light  or any of the interactions (P>0.2 for the different interactions). Enclosures with terrestrial reduction had a lower proportion of terrestrial organisms in the drift than other enclosures , whereas the terrestrial part was dominated by Hymenoptera, Diptera, Coleoptera and Araneae . Drift abundance showed a strong temporal pattern , with a closures .−1 for young fish and 1.1% day−1 for old fish , but not for the August–September period . There was also a significant effect of light in May–June for young fish . Consequently, the effect of terrestrial reduction varied among time periods for young trout, whereas it was consistent over time for old trout.Growth rates of both young and old trout were, just as total drift biomass, highest in May–June, with mean values of 2.2% dayold fish . During old fish . Repeateold fish . For youold fish . BecauseSimulium larvae and Dysticidae larvae, and the most common terrestrial prey were Lumbricidae, adult terrestrial dipterans and Arachnidae. All of these prey items occurred in 15% or more of the analyzed stomachs.Diet of trout contained a broad diversity of prey of both aquatic and terrestrial origin. In total 79 taxa were identified in the diet during the whole season. The most common aquatic prey were chironomid larvae and pupae, Lumbricus spp in the open enclosures than in the covered enclosures, where instead Chironomidae and Simulium spp larvae dominated the diet , but such an effect was not observed. Instead, we found a higher abundance of grazers in our enclosures with increased light levels, which is consistent with studies showing that increased periphyton productivity may become directly transformed into grazer biomass We had also expected an effect of light on the development of periphyton (i.e. chlorophyll −2 s−1R enclosures) to 55–60 µmol m−2 s−1 (as our L and TRL enclosures) biomass to the increased light levels could be related to that the absolute light levels in the manipulated enclosures were too low, or that the stream was nutrient-limited instead of light-limited. Photosynthetic-irradiance measurements suggest that photosynthesis by stream periphyton is saturated between 100 and 200 µmol mWe had expected that there would be ontogenetic differences in the growth and diet of trout in response to reductions of terrestrial subsidies. This is because numerous field studies have shown that there is a positive correlation between body size and the relative contribution of surface-drifting prey to the diet of salmonids, The diet of trout also showed small differences between young and old trout. In June the diet of old trout was dominated by terrestrial and flying aquatic adult prey in all enclosures, with and without reduced terrestrial subsidies, whereas the diet of young trout was dominated by aquatic prey when terrestrial subsidies were reduced. In August the diet of large trout, and to a somewhat smaller extent the diet of small trout, was dominated by terrestrial prey items only in enclosures with unmanipulated terrestrial subsidies. The response by both age classes in August could be a response to the overall low availability of terrestrial invertebrates at this time. Thus, our results on growth and diet indicate that the expected effects of across-habitat resource subsidies on populations were strong, but ontogenetic differences were not as strong as we expected.Although explored in the field (Nakano & Murakami 2001), the relationship between seasonally variable within-habitat food resources and across-habitat resource subsidies has not received much experimental attention. In streams aquatic invertebrates are typically abundant in spring and early summer, followed by a reduction in availability, largely due to the emergence of many aquatic invertebrate taxa Given that both aquatic resources and terrestrial subsidies vary seasonally, the growth of salmonids in the recipient habitat is also expected to vary seasonally The present study indicates that there is an effect of terrestrial input on trout growth and such effects have not been experimentally examined previously in coniferous forest streams. Furthermore, our study indicates that the effects may be similar to those reported from deciduous forest streams Table S1Trout diet. Seasonal changes in the frequency of occurrence (O%) and mean relative biomass (A%) of the most important food items in the diet of young (Y) and old (O) trout in unmanipulated (U), light (L), terrestrial invertebrate reduction (TR) and terrestrial invertebrate reduction and light (TRL) treatments throughout the study period. Chironomidae* refers to all taxa except Tanypodinae.(DOCX)Click here for additional data file.


Brain derived neurotrophic factor (BDNF) has been implicated in memory, learning, and neurodegenerative diseases. However, the relationship of BDNF with cardiometabolic risk factors is unclear, and the effect of exercise training on BDNF has not been previously explored in individuals with type 2 diabetes.Men and women (N = 150) with type 2 diabetes were randomized to an aerobic exercise (aerobic), resistance exercise (resistance), or a combination of both (combination) for 9 months. Serum BDNF levels were evaluated at baseline and follow-up from archived blood samples.all, p>0.05). Similarly, no significant change in serum BDNF levels was observed following exercise training in the aerobic , resistance , or combination groups  compared to the control group . However, reductions in waist circumference were directly associated with changes in serum BDNF following training .Baseline serum BDNF was not associated with fitness, body composition, anthropometry, glucose control, or strength measures (Serum BDNF was not associated with fitness, body composition, anthropometry, glucose control, or strength measures at baseline. Likewise, serum BDNF measures were not altered by 9 months of aerobic, resistance, or combination training. However, reductions in waist circumference were associated with decreased serum BDNF levels. Future studies should investigate the relevance of BDNF with measures of cognitive function specifically in individuals with type-2 diabetes. Brain derived neurotrophic factor (BDNF) is a protein encoded by the BDNF gene. It is a neutrophin  involved in neuronal plasticity, differentiation, and survival in the central and peripheral nervous system Conversely, other studies have found the opposite relationship, noting elevated serum BDNF levels in individuals with type 2 diabetes Exercise training increases fitness 1C in men and women with type 2 diabetes. The primary outcome measure of this ancillary study is the change in serum BDNF levels following exercise training. From the parent HART-D study of 262 sedentary (30–75 years) adults with type 2 diabetes presenting with an HbA1C of 6.5% to 11.0%, serum BDNF values were examined in a subset of controls and adherent exercisers (n = 168) from which a total of 150 participants were included in the final analysis. A consort diagram is shown in 2, blood pressure 160/100 mmHg or higher, fasting triglycerides 500 mg/dL or higher, use of an insulin pump, urine protein greater than 100 mg/dL, history of stroke, advanced neuropathy, retinopathy, or any serious medical condition that prevented adherence to the study protocol or was a contraindication for exercise training. The institutional review board at Pennington Biomedical Research Center approved the protocol annually and all participants provided written informed consent before initiating the any of the study protocols.The full methodology of the exercise training program used in the Health Benefits of Aerobic and Resistance Training in Individuals with Type 2 diabetes (HART-D) has been reported in the main outcomes paper Participants were randomized to one of four groups: a non-exercise control group, aerobic training only (aerobic), resistance training only (resistance), or a combination of resistance and aerobic training (combination). The non-exercise control group was offered weekly stretching and relaxation classes and were asked to maintain their current activity during the 9-month study period. Accordingly, the compliance of the control group was assessed with step counters to confirm continued sedentary behavior. All exercise sessions were supervised by study staff in our exercise training laboratory.The exercise intensity for aerobic exercise training was 50% to 80% of maximal oxygen consumption. We estimated that 150 minutes of physical activity per week  is equivalent to approximately 10 to 12 kcals/kg of body weight per week (KKW) (5). The selected dose of exercise for the aerobic group was 12 KKW and 10 KKW for the combination exercise group. Participants were weighed weekly to calculate the energy expenditure target for each participant. American College of Sports Medicine equations were used to estimate caloric expenditure rate and time required for each session (1).Participants in the resistance training group exercised 3 days per week with each session consisting of 2 sets of 4 upper body exercises , 3 sets of 3 leg exercises , and 2 sets of each abdominal crunches and back extensions. Each set of resistance exercise consisted of 10 to 12 repetitions. Individuals in the combination exercise group had two sessions per week with each session consisting of 1 set of each of the 9 resistance exercises. The weight lifted on each exercise was progressively increased once the participant was able to complete 12 repetitions for each set of exercises on 2 consecutive exercise sessions.1C level of 1.0% or higher. With the recommendation of the data safety monitoring board, randomization into the control group was discontinued (5). Thus, fewer participants completed the study in the control group compared to the other randomization groups.The control group was unblinded after a significant number of participants (17.1%) had an increase in HbA2 peak (L/min), relative VO2 peak (mL·kg−1·min−1), and treadmill test duration (min). All muscular strength assessments were performed using a Biodex System 3 dynamometer . Total work was determined as the work accomplished over 30 repetitions of maximal effort.Exercise testing was performed on a treadmill  with respiratory gasses analyzed using a True Max 2400 Metabolic Measurement Cart . Participants self-selected a brisk walking pace at a grade of 0% for 2 minutes, and then the grade was increased by 2% every 2 minutes until exhaustion. Fitness data are reported in absolute VOBody composition and percent body fat were determined using dual-energy X-ray absorptiometry scans performed using the QDR 4500A whole body scanner . Weight was measured with on a calibrated electronic scale . Waist circumference was measured by Gulick tape midway between the inferior border of the rib cage and the superior aspect of the iliac crest at the end of normal expiration.1C was evaluated from venipuncture, and samples were run on Beckman Coulter Synchron LX automated analyzer . At baseline and follow-up, blood samples were collected and processed after an overnight fast.Serum BDNF levels were measured by the Pennington Biomedical Research Center Clinical Research Laboratory at baseline and follow-up from archived blood samples stored at −80 degrees Celsius. Serum BDNF levels were analyzed using a Chemkine BDNF sandwich enzyme immunoassay . The sensitivity of the assay was ±7.8 pg/mL with an inter-assay variation of ±8.5% and an intra-assay variation of ±3.5%. All samples for serum BDNF at baseline and follow-up were run on the same analysis plate in order to eliminate inter-assay variation between pre- and post-intervention pairs. HbAHART-D had separate intervention and assessment teams, and all assessment staff were blinded to participant randomization. The clinical testing and exercise training laboratories were in separate buildings, and participants were reminded frequently not to disclose their group assignment to assessment staff.2) was used to compare baseline characteristics between categorical variables. Multiple linear regression was used to test the trend of baseline serum BDNF levels across quartiles of fitness, body composition, anthropometry, glucose control, and strength measures.All data was analyzed using SAS version 9.1 . The final analysis of the present study was performed in adherent participants  was used to determine the effect of exercise modality on serum BDNF levels with baseline value, age, sex, duration of diabetes, and ethnic group entered as covariates with outliers removed (N = 150). Results are presented as adjusted least square means with 95% confidence intervals (CIs). In addition, we performed the same analysis in all participants (outliers included) (N = 168). Spearman correlations were used to determine the association between the change in serum BDNF following exercise training and fitness, body composition, anthropometry, glucose control, and strength measures in exercisers only (N = 127). A 2; was 57% female; and was ethnically diverse as approximately 35% of our study sample was Black. The average duration of diabetes in the sample was 7.3 yrs. (5.2), and approximately 99% of the population was currently taking diabetes related medication. No significant differences between randomization groups were present at baseline. Additionally, no significant differences in baseline serum BDNF levels between men (18034.5 pg/ml) and women (18048.0 pg/ml) (p = 0.991) or Blacks (18744.8 pg/ml) and Whites (17591.1 pg/ml) (p = 0.407) were observed.Demographic data are presented in all, p>0.05). However, the trend for lower serum BDNF levels across quartiles of treadmill test duration approached significance (p-trend = 0.06). When analyses were performed separately in men and women, similar results were observed.Serum BDNF levels across quartiles of fitness, body composition, anthropometry, glucose control, and strength measures are shown in The effects of exercise modality on serum BDNF levels are shown in Spearman correlations for change in serum BDNF levels following exercise training (exercisers only) with cardiometabolic risk factors are shown in The primary finding from the present study was cardiometabolic risk factors were not associated with serum BDNF levels at baseline in individuals with type 2 diabetes. Furthermore, 9 months of aerobic, resistance, or combination training did not alter serum BDNF levels compared to control participants, but reductions in waist circumference were associated with decreased serum BDNF following exercise training. To our knowledge, this is the first large randomized controlled study comparing the effects of aerobic, resistance, and combination training on serum BDNF levels in individuals with type 2 diabetes.Currently, data are limited that examine the implications of cardiometabolic risk factors on BDNF in individuals with type 2 diabetes. Suwa et al. 1C in Japanese men and women with type 2 diabetes. Differences between our study and previous studies finding an association between BDNF levels and cardiometabolic risk factors in individuals with type-2 diabetes may be due to the differences in the type of sample analyzed for BDNF (plasma vs. serum), differences in study population (HART-D: 34% Black), and possible differences in the use of diabetes medications.In the present study, no associations were found at baseline between fitness, anthropometry, body composition, strength, or glucose control measures and serum BDNF levels in individuals with type 2 diabetes. However, the relationship between serum BDNF levels and longer treadmill test duration approached significance. Our results are supported by data from Fujinami et al. Several studies have demonstrated an increase in serum BDNF levels following acute exercise The present study is the first to our knowledge to evaluate the effects of exercise training on serum BDNF specifically in individuals with type 2 diabetes or to directly compare the effect of different exercise modalities in a large randomized trial. We found no significant change in serum BDNF following aerobic, resistance or combination exercise training compared to control participants. Additionally, we found that reductions in waist circumference following exercise training in exercisers were positively associated with serum BDNF levels, suggesting that reductions in central adiposity may be involved in the regulation of BDNF. The physiologic importance of these findings on cognitive function should be addressed in future investigations since cognitive measures were not collected in HART-D. Changes in fitness, body composition, glucose control, and strength measures were not associated with changes in serum BDNF following exercise training in the present study.The strengths of this investigation include a randomized study design with a large diverse study population. Additionally, all exercise training sessions were supervised by study staff, controlled for caloric expenditure, and all exercise groups exercised for a similar amount of time. Limitations of the present study are that it is a post-hoc analysis from archived blood samples, and we did not collect any measures of cognitive function. Last, since blood samples were collected at only baseline and follow-up, we cannot evaluate the time course of the changes in serum BDNF during the exercise intervention.In conclusion, the results of the present study found that serum BDNF levels at baseline were not associated with fitness, body composition, anthropometry, glucose control, and strength measures in individuals with type 2 diabetes. In addition, no significant change in serum BDNF levels was observed compared to control participants following 9 months of exercise irrespective of training modality. Future research should evaluate the relationship between serum BDNF levels with memory and cognitive function specifically in individuals with type 2 diabetes, and the physiological mechanisms responsible for the association between exercise training related changes in central adiposity and serum BDNF levels.


The effect of subsoiling in fallow period on soil water storage, accumulation of proline, and formation of grain protein after anthesis were determined. Our results showed that subsoiling in fallow period could increase water storage in the 0–300 cm soil at pre-sowing stage and at anthesis stage with low or medium N application, especially for the 60–160 cm soil. However, the proline content, glutamine synthetase (GS) activity, glutamate dehydrogenase (GDH) activity in flag leaves and grains were all decreased by subsoiling in fallow period. In addition, the content of albumin, gliadin, and total protein in grains were also decreased while globulin content, Glu/Gli, protein yield, and glutelin content were increased. With N application increasing, water storage of soil layers from 20 to 200 cm was decreased at anthesis stage. High N application resulted in the increment of proline content and GS activity in grains. Besides, correlation analysis showed that soil storage in 40–160 cm soil was negatively correlated with proline content in grains; proline content in grains was positively correlated with GS and GDH activity in flag leaves. Contents of albumin, globulin and total protein in grains were positively correlated with proline content in grains and GDH activity in flag leaves. In conclusion, subsoiling in fallow period, together with N application at 150 kg·hm Natural precipitation is the only water resource of dryland wheat. Rainfalls always concentrate from July to September at region producing dryland wheat in loess plateau, which is called fallow period. During the fallow period, soil water is replenished and restored. It is the water stored in this period that determines the soil moisture before dryland wheat is sown which has a great influence on the production of dryland wheat Many researches showed protein content in wheat grains would increase when the water was deficient in soil Up to date, problems in production of wheat in dry land were enumerated as follows: 1) Low water-using efficiency caused by poor soil fertility and low ability of soil water storage and retention; 2) Late straw rotting fails to increase the organic matter of soil, and thus, affect the quality of sowing; 3) Although precipitation in fallow period is considerably abundant, a great evaporation loss of surface water fails to store enough water for spring because of the high temperature in summer; 4) Wheat is often sown under enough moisture or rainfall in dry land, so the sowing time is inconsistent. Many researches showed drought can improve the quality of wheat to a certain degree The objective of the present paper was to provide a new way to increase the water storage and retention for dryland wheat. This was realized by subsoiling after wheat harvest and conducting rotary tillage in the late August. The proline content and physiological mechanism about formation of grain protein in dryland wheat were examined. To have a comprehensive understanding for the issue of dryland wheat quality, the effects of fertilizer application were also studied.st July, 2010. Content of organic matter was measured as 8.65 g/kg using potassium dichromate volumetric method. Total nitrogen was measured as 0.74 g/kg using Kjeldahl determination, alkali-hydrolyzable nitrogen was measured as 32.93 mg/kg using alkali-hydrolyzable proliferation method, and rapidly available phosphorus was measured as 20.08 mg/kg using sodium bicarbonate method.The field experiment was conducted at Wenxi experimental site (summer fellow) of Shanxi Agricultural University from 2009 to 2011. The soil fertility was measured on 1The experimental site is very arid and 60%–70% of the annual precipitation is concentrated in summer and early-fall . Precipitation conditions at the experimental site in 2005–2011 were shown in −2 (LN), 150 kg·hm−2 (MN) and 225 kg·hm−2 (HN)). For subsoil plot, the preceding wheat residues were shredded on the fifth day after harvest , followed by tillage that consisted of disking 30–40 cm deep before planting wheat. Then soil moisture was retained by rotary tillage on 20th August, 2010.The experiment was laid out as split-split plot arrangement and three replications. Two levels (subsoil (SS); and no-tillage (CK)) were allocated to the main plots. Sub plots were three levels of N fertilizer application . Phosphate and potash fertilizer had been applied to the field with P2O5 and K2O at a concentration of 150 kg·hm−2.Dryland wheat cultivar Yunhan 20410, which provided by agricultural bureau of wenxi county, was planted on 29Soil water storage was measured by drying method. Soil samples in the 0–300 cm depth (20 cm per soil layer) were obtained at pre-sowing stage, pre-wintering stage, seedling establishment stage, elongation stage, booting stage, anthesis stage, and maturation stage using a soil auger. The calculation formula of soil water storage is as follows:Soil water storage (mm) = [(mass of damp soil − mass of dried soil)/mass of dried soil×100]×soil thickness (mm)×volume weight per layer.Wheatears with consistent growth and blooming on the same day were tagged at anthesis stage and fifteen ears were sampled every five days after anthesis. Flag leaves and grains were kept at −40°C after being quickly frozen by liquid nitrogen for measuring the activity of enzyme. Another five fresh samples, i.e. flag leaves and grains, were taken to measure proline content. The measurement of proline content was referred to Wheatears with consistent growth and blooming on the same day were tagged at anethsis stage and fifteen ears were sampled every five days after anethsis. Grains were separated in the oven where moisture was removed by heating at 105,°C for 30 min and then weighted after drying at 80°C. Grains were used to measure the content of protein and its component after being crushed by micro high-speed universal grinder. Nitrogen content was measured using the method of semi micro Kjeldahl determination and wheat protein content was obtained by multiplying N content (%) by 5.7 Number of grains ears per unit area, average number of grains per wheat ear, and 1000-grain weight were investigated at maturation stage. Fifty plants of wheat were taken from each plot to measure biological yield, and twenty square meters of wheat were rope to measure economical yield.Statistical analysis was conducted using SAS version 8.0 . Analysis of variance (ANOVA) using the General Linear Model procedure and the difference between means using the Duncan test were determined at the α level of 0.05.Generally, as the depth of soil increasing, water storage in the 0–300 cm depth soil before sowing increased at first but then decreased to the minimum at 200 cm and finally tended to increase again . CompareGenerally, as the depth of soil increasing, water storage of soil in anthesis in the 0–300 cm depth increased to maximum at 80 cm, and then decreased between 80 cm to 120 cm, and finally tended to increase slowly again . Table 2In recent years, dryland cultivators have conducted many researches to study the water storage and retention of dryland wheat, and great progress had been achieved In agreement with previous studies, our results also showed that subsoiling (depth 30–40 cm) significantly increase water storage and retention . Subsoil−2. Accordingly, subsoiling in fallow period together with N application at 150 kg·hm−2 were beneficial to root downward growth and absorption of water in deep soil.The present experiment was conducted on a field which was no-tillage in fallow period for many years and performed rotary tillage before sowing. Subsoiling in fallow period broke the bottom of plough layer formed many years which was crucial to water storage and retention, and could increase water storage of 0–300 depth soil at each stage. Especially, the increment in the 60–160 cm depth amounted to more than 10% of the total water storage. It also increased water storage in the 0–200 cm depth soil with low and medium N application until anethesis. However, water storage in the 0–200 cm depth with high N application decreased which was possibly caused by great water consumption due to high biomass of plants. In addition, whether the dryland wheat was subsoiled in fallow period affected the regulatory effect of N application. Under the condition of subsoiling, N application obviously regulated the soil layer with the depth of 40 cm, 60 cm and 120–200 cm. Water storage of soil in the 200–300 cm depth was the least when N application was 150 kg·hmAs subsoiling breaking the plough layer and loosing the soil, fertilizer could also easily reach to the deep soil and used by the plant root in the same layer. And this may heighten the utilization of N to synthesize protein/amino acids and lead to a better wheat productivity. On the other hand, the developed root system would accelerate the absorption of water and nutrition which result in the flourish of wheat leaves. Thus, the photosynthesis is enhanced and more biomass would be accumulated together with a high grain yield.Osmoregulation is an important mechanism of plant adapting to drought −2. In summary, subsoiling in fallow period was proved as an effective technology of water storage and retention. In addition, it was more favorable to improve the quality of dryland wheat grain accompanied with an N application of 150 kg·hm−2.Our study showed subsoiling could increase glublin content, Glu/Gli, protein yield, glutelin content. Grain protein is an important index to evaluate the quality of wheat. Although subsoiling decreased total protein content, the ratio between glutelin and gliadin was increased. Hence, the quality was improved. And the maximum value was obtained under N application 150 kg·hm


Among the impacts of coastal settlements to estuaries, nutrient pollution is often singled out as a leading cause of modification to the ecological communities of soft sediments. Through sampling of 48 sites, distributed among 16 estuaries of New South Wales, Australia, we tested the hypotheses that (1) anthropogenic nutrient loads would be a better predictor of macrofaunal communities than estuarine geomorphology or local sediment characteristics; and (2) local environmental context, as determined largely by sediment characteristics, would modify the relationship between nutrient loading and community composition. Contrary to the hypothesis, multivariate multiple regression analyses revealed that sediment grain size was the best predictor of macrofaunal assemblage composition. When samples were stratified according to median grain size, relationships between faunal communities and nitrogen loading and latitude emerged, but only among estuaries with sandier sediments. In these estuaries, capitellid and nereid polychaetes and chironomid larvae were the taxa that showed the strongest correlations with nutrient loading. Overall, this study failed to provide evidence of a differential relationship between diffuse nutrient enrichment and benthic macrofauna across a gradient of 7° of latitude and 4°C temperature. Nevertheless, as human population growth continues to place increasing pressure on southeast Australian estuaries, manipulative field studies examining when and where nutrient loading will lead to significant changes in estuarine community structure are needed. The sediment-dwelling invertebrates of estuarine and coastal environments provide important ecosystem services. Suspension feeders remove particles and pollutants from the water, helping to improve clarity Due to the importance of sediment-dwelling communities, a large number of studies have sought to determine those factors that influence their density and diversity. These have shown that at small scales of centimetres to meters, sediment granulometry, sediment organics and flow can influence invertebrate communities Within temperate estuaries, anthropogenic nutrient enrichment is broadly regarded as one of the greatest modifiers of sediment-dwelling communities, and their dependent ecosystems Both local and broad-scale environmental conditions might modify the impact of nutrients on benthic communities. At large scales, climatic setting and estuarine geomorphology might influence whether estuarine waters stratify or not, and hence whether bottom-waters can be reoxygenated following bacterial decomposition of excess primary production, stimulated by nutrient enrichment Here we conduct sampling across 16 estuaries of New South Wales, Australia, spanning 7° of latitude, to test the hypotheses that: (1) nutrient enrichment will explain more variation in macrofaunal community composition than other environmental variables, including sediment characteristics, estuarine geomorphology and latitude; but (2) the relationship between nutrient enrichment and community composition will be modified by local environmental variables.To assess how the local environmental context modulates the effects of nutrient loading on the abundance and diversity of macrobenthic invertebrates, we sampled 16 estuaries along the coast of New South Wales, Australia , Fig. 1.2 intertidal sites , situated 50–100 metres apart. All sites were fully marine  and sampling was done within 14 days during low tides in late spring (November 2009). Seven replicate sediment cores (10 cm in diameter and 15 cm deep) were randomly collected from unvegetated sediment at each site for faunal analysis. The number of macrofaunal samples was chosen following results from a pilot study in which the species accumulation plot (PRIMER v6.0) reached a plateau after six replicate cores and following previous studies in New South Wales estuaries that have suggested that this level of replication is sufficient to detect treatment effects of manipulations of organic enrichment Within each of the estuaries, we collected samples of macrofauna and sediment from three 100 mTo assess how the relationship between nutrient enrichment and macrofaunal communities is influenced by the local environmental context, we quantified sediment organic content and grain size. To assess sediment organic content, a subsample of about 4 g was taken from each of the four small sediment samples after homogenization. Coarse woody debris and shell fragments, where found, were excluded prior to analyses. Consequently, samples contained sediment mixed with organic matter. The subsamples were dried at 105°C for 48 h and weighed prior to combustion at 550°C for 4 h. The organic content was calculated as the percentage difference in weight from before to after combustion. Sediment grain size was determined for two randomly selected replicates at each site. The silt/clay fraction was determined by wet sieving through a 63 µm screen. The remaining sediment was dry sieved through a stack of sieves of decreasing mesh size  and the weight of each fraction was measured. Median grain size and sorting were calculated using the software GRADISTAT 4.0 To test the hypotheses that nutrient enrichment would (1) be correlated to macrofaunal community structure and (2) the strength of this relationship would be determined by the climatic and local environmental context, we collated a matrix of environmental variables. This included: 1) site averages of environmental data collected during this study , and 2) estuary physical and chemical attributes  soldier crabs, capitellid and nereid polychaetes, chironomids and lysianassids were the taxa that mostly correlated with the multivariate abiotic data  abundance of macrofauna per site (i.e. 7 sediment cores) was 397±67 and the species richness was 13±1. The most abundant group was the polychaete worms, followed by bivalves, chironomid larvae, crustaceans and gastropods. Macrofaunal community structure was weakly correlated with individual environmental variables . Those itic data . When antic data . The strrrelated . Only soronomids .Estuaries followed a bimodal distribution in MGS, with two peaks at about 170 and 330 µm and a median value of 255 µm. Therefore, in order to minimize the correlation between latitude and MGS, we separated the dataset into two groups of eight estuaries, based on their MGS value , and ran the same multivariate analyses for each subgroup as for the complete data set. The set of low MGS estuaries included seven of the eight northern-most estuaries, plus one in the south. The set of high MGS estuaries included seven of the eight southern-most estuaries plus one in the north.Casco sp., soldier crabs Mictyris spp. and the gastropod Hydrobia buccinoides were the taxa that most closely correlated with the environmental variables of the BEST model (Mictyris spp. were negatively correlated with sediment silt/clay content (Mictyris spp. (Across the low MGS ( = lower latitude) estuaries, the mean (± SE) abundance of macrofauna per site (i.e. 7 sediment cores) was 112±25 and the species richness was 12±1. Multiple regression analysis on low MGS estuaries indicated that sediment variables other than MGS explained most of the variability in macrobenthic assemblages . These wST model . CorrelaST model . NereidiST model . Capitel content . The aburis spp. .Across the high MGS ( = higher latitude) estuaries, the mean (± SE) abundance of macrofauna per site (i.e. 7 sediment cores) was 682±101 and the species richness was 15±1. Macrofaunal assemblage structure was significantly correlated with each of the environmental variables included in the model, except MGS . LatitudCapitella sp. and nereid polychaetes, chironomid larvae and the bivalves Arthritica helmsi and Soletellina alba  that would affect the fate and utilization of N in the system In contrast to studies done in North America or Europe In conclusion, the relationship between nutrient loading and benthic macrofaunal communities varied among sites according to the grain-size of their sediment and was, on the whole, weak. Instead characteristics of the sediment itself, such as grain size and the amount of silts and clays, were much better predictors of macrofaunal communities. Nevertheless, as human population growth continues to place increasing pressure on southeast Australian estuaries, manipulative studies that can ascertain cause-effect relationships are required to examine when and where nutrient loading will lead to significant changes in estuarine community structure, and how changes in flow regime and sedimentation might influence fauna through effects on sediment properties


KIF1B), strongly associated with progression from chronic hepatitis B (CHB) to hepatitis B virus-related hepatocellular carcinoma (HCC) in Chinese population, this study was carried out to explore the role of the genetic variants in KIF1B in the development of chronic hepatitis B.A recent genome-wide association study has identified a new susceptibility locus, kinesin family member 1B gene  were selected and genotyped in 473 CHB patients and 580 controls with no history of CHB. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated by logistic regression model. None of these three SNPs showed association with CHBs after adjusting for age and gender. Equivalence-based method analysis confirmed the absence of association. In the further haplotype analysis, three common haplotypes were observed in this study population, but no significant effect was also found for haplotypes in the progression to CHB.Three KIF1B, was not associated with progression to CHB, implying distinct genetic susceptibility factor contributes to the progression from hepatitis B virus infection to HCC. Nevertheless, further comprehensive analyses are warranted to dissect the mechanism.This study showed the new locus identified for HCC,  IFNG, TNF, VDR, and HLA loci, have been extensively investigated in the progression to CHB, but results were inclusive Hepatitis B virus (HBV) infection is one of most challenging global public health problems, with more than 2 billion people infected KIF1B) at chromosome 1p36.22, which was reported to be associated with HBV-related HCC in six independent populations KIF1B, one SNP rs17401966 from GWAS study Interestingly, a GWAS conducted in Chinese population by Zhang H et al. raised a new susceptibility locus (rs17401966) in kinesin family member 1B gene  and asparate aminotransferase (AST) for more than six months. Controls matched in age and gender were selected from a physical examination in Peking Union Medical College Hospital in 2004, who had no medical history of chronic hepatitis B and HBV vaccination. Subjects who were positive for HBsAg were excluded from the controls. At recruitment, written informed consent was obtained from each subject and personal information from each participant regarding demographic characteristics such as gender, age, and history of HBV vaccination were collected by questionnaire. This study was approved by the institutional review boards of the Chinese Academy of Medical Sciences Cancer Institute and Tongji Medical College of Huazhong University of Science and Technology.Enzyme-linked immunoadsorbent assay (ELISA) was used to detect the serum HBsAg, anti-HBs and anti-HBc .KIF1B gene were identified by bioinformatics analysis. The SNP of rs8019, which may located in the binding site of microRNA, was selected using an integrated bioinformatics tool “SNP Info”(http://manticore.niehs.nih.gov/snpfunc.htm) http://rulai.cshl.edu/cgi-bin/tools/ESE3/esefinder.cgi?process=home) In addition to the SNP of rs17401966 identified by the GWAS in Chinese population, two other candidate SNPs in the intron of Genomic DNA was extracted from peripheral blood lymphocytes. All subjects were genotyped using TaqMan real-time polymerase chain reaction  without knowledge of subjects' infection status. The program was heating to 95°C for 10 minutes followed by 40 cycles of 95°C for 15 seconds and 60°C for 1 minute. The ABI Prism 7900HT Sequence Detection System was used to read the reacted plates and analyze the endpoint fluorescence. To ensure the accuracy of genotyping, a 15% random sample was tested twice by different investigators. The results were concordant for all of the duplicate sets.2 test. Logistic regression was used to evaluate the associations between KIF1B genotypes or haplotypes and the outcome of HBV infection. Odds ratios (ORs) and their 95% confidence intervals (CIs) were calculated after adjustment for age and sex where appropriate. Nonsuperiority test was used to confirm the absence of association between all SNPs detected and progression to CHB The differences in distribution of genotype between CHB cases and controls were tested by χA total of 473 chronic HB patients  and 580 controls  were enrolled in this study, with mean age of 34.13±13.56 years. Both age and gender distribution was comparable between CHB and controls. The baseline characteristic of CHB patients and controls were shown in KIF1B candidate SNPs are shown in The genotype distributions of 2 = 0.595) for rs8019 and rs17401924, 0.921 (r2 = 0.524) for rs8019 and rs17401966, and 0.931 (r2 = 0.853) for rs17401924 and rs17401966 in our study population. Eight haplotypes were observed in the present study, of which, only three haplotypes AAA, CGG and CAA were the most prevalent haplotypes both in CHB patients and controls owing to strong LD in these three SNPs. Haplotype analysis found that individuals carrying the CAA or CGG haplotype showed no increased risk of progression to CHB with reference of the AAA haplotype. Meanwhile other five haplotypes composing these three SNPs with frequencies smaller than 5% would not be evaluated in the haplotype association study.We further performed haplotypes analysis of these three SNPs  in KIF1B was significantly associated with the progression to CHB, and equivalence-based test confirmed the absence of association.In the study, we conducted a case-control study to firstly investigate whether a new genetic susceptibility locus associated with HBV-related HCC identified by GWAS, KIF1B rs17401966 at 1p36.22 region, was identified to be involved in the progression from CHB to HCC by a GWAS study in Chinese population KIFIB in 473 CHB cases and 580 controls with no medical history of CHB, and no significantly associations were observed between these three SNPs and the progression to CHB after adjusting age and gender. The results indicate that the genetic variants in KIF1B may only play role in the progression from CHB to HCC, but not in the progression to CHB. Similarly result was found by a recent GWAS in Japanese that a identified genetic locus for hepatitis C virus-related HCC was not associated with chronic hepatitis C susceptibility KIF1B locus for HCC is located at 1p36 region, which encodes two spliced isoforms of kinesin protein (KIF1Bα and KIF1Bβ) involved in the transport of organelles and vesicles P-values of nonsuperiority test for rs8019, rs17401966 and rs17401924 is 0.002, 0.050 and 0.017, respectively, and the nonexistence of association between KIF1B and CHB progression is confirmed. Taken together, our current result, plus these published evidence, outlined a hypothetical theory that variable genetic susceptibility mechanisms underlie different procedural steps from HBV infection to hepatocarcinogenesis, and strongly indicated that the genetic variants in KIF1B significantly contributed to the progression of CHB to HCC, but not the CHB susceptibility.Since CHB has been verified to be closely associated with HCC, identification of the genetic predictors of CHB development is very important for fighting against HBV-related HCC. Although researchers have paid much attention to identify genetic susceptibility loci to CHB during past decades, the specific genetic alteration underlying progression to CHB was far from clear. Recently, a new susceptibility locus, KIF1B genetic variants for the progression from CHB to HCC, and the genetic susceptibility mechanism underlying the progression to CHB emergently needs further comprehensive analyses to elucidate.In conclusion, this current study highlights the specificity of the


Banksia menziesii (Proteaceae), a widely distributed Western Australian tree of significance in local ecological restoration programs. An analysis of molecular variance (AMOVA) of 100 amplified fragment length polymorphism (AFLP) markers revealed significant genetic differentiation among populations (ΦPT = 0.18). Pairwise population genetic dissimilarity was correlated with geographic distance, but not environmental distance derived from 15 climate variables, suggesting overall neutrality of these markers with regard to these climate variables. Nevertheless, Bayesian outlier analysis identified four markers potentially under selection, although these were not correlated with the climate variables. We calculated a global R-statistic using analysis of similarities (ANOSIM) to test the statistical significance of population differentiation and to infer a threshold seed collection zone distance of ∼60 km  and 100 km (outlier markers) when genetic distance was regressed against geographic distance. Population pairs separated by >60 km were, on average, twice as likely to be significantly genetically differentiated than population pairs separated by <60 km, suggesting that habitat-matched sites within a 30-km radius around a restoration site genetically defines a local provenance seed collection zone for B. menziesii. Our approach is a novel probability-based practical solution for the delineation of a local seed collection zone to minimize negative genetic impacts in ecological restoration.An increasingly important practical application of the analysis of spatial genetic structure within plant species is to help define the extent of local provenance seed collection zones that minimize negative impacts in ecological restoration programs. Here, we derive seed sourcing guidelines from a novel range-wide assessment of spatial genetic structure of 24 populations of  Ecological restoration – the process of assisting the recovery of an ecosystem that has been degraded, damaged, or destroyed – is a rapidly emerging scientific discipline . Ideally the markers should capture the main sources of neutral and non-neutral population genetic variation – local adaptation, genetic drift, gene flow, and mutation – and the interactions between these sources. Given the complexity of these genetic objectives, there is no ideal single molecular marker technique. However, the polymerase chain reaction (PCR)-based multi-locus DNA fingerprinting technique amplified fragment length polymorphism (AFLP) is perhaps the most suitable marker to meet these objectives. AFLP efficiently delivers information for many markers from across the genome, and typically at least some of these markers detect signatures of natural selection and can strongly influence the overall patterns of genetic differentiation detected . Banksia menziesii is a key species used in the ecological restoration of disturbed and degraded sites across the Swan Coastal Plain (SCP), a region of high species diversity within the South-West Australian Floristic Region (SWAFR), an international biodiversity hotspot  estimate population genetic diversity and differentiation parameters, (ii) provide genetic guidelines for local provenance seed sourcing for the establishment of ecologically restored populations that have high evolutionary potential and are genetically integrated with existing local populations, and (iii) more generally, define a threshold distance that has biological significance, beyond which there are potentially negative restoration consequences from collecting seed for restoration. Additionally, we employ a population genomics approach to look for signatures of natural selection in the markers we have employed, through the identification of molecular marker outliers as well as associations between markers and environmental data obtained for each site.Here, we seek to delimit local provenance seed-source zones in an iconic Western Australian species, Banksia menziesii is a dominant over-story species, widespread along the deep sandy soils of the SCP  within populations, and overall, using GenAlEx v6.5b3  and Shannon's diversity index . Significance of pairwise population differentiation was also determined by permutation testing, and by comparison to the global R-statistic  to 1  (Chapman and Underwood R) and linear geographic distance visually, and by a Mantel test (with 9999 iterations) in Genalex v6.5b3. Sampling locations were plotted using GPS points on OziExplorer (http://www.oziexplorer.com) to generate accurate geographic distances (km) between each pair of locations.An analysis of pairwise population similarities was used to test for significant differences between all pairs of populations, using the ANOSIM  function in Primer V6 (Clarke and Gorley http://www.diva-gis.org/).Climate data  were obtained from the Australian Bureau of Rural Sciences Natural Resources Data Library. These data were then extrapolated to sample population coordinates with Diva geographic information system (http://manteltester.berlios.de/), which uses the Zt software tool  of the residuals of a null model  that are specific to each population and to each locus. The method uses a hierarchical Bayesian approach to estimate the posterior probabilities of two alternative models, one including the effects of selection and one excluding it. The results are expressed as posterior odds (PO), which indicate for each locus the ratio of posterior probability of the selection model against the neutral model and interpreted as different levels of evidence of selection according to Jeffreys scale. We used a threshold of PO >100 (“decisive”) for a marker to be considered under selection. This corresponds to a posterior probability of >0.99 for the model accounting for selection. BayeScan was run with 20 pilot runs with a burn-in of 50,000 followed by 50,000 iterations each, a sample size of 5000, a thinning interval of 10, and an FIS set at a mean of 0.05 based on known data for complete outcrossing in B. menziesii  = −2) means that 1% of corresponding outlier markers  are expected to be false positives. We then conducted simple and partial Mantel tests, as outlined above, on individual markers identified by the Bayesian allele frequency test.To detect signatures of natural selection on individual genetic markers, a Bayesian approach generalized from the method of Beaumont and Balding  was implii Scott . In addiB. menziesii plants sampled from 24 locations across the geographic range of the species, of which 79 were polymorphic (Table I) ranged between 0.12 and 0.21 (mean = 0.16). AMOVA partitioned 82% of the total variation among individuals within populations (Table PT = 0.18), which was significantly different from zero .MDS ordination of the genetic data showed an arrangement of populations that was largely associated with geographic proximity Fig. . A notabR of 0.39, which was significantly different from 0 (P < 0.001). Approximately half (149/276) of all pairwise population tests were significant at P < 0.001 as determined by 999 random permutations. There was a significant positive correlation between genetic distance (as measured by R) and geographic distance . The global R of 0.39 intersected the line of best fit through these points that corresponded to a geographic distance of ca. 60 km  generated an R > 0.39. This result was similar when calculated for the 0–20 km and 0–30 km distance classes. Beyond 60 km, there were 158 pairwise population comparisons, of which 49 (31%) generated an R < 0.39, and 109 (69%) generated an R > 0.39. Beyond 150 km, these values were 12% and 88%, respectively.ANOSIM for all markers generated a global Simple Mantel tests showed significant relationships between all pairs of distance matrices Table . PartialFST values that were 2–3 times those of non-outliers, with strongly positive alpha coefficients (1.5–2.3), “decisive” posterior probabilities (log10PO >2), and a FDR of <0.01% (log10q <−4.0) , one positive (G55 and B93) and one negative (G74 and B93). An MDS ordination of genetic dissimilarity from these four markers (not shown) largely reflected that generated from all markers  between these two genetic dissimilarity matrices.Four markers were identified as significant outliers by BayeScan analysis, with log10q <−.0  genetic distance (as measured by R based on these four markers) against geographic distance resulted in a weaker positive correlation  than for all markers combined  was found between the SCP populations and non-SCP populations  Fig. . The souB. menziesii. In particular, we used the relationship between the R-statistic  and geographic distance, to infer a biologically significant threshold seed collection distance. Beyond this threshold distance we suggest that there are potentially negative consequences from collecting seed for ecological restoration, although this ultimately requires experimental confirmation. This objective materially enhances the generic seed collecting guideline extremes of composite provenancing , which corresponded to a geographic distance of 60 km, or equivalently a 30-km radius around a restoration site. The unique strength of this analysis is that it enabled us to assign probabilities of genetically matching pairs of populations at various distance classes, depending on the proportion of points falling below and above the global R. Within 60 km, 66% of all population pairs fell below the global R, and 34% above. Beyond 60 km, 31% of all population pairs fell below the global R, and 69% above. Consequently, the probability of genetically matching population pairs  by chance when separated by >60 km is half that for populations separated by <60 km. Importantly, the proportion under 60 km does not change for the smaller distance classes of 30 km, nor 20 km. In contrast, above 150 km, there is only a 1 in 10 chance of genetically matching population pairs.For 79 variable AFLP markers in R values, from ∼0 to almost 0.8. For example, Bold Park is a 437 Ha remnant of coastal bushland in the western suburbs of metropolitan Perth that was declared an A-class reserve in 1998 and has been the subject of the most extensive ecological restoration effort in the Perth metropolitan area (BGPA B. menziesii for restoration. For these populations, R = 0.65. The relatively weak genetic similarity between populations in Bold Park and Kings Park has been observed in many other species (e.g., Krauss and He R and geographic distance remains when specifically comparing Bold Park to all other sites, with R > 0.39 for 63% of population pairs separated by >60 km, and 42% of population pairs separated by <60 km.However, even within 60 km, there was a large range in pairwise population rea BGPA . GeograpThis example serves to demonstrate the need to qualify the genetically determined threshold seed collection zone distance of 60 km with the recommendation that habitat and climate matching of the restoration site and potential seed-source populations within the threshold distance should be conducted wherever possible to increase the probability of matching genetically adapted seed-source populations with those in or around the restoration site (McKay et al. Four markers were identified as outliers, and therefore candidates for positive selection. None of these markers, however, were associated with the climate variables assessed here, and overall they were only weakly associated with geographic distance. The weaker association with geographic distance and the higher threshold distance, compared to analyses with all markers, suggests an underlying, yet undefined, complexity of selection driving variation at these markers, and that seed sourcing decisions made purely on the basis of geographic distance may have greater error for these putatively non-neutral than putatively neutral markers. This conclusion serves to emphasize the importance of habitat matching where possible within the defined threshold geographic distance based on overall marker variation. Detection of the variation at these outlier markers by chance alone seems unlikely given the attention to this issue by the BayeScan analysis through the robust FDR (Fischer et al. R-statistic threshold for significance of genetic distance of AFLP markers enables the quantification of probabilities of genetically matching populations as a function of geographic distance. This then establishes a solid quantitative framework for the testing of biological significance of the genetic marker results through the use of reciprocal transplant experiments (O'Brien et al. In conclusion, our novel consideration of the


MS is a rare extramedullary tumor, which should be considered in the differential diagnosis of a soft tissue mass involving the duodenum, especially if there is a coexisting hematological disorder. De novo cases often progress to AML, and current therapy involves Daunorubicin- and Cytarabine-based chemotherapy. The wide cytogenetic and molecular heterogeneity of MS implies a potential role for more targeted MS therapies, which may offer a curative strategy.Myeloid Sarcoma (MS), a rare extra hematopoietic carcinoma composed of blast cells, is located primarily in extramedullary sites such as skin, soft tissue, lymph nodes, and bone. MS usually presents in the setting of coexisting acute myeloid leukemia (AML) and myeloproliferative disorders. Gastrointestinal involvement (GI) is extremely rare from nonspecific abdominal symptoms to obstruction. Eight cases of myeloid sarcoma involving the duodenum including the current case have been reported, overall mean age being 40 years (range 17–71) and M : F ratio 7 : 1. The prognosis of patients with  Myeloid sarcomas (MS) are rare and potentially destructive extramedullary tumors consisting of immature myeloid cells that most often present in the skin, soft tissues, bone, and lymph nodes , 2. Althde novo with no evidence of bone marrow involvement as seen in the current case. We report the case of a de novo MS presenting as a compressive mass involving the 3rd portion of the duodenum in a 48-year-old man who presented with nausea and vomiting. A comprehensive review of the literature with similar clinical presentation, diagnosis, management, and prognosis of patients with these rare GI de novo tumors are discussed.Many MS patients (if not most) have either a coexisting acute myeloid leukemia (AML), myeloproliferative, or myelodysplastic disorder at the time of diagnosis, or it appears at the first sign of relapse from one of these disorders. In rare cases, MS occurs  A 48-year-old man presented to the emergency room at Saint Barnabas Medical Center in Livingston, NJ, USA, complaining of nausea and colicky nonradiating epigastric pain of a 2-weeks duration that was not associated with food ingestion. He also reported intermittent constipation over the past two weeks. Past medical history was significant for bipolar disorder that was well controlled on Lithium. General physical examination revealed no abnormalities; the patient was anicteric and had no palpable lymphadenopathy. The abdominal examination revealed a mildly distended abdomen with moderate tenderness over the epigastrium. No rebound tenderness or guarding was present. Bowel sounds were normal and no organomegaly or masses were noted. Laboratory evaluation, including complete blood count, liver enzymes, renal function, and electrolytes, was all within normal limits. A computed tomography (CT) scan of the abdomen showed concentric wall thickening of the 3rd and 4th portions of the duodenum with adjacent soft-tissue encasement of the superior mesenteric artery and prominent mesenteric lymph nodes. The mass measured 6.1 cm × 5.9 cm × 8.0 cm . At that2) twice a day on days 1–3 and Idarubicin (12 mg/m2) on days 4, 5, and 6. Treatment was tolerated well, with rapid resolution of abdominal pain and obstructive symptoms. A follow up abdominal CT scan done two months later revealed a residual 2.9 cm × 3.5 cm × 4.4 cm tissue mass in the distal duodenum that was notably smaller than on original CT. There was no evidence of bowel obstruction . At the time of report the patient remained symptom-free and has shown no evidence of additional hematological involvement after 18 months.The patient received 1 cycle of Cytarabine ; however, 1 patient in this group eventually developed AML at the time of report (12.5%). Six cases presented with MS were limited to the duodenum (75%), while two cases involved additional structures (25%). The prognosis of patients with de novo MS has been reported to be better than those who have a coexisting leukemia, which was also seen in the reported duodenal MS cases. Both patients with de novo MS who remained leukemia-free after treatment remained in remission, while 50% of those with leukemia eventually expired, 1 remained in remission (25%), and the outcome of one patient was not reported.Eight cases of myeloid sarcoma with duodenal involvement including the current case have been reported since 1998. The clinical data, treatment, and patient prognosis in these cases are detailed in Myeloid sarcoma is a rare extramedullary malignant tumor composed of immature myeloid cells and can be identified by the type of blast cells that compose the mass . The World Health Organization (WHO) has further classified granulocytic sarcomas into 3 main types based on the degree of maturation of the tumor: blastic (myeloblasts), immature (myeloblasts and promyelocytes), and differentiated (promyelocytes and more mature myeloid cells) . These t MS has a predilection for males 2 : 1) but no apparent age preference as all age groups are represented in the literature  but no a, 19.  MS may occur in a variety of locations; however, the most common sites are skin 13%–22%), bone/spine (9%–25%), and lymph nodes (15%–25%), and less often the central nervous system and the orbits . Gastroi%–22%, bode novo MS were initially diagnosed as diffuse large B-cell lymphoma, small lymphocytic lymphoma, peripheral T-cell lymphoma, T-cell precursor lymphoma, and myeloid metaplasia when adequate immunohistochemistry studies were not performed [46%–75% of patients with isolated MS are initially diagnosed with other conditions such as non-Hodgkin's lymphoma , 21. Pilerformed . Immunoherformed , 22. Poserformed , 22. Theerformed . Intereserformed . Our patde novo or in conjunction with AML or other hematologic entity even when age, sex, and anatomical site are taken into consideration. Patients who undergo either allogenic or autologous bone marrow transplants seem to have a higher probability of survival and complete remission [KIT inhibitors Imatinib and Dasatinib, and monoclonal antibodies) aimed at inhibiting the pivotal pathways of leukemogenesis and restoring normal hemopoiesis. In the future, a combination of different new drugs along with conventional chemotherapy may represent a more effective and potentially curative strategy [ Although there is no clear consensus or guidelines on how to treat isolated MS, delays in treatment almost always progress to AML , 22. Theemission . The medemission . Howeverstrategy . Diagnosing duodenal MS can be a diagnostic challenge especially in patients with no previous leukemic disease. The symptoms related to the tumor mass are most often nonspecific, with abdominal pain sometimes being the only complaint, and as a result a wide variety of differential diagnoses are often entertained. In the evaluation of a solid GI mass, myeloid sarcoma should be considered in the differential diagnosis with confirmation by biopsy and immunohistochemical analysis of the tumor. Patients with MS should be thoroughly tested for the presence of concurrent leukemias or other myelodysplastic disorders. The rapid initial diagnosis of MS and immediate treatment with either chemotherapy, radiation, surgery, or a combination of any of these modalities are associated with a better prognosis, as misdiagnosis or treatment delays can result in disease progression to AML. Further studies linking cytogenic abnormalities with the location of the tumors may permit more targeted therapies.


Lamprologus callipterus, large nest males collect and present empty snail shells to females that use these shells for egg deposition and brood care. Small dwarf males attempt to fertilize eggs by entering shells in which females are spawning. The bourgeois nest males exceed parasitic dwarf males in size by nearly two orders of magnitude, which is likely to result from greatly diverging growth patterns. Here, we ask whether growth patterns are heritable in this species, or whether and to which extent they are determined by environmental factors. Standardized breeding experiments using unrelated offspring and maternal half-sibs revealed highly divergent growth patterns of male young sired by nest or dwarf males, whereas the growth of female offspring of both male types did not differ. As expected, food had a significant modifying effect on growth, but neither the quantity of breeding substrate in the environment nor ambient temperature affected growth. None of the environmental factors tested influenced the choice of male life histories. We conclude that in L. callipterus growth rates of bourgeois and parasitic males are paternally inherited, and that male and female growth is phenotypically plastic to only a small degree.In species with indeterminate growth, age-related size variation of reproductive competitors within each sex is often high. This selects for divergence in reproductive tactics of same-sex competitors, particularly in males. Where alternative tactics are fixed for life, the causality of tactic choice is often unclear. In the African cichlid  Indeterminate growth generates age-related size variation of reproductive competitors, which may lead to enormous intrasexual size dimorphism .In the African cichlid lipterus , males plipterus . Large nlipterus . Nest malipterus . In contlipterus  and henclipterus . Dwarf mlipterus . Apart fL. callipterus, because the extremely divergent size optima for males pursuing the nest male and dwarf male tactics in this species  suggested two different growth pathways in Taborsky . In thisTo test for potential genetic paternal effects on the determination of life-history pathways in males and females, we raised offspring of 21 broods sired by either a territorial or a dwarf male, respectively  over time (t)  was modeled using the simplified vBGF ), with Lt: size, here SL of the fish at time t (years); Linf: asymptote or maximum size achievable (length of a fish at infinite age); K: growth coefficient (rate which determines how fast the fish approaches its infinite length); and t0: a theoretical age at which size is theoretically zero  describes the growth process as a differential equation of length , were raised in tanks with different combinations of food availability, temperature, and numbers of shells  and three times this amount (6 mg) for the high food treatment. When the size measurements started, food was changed to Sera Vipan flake food, and the amount per day was adjusted to the total biomass of all fish (% of weight) in each tank. It was offered in high (9%) and low (3%) abundance, where 9% of the fish's biomass is equal to feeding them ad libitum and 3% allows to slow and steady growth Taborsky . Food waShells were offered in high (30) and low (3) numbers resembling high and low densities in the field (own observations).L. callipterus; Temperature was kept at either 24 or 27°C, which reflects the natural range at different depths and seasons in nature (own observations). Temperature was checked continuously by using automatic temperature loggers (“Hamster”) and maximum deviations were ±0.5°C. Due to mortality, the numbers of fish in each tank decreased considerably during the first 9 months and also during the measurement period. In one tank only one fish survived. The baseline mortality rate of juveniles in substrate breeding cichlids is generally high, which corresponds to the large clutch sizes they usually produce (mean = 95 eggs in P < 0.1) were analyzed with nonparametric statistics. For all analyses, two-tailed tests were applied. To test for the genetic paternal influence on growth, we compared for all 21 broods whether the sizes of male/female offspring of nest males differ from those of dwarf males at the age of 12 months with t-tests. We used the Fisher's exact test for each brood to check for deviations from an expected 1:1 sex ratio. With Mann–Whitney U-tests, we checked for differences in the survival rate of offspring sired by dwarf males or nest males. For each sex, we tested for differences in the estimated growth rate, K and the infinite body size, Linf between offspring sired by nest or dwarf males with t-tests. To check for differences in offspring body sizes sired by territorial and dwarf males from a very young age onwards, we used a univariate general linear model that compared body sizes  of nest and dwarf male offspring up to an age of 0.9 years  in dependence of their age (covariate), sire type (fixed factor), and the interaction between age and sire type, all nested for brood number. With Wilcoxon matched-pairs signed-ranks tests, we checked whether male and female offspring of maternal half-sib broods differ in the estimated growth rate, K and the infinite body size, Linf. To test for the influence of environmental factors on growth, data of the first 6 months of measurement were analyzed. We used a univariate general linear model (GLM) with SL at 6 months as a measure of growth as dependent variable and temperature, the amount of food and shells as fixed factors.The distributions of data were tested for normality with the Kolmogorov–Smirnov normality test. Data that differed significantly from normal distributions (N = 38 fish from 7 broods) and male offspring of dwarf males 28.31 ± 2.54 mm , but there was no difference for female offspring . In none of the broods, the sex ratio differed from 1:1 . We found no difference between the survival rates of offspring sired by nest and dwarf males .Male offspring of nest males and dwarf males showed highly divergent growth rates, whereas female offspring of both male types grew similarly quickly . At an ar2 = 0.613, df = 3, F = 526.33, P < 0.001). The age of fish , as well as the sire type  and the interaction between age and male type  significantly determined the SL of offspring  for nest males exceeded that of dwarf males more than threefold . We found no differences in any vBGF parameters between female offspring sired by nest male or dwarf male fathers , but neither temperature  nor the number of snail shells  or any interactions between these parameters affected growth significantly  for dwarf males (SL = 32.3 ± 4.4 mm) that matches the average SL of 32.2 ± 4.7 mm of dwarf males employing the wriggling tactic in the field  exceeds that of nest males (0.37 ± 0.11) more than fivefold  than males from a southern population in a rocky and sandy habitat (maximum SL = 115.9 mm), whereas females showed a similar size range , although no longer (P = 0.54), than females housed with small shells during a period of 18 months , for example, depending on the population small parasitic males may be as old as large nesting males  where some males reach sexual maturity at least 1 year before other males (“jacking”), the father component of the additive genetic variance yielded a significantly higher heritability estimate than the mother component (Heath et al. Symphodus melops, with males either behaving territorial or as sneakers, has been reported to be fixed for life (Dipper In  Dominey  or only  Dominey , but likld Gross , which hld Gross . This anld Gross . In sevee Dipper . Howevere Dipper .L. callipterus growth rates of nest males and dwarf males are heritable in the male sex, which determines two alternative reproductive phenotypes that are fixed for life. Male and female body sizes are phenotypically plastic to only a little degree, and the different body sizes reported from different populations of this species are probably reflecting adaptations to divergent optima caused by local differences in predation pressure and breeding substrate. However, the genetic and environmental influence on body size differences between populations needs further experimental scrutiny.We conclude from our results that in


Osteoporosis is a multifactorial disease in which genetic determinants are modulated by hormonal, environmental and nutritional factors. An important clinical risk factor in the pathogenesis of osteoporosis is the presence of genetics polymorphism in/around susceptibility genes/regions. This study explored whether the region of 4q22.1, which confers risk of developing osteoporosis in some populations, associated with bone mineral density and osteoporosis susceptibility in postmenopausal women of Han Chinese. We investigated 32 SNPs with minor allele frequencies ≥0.05 between 20 kb upstream and 20 kb downstream (40 kb window) of rs6532023, mapping in the 4q22.1 region, which was reported to be significantly associated with osteoporosis in previous studies. We found that rs6532023 was significantly associated with bone mineral density and osteoporosis (corrected p = 0.015) in our sample, including 440 cases and 640 controls, and allele G was supposed as a risk factor while T worked as a protective factor. Further genotype association analyses suggested a similar pattern (corrected p = 0.040). Additionally, analyses by haplotypes indicated that a haplotype block rs7683315-rs6532023-rs1471400-rs1471403 in the region associated with bone mineral density and osteoporosis , and risk haplotype A-G-G-C had almost 1.5-fold increased in the cases. To our knowledge, this is the first report to examine 4q22.1 region polymorphisms and osteoporosis in Han Chinese. Our results provide further evidence for an effect of the region of 4q22.1 on the etiology of osteoporosis and suggest that 4q22.1 may be a genetic risk factor for bone mineral density and osteoporosis. Osteoporosis (OMIM166710) is a common skeletal disorder and can be severe, which is characterized by bone strength weak, and reduced bone mineral density (BMD) So far, a number of loci that contribute to osteoporosis and BMD regulation have been identified. Styrkarsdottir et al. Estrada and others Researches also proved that osteoporosis showed the distinct age The protocol of this study was approved by the Medical Ethics Committee of Xi'an Jiaotong University. All participants have completed written informed consent forms. All subjects used in this study were random unrelated Chinese Han individuals from Shaanxi Province, with no migration history within the previous three generations. Subjects with a history of bone disease, metabolic or endocrine disorders such as hyperthyroidism, hyperparathyroidism, diabetes mellitus, liver disease, renal disease, medications known to affect bone metabolism  were excluded. Meanwhile, given that the possibility of osteoporosis resulted from obesity and the overall level of Asian body mass index (BMI), we ruled out subjects of BMI ≥27 in the study. None of the women had a history of taking medicines for the treatment of osteoporosis, such as active vitamin D3, bisphosphonates, SERM, or calcium. In total, 440 women (aged 49–78 years) with primary postmenopausal osteopenia or osteoporosis and 640 healthy age-matched women (aged 48–77 years) were recruited from the Second Affiliated Hospital of Xi'an Jiaotong University and Xi'an Honghui Hospital in this study.Dual-energy X-Ray absorptiometry  was used to assess BMD at the lumbar spine (L2–4) and femur neck. Bone mineral density was determined according to standard Lunar protocols. Bone mineral density was expressed in g/cm2 and as peak bone mass percentage in normal subjects (T-score) depending on the software used in the device. The anthropometric baseline data of all subjects were obtained by measurement and questionnaire. All continuous variables were tested for skewness. The distribution of all variables was normal. The continuous parameters were presented as mean ± standard deviation, and comparisons of anthropometric characteristics of the subjects were performed using independent sample t-test for continuous variables .We searched for all SNPs with minor allele frequencies (MAF) >0.05 between 20 kb upstream and 20 kb downstream (40 kb window) of rs6532023 in the HapMap HCB database by Haploview Peripheral blood was drawn from a vein into a sterile tube containing ethylenediamine tetraacetic acid (EDTA). Plasma samples were stored at −80°C. Genomic DNA was extracted from peripheral blood leukocytes according to the manufacturer's protocol . DNA was stored at −20°C for SNP analysis. Genotyping was performed for all SNPs using the MassARRAY platform . Briefly, SNPs were genotyped using high-throughput, matrix-assisted laser desorption ionization–time-of-flight (MALDI–TOF) mass spectrometry. Next, the resulting spectra were processed using Typer Analyzer software (Sequenom), and genotype data were generated from the samples. As the final genotype call rate of each SNP was greater than 98% and the overall genotyping call rate was 98.8%, the reliability of further statistical analysis was ensured.Hardy–Weinberg equilibrium (HWE) for each SNP was assessed using GENEPOP v4.0 32 SNPs in the 4q22.1 region were genotyped in 440 cases and 640 controls. The allele and genotype frequencies of all SNPs in cases and controls, including the results of the HWE test, are shown in Because rs7683315, rs1471400 and rs1471403 were in the same Linkage Disequilibrium (LD) block containing rs6532023 from analysis of HapMap dataset, we selected the four SNPs  for further analysis. Taking into account that rs6532023 in the region of 4q22.1 yielded a significant association, we performed linkage disequilibrium (LD) estimation for the 4 SNPs  in the region of 4q22.1. The purpose of the present study was to investigate the relationship between the region of 4q22.1 containing rs6532023 and osteoporosis susceptibility. With the fast development and extensive use of Genome-wide association study, more and more osteoporosis susceptibility loci will be reported. However, their observations will need further evidence from other independent populations. In this study, we present evidence of association between rs6532023 within 4q22.1 and osteoporosis, where some significantly associated haplotypes also appeared. These lend additional support to the positive association results for osteoporosis and 4q22.1. Several lines of evidence suggest that the observed association is unlikely to be an artifact. First, both the single SNP and the haplotype-based association analyses support the association. Second, population stratification is the unlikely explanation because all of our samples are from the same geographical region. Finally, similar results were obtained from the studies of Estrada et al.'s and ours, reaffirming the observed association.To examine if the common risk variants exist in genetically independent populations, we compared our results with those of Estrada et al. The ability to draw conclusions regarding associations based on the analysis of individual SNPs is limited 4q22.1 implicates the region containing multiple genes, where the SNP (rs6532023) was found to be strongly significant association with osteoporosis and BMD in some populations. Actually, the 4q21.1 region contains a cluster of structurally and phylogenetically related genes encoding matricellular phosphoglycoproteins with function in bone formation and growth A major limitation of the current study is that the sample size of our population, especially the case number, seems to be modest. Additionally, it is important to note that we have not attempted to genotype a larger set of SNPs in the region; rather we have selected a subset of 31 SNPs neighboring rs6532023 reported by Estrada et al. In conclusion, our work provides supportive evidence for the association of 4q22.1 with osteoporosis susceptibility. Moreover, we have also confirmed the previous reports suggesting that the region of 4q22.1 is likely responsible for osteoporosis and may play an important role in the etiology of osteoporosis and BMD regulation. Given the complex patterns of association findings in complex disorders such as sex specificity and genetic heterogeneity, further inquiries and wider replications are still needed in men, especially in studies using different ethnic samples.Table S1Allele and genotype frequency of all SNPs association analyses.(DOC)Click here for additional data file.