Reaction times to targets presented in the same location as a preceding cue are greater than those to targets presented opposite the cued location. This observation can be explained as a result of inhibition at the attended location (IOR), or as facilitation at the location opposite the cue (opposite facilitation effect or OFE). Past research has demonstrated that IOR is observed reliably, whereas OFE is observed only occasionally. The present series of four experiments allows us to determine whether or not OFE can be explained by eye movements as suggested by previous authors. Participants' eye movements were monitored as they were presented with an array of four placeholders aligned with the four cardinal axes. Exogenous cues and targets were presented successively. Participants (N=37) completed either: i.) cue-manual and cue-saccade experiments, ignoring the cue and then responding with a keypress or saccade, respectively, or ii.) manual-manual and saccade-saccade experiments, responding to both the cue and the target with a keypress or saccade respectively. Results demonstrated a reliable IOR effect in each of the four experiments . None of the four experiments demonstrated evidence of an OFE . These results are inconsistent with a momentum-based account of cue-target task performance, and furthermore suggest that the OFE cannot be attributed to occasional eye movements to the cue and/or target in previous studies. There is a reliable bias against responding to targets appearing at recently attended locations. In the first study to reveal this effect, Posner and Cohen  presenteBefore settling on an inhibition-based account, Posner and Cohen  considerPosner and Cohen  expandedDespite the fact that attentional momentum had been proposed, tested and rejected by Posner & Cohen , the ideSnyder, Schmidt and Kingstone , for exaThe "eye movement" suggestion is made possible because, with few exceptions, past research examining attentional momentum has failed to monitor eye movements. Thus, as suggested by Snyder et al.  it is poperipheral cues.The present experiment allows us to directly test Snyder et al.'s  suggestiIf the generation of attentional momentum depends on the execution of an eye movement to the cue stimulus and subsequently back to the central location , then w37 undergraduate students  at Dalhousie University participated in the current study in exchange for course credit. All participants reported normal or corrected-to-normal vision.This project received ethical approval from the Dalhousie University Research Ethics Board (project number 2011–2589), which follows the standards laid out in the Tri-Council Policy Statement 2 (Canada). All participants provided informed written consent prior to participation.SR Research Experiment Builder (SREB) was used in combination with EyeLink II  eye tracking system to create and carry out this study. The flow of the experiment and eye movements of the participants were measured using the EyeLink II  head-mounted, video-based eye-tracking device . Calibration of the EyeLink II was carried out in the same picture-plane used to display the experimental stimuli. EyeLink DataViewer software  digitizes the pupil in order to describe the location of the visual gaze fixations. Respondents’ eye movement data (saccadic reaction times) and key presses  were recorded to a text file which was exported into an excel file and ultimately uploaded to SPSS v.15.0 for further statistical analysis.The current study consisted of a four experiments: cue-saccade (E1), saccade-saccade (E2), cue-manual (E3) and manual-manual (E4) response. The same peripheral cue and target stimuli and procedure was used in each of the four experiments. What differed between experiments were the responses made to the cue (no response vs. response) and the nature of responses ; note that a response was always required to the target. Each participant was assigned to either cue-respond group (N = 19), where participants completed both cue-manual (E3) and cue-saccade (E1) experiments, or respond-respond group (N = 18), where participants completed both manual-manual (E4) and saccade-saccade (E2) experiments. The order of group assignment was randomized and the order in which participants completed experiments was counterbalanced.For the duration of each experiment, participants were seated 57 cm from a computer monitor upon which all experimental stimuli were presented. Participants were presented with a central square and four peripheral squares, all equal in size . The peripheral squares were positioned 12.2 degrees from the center of the center square, one above and one below the horizontal meridian and one to the right and one to the left of the vertical meridian. Thus, the 5-square placeholder arrangement formed a “+”. Target dimensions and relative orientation remained the same for all trials.Each trial began with the presentation of a small circle at the centre of the screen to perform a drift correction procedure. Participants were required to fixate this central circle and press spacebar to initiate the trial. If the participant’s gaze was not located within 10.68 degrees of visual angle of the centre circle, a tone was presented, indicating that the participant should realign their gaze with the centre circle and press spacebar. This process continued until an acceptable calibration had been achieved. Participants were then presented with the four-placeholder array for 500 ms. Participants were instructed to maintain gaze at centre. Next, one of the four peripheral placeholders was bolded  for 300 ms (Cue) after which it was restored to its usual line width for 200 ms , after which it was restored to its usual line width for 200 ms  after which its original line width was restored for 200 ms , participants were required to keep their gaze at the centre for the duration of each trial . Although a circular region of interest with a diameter of 10.68 degrees might seem overly generous, the purpose was simply to exclude trials in which eye movements were made to targets during a required fixation period, or trials in which required saccades were made to incorrect targets. Given that targets were located 12.2 degrees away from central fixation, the regions surrounding fixation (5.34 degrees in any direction) and surrounding each target (5.34 degrees in any direction) did not overlap and therefore achieved the desired purpose. As noted by an anonymous reviewer, even if small eye movements had been made this would only strengthen our conclusion that opposite facilitation effects (OFE) are not associated with eye movements.In E3, participants responded to the appearance of only the target , while in E4, participants responded to both the cue and the target  by pressing the spacebar. Any manual response made to the cue in the cue-manual condition resulted in the presentation of an error screen.In the two saccade experiments (E1 and E2), participants made no manual responses. In E1, participants were required to saccade to the target but not the cue (cue-saccade) while in E2, participants saccaded to both the cue and the target (saccade-saccade). In the saccade-saccade experiment, participants moved their eyes back to centre after responding to the cue.In each of the four experiments, failure to generate the required response to a target within 500 ms caused the trial to be aborted, and an error message to be displayed: “You failed to respond in time. 3000 ms penalty”. In the saccade experiments, correct responses were verified by checking whether the participant’s gaze was within 10.68 degrees of visual angle of the centre of the specified target location at the end of the allotted 500 ms period.Manual reaction times were computed as the time difference between the onset of the target and the time at which the participant pressed the space key. Saccadic reaction times were computed as the time difference between the onset of the target and the time at which the participant initiated a saccade whose amplitude was greater than 2.0 degrees of visual angle. Reaction times from all error trials were excluded from further analysis, as were reaction times less than 100 ms (as these were assumed to result from participants anticipating the appearance of the target).We analyzed RT data according to target location and the angular distance between the cue and target which will be referred to as "cue-target offset". Reaction time data from each of the four experiments were submitted to a 4  by 3  repeated measures ANOVA. Data from -90 and 90 degree cue-target offsets were pooled in order to generate a single, 90-degree offset  category. Pairwise comparisons were used to test for differences that directly assess IOR  and OFE . Post-hoc comparisons were used to explore effects due to target location. Mauchly’s test was used to test the assumption of sphericity. In cases where sphericity was violated, Greenhouse-Geisser corrected df, MSE and p-values are reported.In the 4  by 3  repeated measures ANOVA, there was a significant main effect of target location  = 6.8, MSE = 1678.94, p = 0.001): mean RTs for down, left, right, and up targets were 222, 192, 195, and 197 ms, respectively. Bonferroni-corrected pairwise comparisons indicated significant differences between down (222 ms) and left (192 ms) (p = 0.0055), down (222 ms) and right (195 ms) (p = 0.0023) and down (222 ms) and up (197 ms) (p = 0.0075). No other comparisons reached statistical significance.There was a significant main effect of cue-target offset  = 32.8, MSE = 963, p<0.001): mean RTs for 0, 90 and 180-degree offsets were 225, 188, and 191 ms, respectively. These data are presented in In the 4  by 3  repeated measures ANOVA, there was a significant main effect of target location on RT  = 7.82, MSE = 1830, p = 0.002): mean RTs for down, left, right, and up targets were 206, 179, 180, and 181 ms, respectively. Bonferroni-corrected multiple comparisons indicated significant differences between down (206 ms) and left (179 ms) (p = 0.0017) and down (206 ms) and right (180 ms) (p<0.001). No other comparisons reached significance.There was a significant main effect of cue-target offset  = 16.4, MSE = 1792, p<0.001): mean RTs for 0, 90 and 180-degree offsets were 204, 173, and 181 ms, respectively. These data are shown in In the 4  by 3  repeated measures ANOVA, the main effect of target location was not significant  = 1.22, MSE = 641, p = 0.311). There was a significant main effect of cue-target offset  = 4.65, MSE = 550, p = 0.016): mean RTs for 0, 90 and 180-degree offsets were 258, 248, and 249 ms, respectively. These data are presented in In the 4  by 3  repeated measures ANOVA, the main effect of target location on RT was not significant  = 0.177, MSE = 535, p = 0.911). There was a significant main effect of cue-target offset  = 9.077, MSE = 517, p = 0.001): mean RT for 0, 90 and 180-degree offsets were 233, 222, and 218 ms, respectively. These data are presented in The results from our 4 experiments are summarized in A previous study contrasted these two mechanisms using target-target methods comparable to those employed in Experiment 4, yielding results which are remarkably similar to those reported in the current study . In thatIn another study , particiEssentially the results of our E4 were identical to those of E3 (a significant 11 ms of IOR and a non-significant 4 ms of OFE). These findings  are in In summary, whereas RTs were significantly slower in the 0 degree offset condition compared to the 90 degree offset condition in all four experiments , in no experiment did we find significantly faster RTs to targets opposite from the cued location (180 degree offset condition) compared to the 90 degree offset condition. Hence, the results from all four experiments confirm the predictions made by an inhibitory gradient account, and are inconsistent with those made by attentional momentum. Let us return to the speculative conclusion offered by Snyder et al.  that the


The association between smoking and suicide is still controversial, particular for early life cigarette smoking exposure. Few studies have investigated this association in adolescents using population-based cohorts, and the relationship with second hand smoking (SHS) exposure has not been addressed.In this study, we followed a large population-based sample of younger people to investigate the association between smoking, SHS exposure and suicide mortality. Between October 1995 and June 1996, 162,682 junior high school students ages 11 to 16 years old living in a geographic catchment area in Taiwan were enrolled and then followed till December 2007  through linkage to the National Death Certification System. Participants who were currently smoking at baseline had a greater than six-fold higher suicide mortality than those who did not smoke  as well as higher natural mortality . After controlling for gender, age, parental education, asthma, allergic rhinitis, and alcohol consumption, the adjusted hazard ratios for suicide were 3.69 (95% CI 1.85-7.39) in current smokers, and 1.47 (95% CI 0.94-2.30) and 2.83 (95% CI 1.54-5.20) respectively in adolescents exposed to SHS of 1-20 cigarettes and >20 cigarettes/per day. The estimated depression-adjusted odds ratio did not change substantially. The population attributable fractions for suicide associated with smoking and heavy SHS exposure (>20 cigarettes/per day) were 9.6% and 10.6%, respectively.This study showed evidence of excess suicide mortality among young adults exposed to active or passive early life cigarette smoking. Adolescent smoking is a pressing public health issue especially in areas of rapid economic growth, and more than 80% of adult smokers have their first experience of smoking before the age of 18 . The gloAs well as all-cause mortality, some prospective studies have also shown a significant association between smoking and suicide in adults . ExplanaThe prospective analysis described here investigated the smoking-suicide connection in a young population taking into account a range of potential confounders. Secondhand smoke was also investigated as an exposure.Between October 1995 and June 1996, all 170,457 junior high school students aged 11 to 16 years old (grades 7 to 9) in all 123 public and private junior high schools in the Kaohsiung and Pingtung areas in southern Taiwan were invited to participate a heath investigation . The oriEach student and his/her parents completed a structured questionnaire (including video instruction) concerning asthma symptom severity, allergic rhinitis, the habits of alcohol consumption and cigarette smoking. Items in the parental questionnaire included demographic characteristics , family smoking status (how many family members living with the student were regular smokers), the total amount of cigarettes per day used by the family, the student’s exercise habits , and a history in the student of allergic rhinitis. SHS was defined on the basis of at least one family member living with the student who was a regular smoker, with the total daily cigarette consumption by family members calculated. We grouped SHS status into three groups of no, 1–20 cigarettes and >20 cigarettes exposure. Information on students’ cigarette smoking and alcohol consumption was obtained from the students themselves and the asthma status and symptom profile of each student were obtained through a video guided questionnaire .The cohort was followed by record linkage to the Taiwan National Death Certification System from January 1, 1995, to December 31, 2007, as previously described . Using uMortality rates were calculated for specific causes in the smoking and nonsmoking groups, with lifetable survival analysis and Gehan’s generalized Wilcoxon tests used to assess differences in incidence. Cox proportional hazards models were used to investigate associations with smoking and SHS exposures, adjusting for gender, age, parental highest education, asthma, allergic rhinitis and alcohol consumption. Crude and adjusted hazard ratios and 95% confidence intervals were calculated with covariates chosen on the basis of significant or borderline association with suicide (p<0.15) in the unadjusted models. Population attributable fractions were calculated  using adFirst, previous studies ,20 have Second, it was considered that some suicide deaths might have been misclassified as undetermined unnatural deaths . Thus, wThird, for obtaining unbiased proportional hazards regression coefficients, a prior study  had recoA total of 162,682 students were recruited (response rate: 95%), 49.8% of whom were male. The mean (SD) age was 13.8 (0.9) years, smoking behavior was reported by 2.4%, and exposure to SHS of 1–20 cigarettes and >20 cigarettes were 48.5% and 6.9%, respectively .Incidences of both natural and unnatural mortality were significantly higher in smokers compared to non-smokers . SpecifiThe proportional hazards assumption over the entire follow-up period was confirmed for all estimated variables used in the Cox regression. The crude hazard ratio for suicide mortality in the smoking compared to non-smoking group was 6.1. After controlling for other covariates, the adjusted hazard ratio was 3.69 . Family In sensitivity analyses, following further external adjustment for depression as an unmeasured confounder, the odds ratio between smoking and suicide was 4.80 see , comparaCombining suicide and undetermined unnatural death as the outcome, the adjusted hazard ratio for suicide mortality in the smoking compared to non-smoking group was 3.81, which was equivalent to the effect size from the main analysis (3.69), as was that for family SHS . FinallyIn a large prospective cohort study, assembled using data linkage, we investigated the association of active and passive smoking exposure with suicide specifically in the context of other-cause mortality more widely. To our knowledge, ours is the first study to extend previous findings potentially linking smoking and suicide to an investigation in adolescents specifically. Additionally, we believe ours to be the first investigation of the association between SHS exposure and suicide. Both smoking and SHS exposure were found to be independently associated with increased risk of suicide, findings for the latter indicating a dose-response relationship.Previous studies of smoking and suicide have reported inconsistent findings. Some epidemiological studies have found an independent association ,9,24, whA range of possible explanations for the association between smoking and suicide have been proposed. To begin with, smoking may be a non-causal marker of other risk factors for suicide such as lower income, lower education, unmarried status, unemployment, or lack of religious affiliation. It may also be a marker for mental disorders such as anxiety, depression, psychoses, and substance use disorders, or indicate other adverse mental health states such as low self-esteem, a propensity to risk-taking behaviour, impulsivity, aggression, antisocial personality, fatalism, or emotional instability. People with pre-existing or “latent” psychiatric or psychological problems may use nicotine to “self-medicate” to abate these problems ,27. HoweSome large prospective studies have controlled for alcohol consumption, and continued to find an association between smoking and suicide ,33; but We found an association between SHS exposure and suicide in adolescents which, to our knowledge, has not been previously investigated. Clearly the findings require replication, although the association is plausible. In non-smoking adolescents, duration of exposure to SHS in the home was found to have a positive dose-response relationship with psychological distress as measured by the 12-item General Health Questionnaire . AdolescKey strengths of the study were the large, representative sample and comprehensive follow-up. Limitations primarily reflect the use of historical data collected for purposes other than the objectives of this analysis. The information from questionnaire included measures of demographic characteristics, whether alcohol was consumed or not, smoking status and SHS exposure, although these relied on self-reporting and may be subject to measurement error. More importantly, information was lacking on certain risk factors for suicide such as mental disorders, personality traits, illicit drug use and family history of mental disorders. Although, a sensitivity analysis did not suggest that depression was likely to be a major confounding factor, the influence of mental health on the associations of interest could not be comprehensively evaluated. Besides mental disorders, other factors such as parental divorce, low emotional control, medication for nervous problems, contact with police and childcare services, drug use, and education may confound associations between smoking and suicide . BecauseDespite the relatively low smoking prevalence (2.4%) in our cohort, the prospect of longer-term exposure and consequent adverse outcomes are important issues for adolescent smoking. Furthermore, more than half of the adolescent population had been involuntarily exposed to SHS in this 1995 initiative cohort study. Protecting younger populations from early life cigarette smoking exposure is an important issue and needs a public health approach. Initial contact advice and reference materials provided by the pediatrician, followed by frequent, regular contacts and home visits was found to be an ideal approach , as wellS1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S1 Text(DOCX)Click here for additional data file.


Now, chronic psychological stress (CPS) related diseases are increasing. Many CPS patients have gastrointestinal complaints, immune suppression, and immune imbalance. Increasing evidence is indicating that acupuncture (AP) at the Zusanli point (ST36) can alleviate functional gastrointestinal disorders (FGID), immune suppression, and immune imbalance. However, few studies have investigated the potential mechanisms. In this study, CPS rat models were established, and electroacupuncture (EA) at ST36 was done for CPS rats. Daily food intake, weight, intestinal sensitivity, the morphology of interstitial cell of Cajal (ICC) in the small intestine, and serum indexes were measured. The study found that, in CPS rats, EA at ST36 could improve food intake, weight, visceral hypersensitivity, and immunity; in CPS rats, in small intestine, the morphology of ICCs was abnormal and the number was decreased, which may be part causes of gastrointestinal motility dysfunction. EA at ST36 showed useful therapeutic effects. The mechanisms may be partially related to its repairing effects on ICCs damages; in CPS rats, there were immune suppression and immune imbalance, which may be part causes of visceral hypersensitivity. EA at ST36 showed useful therapeutic effects. The mechanisms may be partially related to its regulation on immunity. With the changes of the social environment and ways of life, job stress is increasing and the pace of life is accelerating. Meanwhile, chronic psychological stress (CPS) related diseases are growing rapidly.The digestive system is very vulnerable to the influence of CPS. Functional gastrointestinal disorders (FGID) are very common in daily clinical practice, and they are characterized by disturbances in the motility patterns and/or visceral hypersensitivity , 2. CPS Acupuncture (AP), a traditional Chinese medicine, has garnered significant attention due to its role in regulation of gastrointestinal function. In the clinic, growing evidences have shown that it could alleviate FGID –9. Also,The interstitial cell of Cajal (ICC) exhibits a highly branched morphology and forms unique network in the gastrointestinal tract (GI). ICCs serve as electrical pacemakers, active propagation pathways for slow waves, and mediators of enteric motor neurotransmission. They play an important role in generating and regulating gastrointestinal motility . The entVisceral hypersensitivity and gastrointestinal inflammation are important pathophysiological factors for gastrointestinal motility dysfunction diseases, and they closely correlate with immunity –19. In tRepeated exposure to water avoidance stress has been shown to successfully establish a CPS rat model for sustained visceral hyperalgesia , 21. In Thirty seven-week-old male Wistar rats were provided by the experimental animal center of Dalian Medical University. The rats were maintained in a normal light-dark cycle, housed 3-4 rats per cage, and provided with free access to food and water. All protocols were approved by the Ethics Committee of The First Affiliated Hospital of Dalian Medical University.2.2.The CPS rat models were established by the method of Bradesi et al. . AccordiAt day 11, rats in the control group were housed separately for one day, and daily food intake (24 h) was measured. At day 12, the rats were fasted starting at 3 p.m. but provided free access to water. At day 13, the rats were weighed and intestinal sensitivity was evaluated. Then, the rats were sacrificed under anesthesia. Blood was collected from the inferior vena cava, and the proximal jejunum was cut and collected. The blood was centrifuged at 3000 r/min for 15 min, and the serum was segregated for subsequent analyses. The proximal jejunum was washed quickly with normal saline and then dissected into 1-2 cm fragments, fixed in 4% paraformaldehyde, and placed in the refrigerator. At day 11, 6 rats in the model group were selected according to the random number table method. The rats were processed the same way as the control group.At day 11, rats in the model group (the remaining 6 rats), AP group, and sham-AP group were fed for another 15 days. In the AP group, every day, rats underwent binding and EA at ST36 for 20 minutes between 7 a.m. and 9 a.m. The acupoint was located at the posterior and lateral side of the knee joint, 5 mm below capitulum fibulae . The AP At day 24, in each group, rats were housed separately for one day, and daily food intake (24 h) was measured. At day 25, the rats were fasted starting at 3 p.m. but provided free access to water. At day 26, each rat was weighed and intestinal sensitivity was evaluated. Then, the blood and jejunum were processed as described above.Abdominal withdrawal reflex (AWR) in response to acute visceral pain was used to assess the intestinal sensitivity following the method of Al-Chaer et al. . Acute vCRD. The distention was applied using angioplasty balloons  inserted rectally into the descending colon of mildly sedated rats  and secured by taping the attached tubing to the rat's tail. The rats were then housed in small Lucite cubicles (20 × 8 × 8 cm) on an elevated Plexiglas and allowed to wake up and adapt (1 hour). Measurement of the AWR consisted of visual observation of the animal response to graded CRD  by blinded observers and assignment of an AWR score. The rats were given CRD for 20 seconds every 4 minutes. To achieve an accurate measure, the distention was repeated 5 times for each intensity. The data for each animal was averaged for analysis. The results obtained were compared across groups.The Score of AWR. 0 indicates no behavioral response to CRD; 1, brief head movement followed by immobility; 2, contraction of abdominal muscles; 3, lifting of abdomen; 4, body arching and lifting of pelvic structures. Behavioral measurements were reproduced by 2 different blinded observers. CRD was applied in increments of 10 mmHg starting at 10 mmHg.A part of proximal jejunum was cut into 1-2 cm fragments, fixed in 4% paraformaldehyde, and placed in the refrigerator at 4°C overnight. The tissues were dissected with an anatomic microscope to strip the mucosa and submucosa as one layer and retain the whole intestinal muscularis.Next, (1) the specimens were incubated in 0.5% Triton-X in 0.05 mol/L Tris-HCl buffer (pH 7.6) solution at 37°C for 4 h; (2) after rinsing twice in PBS, specimens were incubated for 1 h at room temperature in 1% BSA to prevent nonspecific binding; (3) after rinsing twice in PBS, specimens were incubated for 48 h at 4°C in rabbit anti-rat c-kit polyclonal antibody ; (4) specimens were rinsed twice and incubated 2 h at 4°C in the dark in Cy3-mouse anti-rabbit IgG antibody ; (5) specimens were embedded in fluorescence mounting medium and examined under a confocal laser scanning microscope .In step (3), the control group was incubated in only antibody diluent instead of antibody.Specimens were observed using a TCS-SP2 laser scanning confocal microscope  with immersion objectives . Specimens were excited using a krypton/argon laser with excitation and barrier filters set for fluorophores according to its excitation-emission spectra (Cy3 = 550 nm). The emitted light was measured by a photomultiplier tube and converted into a digital pixelated image by an analog-to-digital converter. The detection pinhole was set for use with different objectives accordingly. C-kit positive fluorescence was red. Six specimens were observed in each group, and three fields of vision were observed randomly in each specimen. Three-dimensional images were constructed and integrated optical density (IOD) was performed by the Leica confocal software.The serum immunoglobulin G (IgG) and immunoglobulin M (IgM) levels were measured using commercially available sandwich ELISA kits  according to the manufacturer's instructions. Absorbance was read at 492 nm. The serum interleukin-2 (IL-2) and interleukin-6 (IL-6) levels were measured using commercially available sandwich ELISA kits  in accordance with the manufacturer's instructions. Absorbance was read at 450 nm.The measurements of weight and food intake, the assessment of intestinal sensitivity, and the measurement of ICCs and serum immune indexes were performed by blinded researchers.t-test. Differences between multigroup were compared by the one-way ANOVA; then differences between two groups were compared by S-N-K test, and P < 0.05 denoted the difference possessing statistical significance.The food intake, weight, scores of AWR, IOD of ICCs, and immune indexes data were analyzed by a commercial software package (SPSS13.0). Data was expressed as the mean ± standard error. Differences between two groups were compared by the In the model group, food intake and weight were significantly lower than those in the control group .In the model group, when the pressure of CRD was 20, 40, 60, or 80 mmHg, score of AWR was significantly higher than that in the control group .The control group  shows thThe model group  shows thIn the model group, the serum IgG, IgM, and IL-2 levels were significantly lower than those in the control group Figures , and theIn the AP group, food intake  and weigWhen the pressure of CRD was 20 mmHg, scores of AWR were not significantly different among the model group, AP group, and sham-AP group; when the pressure of CRD was 40, 60, or 80 mmHg, in the AP group, score of AWR was significantly lower than that in the model group, and there was no significant difference between the model group and the sham-AP group .The model group  shows thThe AP group  shows thThe sham-AP group  shows thIn the AP group, the serum IgG, IgM, and IL-2 levels were significantly higher than those in the model group Figures , and theNowadays, with the accelerated pace of life, we tend to bear more pressure in our profession and family life. Meanwhile, chronic psychological stress (CPS) related diseases are growing.CPS can cause various symptoms related to the digestive system. FGID are digestive diseases with high morbidity rates , 2, and AP, a physical intervention which involves placement of small needles in the skin at different acupoints, has been practiced in China for 2000 years. In the clinic, growing evidences have shown that it could alleviate FGID –9. A ranIn this study, CPS rat models were established. Food intake, weight, intestinal sensitivity, ICC of small intestine, and serum immune indexes were measured, and therapeutic effects of EA at ST36 were investigated.In clinic, most CPS patients eat less, and some patients show significant weight loss. In this study, food intake and weight were significantly decreased in CPS rats than in the control group, and this is consistent with the clinic. This partially indicates that the models were established successfully.In clinic, many CPS patients make complaints about the gastrointestinal disorder. In this study, was there abnormality of gastrointestinal function in CPS rats? AWR in response to acute visceral pain was used to assess the intestinal sensitivity following the method of Al-Chaer et al. . The resStudies have shown that CPS is associated with suppression of both cellular and humoral measures , 6. In tIn this study, after EA at ST36, food intake and weight were significantly increased; the scores of AWR were significantly decreased, which indicates the visceral hypersensitivity was improved; the serum IgG, IgM, and IL-2 levels of CPS rats increased markedly, and the serum IL-6 level decreased markedly. It indicates that immune suppression and immune imbalance were improved.In conclusion, EA at ST36 could improve food intake and weight, could improve the symptom of visceral hypersensitivity, and could regulate immune suppression and immune imbalance in CPS rats. It may be a useful therapeutic approach.Studies have shown that CPS plays an important role in the development and exacerbation of symptoms in FGID . FGID arEA at ST36 showed useful therapeutic effects on visceral hypersensitivity of CPS rats. After treatment, ICCs in intestinal tissues were investigated using confocal laser scanning microscopy, and the results showed that the number of ICCs was increased and cellular morphology resumed normal. This indicated that the mechanisms underlying these effects may be partially related to its repairing effects on ICCs damage.Studies have shown that brain-gut interactions play an important role in CPS related FGID. The brain receives a constant stream of interoceptive input from the GI, integrates this information with the other interoceptive information from the body and with contextual information from the environment, and sends an integrated response back to various target cells within the GI , 35. In Studies have shown that neuroendocrine signaling plays a key role in AP treatment. Specifically, AP sends signals to the CNS and the CNS then sends signals to various target cells , 38. We In CPS rats, there were visceral hypersensitivity, immune suppression, and immune imbalance. After EA at ST36, the immune suppression and immune imbalance were improved, and the visceral hypersensitivity was improved either. Studies have shown that visceral hypersensitivity closely correlates with immunity –19. We sIn CPS rats, neuroendocrine signaling may also play an important role in changes in immunity. Repeated stress will send signals to the CNS. Correspondingly, the CNS will send signals to the thymus, spleen, and gastrointestinal tissue via the neuroendocrine system to modulate the immune system. This process is very complex. At present, the hypothalamic-pituitary-adrenal axis, sympathetic nerve-adrenal medulla axis, and some hormones, peptides, and cytokines are thought to be involved –42. In tThe study indicates that therapeutic mechanisms of EA at ST36 may be multifaceted and target multiple components. Now, the exact mechanisms by which CNS affects ICCs in GI are not fully understood, and the pathophysiologic immunity changes in CPS rats are complex. They remain to be further investigated.In CPS rats, EA at ST36 could improve daily food intake and weight as well as the symptom of visceral hypersensitivity. Also, EA at ST36 can regulate immune suppression and immune imbalance. It may be a useful therapeutic approach.In CPS rats, in small intestine, the morphology of ICCs was abnormal and the number was decreased, which may be part causes of gastrointestinal motility dysfunction. EA at ST36 showed useful therapeutic effects on gastrointestinal motility dysfunction. The mechanisms may be partially related to its repairing effects on ICCs damage.In CPS rats, there were immune suppression and immune imbalance, which may be part of reasons of visceral hypersensitivity. EA at ST36 showed useful therapeutic effects on visceral hypersensitivity. The mechanisms may be partially related to its regulation on immune suppression and immune imbalance.


The aim of this article is to study the effect of goat milk–based formula (GMF) on development in weaned rats.One hundred Sprague-Dawley rats were randomly divided into five groups: control, 20% cow milk–based formula (CMF), and 5%-GMF, 10%-GMF, and 20%-GMF groups.GMF did play an active role in accelerating body and femur length, but not body weight growth. Compared with the control, GMF had better cognitive, space, and locomotor activity. The level of IFN was increased in GMF groups, as well as the level of IL-2 and TNF was decreased in GMF groups.These results indicate that GMF has an effect on development and immunity improvability in weaned rats. Goat milk is a food of high nutritional value, with high biological value protein, and a good source of short- and medium-chain fatty acids, minerals, and vitamins . The fatWhen discussing GMF according to the existing literature, we found that they focused mainly on following points: First, the topics are usually on the comparison of nutrition content between goat, cow, and human milk –10. SecoTherefore, the present article systematically discussed the effect of GMF on development in weaned rats. When rats were treated with GMF in the weaning period, immune effect should also be considered.GMF and cow milk–based formula (CMF) were provided by Ausntria Hyproca Dairy Group BV . The experimental diet was supplemented with ‘AIN93G’ to give 50 mg GMF, 100 mg GMF, 200 mg GMF, and 200 mg CMF of mixed MF g-1 diet. Dietary ingredients were thoroughly mixed in a mixer, made into pellets, and air-dried at room temperature. The rats Th1  bead-based multiplexed assay kits were purchased from Bender MedSystems Inc. .www.lab.pku.edu.cn).In total, 100 male and female Sprague-Dawley rats (21 days old), weighing 40–60 g, were obtained from the Animal Service of Health Science Center, Peking University, Beijing. Rats were housed two per plastic cage with free access to chow and tap water in a specific pathogen-free filter-protected air-conditioned room with controlled temperature (21±23°C), relative air humidity (55±5%), and 12 h light/dark cycles (light on 07:30–19:30 h). All animals were handled in accordance with the guidelines of the National Institutes of Health (NIH Publication No. 85-23 revised 1985) and the guidelines of the Peking University Animal Research Committee : one control group, and four experimental groups. Control rats were fed with ‘AIN93G’ as rodent diet . Rats in the four experimental groups were fed with 5% GMF, 10% GMF, 20% GMF, or 20% CMF (wt/wt) in the diet . Animals were observed daily and weekly for adverse clinical signs. A detailed physical examination was conducted at least once during the pretreatment period, once weekly during test substance administration, and on the day of necropsy. Body weight, body length, and food consumption were recorded weekly. Diet utilization ratio was calculated using the following formula: Diet utilization ratio=[weight gain (g)/food consumption (g)]×100%.2 inhalation and then sacrificed by cervical dislocation.On the 45th day, the urine and feces were collected in 24 h. The volume of urine and the weight of feces were recorded. Animals were fed with diets for 90 days. After 90 days, rats were anesthetized with CO2 and black walls of 30 cm high. The test room had a controlled illumination (dim light condition). The floor of the arena was divided into 16 equal squares. Each rat was placed in the center of the open field, and the latency time, the number of squares crossed with all paws (crossings), rearing, modification frequency, and modification were manually counted in a 3 min session. Locomotor activity was recorded between 13:00 and 17:00 h. The apparatus were cleaned with a solution of 10% ethanol between tests in order to hide animal clues.On day 45, spontaneous locomotor activity of rats was measured using the open field test (OFT). The apparatus, made of wood covered with impermeable formica, had a black floor of 100×100 cmBlood samples were collected from femoral arteries for clinical pathology evaluations  from rats immediately prior to the scheduled necropsy on day 90. All animals were fasted overnight prior to blood collection.The following hematology parameters were analyzed using MLA Electra 1,400 CTM automated Coagulation Analyzer: white blood cells, lymphocyte count (LY#), monocytes count (MO#), neutrophilicgranulocyte count (GR#), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), hemoglobin (HGB), and eosinophil ratio (EO%).The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total protein (TP), albumin (ALB), blood urea nitrogen (BUN), creatinine (CR), triglyceride (TG), globulins (GLB), ALB/GLB (A/G) and transferrin (TRF) in serum were detected by automatic biochemistry analyzer .An ELISA assay was used to measure serum immunoglobulin  and sIgA of intestinal mucosa immunoglobulin. Serum, containing Immunoglobulin A (IgA), T IgE, IgG, and IgM, is diluted with buffer containing polyethylene glycol (PEG) and mixed with specific polyclonal goat anti-IgA, anti-IgE, anti-IgG, or anti-IgM antiserum, forming a complex. This complex can be quantitated by measuring light absorbance at 700 nm. The sensitivity and the rate of forming the immune-complex can be increased by the addition of the polymer, PEG. sIgA followed the same method as serum immunoglobulin.g, and the supernatant discarded carefully, leaving 100 µl of liquid in each tube. This was repeated, and samples were further incubated with 50 µl streptavidin–phycoerythrin for 1 h at room temperature in a dark room. After two further centrifugation steps as mentioned above, the beads were re-suspended in 300 µl assay buffer and read on the BD FACSCalibur flow cytometer to determine cytokine concentration. The lower levels of detection for the 10 cytokines were between 0.7 and 15.7 pg/ml.A bead-based multiplexed assay , 20 was All animals were euthanized by carbon dioxide inhalation followed by exsanguination. Animals were necropsied and the external surface, all orifices, and the cranial, thoracic, abdominal, and pelvic cavities including viscera were examined. Organ weights were measured for the adrenal glands, brain, epididymides, heart, kidneys, liver, femurs, and ovaries with oviducts, spleen, testes, thymus and thyroids/parathyroids from all animals. Approximately 40 tissues were collected from each animal in the study for microscopy. The femur length and midpoint width were measured.P values were two-sided. A value of p<0.05 was considered significant.Statistical analyses were performed using SPSS software . Variances in the measurement data were checked for homogeneity by Bartlett's test. When the data were homogeneous, the one-way analysis of variance test and LSD methods were used. Tamhane's T2 test is used after data are transformed to analyze data among multiple groups if variances are unequal. All reported There was no remarkable difference in general condition and behavior between the control and treated rats of both sexes.p<0.05); meanwhile, the body weight of the GMF-treated female group was lighter than female control at the age of 4 weeks (p<0.05). However, at 12 weeks, the body weight of the 10%-GMF and 20%-GMF female groups was heavier than the control (p<0.05). In weeks 4 and 8, compared with the control, there were significant differences in body length in the 10%-GMF female group, as well as the CMF, 5%-GMF and 10%-GMF male groups . The organs were weighed, and there were no significant differences in organ-to-body ratio among groups of either sex. The femur length in all GMF-treated female groups was longer than the 20%-CMF female and control female groups. There were no remarkable differences in midpoint width among the GMF-treated, 20%-CMF, and control groups (p>0.05).Compared with the control, only in week 4, the diet utilization ratio was increased with significant differences in all GMF and the 20%-CMF group for both female and male rats (p<0.05). Oppositely, the modification time in the 20%-CMF male group was remarkably longer than that in the 10%-GMF male group (p<0.05). Similarly, the rearing frequency in the 20%-CMF female group was lower than that in the 20%-GMF female group; whereas it was higher than that in all GMF-treated male groups and the control.The effect of GMF treatment on locomotor activity, by OFT, was shown in p<0.05). For the male part, there were many remarkable changes in MCV and MCH. Compared with the control, the level of MCV was significantly increased in all GMF-treated groups and the 20%-CMF group; moreover, the level of MCV in the 10%-GMF and 20%-GMF groups was lower than that in the 20%-CMF group (p<0.05). The level of MCH in the 5%-GMF and 20%-CMF groups was significantly higher than that in the control (p<0.05). There was remarkable difference between the 5%-GMF and 20%-CMF groups in GR#. In comparison with the control, the level of LY# was decreased in the GMF-treated groups; moreover, there were significant differences in the 10%-GMF and 20%-GMF groups. There was only one remarkable difference between the 20%-GMF male group with the 20%-CMF male group in MO# (p<0.05).Investigations of the hematology parameters were performed in p<0.05). The significant differences of ALB were observed in the 5%-GMF and 10%-GMF female groups, compared with the control and 20%-CMF.Comparison of mean serum chemistry values from the GMF treated groups with the values from normal control is shown in p<0.05). Similarly, the level of IL-2 were remarkable decreased in 20%-CMF and 10%-GMF male group. The level of IFN was remarkable increased in the 20%-CMF and 10%-GMF groups, compared with the control (p<0.05). Compared with the 20%-CMF group, all the cytokine levels were at the same level in the GMF group. However, for the male part, there were no significant differences between the control and all GMF groups.As shown in p>0.05). In general, there was no remarkable change of GMF in immunoglobulin in either sex (The effect of GMF on immunoglobulin in rats was obtained by testing IgA, T IgE, IgG, and IgM in serum, and sIgA in intestinal mucosa. Although, compared with the 20%-CMF group, the level of IgG in all GMF groups and the level of sIgA in the 20%-GMF group are higher, there were no significant differences (ther sex .This study, on the developmental effect of GMF in the weaned period, aims at augmenting the currently available data. It indicated that GMF had the effect on development and immunity improvability. To the best of our knowledge, this is the first GMF study on development in weaned rats.As a whole, it can be seen in this study that GMF did not play an active role in accelerating body weight growth, but did for body and femur length. At the first 8 weeks, the body weight of the GMF-treated male group was significantly lighter than the control. At week 12, all the body weights and body lengths were at the same level in all GMF-treated, 20%-CMF, and control groups, both female and male. This is consistent with the result of C Grant's study –23. Fat We analyzed the whole hematology parameters. Compared with the control, there was remarkable change in the level of RBC#, MCV, and MCH, which indicated GMF may have an anti-anemia ability . AdditioOFT is now one of the most popular procedures in animal psychology. In such a situation, rodents spontaneously prefer the periphery of the apparatus to activity in the central parts of the open field. Indeed, rats walk close to the walls, a behavior called thigmotaxis. Normally, rats’ instinct will prompt them to avoid empty fields . In the In this study, the level of IFN was increased in GMF groups, as well as the level of IL-2 and TNF was decreased in GMF groups. GMF may have the effect of enhancement on immunity function. However, EO#, allergy-reflection index, did not have a statistical difference. For this part, further studies need to be performed using allergy models.GMF may play roles in the promotion of early growth, development, and immunity after weaning, especially in female rats. Further research is needed to use specific models to elucidate the effect of GMF on neurobehavior or allergy.


Rps, Rpl, Mprs, and Mprl genes were upregulated in the hypothalamus and downregulated in the hippocampus, which may indicate ribosomal dysfunction following chronic social defeat stress. There were no differentially expressed ribosomal genes in the ventral tegmental area, midbrain raphe nuclei, or striatum. This approach may be used to identify a pharmacological treatment of ribosome biogenesis abnormalities in the brain of patients with “ribosomopathies.”Chronic social defeat stress leads to the development of anxiety- and depression-like states in male mice and is accompanied by numerous molecular changes in brain. The influence of 21-day period of social stress on ribosomal gene expression in five brain regions was studied using the RNA-Seq database. Most  Chronic social defeat stress (CSDS) can lead to the development of behavioral psychopathology, which is accompanied by anxiety- and depression-like states in male mice –3 similaRps and Rpl, Mrpl and Mrps) which are responsible for translation, transcription, and proliferation and are involved in neural plasticity in healthy cells.Analyzing the whole transcriptome using RNA-Seq in five brain regions of depressive mice with chronic social defeats experience, we observed changes in the expression of numerous genes. This report is concentrated on the analysis of genes encoding ribosomal and mitochondrial ribosomal proteins . Experiments were performed on 10–12-week-old animals. All procedures were in accordance with the European Communities Council Directive of November 24, 1986 (86/609/EEC). The study was approved by Scientific Council number 9 of the Institute of Cytology and Genetics SB RAS of March, 24, 2010, N 613.Adult male mice of C57BL/6J were obtained from Animal Breeding Center in Pushchino . Animals were housed under standard conditions  and water availableProlonged negative social experiences (defeats) in male mice were induced by daily agonistic interactions with an aggressive partner , 10. Paihttp://mouse.brain-map.org/static/atlas). All of the biological samples were placed to the RNAlater solution  and stored at −70°C until sequencing.All of the mice were decapitated simultaneously: the 21-time defeated mice were sacrificed 24 hours after the last agonistic interaction and the control animals. The brain regions from both experimental groups were dissected by one experimenter according to the map presented in the Allen Mouse Brain Atlas , where the mRNA was extracted using the Dynabeads mRNA Purification Kit . cDNA libraries were constructed using NEBNext mRNA Library PrepReagent Set for Illumina  following the manufacturer's protocol and were subjected to Illumina sequencing. More than 20 million reads were obtained for each sample. The resulting “fastq” format files were used to align all of the reads to the GRCm38.p3 reference genome using the TopHat aligner [P < 0.01.The collected samples were sequenced at JSC Genoanalytica . Thus, the largest number of differentially expressed genes in depressive mice was observed in the hypothalamus, and approximately 4 times fewer genes were observed in the striatum and hippocampus and approximately 7 times fewer genes were observed in the VTA and midbrain raphe nuclei. In the hypothalamus and striatum, the number of upregulated genes was higher than number of downregulated genes. In the VTA and midbrain raphe nuclei area, most of the differentially expressed genes were downregulated. In the hippocampus, the numbers of upregulated and downregulated genes were approximately equal. We can assume that the number of differentially expressed genes and direction of change (up or down) may be used as marker of more or less intensive involvement of any brain area into molecular mechanisms of depression-like state in mice. These changes may depend on function of brain regions and different mechanisms regulating CSDS. Other authors have demonstrated changes of numerous genes expression in the nucleus accumbens under CSDS [Gene expression levels were compared between mouse groups affected by social stress, depressive mice, and the control animals. Analysis of differentially expressed genes showed their dependence on the brain regions : in the der CSDS . Rps, Rpl, Mrpl, and Mrps genes did not change their expression under CSDS. Because ribosomes are responsible for protein synthesis in all cells, we suspect that the translation, transcription, and proliferation of proteins are not significantly disturbed in these brain regions.In the midbrain raphe nuclei, VTA, and striatum, theRps) and the large subunit that connects amino acids to form a polypeptide chain (Rpl)—changed their expression under CSDS. In the hippocampus of depressive mice, the largest number of ribosomal genes  was downregulated and only 2 genes (Rpl35 and Rpl18) were upregulated. We can assume that downregulation of the ribosomal genes may be associated with a decrease of proliferation in the hippocampal dentate gyrus under CSDS in mice, as described by many authors using a similar experimental paradigm [In the hippocampus and hypothalamus, the major components of ribosomes—the small ribosomal subunit that reads the RNA ( Rps and 22 Rpl genes). Rps14, Rps8, Rps6ka1, Rps9, Rps5, Rps19, Rps16, Rps3, Rpsa, Rps2, Rps26, and Rps10 and Rpl37a, Rpl41, Rpl19, Rpl23a, Rpl37, Rpl8, Rpl10a, Rpl36, Rpl7a, Rpl12, Rpl35, Rpl34, Rplp0, Rpl6, Rpl28, Rpl18, Rplp2, Rpl13, Rpl18a, Rpl29, and Rplp1 were upregulated, and Rpl22l1, Rps6ka3, and Rps6ka6 were downregulated  in approximately 43% of patients [ Rps19, Rps14, Rps10, and Rps26 genes, which are involved in Diamond-Blackfan anemia, was observed in depressive mice. We did not find literature data concerning ribosome dysfunction during depression; however, our observation concerning changes in the expression of ribosomal genes in the hippocampus and hypothalamus in mice indicates developing ribosomal dysfunction under CSDS.In recent years a number of human diseases have been identified and categorized as “ribosomopathies” , 14 causpatients , 16. Int Mrpl54, Mrpl12, Mrpl38, Mrpl52, Mrpl28, Mrpl23, Mrpl34, Mrpl4 Mrps18a, and Mrps12 were upregulated in depressive mice, whereas Mrpl1 and Mrpl3 were downregulated ( Tph2, Sert, Maoa, and Htr1a gene expression, which are associated with the synthesis, inactivation, and reception of serotonin, respectively, in the midbrain raphe nuclei [There were no mitochondrial ribosomal genes found that changed expression under CSDS in the hippocampus. However, in the hypothalamus the results obtained indicate the development of possible mitochondrial protein dysfunctions: the mitochondrial ribosome genesegulated . Thus, we nuclei . We can e nuclei  in the re nuclei , 21 in te nuclei . It coule nuclei .Mitochondrial disorders may be caused by either acquired or inherited mutations in the mitochondrial DNA or in nuclear genes that code for mitochondrial components . These dUndoubtedly it is difficult to find a direct association between the overexpression of ribosomal genes and mitochondrial ribosomal genes in the hypothalamus and the depression-like state in mice, which would help to understand causes and consequences of these processes. At this stage of research, it is impossible to elucidate the detailed sequence of neurochemical events involved, and, as a result, the molecular changes that occur due to restructuring brain regulation in male mice under CSDS. However, it is clear that, starting with a change in social behavior and psychoemotional state under CSDS, at certain stages this process launches a cascade of systemic changes at the whole brain level, its regions, and specific neurons following changes in metabolism and reception of neurotransmitter systems. As a result, it leads to the changes in the expression of genes involved in the development of affective disorders. The changes observed in ribosomal and mitochondrial ribosomal gene expression may indicate ribosome dysfunction. Our model, which induces a mixed anxiety/depression-like state , 2 in ma


Avicennia schaueriana (S2) and Rhizophora mangle (S3). Environmental variables in S1 and S2 were more similar than in S3; however, when comparing the bacterial compositions, S2 and S3 shared more OTUs between them, suggesting that the presence of vegetation is an important factor in shaping these bacterial communities. In silico analyses of the fragments revealed a high diversity of the class Gammaproteobacteria in the 3 sites, although in general they presented quite different bacterial composition, which is probably shaped by the specificities of each habitat. This study shows that microhabitats inside of a mangrove ecosystem harbor diverse and distinct microbiota, reinforcing the need to conserve these ecosystems as a whole. We investigated the relationship among environmental variables, composition, and structure of bacterial communities in different habitats in a mangrove located nearby to an oil exploitation area, aiming to retrieve the natural pattern of bacterial communities in this ecosystem. The T-RFLP analysis showed a high diversity of bacterial populations and an increase in the bacterial richness from habitats closer to the sea and without vegetation (S1) to habitats covered by Mangroves are coastal ecosystems that have been seriously threatened by anthropogenic activities. Worldwide, mangrove areas have been used for urban development, tourism, oil exploitation, agriculture, and shrimp farming. Between 1980 and 2005, about 3.6 million hectares of mangrove was lost [Although these ecosystems are well known for their typical flora and associated fauna, comparatively, only a few studies deal with their microbial diversity –8. On thIn the environment microorganisms fulfill various niches and are fundamental for the functioning of mangroves, being particularly important in controlling the geochemistry of these habitats , 13. Rec in silico methods, such as the resources available on the MiCA3  website, make the identification of specific organisms in a community based on the length of Terminal-Restriction Fragments (T-RFs) possible, as they predict T-RFs from known and deposited sequences in databases that can be compared with the submitted T-RFs [Genetic fingerprinting techniques provide a pattern or profile of the genetic diversity in a microbial community, which are important in distinguishing PCR products that have different nucleotide sequences. Terminal-Restriction Fragment Length Polymorphism (T-RFLP) has proven to be a valuable tool to study bacterial community structure in complex environments such as sediments and soils , 18, 19.ed T-RFs , 21.In this context, we hypothesized that the zonation of mangrove species, as well as the daily fluctuations imposed by the tidal regimes, shapes the microbial communities that are present in these habitats, making them unique to each mangrove area. In order to test our hypothesis, we employed T-RFLP to access the composition and structure of bacterial communities of sediments in vegetated and nonvegetated areas in a mangrove located in Northeastern Brazil and its relation to the biotic and abiotic variables in a region known for petroleum exploitation, which is considered a risk area for oil contamination. Avicennia schaueriana forest; and the third site (S3) was located in a region of a robust forest of Rhizophora mangle. The sites were 150 m apart from each other. At each site, five sediment samples (0–10 cm depth) were randomly collected using a cylindrical sampler (30 cm long and 10 cm in diameter) and transferred to sterile jars. The samples were kept in an ice-cooled box for about 2 hours before being transported to the laboratory. In the laboratory, the five replicate samples from each site were homogenized in order to obtain composed and representative samples of each habitat and a portion was stored at −20°C for DNA extraction and the remaining fraction was used for sediment analyses. Granulometry was performed by dry sieving [Barra Grande mangrove is located in Icapuí, on the extreme east coast of the state of Ceará, Northeastern Brazil (37°20′W 4°40′S), in a region comprised of an extensive tidal flat, covering an area of 1,260.31 ha . Due to  sieving , and org sieving . The env HhaI and MspI following the manufacturers' recommendations . Digestion products were dried at 40°C and sent to the Research Technology Support Facility, Department of Plant Biology, Michigan State University  Facility, where T-RF profiles were generated. The analysis was performed using 2 μL of the digestion with 8 μL of a solution containing the internal standard MapMaker™ 1000  labeled with ROX (6-carboxy-X-rhodamine) and the running buffer (deionized formamide). DNA fragments were detected by capillary electrophoresis on an ABI Prism 3100 Genetic Analyzer  automatic sequencer. The T-RFs were visualized using the GeneScan Analysis Software (Applied Biosystems), exported to Excel, and analyzed with the Ibest tool (http://mica.ibest.uidaho.edu/) using the height of 50 units of fluorescence as an initial point for the electropherogram analysis and normalized by calculating the relative abundance of each T-RF from the fluorescence intensity area. T-RFs in the range of 50–990 bp were used for the analysis. T-RFs that differed by less than 1 bp were considered identical. The files were exported from Ibest and analyzed by the T-Align tool . Each individual T-RF was considered an OTU .Bacterial communities were analyzed by T-RFLP, following the protocol described by Marsh . The extH′) by log2, the Simpson diversity (λ), and the Pielou equitability (J′) [The relative abundance of OTUs was used to calculate diversity indices for each sample. The Shannon index (ity (J′)  were calhttp://mica.ibest.uidaho.edu/pat.php) was used to identify OTUs for T-RF peaks, based on the RDP  Release 9.60 16S rRNA gene database [The web-based analysis tool (PAT+) provided by MiCA3  than MspI (87 T-RFs). This means that HhaI best resolved the constituent community in the analyzed samples. Thus, results from the digestion with HhaI were selected for further analyses.Three different community structures with a higher similarity between S2 and S3 were observed, both inside forested areas. The digestion withThe relative abundance of OTUs  revealed Flavobacterium sp., Capnocytophaga sp., Vibrio sp., and Photobacterium sp., whereas many species of Bacteroidetes from the Flavobacteriaceae family and some Alphaproteobacteria were assigned to fragment 65.Using PAT+ in MiCA, we predicted the potential bacterial groups based on digestion pattern of the fragments obtained by T-RFLP. T-RFs 54 and 65, which were shared by the three sites, were mainly represented by uncultured bacteria of the Bacteroidetes and Proteobacteria phyla. Among the possible species that can be attributed to T-RF 54 are Vibrio parahaemolyticus; and T-RF 394 corresponding to an uncultured bacterium. At S2, four dominant fragments were detected: T-RF 57 including Alphaproteobacteria such as uncultured Azospirillum sp. and the Bacteroidetes Mariniflexile fucanivorans and cultured and uncultured Cytophaga spp.; T-RF 56 comprising Alphaproteobacteria as Sneathiella sp., uncultured Mesorhizobium sp., various species of Thalassospira, members of Rhodospirillaceae, some uncultured Gammaproteobacteria from Piscirickettsiaceae, and groups of the phylum Bacteroidetes, such as Fluviicola sp., Aequorivita sp., A. antarctica, A. sublithincola, Subsaxibacter sp., Persicivirga sp., Salinimicrobium sp., Mariniflexile gromovii, Myroides sp., M. odoratimimus, M. profundi, M. pelagicus, Gelidibacter sp., G. algens, and an uncultured Sphingobacterium; T-RF 77 which could not be identified by the web-based tool; and T-RF 94 which was identified as Escherichia coli.Considering the dominant fragments at each site, S1 showed three different fragments: T-RF 100 associated with uncultured halophilic bacteria; T-RF 325 represented by uncultured bacteria, including species of Gammaproteobacteria and the cultured bacterium Capnocytophaga sp.; T-RF 72 represented by uncultured members of the order Bacteroidales; T-RF 167 which consisted of undetermined uncultured bacteria and several uncultured Gammaproteobacteria and cultured representatives such as Pseudomonas sp., Pseudoalteromonas sp., Shewanella sp., Salicola sp., S. salis, S. marasensis, and Halovibrio denitrificans.S3 showed three dominant fragments: T-RF 55 represented by uncultured bacteria andIn this study, we observed differences in the bacterial community structure and composition that could be attributed to the specific characteristics of each sampled mangrove habitat. It is well known that mangroves are under the influence of marine and terrestrial environments, which generate gradients in the texture of sediments and organic matter content and in salinity as a result of sea and freshwater inputs , 28. TakPeixoto et al., 2011 , have sh Avicennia, while a predominance of Actinobacteria was evidenced in nonvegetated sediments. Ramírez-Elías et al. [ Laguncularia rhizosphere harbored the highest microbial population when compared to other mangrove species.Also, microbial populations seem to be influenced by the presence and type of mangrove species. Gomes et al., 2010 , demonsts et al.  studying A. schaueriana and R. mangle, respectively, shared more similarities in terms of microbial composition with each other than with S1, the site without vegetation. The numbers of OTUs detected in S1, S2, and S3 were 34, 43, and 73, respectively. This increase in the richness from S1 to S3 was observed, which suggests that, besides local abiotic variables, the nature of exudates and nutrients provided by each plant species select specific communities [Sites S2 and S3, which are habitats covered bymunities .The observed differences in community structure were reflected in the relative abundance as well as in T-RF composition. Regarding the percentage of unique T-RFs, it is notable that this mangrove harbors quite different bacterial communities, as confirmed by the high value of exclusivity, especially at S1, which was 76.5%, and the low level of similarity among the sites, considering that only two T-RFs were shared by them. Thus, these data confirm that local differences were responsible for distinguishing bacterial populations.A. schaueriana) like S3, but with many abiotic variables similar to those found at S1, due to its proximity to the sea.We detected an increase in the evenness of bacterial communities over the three sites; that is, S1 showed lower evenness and the presence of some dominant OTUs. Due to the proximity of the sea, the microbiota in S1 is under the influence of tidal hydrodynamics, which probably led to the selection of several species that are more adapted to marine environments. At S3, a wider distribution of OTUs was observed, with a lower occurrence of dominant OTUs, demonstrating that the environmental conditions have not favored any particular OTU. Intermediate characteristics were shown at S2, which can be explained by its location in an area inside the vegetation , there was prevalence of uncultured bacteria, which shows the wide gap in extant data on microbial diversity, considering the large number of unknown organisms [Putative community compositions were determined using a phylogenetic assignment tool (PAT) developed by Kent et al. , using Mrganisms .Taking into account the possible groups associated with T-RFs, it can be observed that, besides the uncultured bacteria, the main common OTUs were phylogenetically affiliated with the Bacteroidetes, with a large number belonging to the Flavobacteriaceae. In Brazilian mangrove sediments, some bacteria affiliated with the Bacteroidetes were also observed in clone libraries, but at a low number compared to the phylum Proteobacteria, which appears to be dominant in these environments , 33, 34. Vibrio parahaemolyticus. At S2, located in the root zones of A. schaueriana, several uncultured bacteria were found, with a predominance of Alphaproteobacteria, some of them known nitrogen fixers, such as an uncultured Azospirillum, which was previously reported in roots of A. marina and in the rhizosphere of Suaeda monoecious in a mangrove from India [ Thalassospira were detected, which were previously isolated from oil-contaminated seawater. It was found that these could degrade several polycyclic aromatic hydrocarbons (PAHs), including naphthalene, dibenzothiophene, phenanthrene, and fluorene [At site S1, which lies in a region close to the sea, without vegetation, there was prevalence of uncultured halophilic bacteria, uncultured Gammaproteobacteria, andom India . Severalfluorene . These s Vibrio sp. and Photobacterium (Proteobacteria) and Rhodovulum sulfidophilum, a marine photosynthetic bacterium (Alphaproteobacteria). Some members of the genus Pseudomonas which are able to metabolize petroleum hydrocarbons [Among the OTUs found at S3, some were identified as belonging to the phylum Proteobacteria, such asocarbons  were alsAltogether, the data showed distinct bacterial communities among the three mangrove habitats due to the presence and type of vegetation and the divergence environmental variables in which these habitats are submitted. In general, all the potential bacteria corresponding to the identified T-RFs are typical of marine environments and play important roles in maintaining the dynamic balance of the ecosystem. This study highlights the importance of preserving mangrove ecosystems as a whole, due to the uniqueness of each habitat.The main contribution of this study was to demonstrate that mangrove soils hold highly diverse bacterial populations with increasing richness from the sea to forested areas, selected by the local differences. The existence of unique bacterial communities to each mangrove habitat covered by distinct plant species clearly demonstrates the importance of the conservation of this ecosystem as a whole.


Magnetic resonance imaging (MRI) is a useful diagnostic tool for the evaluation of congenital or acquired brain lesions. But, in all of less than 8-year-old children, pharmacological agents and procedural sedation should be used to induce motionless conditions for imaging studies. The purpose of this study was to compare the efficacy and safety of combination of chloral hydrate-hydroxyzine (CH+H) and chloral hydrate-midazolam (CH+M) in pediatric MRI sedation. In a parallel single-blinded randomized clinical trial, sixty 1-7-year-old children who underwent brain MRI, were randomly assigned to receive chloral hydrate in a minimum dosage of 40 mg/kg in combination with either 2 mg/kg of hydroxyzine or 0.5 mg/kg of midazolam. The primary outcomes were efficacy of adequate sedation (Ramsay sedation score of five) and completion of MRI examination. The secondary outcome was clinical side-effects.Twenty-eight girls (46.7%) and 32 boys (53.3%) with the mean age of 2.72±1.58 years were studied. Adequate sedation and completion of MRI were achieved in 76.7% of CH+H group. Mild and transient clinical side-effects, such as vomiting of one child in each group and agitation in 2 (6.6 %) children of CH+M group, were also seen. The adverse events were more frequent in CH+M group.Combinations of chloral hydrate-hydroxyzine and chloral hydrate-midazolam were effective in pediatric MRI sedation; however, chloral hydrate-hydroxyzine was safer. Magnetic resonance imaging (MRI) is a useful diagnostic tool for evaluation of congenital or acquired brain lesions, dysmyelination or demyelination, gliosis, primary and metastatic brain tumors, cerebral edema, and acute stroke. But, in all of less than 8-year-old children, pharmacological agents and procedural sedation should be used to induce motionless conditions for imaging studies , for whiSchulte-Uentrop and Goepfert  in theirChloral hydrate (CH) is a non-opiate, nonbenzodiazepine sedative-hypnotic drug, which has been used for pediatric sedation in a dosage of 40-100 mg/kg for many years ,5. But, Chloral hydrate at a dose of 40 mg/kg is safer and its combination with antihistamines might decrease chloral hydrate dosage ,8.Combination of chloral hydrate and hydroxyzine has been used for sedation of children in dental procedures, and it can decrease the required dosage of chloral hydrate and also cause improvement in safer sleeping of patient and decrease the risk of chloral hydrate related nausea and vomiting .Midazolam is a water-soluble benzodiazepine, which can be used in different routes  for sedation induction in children ,11.Oral midazolam is a non-parenteral route, which does not cause pain of injection and is used at dosages of 0.5-1 mg/kg in pediatric sedation induction .The purpose of this study was to compare the efficacy and safety of chloral hydrate at minimum dosage in combination with hydroxyzine or midazolam in pediatric MRI sedation induction.We followed a randomized single-blind study on children who underwent sedation for elective MRI at the Radiology Department of a tertiary-care hospital  in Yazd, Iran from November 2012 to March 2013. An Informed consent was taken from patients’ parents before the administration of the drugs, and the study was approved by the Ethics Committee of Shahid Sadoughi University of Medical Sciences, Yazd, Iran.The sample size was calculated to be 30 children in each group to detect a 20% difference in efficacy between the two groups with type one error  of 0.05 and 80% power. Eligible participants included children aged 1-7 years, who were in American Society of Anesthesiology (ASA) class 1  or 2  . Exclusion criteria consisted of presence of gastritis or any other serious systemic diseases, severe systemic reaction, head injury, and receiving a sedative hypnotic agent within the past 48 hours.The trial used computer generated equal simple randomization by random numbers, and allocation ratio was 1:1 for the two groups. Randomization and blinding were performed by an investigator with no clinical involvement in the trial. Data collectors, outcome assessors, and data analysts were all kept blinded to the allocation. But, patients and an MRI nurse allocated to the intervention group, were aware of the allocated arm. A pharmacist prepared the drugs, which were given in a suspension of 1 cc/kg. The drug combination was delivered by MRI nurses who had certificates in pediatric advanced life support and basic life support, when the participants entered the preparation room. Primary and secondary outcomes were assessed by a pediatric resident who was not informed of the group assignment of drug combination.The children were randomly assigned to two groups to receive either 40 mg/kg of chloral hydrate and 2 mg/kg of hydroxyzine (Group I) or 40 mg/kg of chloral hydrate and 0.5 mg/kg of midazolam (Group II).Ramsay sedation scale was used for assessment of the sedation level , and it The primary outcomes were efficacy in adequate deep sedation and completion of MRI examination.The secondary outcomes included clinical side-effects, serious adverse events , time from administration of the drug combination to adequate sedation, caregiver’s satisfaction on a likert scale of 1-5 , and total stay time in MRI center. Respiratory depression requiring assisted ventilation, oxygen saturation of less than 90%, or a 25% or greater decrease in pre sedation mean arterial blood pressure were considered as serious side-effects. Failure to achieve adequate sedation  and procedure abortion due to serious adverse events, were considered as failure of sedation regimen.The developmental status of the patient was assessed by a pediatric neurologist based on Denver II Developmental screening test .The strength of the MRI machine of our hospital was 1.5 Tesla, and it had been manufactured by Siemens in 2010. Non-contrast brain MRI of these children was interpreted by a pediatric neurologist and a radiologist using a threepoint scale: 1= good quality and no motion, 2= reportable and minor movement, and 3= non reportable and major movement.The data were analyzed using SPSS statistical software (version 17). Chi-square and Fisher’s exact tests were used for data analysis of qualitative variables, and mean values were compared by independent t-test. Kaplan–Meier survival analysis was used to calculate probability of adequate sedation during the observation period. The differences were considered significant at p<0.05.This study was registered in Iranian clinical trials with registration number IRCT201302092639N10.The design and conduct of this trial was straightforward, and we did not have any losses to follow-up or exclusions. Twenty-eight girls (46.7%) and 32 boys (53.3%) with the mean age of 2.72±1.58 years were evaluated. No statistically significant differences were observed between the two groups in terms of mean of age, mean of weight, sex distribution, developmental status, and age group of children in two groups . AdequatAlso, the quality of MRI was not significantly different between the two groups .Table 2 shows comparison of the mean of acquired Ramsay sedation score, time between drugs taking and reaching the Ramsey score of five, caregiver’s satisfaction scale, and total stay time in radiology department which indicates that in chloral hydrate-hydroxyzine group, parents waited less in the radiology department.Comparison of completion of MRI examination in both groups based on developmental status and age group is shown in Table 3, which indicates that the efficacy of both drugs combination in sedation induction was not statistically different in children with and without developmental delay, in infants (less than two years old), and also in children.Probability of being adequately sedated vs. time after taking the drugs is shown in Kaplan–Meier plots in Fig.1, which indicates that Ramsay sedation score of five was obtained in all children who achieved adequate sedation 40 minutes after taking the drugs combination. No serious adverse events were seen in the two groups. Mild and transient clinical side-effects were seen, such as vomiting in one child in each group and agitation in 2 (6.6%) children of CH+M group. The adverse events were more frequent in CH+M group (p=0.04).Various drugs have been used for pediatric MRI sedation. The Results of this randomized clinical trial showed that combination of chloral hydrate at minimum dosage and hydroxyzine or midazolam were equally effective in children who underwent MRI.In another Iranian study, adequate sedation and completion of CT scan examination were achieved in 76.7% of children who aged 1-10 years and received 100 mg/kg chloral hydrate orally .In Mason et al.’s study, efficacy of oral chloral hydrate at a dosage of 50 mg/kg and oral pentobarbital were equal in MRI sedation of younger than 1-year-old infants; however, side-effects frequency was significantly lower in pentobarbital group .However, Schulte-Uentrop and Goepfert concluded that in sedation induction for MRI, chloral hydrate, pentobarbital, and midazolam are not proper and dexmedetomidine may be a more effective drug in sedation induction in children without cardiac risk, and if anesthesiologists or pediatric intensivists are present, propofol can be used and general anesthesia is a preferred technique in preterm or small children .In a study in Iowa, USA, a combination of 25 mg/kg chloral hydrate, 1 mg/kg hydroxyzine, and 1 mg/kg meperidine was more effective than 0.65 mg/kg of oral midazolam in sedation for dental procedures .In a study in Miami, USA, efficacy of chloral hydrate, combination of chloral hydrate and diphenhydramine, chloral hydrate-hydroxyzine hydrochloride combination and midazolam alone were compared in case of sedation induction for echocardiography. In chloral hydrate group, children fell asleep the most quickly and chloral hydrate and diphenhydramine group had the most prolonged sedations .In a study in Mexico City, combination of 70 mg/kg chloral hydrate and 2 mg/kg hydroxyzine in comparison with 70 mg/kg chloral hydrate alone caused a significantly more decrease in crying and movement within 45-60 minutes after a rubber dam insertion. But, in both groups, overall behavior was not different during the dental procedures .In a study in Mexico, a combination of 0.50 mg/kg midazolam and 1.5 mg/kg hydroxyzine or 50 mg/kg chloral hydrate and 1.5 mg/kg hydroxyzine were more effective than 2 mg/kg hydroxyzine alone in sedation induction for dental procedures .In the present study, the efficacy of both combination of the drugs in sedation induction were not statistically different in infants (less than two years old) and also in children. But, in a Korean study, chloral hydrate was more effective in MRI sedation of younger than 18-month-old children .In our study, the efficacy of both combinations of the drugs in sedation induction for MRI was not statistically different in children with and without developmental delay, which is in agreement with a study in the USA .Possible explanations for these discrepancies include differences in age, drugs combination and dosage, race, sample size, type of procedure, etc.In the present study, both sedation regimens were safe and no serious clinical adverse events were seen in the two groups. But, in Fávero et al.’s study, respiratory complications occurred in two of 41 children who received 50 mg/kg of chloral hydrate  and in HIn this research, achieving sleep in majority of children who were adequately sedated with the two sedation regimens appeared up to 40 minutes after administration of the drugs combination, therefore, administration of these sedative drugs in 40 minutes before the procedure could be more effective.In conclusion, results of the present study showed that a combination of chloral hydrate at minimum dosage and hydroxyzine or midazolam are equally effective in pediatric MRI sedation. But, a combination of chloral hydrate and hydroxyzine is safer.


Our geometric morphometric analysis of a 15 landmark-based configuration of platyrrhines’ first and second lower molars suggest that morphological stasis may explain the reduced molar shape variation observed. Platyrrhine lower molar shape might be a primitive retention of the ancestral state affected by strong ecological constraints throughout the radiation of the main platyrrhine families. The Patagonian fossil specimens showed two distinct morphological patterns of lower molars, Callicebus—like and Saguinus—like, which might be the precursors of the extant forms, whereas the Middle Miocene specimens, though showing morphological resemblances with the Patagonian fossils, also displayed new, derived molar patterns, Alouatta—like and Pitheciinae—like, thereby suggesting that despite the overall morphological stasis of molars, phenotypic diversification of molar shape was already settled during the Middle Miocene.The phylogenetic position of many fossil platyrrhines with respect to extant ones is not yet clear. Two main hypotheses have been proposed: the  Chilecebus (20 Ma), a fossil specimen  and Dolichocebus  have been included in Cebinae , eight in the Argentinian Patagonia, five in the Greater Antilles, five in Brazil, and one each in Chile, Bolivia and Peru . Neosaim Cebinae . Neosaimar shape . Its molventiana . Laventiosaimiri , althougosaimiri . Lavential flare . Dolicho monkeys . However monkeys  argued tAotus dindensis was first described as a sister taxon of extant Aotus  bear resemblance also with the callitrichines. Indeed, Soriacebus and pitheciines or with callitrichines are due to homoplasy, rather than phylogenetic relationships among such lineages . Cebupithecia, although considered to be more derived than Nuciruptor  exhibits similar dental size and morphology to extant Alouatta; showing molar teeth with sharp and well-formed crests, a long cristid oblique, small trigonid, and spacious talonid basin  of molar shape for studying patterns of inter-specific variation and their implication in phylogeny and ecological adaptations e.g., . HoweverM1 and M2) of extant Platyrrhini primates to asesses the affinities of the Patagonian, Colombian, and Antillanean fossil taxa with the extant forms and to estimating the efficiency of molar shape for discriminating fossil specimens.The aim of the present study is to use two-dimensional (2D) GM to quantify and analyze occulsal shape variation of lower molars (Proteopithecus sylviae), were obtained from the literature , discovered in Argentina, Colombia, and Jamaica, and dated to between Holocene and early Miocene  quantifies shape differences between biological structures using a set of digitized homologous points (landmarks) in two-dimensional or three-dimensional spaces . LandmarAlouatta belzebul, Aotus dindensis, Callicebus personatus, Callithrix geoffroyi, Pithecia irrorata). The landmarks were digitized nine times during three non-consecutive days. Mean Procrustes distance between paired repetitions was 0.13328, with a standard deviation of 0.04644, and the average Pearson correlation between Procrustes distance matrices (Mantel test) of the repeated measurements was 0.9887. No significant differences in shape configurations among repetitions were obtained with a non-parametric MANOVA test . The inter-observed error rates were not computed since a single researcher (MND) made all the analyses.Intra-observer landmark digitizing error was measured in a subsample of five specimens representative of five different species including one fossil taxon  to classify fossil specimens, since PCA removes the irrelevant and redundant dimensions , and the a posteriori probability score was used as the probability that a fossil belongs to a particular group. Several LDAs were made considering different discriminant factors: (1) family , (2) the subfamily-level classification proposed by Cebus, Sapajus, Saimiri, Callithrix, Mico, Cebuella, Callimico, Leontopithecus, Saguinus, Aotus, Callicebus, Cacajao, Chiropotes, Pithecis, Lagothrix, Brachyteles, Atelles, Allouatta). The LDA analyses were carried out with SPSS v.15 .After the procrustes superimposition, the covariance matrix of all the compared shapes was used to derive a Principal Components Analysis (PCA) . The PCAmensions . LDA maxmensions . The relM1 for all platyrrhines  and especially Nuciruptor (F11) and Cebupithecia (F12) that had negative PC1 and positive PC2 scores. Most extinct forms overlapped with the extant platyrrhines, within Callicebinae, Callitrichinae, and Atellinae, except Xenothrix (F13), Nuciruptor and Cebupithecia, which do not.Even though the PCA does not distinguish subfamilies, the plot of PC1 M2 , and some specimens of Neosaimiri clearly fell within the Alouattinae clade, Dolichocebus (F3) within Saimiriinae, and Nuciruptor (F11) was closer to Cebinae and Pitheciinae on the negative scores of PC1. Homunculus (F4) did not fell at all within any extant taxa, showing highly positive PC2 scores.The first two PCs for M2  accountePC2 plot  showed spost-hoc percentages of correct classification after cross-validation (pcc) were high both for M1  and Rosenberger’s  pcc values were 2.3% for M1 and 1.6% for M2 (genus) to 100% (family) for M1, and from 66.1% (genus) to 100% (family) for M2. The highest percentage of total variance explained by DF1 was 56.0% (family) for M1 and 68.3% (family) for M2, and the highest one for DF2 was 44.0% (family) for M1 and 32.8% (subfamily R) for M2.The % for M2 . The per% for M2  for all a priori classification probabilities varied depending on the discriminant factors used  and Nuciruptor neotypes showed a high probability of belonging to the pitheciid clade in LDAs. In contrast, Xenothrix (M2 not available) likely belonged to Callithrix, despite in the PCA this fossil specimen did not fall within Callitrichinae range. Stirtonia was assigned to the Atelidae clade, and to Alouatta at the genus level, except for Rosenberger’ subfamily factor for M2. Laventiana was also classified into the atelids for M1, but was more closely related to callitrichines for M2. Aotus dindensis showed a high probability of belonging to Aotustaxa for M1, but Callicebus was the group with the greatest affinity for M2. Finally, Proteopithecus showed a high resemblance with Saimiri for M1, but with Callimico for M2.Regarding the classification of the fossil specimens, the ranges of the Proteopithecus sylviae (F1) in the morphospace and its molar shapes showed pattern resemblance to that of platyrrhines. However, because, many dental and postcranial features of P. sylviae are considered to be symplesiomorphic characters of all anthropoids, it is placed as the stem anthropoid  . Thus, if the Fayum form likely was a sister taxon to platyrrhines, the interspecific variation of shape would have shown relatively little change. This could mean that the main traits of molars shapes in platyrrhines represent retention of a primitive ancestral form. Moreover, the LDA showed a high probability of P. sylviae belonging to the Cebidae clade, suggesting that the molar of the earliest ancestors of platyrrhines must have exhibited close similarity to Saimiri-Callimico. This resemblance matches with the description of an Oligocene primate fossil found in South America, Branisella , showing phenetic similarities with these two extant subfamiliesMost of the traits used to identify phylogenetic affinities among Early Miocene platyrine fossils show high levels of homoplasy Callicebus-shaped and a Sagunus-shaped. For example, Dolichocebus (F3) was classified as a pitheciid, mainly by having a square occlusal polygon  was related mainly to the callitrichine clade, but for M2 the probability of belonging to this group was small . Soriacebus were convergent. In contrast, Soriacebus represents the first branch of pitheciines. Although marmosets are considered derived lineages , it was classified as a Callitrichinae in the DFA. However, it has been shown to be more similar with Callicebus than marmosets, such as the shape contour and quadrate alignment of cusps in both molars. Homunculus (F4), was placed outside the range of Patagonian forms in the PCA  was the rectangular-shaped molar (see Xenothrix below). This phenetic similarity among phyletically distinct groups of extinct primates indicates that a rectangular-shaped molar almost certainly represents a plesiomorphy in the Patagonian fossils. Thus, the trend toward ovoid molar shape might be a derived feature in many living forms. Laventania (F7) exhibited distally oriented cusps on M1, showing considerable resemblance with some atelid groups, which provided a confusing classification between atelids and Callicebus in the LDA  were classified in completely distant taxonomic groups  and Aotus dindensis (F9) have been considered by Kay and collaborators  and Cebupithecia (F12), the occlusal views in both species were relatively rounded, with a slightly rectangular alignment of cusps, and buccally oriented, which resembles the condition in most extant Pitheciinae. Moreover, the LDA indicated that Cebupithecia and Nuciruptor had a close affinity with the Pitheciidae clade (Nuciruptor on M2)  . Severalr on M2) .Stirtonia and Alouatta was classified in the LDA with a 99.9% probability for M1 and 94.0% for M2. Likewise, the PCA showed that Stirtonia was placed close to howler monkeys , the Caribbean platyrrhine form, has been allied with pitheciids  differs from that of the pitheciids. It is likely that Xenothrix could be a single branch that evolved independently of crown platyrrhines, as was suggested by some investigations that proposed an early Antillen arrival  and forms from La Venta. This small phenotypic variation could be due to developmental and functional constraints, given the role in occlusion and mastication  explains the low interspecific variation between extinct and extant linages and between Early Miocene platyrrhines (including tication  and the tication . Followipithecus , it is lic range . The Calvolution . Thus, iM1 and M2 to explore phenotypic variation in extinct and extanct platyrrhines. Our results showed that morphological stasis explains the low phenotypic changes in extinct and exctant platyrrhines, probably due to ecological constraints, caused by phenotypic adaptation of platyrrhines to a relatively narrow ecological niche. Early and Middle Miocene platyrrhines shared some relatively similar molar shape patterns, whereas the Colombian fossils more closely resemble Alouatta and the Pitheciinae. The relation between both fossil samples could be due to: 1. All platyrrhine molar shapes share a primitive retention of the ancestral state. 2. An early divergence between two parallel shapes; a Callicebus-like and a Saguinus-like, which would be the ancestral precursors to all other forms. 3. Callicebus-like and Saguinus-like morphologies independently occurred in the early stem platyrrhines.This study develops a dental model based on molar shapes of 10.7717/peerj.1967/supp-1Supplemental Information 1Click here for additional data file.10.7717/peerj.1967/supp-2Supplemental Information 2Click here for additional data file.