Leishmania (L.) infantum (rA2-Ag) and evaluated for detection of anti-Leishmania antibodies in dogs compared to standard direct agglutination test (DAT).This study aimed to set-up latex agglutination test (LAT) and ELISA based on recombinant A2 from Iranian strain of The rA2-Ag was synthesized under a part of the A2 gene sequences which contain immune dominant sequences and less number of repetitive sequences. Latex beads, 0.8 μm  were sensitized with rA2-Ag. The tests were carried out on sera collected from 350 ownership dogs including symptomatic (n=67), asymptomatic (n=230) canine visceral leishmaniasis (CVL), and (n=53) uninfected domestic dogs as control group.Anti-leishmanial antibodies were detected in 97 (27.7%), 96 (27.4%) and 29 (%9) of the serum samples by using DAT, rA2-ELISA, and rA2-latex, respectively with ≥1:320 as a cut-off titer when DAT-confirmed cases were compared with the control groups. A combined sensitivity of 52% and specificity of 82.40% for rA2-ELISA and 23.8% and specificity 95.38%, respectively were found with ≥1:320 as a cut-off titer when DAT-confirmed cases were compared with the control groups. The concordance between rA2-ELISA and rA2 latex compared with DAT as a gold standard serological test for VL were found 73.7% and 77.5%, respectively.A good degree of agreement was found between rA2-ELISA and DAT (73.7%). rA2-ELISA could detect more seropositive serum samples than rA2-LAT and it may be recommended as an alternative tool for the diagnosis of CVL. L. Infantum transmitted by female sand flies and caused by Leishmania species which is fatal if left untreated (Canis familiaris) are the principal reservoir host of Mediterranean type of VL , or Kala-Azar, is a zoonotic disease due to ntreated . Domestipe of VL , 3.L. infantum infection as well as symptomatic dogs , the dogs represent cachexia and weakness, baldness, loss of appetite, weight loss, keratoconjunctivitis, and death . Asymptotic dogs , 4.Leishmania circulating antibodies by the leishmanial recombinant antigen, such as rK39 . The recombinant plasmid (pEASY-A2) and expression vector (pET22) were digested by EcoRI and NcoI enzymes. The products of digestion were analyzed by electrophoresis on 1% agarose gel. In order to express recombinant protein (pET-A2), E-coli BL21/DE3 cells were used as host cell. The expression of A2 was induced at OD 600 of 0.6 by adding of 1mM of Isopropyl-β-D-Thiogalactopyranoside (IPTG) and incubation at 37 °C. The A2 expression was analyzed by 12% SDS-Polyacrylamide Gel Electrophoresis (SDS-PAGE)  was transformed to Top10 DS-PAGE) . The purDS-PAGE) . The antThis study was conducted over a period of 2 yr from 2011 to 2013 in the Meshkin-Shahr district in the northwest part of Iran, where visceral leishmaniasis is endemic ,18.Leishmania antibody ≥ 1:320 by DAT was considered as symptomatic dogs. Positive samples with titer 1:80 and 1:160 by DAT with no signs and negative samples were considered to correspond to asymptomatic and uninfected dogs, respectively.Overall, 350 dog sera were collected and screened from both male and female domestic dogs of various breeds and ages. The animals were tested after obtaining ethical permission from the dog owners. An expert veterinarian was clinically examined the dogs. Dogs blood samples (2.5 ml) were collected by venipuncture, centrifuged at 800 g for 5–10 min, and the sera were separated and stored at −20 °C until tested. Each serum sample was tested for anti-leishmanial antibodies by using the standard DAT in the Leishmaniasis Laboratory of the School of Public Health at Tehran University of Medical Sciences . SuspectL. infantum antigens for this study were prepared in the protozoology unit of the School of Public Health at the Tehran University of Medical Sciences. Briefly, the procedure for preparing the DAT antigen was mass production of promastigotes of L. infantum in RPMI-1640 medium supplemented with fetal bovine serum (FBS), trypsinization of the parasites, staining with Coomassie Brilliant blue and fixing with formaldehyde. Serial dilutions of serum were prepared to start at a dilution of 1:10 to a maximum dilution of 1:20480. Antigen was added to each microplate wells. In position of a positive test, the wells represented a cloudy condition, otherwise during a negative result; the point is deposited at the bottom of well. All canine serum samples were tested by DAT according to this method  were sensitized with rA2 antigen according to the methods  with som3. One hundred microliters per well were used to coat 96-well microtiter plates  overnight at 4 °C. Free binding sites were blocked with a 1% BSA solution for 2 h at 37 °C. After three washes with PBS-Tween20 (PBS-T) at 0.05%, the plates were incubated with 100μL of canine sera diluted 1:100 in phosphate buffered saline (PBS-T) for 1 h at 37 °C. The plates were then washed three times with PBS-T and incubated with 1:20000 anti-canine IgG antibody  conjugated with horseradish peroxidase. The reaction was stopped by the addition of 100μL of a 1M solution of HCl per well. The optical density (OD) was measured at 450 nm in an ELISA microplate spectrophotometer . The lower limit of positivity (cut-off) was determined by the OD mean of 20 negative canine control sera plus two standard deviations.Protein concentrations were calculated by the Bradford assay  with bovSensitivity =TP/TP+FN Specificity =TN/TN+FP Concordance=TP+TN/Total samplesRelative sensitivity, specificity of test for detection of anti-leishmanial antibodies in dog sera were determined and the test parameters of the rA2-ELISA and the rA2-LAT were compared to the corresponding parameters of DAT, considered the gold standard for serological diagnosis of CVL.SDS-PAGE analysis of induced bacteria showed expression of rA2 protein, which was absent in un--induced bacteria. The antigenicity of A2 protein was surveyed in western blotting using both pooled dog sera and C9, an anti-A2 monoclonal antibody , 2.In position of a positive test by DAT, the wells represented a cloudy condition, otherwise during a negative result; the point is deposited at the bottom of well .A rapid and simple LAT was developed using rA2 antigen. While the serum was diluted 1/2 – 1/32 with PBS, strong agglutination with positive serum was observed at dilution of 1/32 and it is suggested for serum dilution. Positive results were read on a +1 to +4 scales depending on the extent of agglutination and if no agglutination was observed within 5 min the test was negative .Among these samples, 29 sera were positive by latex and the relative sensitivity, specificity and concordance of the test were based on DAT test. In symptomatic dogs the relative sensitivity and specificity of rA2-LAT were 25.71% (95% CI 16.01%–37.56%) and 95.53% (95% CI 92.14%–97.75%) respectively and concordance of 80.1%  while in asymptomatic dogs were 5% (95% CI 2.25%–7.86%) and 74.29% (95% CI 62.44%–83.99%) and concordance of 19.9%  with DAT test.ELISA procedure was optimized with regard to antigen concentrations and the serum dilution optimal antigen concentrations were 0.1μg/ml. When we tested sera by rA2-ELISA, this test could detect anti leishmainial antibody in 96 (27.4%) of sera and the relative sensitivity and specificity of rA2-ELISA in symptomatic dogs were 62.7% (95% CI 50.01% to 74.20%) and 80.92% (95% CI 75.85% to 85.33%) while in asymptomatic infected dogs, were 17.6% (95% CI 13.09% to 22.90%) and 48% (95% CI 37.90% to 58.22%), respectively with ≥1/80 cut-off titer of DAT-confirmed cases  & 2.In symptomatic and asymptomatic dogs the concordance of rA2-ELISA with DAT as a gold standard test were 77.4%  and 26.2% , respectively.In the present study, a rapid rA2- LAT was developed for the first time to detect visceral antibodies from dog samples. The sensitized beads could detect the antibodies against the canine VL (CVL). The results obtained with latex using samples collected from symptomatic and asymptomatic dogs indicated that the sensitivity of the test was 27.27%; however, its specificity was 94.64% in symptomatic dogs. LAT as a useful assay in difficult field conditions because it is simple, easy to perform, inexpensive, does not require any electric equipment, and is read visually for routine screening of VL in endemic areas where laboratory facilities are limited. The sensitivity of the test was low and must be developed in the future works.This is comparable with the only recombinant antigen used for detection of anti-leishmanial antibody by the rk39 latex test. This antigen is generally used in LAT at a concentration of 1 mg/ml in glycine-buffered saline. The sensitivity of the latex test among parasitologically proven index cases was only 80%, which was due to a lower power of antibody detection by this test . FractioL. infantum visceral infections in dogs. Sensitivity of rA2-ELISA was 76% in symptomatic dogs and 89% in asymptomatic dogs but the rA2 specificity was 96%  used for detection of rA2 antibody in dog’s sera. In their study, the sensitivity and specificity of this test were 52.58% and 82.21% respectively in symptomatic dogs . An ELIS was 96% . In contSerologic test performance depends on many factors such as infection status and the type of diagnostic antigen. There might be some differences in immune responses of CVL to VL and explain this disagreement but in serological tests, the problems due to sensitivity and specificity also arise .Leishmania parasites (Although there are several antigens represented by both amastigotes and promastigotes stages of arasites , 23, howarasites , 25.Leishmania antibodies in symptomatic dogs and but rA2-ELISA could detect more seropositive serum samples than rA2-LAT in symptomatic and asymptomatic dogs. However, more study is needed to develop the test by using other concentrations and sizes of latex beads in order to reduce the rate of false-negative reactions.Agreement of the rA2-LAT and rA2-ELISA when compared with the DAT as a gold standard were 77.4% and 26.2% respectively for rA2-ELISA and 80.1% and 19.9% for rA2-LAT in symptomatic and asymptomatic dogs respectively. Both LAT and ELISA test may be applied for detection of Anti-


To determine the outcome of early use of non-invasive positive pressure ventilation (NIPPV) in Pakistani patients with acute exacerbation of chronic obstructive pulmonary disease.2 were monitored and NIPPV was given for six hours to evaluate clinical outcomes and analyze the factors predicting failure . Frequency and percentages were calculated for qualitative variables while Mean and Standard Deviation for quantitative variables. Chi-square and t-test were used to see differences in pre and post NIPPV arterial blood gases.This descriptive study was conducted at Shifa International Hospital Islamabad from April 2015 to January 2017. A total of 120 patients with acute exacerbation of chronic obstructive pulmonary disease receiving NIPPV alongside standard therapy were included in the study. The patients were clinically assessed before starting on NIPPV. The parameters of respiratory rate, pH and paCO2 was 61.87±9.60 mm of Hg before and 57.46±6.79 mm of Hg after NIPPV (P < 0.0003). Twenty Four (20%) patients required invasive ventilation of which 19 (15.8%) patients could not survive.Patients’ mean age was 58.88±10.09 years. Males were 88 (73.3%) and females were 32 (26.7%). The mean respiratory rate was 24±1.45 per minute before and 17.96±1.35 per minute after NIPPV (p < 0.00001). The mean pH before NIPPV was 7.27±0.04 and afterwards 7.38±0.02 (p < 0.00001). The mean pCOThere was remarkable improvement in the arterial blood gases after NIPPV. However, the high mortality rate and significant number of COPD patients requiring mechanical ventilation necessitates further investigation into our population. Patients with ARF can be ventilated by either positive or negative pressure invasively or non-invasively.7The use of NIPPV for improvement in arterial blood gases (ABGs) in patients with acute exacerbation of COPD has also been studied in Pakistan.This was a prospective hospital based descriptive study carried out in Department of Medicine at Shifa International Hospital, Islamabad. This was a longitudinal study conducted over a period of 21 months from April 2015 to January 2017. A total of 120 patients with acute exacerbation of chronic obstructive pulmonary disease were included using non-probability (consecutive) sampling technique. Sample size was calculated as 120 using WHO sample size calculator considering 95% confidence level, absolute precision 6% and anticipated population proportion p=12.5 %.2 > 45mmHg and PaO2 < 60 mmHg. Patients with following conditions were excluded from the study:1) Arterial blood pH < 7.20 2) Unconsciousness 3) Unable to clear their airways 4) Un-cooperative, agitated 5) Hemodynamic instability 6) A central nervous system disorder unrelated to hypercapnia associated encephalopathy or hypoxemia.Patients with COPD exacerbation fulfilling the following criteria were included in the study: 1) All age groups 2) Both males and females 3) Dyspnoea at rest or respiratory rate > 25 breaths per minute and oxygen saturation less than 85%. 4) FEV 1<50% predicted or PEFR < 100 litter per minute. 5) ABGs showing pH<7.35, Pa Co2 < 55 mm of Hg and O2 saturation more than 90% on ABG was required before patients were considered for weaning from NIPPV. In case of deterioration in patient’s clinical condition and/or ABGs, patients were shifted to Intensive Care Unit (ICU) and followed for a week in terms of requirement for mechanical ventilation and any hospital mortality following intervention failure was recorded.Institutional review board approval was obtained before starting data collection.  Written informed consent was taken and a detailed history followed by rigorous clinical examination of the patients were documented. Respiratory rate and ABGs were taken before Non-invasive positive pressure ventilation (NIPPV). NIPPV was given to patients for at least 6 to 8 hours, during which time the patients were assessed for clinical stability. Clinical signs and ABGs were checked again after 6 to 8 hours of NIPPV. The presence of sustained clinical improvement with decrease in respiratory rate < 24/minutes, presence of normal pH, PaCO2 and pCO2 before and after NIPPV. Effect modifiers like age and gender were controlled by stratification. Post stratification paired t-test was applied to see difference in the pre- and post- NIPPV quantitative variables. chi-square test was used for qualitative variables. A p-value ≤ 0.05 was considered as significant.Data was analyzed on SPSS Version 10. Frequency and Percentages were computed for qualitative variables like gender, mortality and intubation. Mean and Standard Deviation were computed for quantitative variables like age, respiratory rate, pH, pO2 are given in 2) were measured before and after NIPPV. The mean respiratory rate of the patients was 24 ± 1.45 per minute before and 17.96 ± 1.35 per minute after NIPPV. This difference was statistically significant with p value of less than < 0.00001. The individual variation in respiratory rate are shown in Out of a total of 120 patients with acute exacerbation of chronic obstructive airway disease, males were 88 (73.3%) and females were 32 (26.7%). Patients age ranged from a minimum of 40 years to a maximum of 75 years with a mean age of 58.88 ± 10.09 (p=0.002). Descriptive statistics for respiratory rate, pH and pCO2 was 61.87 ± 9.60 mm of Hg before and 57.46 ± 6.79 mm of Hg after NIPPV. This difference in pCO2 levels was also statistically significant (P < 0.0003). Overall NIPPV was successful in 80% of the patients where the patients were weaned off from NIPPV and discharged home safely.The mean pH before NIPPV was 7.27 ± 0.04 and afterwards was 7.38 ± 0.02 with statistically significant difference (p < 0.00001) as shown in In our study, twenty four (20%) patients required endotracheal intubation and invasive ventilation compared to 96 (80%) patients who were safely weaned off which was statistically significant (p = 0.011). Out of the 24 patients who were intubated, only five survived making total mortality in hospital as 15.83% (19 out of 120 patients).2 and decreasing PaCO2 and reduction in the diaphragmatic activity.14We studied the effect of NIPPV on respiratory rate, arterial blood gases, the number of patients who could be discharged home safely, the need for mechanical ventilation and mortality rate in patients with acute exacerbation of COPD. There was significant difference in term of improvement in respiratory rate and ABGs from the baseline. Our results coincide with earlier studies which reported similar findings for respiratory rate and ABGs.In our study, 96(80%) patients were discharged home safely while 24 (20%) required mechanical ventilation. All of the 24 patients who did not improve with NIPPV were given mechanical ventilation out of which only 6 (4.2%) recovered. This requires further research into the management of patients who don’t improve with NIPPV. Better hospital survival, fewer intubations, less complications and a shorter hospital stay was found in some studies.2 and respiratory rate and significantly reduced the length of hospital stayWhile the findings of our study were comparable to the previous studies This was a hospital based cross-sectional study. Large scale hospital-based studies are required as the prevalence of COPD is gradually increasing in Pakistan. While we didn’t use a standard control group, we did compare our results to the data available in Pakistan. Long term follow-up was not done in these patients which could have better explained the outcome. Effect of comorbid conditions was not analyzed which could have contributed to mortality.There was significant improvement in biochemical and clinical parameters after non-invasive positive pressure ventilation compared to baseline. However, the number of patients with acute exacerbation of COPD in our study requiring mechanical ventilation and the high mortality rate shows that it was underreported from Pakistan previously necessitating the need for further research in our country.NI: Conceptualization of study, study design, data collection, analysis and write up.NG: Study design, Literature review, data analysis and write up of manuscript.NZ: Study design, data collection and review of the manuscript.


Estimating impacts of offshore windfarm construction on marine mammals requires data on displacement in relation to different noise levels and sources. Using echolocation detectors and noise recorders, we investigated harbour porpoise behavioural responses to piling noise during the 10-month foundation installation of a North Sea windfarm. Current UK guidance assumes total displacement within 26 km of pile driving. By contrast, we recorded a 50% probability of response within 7.4 km (95% CI = 5.7–9.4) at the first location piled, decreasing to 1.3 km (95% CI = 0.2–2.8) by the final location; representing 28% (95% CI = 21–35) and 18% (95% CI = 13–23) displacement of individuals within 26 km. Distance proved as good a predictor of responses as audiogram-weighted received levels, presenting a more practicable variable for environmental assessments. Critically, acoustic deterrent device (ADD) use and vessel activity increased response levels. Policy and management to minimize impacts of renewables on cetaceans have concentrated on pile-driving noise. Our results highlight the need to consider trade-offs between efforts to reduce far-field behavioural disturbance and near-field injury through ADD use. This Opportunities to address these data gaps by tracking behavioural responses of tagged individuals or using arrays of passive acoustic monitoring (PAM) sensors are rare –12. AlteHarbour porpoises are the most common marine mammal in many areas exposed to offshore energy developments ,22. The In this study, we aimed to inform these policy and management decisions by investigating two key questions. (1) How do harbour porpoise behavioural responses to construction noise vary in relation to: (a) received noise levels; (b) distance from piling; (c) time since the start of construction; and, (d) the duration of individual piling events? (2) To what extent is this response modified by: (a) ADD use prior to piling; and (b) vessel activity? The overall aim was to estimate a proxy porpoise dose–response curve to construction noise in order to refine predictions of the number of individuals displaced by pile-driving.2.Following the approach used to study responses of harbour porpoises to a seismic airgun survey , we usedet al. [Underwater noise levels were recorded between March and October 2017 at six locations  using auet al. ; (2) theet al. ,36, and et al. ; and (3)et al. .Figure www.chelonia.co.uk). We assume that variations in echolocation click detections provide a robust index of changes in the occurrence of porpoises, as indicated by previous work in this study area demonstrating relationships between echolocation detections and two independent measures of relative density derived from visual and digital aerial surveys [Spatio-temporal variation in the occurrence of porpoise echolocation clicks was measured using V.O and V.1 CPODs ( surveys . CPODs w surveys ) were es surveys  or seaso surveys . Harbour surveys  and as tTo characterize baseline variation in day to day changes in occurrence, we used data from 7th to 16th March 2017 before piling began and from a second period from 7th to 16th December 2017 after piling had stopped from 12 similar offshore sites located outside the wind farm site at least 25 km from the construction site. These data were used to produce a null distribution of proportional change in occurrence (DPH) by randomly sampling 1000 times from 9th to 15th March 2017 and from 9th to 15th December 2017 for each site and determining the proportion change in the number of DPH in the 24-h period following each randomly selected time relative to the number of DPH in the 24-h period 2 days prior to it . Using tThe probability that porpoise occurrence did (1) or did not (0) show a response to piling was modelled as a binomial response with a probit link function  using ge3.Harbour porpoises were present within the windfarm construction site throughout the construction period in 2017 . The numn = 9) preceded by a longer 192-h break in piling . Based on the relationship with distance from piling, at the start of the construction period in April there was greater than or equal to 50% chance of harbour porpoises responding to piling in the 24-h period after piling at distances up to 7.4 km (95% CI = 5.7–9.4) from piling (a). By the 47th location in July, this threshold had decreased to 4.0 km (95% CI = 2.7–5.2), declining further to 1.3 km (95% CI = 0.2–2.8) by December, when the final (86th) location was piled. Similarly, there was a greater than or equal to 50% chance of porpoises responding in the 24-h period after piling to audiogram-weighted SEL of 54.1 dB re 1 µPa2 s (95% CI = 52.0–56.7) at the first location piled, increasing to 60.0 dB re 1 µPa2 s (95% CI = 57.5–63.4) by the 47th location and 70.9 dB re 1 µPa2 s (95% CI = 63.0–87.0) by the final location (b). For the relationship with unweighted single-pulse SEL, there was a greater than or equal to 50% chance of porpoises responding in the 24-h period after piling to unweighted SEL of 144.3 dB re 1 µPa2 s (95% CI = 142.1–146.8) at the first location piled, increasing to 150.0 dB re 1 µPa2 s (95% CI = 147.5–153.6) by the 47th location and 160.4 dB re 1 µPa2 s (95% CI = 153.2–178.9) by the final location .For both 12-h and 24-h responses, harbour porpoise responses were best explained by the interaction between the cumulative number of locations piled and either distance from piling on a logarithmic scale or audiogram-weighted single-pulse SELs . The samm piling a. By thelocation b. For thp = 0.40), but ADD use was a significant covariate in models of the 12-h response  and did not change the model. There was no support for including ADD use in models of the 24-h response  piling events during construction, when operational or weather conditions had caused delays that provided a suitable baseline period. This also meant that we were unable to examine finer-scale or instantaneous responses. Consequently, the results represent a response to cumulative exposure to pile driving, vessel activities and the use of ADD. Part of the rationale for focusing our analysis on these baseline and reference periods was to minimize potential effects of poor signal-to-noise ratio on detection probability during periods of piling. Porpoise click detections on CPODs decrease with increasing noise around oil and gas platforms, particularly between 20 and 160 kHz . By starAnother consideration is whether decreases in detections resulted from changes in vocalization rates rather than displacement. One of two tagged harbour porpoises exposed to close vessel passes ceased echolocating for several minutes , therefoet al. [Finally, detection probability will be affected by a number of environmental variables including depth, temperature, salinity and bottom substrate. In the Baltic, Carlén et al.  estimate4.2.−2 [Given the widespread distribution and abundance of harbour porpoises in the North Sea, potential disturbance impacts on protected populations must be considered within consent applications for most, if not all, wind farm developments in this region. Several modelling frameworks now exist for predicting the population effects of such disturbance –7, but t−2 , the JNCPolicy instruments in many countries aim to reduce anthropogenic noise that may adversely affect the marine environment . HoweverEfforts to reduce behavioural disturbance must also be balanced against efforts to reduce the risk of near-field injuries as a result of loud impulsive pile-driving noise. In Germany, it is mandatory to deploy an ADD at least 30 min before piling to mitigate the risk of physical injury. Consequently, most previous observational studies of harbour porpoise responses to pile driving noise, represent responses to the cumulative impact of ADDs, pile driving and associated construction vessels. For example, ADDs were used during the construction of all the commercial windfarms in the studies cited in table 1 in . ADDs wePolicy and management measures aimed at minimizing the environmental impacts of wind farm construction on marine mammals have tended to focus on high-energy pile-driving noise. These impulse noise sources are a more significant risk with respect to near-field injuries, but there is more uncertainty over how noise influences behavioural reactions. If animals are responding to a perceived threat, then their reactions may be more closely related to an individual's distance to the source rather than received noise levels. Either way, their perception of loudness or other signal characteristics will depend on the shape of their audiogram  leading


The aim of this study was to investigate the effects of aquatic exercise on mental health, functional autonomy and oxidative stress parameters in depressed elderly individuals.Initially, ninety-two elderly individuals were included in the study and were allocated into the depression group (n=16) and nondepression group (n=14). Both groups engaged in the aquatic exercise program for 12 weeks, including two weekly sessions (45 min/session) at a low intensity  throughout the intervention. All outcomes were evaluated at baseline and 12 weeks later.p<0.005).The patients were 63.5±8.8 years old. The following scores were decreased after training in the depressed group: depression (53%), anxiety (48%), and Timed Up & Go (33%). The following scores increased: Berg Balance Scale (9%) and flexibility (44%). Regarding the blood-based parameters, there were decreases in protein carbonylation (46%) and nitric oxide (60%) and increases in glutathione (170%) and superoxide dismutase (160%) in the depression group (The aquatic exercise program reduces depression and anxiety, improves functional autonomy and decreases oxidative stress in depressed elderly individuals. Decreased physical activity can contribute significantly to increased levels of depression Recently, research in this area has been focused on mental health markers, which reflect common pathophysiological processes, related to the loss of functional autonomy and increased oxidative stress that are thought to be involved in depression Although studies have shown significant improvement in the depressive symptoms of elderly individuals after the regular practice of physical exercises This study was approved by the National Health Council, National Research Ethics Commission resolution CNS 466/12 for research involving humans and approved by the local Ethics Committee University Extreme Sul Catarinese (CAEE 47120815.3.0000.0119).All participants provided written consent prior to participation.This longitudinal clinical study, conducted for 12 weeks, subjected both the depression group and the nondepression group to the aquatic physical training program. Two days before and 48h after the program, parameters related to mental health, oxidative stress and physical fitness were analyzed.Initially, 56 patients were screened with a medical diagnosis of major depression and confirmed by the medical clinic psychiatrist at the local University, and 36 nondepressed elderly individuals participated in the study. Of the 56 patients, 29 did not meet the inclusion criteria, and 7 declined to participate. Thus, 20 patients (9 men and 11 women) with depression, aged between 50 and 80 years old, were included in the study and formed the depression group. Of the 36 nondepressed individuals, 12 did not meet the inclusion criteria, and 4 declined to participate. Thus, 20 (13 men and 7 women) elderly individuals without depression, aged between 50 and 80 years old, were included in the study and formed the nondepression group. Both groups were subjected to the same physical training program at the same place, time and days of the week. (Patients with a diagnosis of major depressive disorder (MDD), according to established criteria , and healthy elderly individuals (according to WHO criteria) were recruited through advertising in local newspapers. The criteria for inclusion in the depression group included the following: had MDD, were not engaged in regular exercise , were 50 years old or over, and had a recent medical release for exercise practice (in the 6 months preceding the study). The exclusion criteria were current drug abuse, psychotherapeutic treatment, contraindications to physical exercise, suicidal behavior according to the Hamilton depression rating scale Patients underwent a 12-week aquatic training program with two weekly sessions, lasting 45 min. In both groups, the intensity was similar. Heart rate (HR) was monitored every 10 sec , and the rate of perceived exertion  was measured in the final 20 sec of each stage. The aquatic training classes were always held in the afternoon in a pool with a depth of 1.20 m, measuring 25 m×12.5 m, with water in the approximate temperature range of 26 to 28°C.Each 45-min session was divided into a warm-up period (5 min) and the main training program (40 min), followed by a cool-down period (5 min). Both groups underwent interval-training programs with the following schedule: 9 exercises in each session; each exercise was performed as 4 sets of 30 sec with 10-sec intervals; two weekly sessions (Tuesdays/Thursdays). Exercise intensity was measured by a heart rate monitor and maintained in a low-intensity range (50% and 60% of maximum HR) or a Borg scale score of 13 to 14 points. The exercises involved the large muscle groups, with upper and lower limbs together. Both groups had identical training protocols .These tests were conducted 48h before the first session and 48 h after the last training session in the patients for a clinical evaluation and included assessments of mental health, functional mobility, and oxidative stress. Before and after the intervention, an alimentary assessment was performed over 3 consecutive days.The Beck Depression Inventory (BDI) For the Timed Up & Go (TUG) test, individuals were seated in a chair (45 cm high) with their back against the chair. They were instructed to stand, walk 3 m following a straight line on the ground as fast and safely as possible, return to the chair, and sit in the starting position Protein oxidation was measured by the formation of carbonylated protein derivatives using 2.4-dinitrophenylhydrazine read spectrophotometrically at 370 nm Eight-milliliter samples of blood were obtained from the antecubital vein. Blood was collected in vacutainers without additives and centrifuged at 1500 rpm for 10 min at 4°C. Aliquots of red blood cells and serum were stored at -70°C until used in the biochemical assays.2 test for nonparametric analyses was also used and followed by the Bonferroni post hoc test. The a priori sample size was calculated based on a predicted difference of 0.5% in the levels of SOD=0.05 and power of 0.90, performed with IBM Statistical Package for the Social Sciences (SPSS) software , that was based on the study of Silva et al. p<0.05. SPSS version 21.0 was used as the statistical software.All analyses were performed by blinded evaluation. The data are expressed as the means ± standard errors of the mean (SEM). The Kolmogorov–Smirnov test was used to confirm normality. The χThe study participants were divided into two groups: depression and nondepression. In the depression group, 56 patients were assessed for eligibility, 20 were allocated to the group, and 16 were included in the exercise program and analyses. The four individuals who were not included in this analysis were 2 who dropped out of the study and two who had inadequate adherence (<90%). In the nondepression group, of the 36 patients assessed for eligibility, 20 were randomized and 14 were included in the analyses and exercise program. The six individuals who were not included in this analysis included 2 who dropped out of the study and four who had inadequate adherence (<90%). To perform an intention-to-treat analysis, all participants were invited to undertake before and posttraining evaluations. The distribution of individuals can be observed in p>0.05). The values are presented as the means ± SEM.Age, duration of depression, body mass, BMI, sex, and medical treatments are presented in p<0.05).As observed in p<0.01). The control group did not show significantly altered results on the depression scale after (4.2±1.1) compared to before (5.2±1.9) the program (p>0.05). Our results indicated that clinically, there was a 53% decrease in the scores for depression in the depressed elderly individuals.The results shown in p<0.05). The control group did not show significantly altered results for anxiety after the program (4.5±3) compared to before the program (4.7±1) (p>0.05). Our results indicated that clinically, there was a 48% decrease in the anxiety scores in the depressed elderly individuals.As shown in p<0.05). The nondepression group did not have significantly altered results on the TUG test after (7.4±0.3 sec) compared to before (7.1±0.4 sec) the program (p>0.05). In the Berg test, the results showed a significant increase after the aquatic exercise program in the depression group (55.5±0.2) compared to before the program (50.8±1.3) (p<0.05). The nondepression group did not show altered results after (56±0.1) compared to before (56±0.3) the program. In addition, the results demonstrated a significant increase in the flexibility of the hamstring muscles after the aquatic exercise program in the depression group (26±2 cm) compared to before the program (18±1.8 cm) (p<0.05). The nondepression group did not have altered flexibility after (27±1.6 cm) compared to before (26±3.8 cm) the program. Our results indicated that clinically, there was an increase of 33% in dynamic mobility, 9% in static mobility and 44% in levels of flexibility in the depressed elderly individuals.The results in p<0.01). The nondepression group did not show significantly altered oxidative damage after (0.13±0.02 nmol/mg protein) compared to before (0.14±0.03 nmol/mg protein) the program (p>0.05). Regarding nitric oxide, the results showed a significant decrease in its production after the aquatic exercise program in the depression group (11.7±3.5 nmol/mg protein) compared to before the program (28.2 ±2.5 nmol/mg protein) (p<0.001). The nondepression group had significantly increased levels of this marker after (12.8±2.5 nmol/mg protein) compared to before (5.56±2.8 nmol/mg protein) the program (p<0.05). Regarding superoxide dismutase, the results showed significantly increased SOD after the aquatic exercise program in the depression group (1.3±0.3 nmol/mg protein) compared to before the program (0.5±0.01 nmol/mg protein) (p<0.05). The nondepression group did not show altered SOD after (1.44±0.3 nmol/mg protein) compared to before (1.3±0.2 nmol/mg protein) the program (p>0.05). In addition, glutathione significantly increased after the aquatic exercise program in the depression group (4.17±0.8 nmol/mg protein) compared to before the program (1.54±0.07 nmol/mg protein) (p<0.01). The nondepression group did not show altered GSH results after (2.88±0.03 nmol/mg protein) compared to before (2.97±0.08 nmol/mg protein) the program (p>0.05). Our results in the depressed group indicated that clinically, there was a reduction of 46% in protein carbonylation and 60% in nitric oxide, and there was an increase of 170% in GSH and 160% in SOD.As observed in Food intake, macronutrient distribution  and daily calorie intake were not altered by the 12-week training period (data not shown).The present study demonstrates that a low-intensity aerobic training program in the aquatic environment can contribute to the treatment of depression by reducing anxiety and depression scores, improving functional autonomy and decreasing oxidative stress.The reduction in the depression scores found in the present study demonstrated the efficacy of aquatic training on depression in elderly individuals and corroborates previous meta-analyses In the current study, anxiety levels were reduced among the depression group; these findings were consistent with previous studies The anatomical and physiological changes observed in the elderly population have been associated with increased disability, frailty and falls Depression may be accompanied by increased oxidative stress and a decreased antioxidant system A recent study suggested the involvement of NO mechanisms in the pathogenesis of depression There is also evidence to suggest that antioxidant enzymes are decreased in people with depressive symptoms On the other hand, there is some evidence to suggest that antidepressants have antioxidant properties and may act by reducing ROS production and improving antioxidant levels, thereby reducing oxidative stress All of the authors participated in the design, interpretation of studies, data analyses and manuscript review. Silva LA, Tortelli L, Motta J, Menguer L, Mariano S, Tasca G and Silveira GB performed the experiments and were responsible for the laboratory analysis of mental health, functional autonomy and oxidative stress. Silva LA and Silveira PCL wrote the manuscript and performed the statistical analysis.


This study’s objective was to assess, on a national scale, residual risks of death, major disease-related events, and quality of care during the first five years in children diagnosed at birth with sickle cell disease (SCD). Data were retrospectively collected from medical files of all children with SCD born between 2006–2010 in France. Out of 1792 eligible subjects, 1620 patients  had available follow-up data, across 69 centers. Overall probability of survival by five years was 98.9%, with 12/18 deaths related to SCD. Probability of overt stroke by five years in SS/SB° patients was 1.1%, while transcranial Doppler (TCD) was performed in 81% before three years of age. A total of 26 patients had meningitis/septicemia . Prophylactic penicillin was started at a median age of 2.2 months and 87% of children had received appropriate conjugate pneumococcal vaccination at one year. By five years, the probability of survival without SCD-related events was 10.7% for SS/SB° patients. In contrast, hydroxyurea was prescribed in 13.7% and bone marrow transplant performed in nine patients only. In this study, residual risks of severe complications were low, probably resulting from a good national TCD, vaccination, and healthcare system coverage. Nonetheless, burden of disease remained high, stressing the need for disease-modifying or curative therapy. Newborn screening (NBS) programs for sickle cell disease (SCD) aim to reduce early mortality and morbidity by introducing preventive measures directed towards major disease-related identified risks: Pneumococcal infection, acute splenic sequestration, and neurovascular injury. Prophylactic intervention such as penicillin and pneumococcal vaccination, parental education, and neurological screening by transcranial Doppler (TCD) have demonstrated significant efficacy in reducing these risks in children ,2,3,4. ROnly very few nation-wide data are available, however, regarding both medical care and residual early morbidity and mortality in countries where newborn screening and prophylactic measures are implemented. Only such studies may in fact address disparities across regions, centers, or countries and provide a true-to-life evaluation.A SCD NBS program was fully implemented since 2000 in France, targeting newborns identified at risk in mainland France and all newborns in French overseas territories. More specifically, regarding the targeted SCD NBS program, newborns are screened when parents originate from a country where SCD is endemic  and/or in case of family history of SCD and/or in case of uncertainty regarding both previous issues, in accordance with national guidelines . FollowiThe main objective of this retrospective study was to assess, on a national scale and during the first five years of life, residual risks of death, overt stroke, and bacterial meningitis/septicemia resulting from current transcranial Doppler (TCD) and pneumococcal prophylactic utilization in SCD children diagnosed at birth. The secondary objective was to evaluate in these young children the frequency of other main SCD-related events and the use of disease-modifying or curative therapy (hydroxyurea (HU), transfusion (TF) programs, and hematopoietic stem cell transplantation (HSCT)).We retrospectively identified all patients born between 1 January 2006 and 31 December 2010 diagnosed with SCD through the national NBS program in mainland France, French West Indies , and Réunion-Mayotte. Results of NBS indicating SCD were fed back to a referral sickle cell center where confirmatory tests were performed, and results were explained to parents. The follow-up was thereafter organized in the closest sickle cell center according to the child’s residence. Sickle cell centers were formally identified in a national network in 2004. Patients diagnosed in French Guyana were not included in the study because at the time of data collection, cross identification of patients following NBS was not possible in that region.Data were collected in 2014/2015 from the patients’ medical files up to the age of 5 years regarding age at first prescription of penicillin prophylactic therapy, occurrence of death/overt, stroke/bacterial meningitis or septicemia, first event of acute splenic sequestration, acute chest syndrome/pneumonia, vaso-occlusive crisis (VOC), age at first transfusion, as well as use of pneumococcal vaccines/first chronic TF program/hydroxyurea/hematopoietic stem cell transplantation and TCD results.Patients were considered lost to follow-up when no data were available during the 2-year period of data collection i.e., 2014/2015 in any center in France. In addition, patients residing abroad and who were seen only occasionally in France were excluded from analysis.Fatal cases were analyzed in detail and the numbers were checked with the National French Death Registry for exhaustivity purpose.TCD velocities were considered abnormal when ≥2 m/s and conditional when ≥1.7 m/s but <2 m/s (according to the STOP criteria ), as recVaso-occlusive crisis (including dactylitis) was defined as an acute non-infectious, non-traumatic pain requiring analgesics for more than 12 h and/or hospital admission.Acute chest syndrome (ACS) was defined as a new pulmonary infiltrate on chest X-ray, with or without pain, cough, fever (≥38.5 °C), or hypoxemia. Given the overlapping definition of pneumoniae in young children, the latter events were pooled with ACS.Acute anemic events were defined by reductions in hemoglobin ≥20% versus steady state.Acute splenic sequestration (ASS) was defined as splenic enlargement increased ≥2 cm from baseline associated with acute anemia.Stroke was defined as an ischemic or hemorrhagic event lasting >24 h and resulting in focal neurologic deficit.Other clinical outcomes were collected as reported in the medical file and no further verification was performed. However, the following definitions are consensual in France:During the study period, national guidelines recommended prophylactic penicillin to all children with SCD at least until the age of 5, pneumococcal vaccinations using conjugate vaccines  in the first year of life  from April 2010) followed by a boost during 2nd year, and a dose of polysaccharidic 23 valent pneumococcal vaccine (P23) at 2 years. Annual TCD screening was recommended from 2 to 16 years in patients with SS or SB°-thalassemia (SB°) disease only  and was Chronic transfusion program was defined as any transfusion program that lasted >3 months. Hydroxyurea was recommended by national guidelines in 2005 for adults and children over 2 years with recurrent vaso-occlusive crises and/or acute chest syndrome but was specifically licensed with adapted pediatric dosage for patients with SCD in 2007 only.This study (NCT 03119922) was approved by the Inserm Ethics Committee/Institutional Review board (13–118), the French National Committee for Computerized Databases (CNIL-127020), and by EC/CCTIRS (13–118/12319). In accordance with the French regulation authorities, given the observational retrospective nature of the study, families were informed about the study by their physician including the possibility to decline participation but were not required to give written informed consent.The results are expressed as numbers and percentages for categorical variables and as median  for continuous variables. The distribution of variables was analyzed using the chi-square test for categorical variables and the Mann–Whitney test for continuous variables.+thalassemia (SB+), we stratified the analysis into two groups, SS/SB° (including SDPundjab) and SC/SB+ (including HbSE) patients, whenever appropriate. Survival curves were compared using the log-rank test. Statistical analyses were performed using STATA version 14.1 . Overall and age-specific rates of mortality and stroke were calculated as the number of deaths/strokes divided by total person–years at risk.Entry into the study cohort was defined by the date of the birth. Data were censored at the date of 5 years or last follow-up in 2014/2015 or death. Assessment of overall survival and survival without specific SCD-related complications were generated by the time to first event. Kaplan–Meier survival estimates and the 95% confidence interval (95% CI) were calculated for all SCD-related complications. Given that patients with SS or SB° tend to have a more severe clinical course than those with SC or S-betaBetween 01/01/2006 and 31/12/2010, 1801 consecutive newborns with SCD were identified by the National NBS program . Nine faOf note, the targeted nature of the NBS program in mainland France during our study period most likely did not leave out a significant number of SCD cases: A large retrospective study over the period ranging from 2005–2017 found 24 missed cases, representing 0.6% of the screened cohort during that same period . In addition, during our study period (2006–2010), the mean national proportion of babies targeted for SCD at birth in mainland France was 30%, a rather large proportion.+ disease. Main characteristics of the cohort are shown in A majority of children (71.8%) was diagnosed with SS or SB° and 20.3% with SC or SBFollow up was performed in 69 centers across four main French regions: Paris area, province, French West Indies (Martinique-Guadeloupe-St Martin), and Réunion-Mayotte. Over a half of patients (59.6%) were residents in the greater Paris area, while 9.4% resided in the French West Indies where a substantially greater fraction of patients had SC disease (42%).Among the 69 centers, a majority  followed less than 20 patients included in the study: 17 (24.6%) centers followed between 20–50 patients; 10 centers (14.5%) followed more than 50 patients .Altogether, among the patients included in EVADREP cohort, 1540 (95.1%) had a regular follow-up while 80 (4.9%) were temporarily lost to follow-up at some point during the study period—during the first 6 months in 31(1.9%) or later during at least a 2 year period in 49 (3%). These figures did not change significantly across regions of birth, but did according to the size of the centers, the largest centers having the lowest percentage of patients lost to follow up, both following NBS or thereafter . In addi• Survival and mortality rates and causes+ patients, respectively  and was 98.6% (97.7–99.1) and 99.6% (98.2–99.9) in SS/SB° and SC/SBectively .+ children.Mortality rate for all patients was 0.31/100 person–years (0.17–0.58) and 0.23/100 person–years (0.15–0.37) at two years and at five years, respectively. When analyzing by disease severity, mortality rate was 0.29/100 person–years (0.18–0.47) at 5 years in children with SS/SB° as opposed to 0.09/100 person–years (0.02–0.36) in SC/SBA total of 18 patients died at a median age of 1.6 years (range: 0.01–4.2) with 10 of these deaths occurring before age 2. Among all deaths, 12 (66.7%) were SCD-related, all but one in patients with SS/SB° . Over ha• Stroke and TCD coverageProbability of overt stroke by five years of age was 1.1% (0.6%–1.9%) in SS/SB° patients only, as illustrated in n = 1164) was performed before two and three years of age in 56% and 81% of patients, respectively. A total of 81 (7%) children had not benefitted from TCD screening at five years, with comparable numbers of unscreened children across regions. However, the proportion of unscreened children varied according to the center size with the lowest% of unscreened children in the two largest centers (5 (2.4%) versus 15 (16%) in the smallest centers)  . The medThe overall probability of abnormal TCD (velocities ≥200 cm/s) at five years of age was 10.4% (8.6–12.4). This probability varied across regions with the highest probability in the Paris area (13.3% (10.8–16.2)) and was significantly higher in the two largest centers (20.0% (15.0–26.5)) . The pro• Severe Infection and infection prophylaxis coverageA severe infection (meningitis or septicemia) developed in 26 patients at a median age of 2.3 years (1.3–2.6), in all but two patients with SS/SB° disease. Bacteria was identified in 21 patients, with Streptococcus pneumoniae in nine patients (including one patient with SC disease). Of them, 8/9 had been fully vaccinated with PCV7 and 3/5 aged over two years with 23-valent pneumococcal polysaccharide vaccine (P23).The outcome of severe infection was fatal in eight cases (30.8%), following pneumococcal infection in half the cases.Overall probability of severe infection at five years was 1.6% (1.1%–2.4%) in children with SS/SB° while it was significantly lower 0.4% (0.1–1.8) in patients with SC/SB+.• Penicillin prophylaxis and vaccination coverageOver 99% of all SCD patients had been prescribed prophylactic penicillin, which started at a median age of 2.2 months of life (1.7–3.0). Regarding vaccination, 87% of children received at least three doses of either PCV7 or PCV13 during their first year of life. However, only 768 (47.4%) children had been fully vaccinated before age 3, i.e., had received at least four PCV and one P23. P23 was administered in 65% of patients before three years and the probability of receiving P23 was of 90% at five years. The pneumococcal immunization coverage increased according to the size of the center, ranging from 24.8% to 78.8% in centers with <10 patients to centers >100 patients . Convers+, respectively, illustrated in At three years, 42.9% of the EVADREP cohort had experienced a first VOC and the probability of survival without VOC was 37.6% (35.2–40.0) at five years. Expectedly, this probability differed significantly according to genotypes and was 29.0% (26.3–31.7) and 59.6% (54.8–64.1) in SS/SB° and SC/SBp < 0.001); see also A total of 549 children developed at least one episode of either pneumonia or ACS or both leading to a five-years probability of survival without experiencing a first episode of pneumonia/ACS of 64.8% (62.4–67.2). This probability was significantly higher for patients with SC/SB+ genotype than for those with for SS/SB° . This complication was fatal in two cases. This probability differed significantly (97.1% (95–98.3)) in patients with SC/SBWhen pooling all SCD-related events , 58.6% of children had experienced at least one event at three years. The probability of survival without SCD-related events at five years was 10.7% (8.9–12.6) for SS/SB° patients versus 46.3% (41.5–51) for SC/SB+.At three years, 78.1% of all children had experienced a first hospitalization and the probability rose to 89.9% (88.3–91.3) by five years of age.• Transfusion+ genotypes (+ disease was splenectomized at three years). Cholecystectomy concerned only children with SS/SB° with a probability of 3.7% (2.7–5.1) by five years of age.The probability of being transfused at least once at five years was 65.2% (62.3–68.0) in SS/SB° patients and 11.8% (9.1–15.2) in patients with SC/SBenotypes . Splenec• Chronic transfusion:Overall, the probability by five years of age to have benefitted from a chronic TF program was 13.2% (11.6–15) and concerned a total of 204 patients, all but one with SS/SB°. Indication for initiation of the first TF program in SCA patients was stroke in 10 cases (4.9%); acute splenic sequestration in 75 (36.8%); abnormal TCD in 76 (37.2%), and other reasons in 42 (20.6%), including vaso-occlusive events in 22 cases, additional neurovascular reasons in 14, and severe anemia in 6. At the time of data collection, the TF program was ongoing in 101 (49.5%) patients and had been halted for the remaining 103 children. Out of 60 children who were initially transfused for acute splenic sequestration and discontinued, 39 (65%) were thereafter splenectomized; among 23 children who discontinued TF initiated for abnormal TCD, 12 were thereafter switched to HU, 4 underwent HSCT, and 5 had no specified subsequent treatment. Among 15 who stopped TF initiated for ACS/VOC, 14 were switched to HU, while the last patient had no subsequent treatment. The remaining five children had miscellaneous indications for both initiating and stopping TF. The mean duration for those in whom the program was discontinued was 1.1 year (0.7–1.9). Probability of receiving a first transfusion program by five years of age was significantly increased in Paris area versus other regions and significantly increased in the largest centers .• Hydroxyurea (HU):+ disease was treated with HU. At five years, among the 145 children who were initiated on HU, 137 (94.5%) were still on treatment. There was no major difference in the proportion of treated children according to regions, but there was a significant difference according to the size of the centers, with an increased probability in the largest centers  and the probability was 13.7% (11.7–15.9) at five years. No patient with SC/SB centers .• Hematopoietic stem cell transplantation (HSCT):Only nine patients with SS/SB° underwent HSCT before the age of five, at a median age of 4.1 (4.0–4.4). Indication for HSCT was alloimmunization in one case, VOC despite HU in one case, acute splenic sequestrations in one case, and cerebral vasculopathy in six cases .This study allows for the first time, to the best of our knowledge, a “real world” analysis on a national scale of both the early burden and mortality of sickle cell disease in a high-income country where up-to-date standard of care is easily available. Outcomes regarding 1620 children born between 2006 and 2010 and evaluated at five years show a very low SCD-related mortality and demonstrates that frequency of serious SCD-related complications such as severe infection and stroke have drastically declined. In contrast, the overall burden of the disease remained very high with a sustained elevated proportion of abnormal TCD results, VOC, ASS, ACS, transfusion requirement, hospital admissions, and chronic transfusion programs, contrasting sharply with a very low fraction of children benefitting from disease modifying or curative therapy, namely hydroxyurea and HSCT. Overall, 8.5% of the children identified following NBS were lost to follow-up at the time of data collection, with a fraction of them known as not residing in France. In this study, overall mortality rate for all patients was 0.23/100 person–years during the first five years of life and overall probability of survival was of 98.9%. Although the rate of death between 1–4 years of age remained higher than in the French general population for this age-group during the same period (0.19 versus 0.03/100),  these reIn addition to the known beneficial effect of current management guidelines, these results also reflect a real-life efficient access to medical care for patients in the French healthcare system, including “vulnerable” populations like patients with SCD who are, for the majority, children of recently immigrated families with significant socio-economic difficulties . In FranDespite these favorable conditions, it is noteworthy to mention that 6 of the 12 strokes occurred in children who had not been screened. Likewise, severe infections  still accounted for the main cause of death. Although severe pneumococcal infections most often occurred in patients fully vaccinated with PVC as previously reported,  further This study showed that 8.4% of children screened at birth were lost to follow-up in their first five years of life at the time of data collection. While there is no possibility of speculating on the outcomes, because this a nation-wide study, it is probable that none of these children died nor had a severe event such as stroke or severe infection in France. While such events may still have occurred abroad, it is probable that mortality and severe morbidity rates were not significantly biased by this missing data. Although the rate of patients lost to follow-up is similar on a national scale to results from regional or single center European cohorts, which range from 6%–11% , the figResults on less severe outcomes such as abnormal TCD, VOC, ASS, ACS, and hospitalization rates showed a very high burden of the disease with no improvement in their frequency when compared to cohorts in the previous decade . The oveIn conclusion, this study shows, on a national scale, the benefits of NBS and preventive measures, confirms the good implementation of these measures, and does not reveal major disparity across regions or centers, although the proportion of severe patients is highest in larger centers, most often located in the Paris area. Notwithstanding, this study underscores the importance of referral of patients to tertiary centers where systematic annual specialized work-up may be more readily performed and general educational or immunization issues double-checked by dedicated healthcare providers in order to further improve vaccination and TCD coverage, notably. In addition, this study reveals a very low proportion of children treated with HU or HSCT despite a sustained burden of the disease. Predictably, future studies will demonstrate an increase in disease modifying therapy use. In addition, given the ongoing progress in HSCT outcomes, indications will broaden so that altogether curative treatments  will ho


Hypogonadism refers to a syndrome that results from failure of gonads to function properly. The main concern is considerable rise in morbidity, as shown by increased cardiovascular risk, infertility, osteoporosis and above all, the psychological impact on the life of the patients with hypogonadism. Judicious steroid replacement and culturally-sensitive psychological support before and during steroid therapy remains the key tool in the management of this condition. The present study aimed at filling the knowledge gap on hypogonadism in Cameroon.We conducted a cross-sectional study over a period of 12 months, in 3 reference hospitals of Cameroon. We included males and females diagnosed with hypogonadism, aged 16 to 50 years and 16 to 45 years respectively. After a complete clinical examination, patients were invited to fill the modified middlesex hospital questionnaire for psychoneurotic evaluation.We recruited 59 patients with a sex ratio of 1:1. The mean age of the females and males were 27.7 ± 9.1years and 30.8 ± 11.7 years respectively. Normosmic Idiopathic Hypogonadotropic Hypogonadism (NIHH) was the most common presentation. Compulsive obsessive traits, phobic anxiety and hysterical trait, were most pronounced in these patients. Testosterone titers significantly correlated positively with testicular size and negatively with body mass index (BMI). A significant positive correlation was found between the testicular volumes measured with ultrasound (US) and with the orchidometer.Normosmic idiopathic hypogonadotropic hypogonadism is the most common presentation of hypogonadism in the study population. There is a significant psychosocial impact requiring further investigation and attention. Hypogonadism refers to a syndrome that results from failure of gonadal function. It is a relatively rare disease that is underdiagnosed, with an unknown combined prevalence in both males and females worldwide . MoreoveWe conducted a cross-sectional study, over a period of 12 months, from January to December 2016. Our study was conducted in the Yaoundé Central and General hospitals; and at the Douala General hospital where patients with hypogonadism seen during outpatient visits were included. We invited people with hypogonadism-related symptoms through an announcement to consult an endocrinologist in one of the 3 study sites. For patients who already had follow-up in each unit, we recorded data from their files. All consenting patients received at outpatient endocrinology consultation with a diagnosis of hypogonadism and aged 16-50 years for males and or 16-45 years for females , 8 were Clinical data: for all patients we recorded medical history including cryptorchidism, mumps orchitis, impuberism, amenorrhea and their physiological age at the first consultation for hypogonadism-related symptoms. We also looked for symptoms such as anosmia and visual field defects. Family history of infertility or impuberism was also noted. Patients underwent a clinical exam where anthropometric parameters, eunuchoid presentation and specific signs of malformation such as web neck, narrow shoulders and cleft palates were assessed. Concerning external genital evaluation in men, we verified the presence of testes in the scrotum, measured testicular size with an orchidometer and the resting penile length using a tape then both were staged using the Tanner staging system. We evaluated and staged the female breast using the Tanner staging system. In both genders, pubic and axillary hair was evaluated for type and distribution and staged using the same staging system. Complete physical examination was conducted topographically.Psychosocial aspect: we invited patients to fill a form that evaluated the direct effect of hypogonadism on their psychological state and the resultant psychosomatic disorder that could be associated with hypogonadism. We used the Middlesex Hospital Questionnaire (MHQ) that has been approved and considered valid, reliable [reliable , 10 and Data analysis: data were collected and analyzed using SPSS software version 23. Student T-test was used to compare means and Spearman's correlation to establish relationship between qualitative variables. The results were expressed in terms of percentages for qualitative variables, medians and means for quantitative variables. The threshold of significance was set at a value p < 0.05.Ethical consideration: institutional ethical clearance was obtained from the Ethical committee of the Faculty of Medicine and Biomedical Sciences of the University of Yaoundé I to carry out this study. Each participant gave his informed written consent. They were free to withdraw from the study at any time they wished. All information obtained or used in the study was kept confidential.2 and 33.3% (10) also had BMI between 30 and 35 kg/m2. Whereas in the male population 34.5% (10) had BMI between 25 and 30 kg/m2 and 34.5% (10) also had BMI between 30 and 35 kg/m2. The main presenting complaints in the females were primary amenorrhoea 53.3% (16) and secondary amenorrhoea 33.3% (10)  of our patients were single. The majority of our population had body mass index (BMI) above normal. In females, 33.3% (10) had BMI between 25 and 30 kg/m.3% (10) . In addiIn the male population, micro-penis accounted for 48.3% (14) and small testes for 37.9% (11) of the presenting complaints. In addition, 75.9% (22) had sleep disorder, 72.4% (21) had both erectile dysfunction and low libido, 55.2% (16) had fatigue, 44.8% (13) irritability, 41.4% (12) had a high pitched voice and 14 had infertility. Concerning clinical signs, 79.3% (23) had eunuchoid habitus, 69.0% (20) lacked beard and most were in Tanner stage 3 or 4 for pubic hair development accounting for 34.5% (10) and 31.0% (9) respectively. On the other hand, 72.4% of the males had a penile length in Tanner stage 5, 62.1% (18) had gynecomastia. Kallman and Klinefelter syndromes were identified. In the overall population, the most common diagnosis was congenital hypogonadotropic hypogonadism seen in 44.1% (26) of the patients. Congenital hypergonadotropic hypogonadism was found in 22.0% (13), 18.6% (11) had acquired hypogonadotropic hypogonadism meanwhile 15.3% (9) had acquired hypergonadotropic hypogonadism. In female patients, the main etiology of central hypogonadism was NIHH with a prevalence of 20.0% (6). Peripheral or primary etiologies included primary ovarian failure (POF) accounting for 20.0% (6), Turner syndrome accounting for 16.7% (5), prolactinomas accounting for 13.3% (4) and pure gonadal dysgenesis in 10.0% (3) of the cases. On the other hand, male patients presented with NIHH in 34.5% (10) of the cases and 23.8% (4) of cases were part of the combined pituitary hormone deficiency syndrome .Correlation studies: there was a significant positive correlation between testicular volume measured using the orchidometer and that measured by testicular ultrasound with a correlation coefficient 0.860 and p-value < 0.001 on both sides. There was a positive correlation between the BMI and the level of androgens in both sexes.Psychological profile: concerning the psychological profile in males, compulsive obsessive trait was common with a mean score of 8.6 ± 3.4 followed by phobic anxiety and depression with scores 7.8 ± 2.9. Meanwhile women presented a score of 9.2 ± 2.5 for compulsive obsessive trait, followed by phobic anxiety with a mean score 8.7 ± 2.7 and the minimum mean was 6.9 ± 2.1 for hysterical trait  is the most common presentation of hypogonadism in the study population. The clinical presentation of hypogonadism is heterogeneous and there is a significant psychosocial impact requiring further investigation and attention.Hypogonadism is a frequent complain in endocrine consultation;The etiologies are not different in both genders;Spectrum of anosmia is widely variable in hypogonadism.Hypogonadotropic hypogonadism is the most frequent forms of hypogonadism in our context;The study highlights a delay diagnostic in our context and the clinical presentation at the end stage of the disease, difficulty and the limits of the management of hypogonadism in a low resource setting;It presents specific psychiatric disorders in all types of hypogonadism and importance to systematically implicate psychiatrists and psychologists in the overall management of hypogonadic patients in our context.The authors declare no competing interests.


Z value of 4.6 formed the best spherical droplets at 10,000 Hz and produced perfect color images on cotton fabrics.The jetting performance of dye inks determines the image quality, production efficiency, and lifetime of the print head. In the present study, we explored the jetting performance of mixed solutions of polyethylene glycol (PEG) and reactive dye by testing the visible absorption spectra, rheological properties, and surface tension, in addition to the observation of droplet formation. The results indicate that PEG macromolecules could change the aggregate groups of Red 218 molecules into smaller ones through hydrophobic interactions and separation effect. The addition of PEG into the dye solution increased the viscosity and decreased the surface tension. In the whole shear rate range tested, the 10% and 20% PEG400, as well as the 30% PEG200 dye solutions, showed good Newtonian fluid behavior. PEG macromolecules improved the droplet formation of the dye solutions. Increasing the PEG400 concentration to 30% and 40% resulted in elimination of the formation of satellites and the formation of ideal droplets at 10,000 Hz jetting frequency. A 30% PEG600-dye solution with the  Digital inkjet printing has become an important technology in the textile industry ,6,7,8,9.In order to avoid the satellites, other than designing new nozzle geometries, various aqueous solutions of glycerol, glycols, and polyethylene oxide have been well studied in the past years ,17. For Reactive dye inks are usually composed of humectants, surfactants, solubilizers, bactericides, reactive dyes, and water ,22. In oPolyethylene glycols with different number averaged molecular weights—200 g/mol (PEG200), 400 g/mol (PEG400), and 600 g/mol (PEG600)—were purchased from Sinopharm Chemical Reagent Co., Ltd. . Pure C. I. Reactive Red 218 (Red 218) for ink preparation was purchased from Yongguang Chemical Industry Co., Ltd. , and was used without further purification. The chemical structures of the Red 218 and polyethylene glycols are shown in The weighted reactive dye was slowly dissolved in a mixture of water and PEG under stirring by a HJ-6A digital multi-head magnetic stirrer  at 25 °C for two hours. The mixtures of PEG and water were prepared before use. The dye concentration was 7% by weight, unless otherwise indicated. The concentrations of PEG400 in PEG–dye solutions were 0.0%, 10.0%, 20.0%, 30.0%, and 40.0% by weight, respectively, and the concentrations of PEG200 and PEG600 were 30.0%.The visible absorption spectra of the Red 218 inks with various PEG concentrations were measured using a U-3900H ultraviolet spectrophotometer  with a 0.01 mm thickness cuvette at 25 °C.The variation of Red 218 and PEG solution viscosity vs. relatively high shear rate was measured using a FLUDICAM RHEO microfluidic visual rheometer  at 25 °C. The rheological properties under low shear rates were measured using a DV-III + programmable rheometer  at 25 °C. The surface tensions of Red 218 and PEG solutions were measured with a SITA surface tension meter  at 25 °C.η is the fluid viscosity, and γ is the surface tension.The densities of the PEG–dye solutions were measured in accordance with the China national standard GB/T 4472-2011 at 25 °C. The Z values of the PEG–dye solutions were calculated per Equation (1):The droplet formation in the PEG–dye solutions was observed at 25 °C using an IIA-1502 ink droplet observer  equipped with a KJ4B300 inkjet print head  with a nozzle diameter of 20 µm. The voltage waveform had a rise and fall time of 2 µs, the driving voltage was 26 V, and the pulse width was 6 µs. The inkjet printing was conducted with the resolution of 600 × 600 dpi at 25 °C.In order to understand the dye–PEG interaction, visible absorption spectra were measured as shown in −1. However, when the PEG400 concentration reached 40%, the viscosity began to slowly decrease at a shear rate of 5000 s−1. At the concentration of 30%, PEG600 had the highest viscosity compared to PEG200 and PEG400. The viscosities of 30% PEG600 and PEG400 dye solutions decreased slightly at a shear rate of 40,000 s−1. In the whole shear rate range, the 10% and 20% PEG400, and the 30% PEG200 dye solutions showed excellent Newtonian fluid behavior.Rheological properties determine inks’ jetting ability ,19,25,26The surface tension of ink is an important physicochemical parameter for droplet formation ,27,28. SDroplet formation in inkjet printing is determined by many factors, such as the jetting frequency, the driving voltage wave, the fluid viscosity, and the surface tension. Due to the arrangement of PEG macromolecules at the air/liquid interface, the addition of PEG in the dye solution is certain to change the formation of droplets. For 30% PEG400 and 7% dye solutions, increasing the jetting frequency from 1500 Hz to 5000 Hz led to the formation of more spherical drops a,b. WhenFor 30% PEG200, PEG400, and PEG600 dye solutions, the jetted droplet images are shown in According to the results, we printed cotton fabrics using the PEG600–dye solution. The result is shown in In aqueous solutions, PEG macromolecules could change the aggregate groups of Red 218 molecules into smaller ones through hydrophobic interactions and separation effect. The addition of PEGs into the dye solution increased the viscosity and decreased the surface tension. In the whole shear rate range tested, the 10% and 20% PEG400, and the 30% PEG200 dye solutions showed good Newtonian fluid behavior. PEG macromolecules improved the droplet formation of dye solutions. Increasing the PEG400 concentration to 30% and 40% resulted in elimination of the formation of satellites and the formation of ideal droplets at a 10,000 Hz jetting frequency. Compared with 30% PEG200 and 30% PEG400, 30% PEG600–dye solution with the Z value of 4.6 formed the best spherical drops at 10,000 Hz and produced perfect color images on cotton fabrics.


Patients after transcatheter aortic valve replacement (TAVR) and persistent severe mitral regurgitation (MR) are increasingly treated with percutaneous edge-to-edge mitral valve repair (PMVR). The impact of a former TAVR on PMVR procedures is not clear.We retrospectively analyzed 332 patients undergoing PMVR using the MitraClip system with respect to procedural and clinical outcome. 21 of these 332 patients underwent TAVR before PMVR. Intra-procedural transthoracic (TTE) and transesophageal echocardiograms (TEE) immediately before and after clip implantation as well as invasive hemodynamic measurements were evaluated. At baseline, we found a significantly smaller mitral valve anterior-posterior diameter in the TAVR cohort (p < 0.001). A reduction of MR by at least three grades was achieved in a smaller fraction in the TAVR cohort as compared to the cohort with a native aortic valve (p = 0.02). Accordingly, we observed a smaller post-procedural cardiac output in the TAVR cohort (p = 0.02).PMVR in patients who had a TAVR before, is associated with altered MR anatomy before and a reduced improvement of MR after the procedure. Future larger and prospective studies will have to determine, whether a previous TAVR influences long-term clinical outcome of patients undergoing PMVR. Patients with severe heart valve defects not eligible for conventional surgery can be treated with interventional techniques. For instance, patients with aortic stenosis and aortic valve regurgitation undergo transaortic valve replacement (TAVR) and patients with mitral regurgitation (MR) are treated with percutaneous mitral valve edge-to-edge repair (PMVR) using for example the MitraClip system. In a relevant fraction of these patients both aortic stenosis and MR are present. As a matter of fact, 20% to 30% of patients undergoing TAVR present with moderate to severe MR . It has With respect to PMVR, positive 5-year-outcome results were published recently, showing an increased rate of relapsing MR after one year compared to surgery, while beyond 1 year there was no difference compared to surgery , implyinThis study was initiated to determine, whether PMVR using the MitraClip system in patients with a preceding TAVR is more difficult to carry out or is associated with inferior clinical outcome.For this study, we retrospectively evaluated 339 consecutive patients, who underwent PMVR using the MitraClip system  between May 2014 and July 2017 at the University hospital, Department of Cardiology and Cardiovascular Medicine, University of Tübingen. Seven patients  were excluded from further analysis because of potential preexisting alterations. Of the remaining 332 patients in the study collective, 21 patients had undergone TAVR and 18 surgical aortic valve replacement (SAVR). The study was approved by the local ethics committee . The patients provided informed written consent to have their medical record data used in research. The decision for treatment by PMVR was made by an interdisciplinary heart team of interventional cardiologists and cardiac surgeons based on either the EuroSCORE [®. Bleeding events were identified according to the VARC-2 criteria [The procedure was carried out either in general anesthesia (GA) or in deep sedation (DS) as described before , 8, 9. Acriteria .Statistical analysis was performed with SPSS . Categorical variables are displayed in percentages and absolute numbers. The level of significance in these variables was tested using the Chi-Square test. Ordinally scaled and continuous data are shown as mean ± standard deviation (SD). The Shapiro-Wilk test was used to check for normal distribution. In case of not normally distributed data, the Mann-Whitney-U test was used for inter-group comparisons. For normally distributed data the Student T test was performed. Two-tailed p values were calculated with a p-value < 0.05 being considered as statistically significant.Table 1. The mean age was 76.9 ± 8.9 years, and 57.2% of the patients were male. The etiology of MR was functional in 51.7%. 48.3% had a left ventricular ejection fraction (LVEF) of ≤ 35% and the mean EuroSCORE II was 10.8 ± 10.3. There were significant differences regarding the baseline characteristics between the cohort with a native aortic valve and the cohort with previous TAVR. A significant smaller fraction of patients in the cohort with a native aortic valve had coronary heart disease  and also hyperlipoproteinemia was significantly decreased . In the cohort with previous TAVR, a higher fraction of patients had renal insufficiency as compared to the cohort with a native aortic valve . EuroSCORE II levels were higher in the TAVR cohort . Less patients in the TAVR cohort were treated with betablockers  and with ACE inhibitors/sartans . Besides that, the TAVR and the native aortic valve group were balanced and comparable regarding the baseline characteristics. 73.7% of patients with native aortic valve and 66.6% of patients with TAVR underwent the PMVR procedure in deep sedation . As a point of orientation, we also looked at the cohort with SAVR in which 88.9% had deep sedation . Although the absolute numbers were different, there was no significant difference with respect to procedure time between the cohorts . At baseline, we observed a significantly smaller mitral valve anterior-posterior diameter in the TAVR cohort. The diameter was 32 ± 5 mm in the cohort with a native aortic valve vs 28 ± 5 mm in the cohort with previous a TAVR (p < 0.001). In the cohort with SAVR, the anterior-posterior diameter was 31± 5 mm . There were no significant differences between the cohorts regarding mitral valve ellipticity, defined as anterior-posterior annulus diameter divided by the medial-lateral diameter  and 1.07 ± 0.14 in the SAVR cohort . Fig 1E demonstrates the anatomic situation of the mitral valve with a native aortic valve (left panel) and with a TAVR prosthesis (right panel).In all patients receiving PMVR, we compared the cohort with a native aortic valve and the cohort with previous TAVR with regard to procedural and clinical parameters such as procedural success, reduction of MR and increase in cardiac output. The cohort with previous SAVR served as an additional point of orientation. In total, 332 patients were included in our study. Baseline characteristics are listed in Fig 2A). In Table 2, the implanted TAVR models are listed and the corresponding success rates of the PMVR procedures are given. Procedural success rates were 8/9 in patients with a CoreValve, , 9/9 in patients with an Edwards Sapien , 2/2 in patients with a Lotus , and 1/1 in patients with a Symetis  prosthesis. Detailed data on the different types and sizes of TAVR prosthesis are given in S1 Table.Comparing the procedural outcome data, we found no significant difference between the cohorts with respect to procedural failure. Procedural failure was 1.4% in the native aortic valve cohort vs. 4.8% in the TAVR cohort  and 5.6% in the SAVR cohort  and 8.1 ± 2.6 x 109/l in the SAVR cohort . Postinterventional CRP levels were 310 ± 298 nmol/l in the native aortic valve cohort, 285 ± 158 nmol/l in the TAVR cohort , and 386 ± 384 nmol/l in the SAVR cohort . Postinterventional bleeding complications were also evaluated. As defined by the VARC-2  criteria [Fig 3C.When we evaluated postoperative inflammation, we could not detect a significant difference between the cohorts. Postinterventional leukocyte counts were 8.6 ± 3.0 x 10criteria , those wFig 4A). For better illustration of the intraprocedural situation, Fig 4B depicts a TEE image (left ventricular outflow tract view) of MR before and after PMVR in a patient with a native aortic valve, Fig 4C of a patient with a TAVR prosthesis. Furthermore, the cardiac output after PMVR was significantly lower in the cohort with TAVR as compared to the control cohorts. Cardiac output after PMVR was 5.5 ± 1.9 l/min in the cohort with native aortic valve, 4.1 ± 1.2 l/min in the cohort with TAVR , and 6.2 ± 1.6 l/min in the cohort with SAVR . At baseline, there was no significant difference with respect to cardiac output between the three cohorts. At baseline, cardiac output was 4.7 ± 1.6 l/min in the cohort of patients with a native aortic valve, 4.2 ± 0.7 l/min in the cohort with TAVR , and 4.8 ± 1.4 l/min in the cohort with SAVR .Interestingly, the achieved reduction of MR by PMVR was less in the cohort with previous TAVR as compared to the cohort of patients with a native aortic valve. In the cohort with a native aortic valve, a reduction of MR of at least 3 grades was achieved in 45.7% of patients compared to 19.0% of patients in the cohort with TAVR  and 55.6% in the cohort with SAVR  mitral valve anatomy in patients with previous TAVR is altered, ii) the percentage of procedural success regarding MR is significantly lower and iii) that cardiac output is significantly reduced after PMVR in patients who have previously undergone a TAVR procedure.PMVR is a successful and increasingly performed treatment option for patients with MR who are not eligible for conventional surgery . Both TAIn the literature, mitral valve stenosis was observed in some cases after TAVR , 26. IntPMVR using the MitraClip system is influencing the mitral valve apparatus, too. As we could recently demonstrate, the procedure results in a reduction of mitral annular size, which correlates inversely with residual MR and is stable at follow up . These oBesides affecting mitral valve geometry, PMVR has short-term and long-term effects on hemodynamics. For instance, we found an immediate increase of cardiac output after PMVR from 4.6 ± 1.4 l/min to 5.4 ± 1.6 l/min (p < 0.001) . GaemperThis study has clear limitations. One of the main limitations is the limited sample size, particularly of patients, who had both a previous TAVR prosthesis and the indication for PMVR in our retrospective collective. There were some imbalances between the cohort of patients with a native aortic valve and that with TAVR. Due to the limited sample size of the TAVR cohort, an adjusted analysis was not performed. Moreover, different device types of TAVR prostheses were implanted in this study patient collective, which may differ in their influence on heart geometry . SimilarHere, we assessed whether the presence of a TAVR prosthesis in patients undergoing additionally PMVR therapy using the MitraClip system has any clinical relevance. Interestingly, we observed altered mitral valve annulus diameter geometry depending on the presence of a TAVR prosthesis, and patients, who had previous TAVR treatment showed less improvement of MR and a smaller CO after PMVR. Future analyses on larger patient collectives and prospective trials have to further scrutinize the question, how outcome of PMVR is influenced by a previous TAVR procedure.S1 Table(DOC)Click here for additional data file.S1 FigAt baseline, cardiac output was 4.7 ± 1.6 l/min in the cohort with a native aortic valve, 4.2 ± 0.7 l/min in the cohort with TAVR , and 4.8 ± 1.4 l/min in the cohort with SAVR.(TIF)Click here for additional data file.S1 File(XLSX)Click here for additional data file.